|
1
|
Di H, Rong Z, Mao N, Li H, Chen J, Liu R, Wang A. Transcriptomic landscape of Hras12V oncogene-induced hepatocarcinogenesis with gender disparity. BMC Cancer 2025; 25:94. [PMID: 39819515 PMCID: PMC11737189 DOI: 10.1186/s12885-025-13476-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2024] [Accepted: 01/08/2025] [Indexed: 01/19/2025] Open
Abstract
The genesis of hepatocellular carcinoma (HCC) is closely related to male factors and hyper-activated Ras signals. A transcriptomic database was established via RNA-Seq of HCC (T) and the adjacent precancerous liver tissue (P) of Hras12V transgenic mice (Ras-Tg, HCC model) and the normal liver tissue of wild-type mice (W) of both sexes. Comparative analysis within W, P, and T and correlation expression pattern analysis revealed common/unique cluster-enriched items towards HCC between the sexes. Specifically, the numbers of differentially expressed genes (DEGs) were much higher in females than in males, and tumor suppressor genes, such as p21Waf1/Cip1 and C6, were significantly higher in the female P. This finding denotes the higher sensitivity of female hepatocytes to the Ras oncogene and, therefore, the difficulty in developing HCC. Moreover, convergence in HCC between the sexes suggests the underlying mechanisms for the ineffectiveness of sex hormone therapies. Additionally, expression pattern analysis revealed that the DEGs and their relevant pathways were either positively or negatively associated with the HCC/Ras oncogene. Among them, the vital role of glutathione metabolism in HCC was established. This work provides a basis for future research on elucidating the underlying mechanisms, selecting the diagnostic biomarker, and planning the clinical therapy in HCC.
Collapse
Affiliation(s)
- Huaiyuan Di
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, 116044, China
| | - Zhuona Rong
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, 116044, China
- Department of Pharmacy, Peking University First Hospital, Xishiku Street, Xicheng District, Beijing, 100034, China
| | - Nan Mao
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, 116044, China
| | - Huiling Li
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, 116044, China
| | - Jun Chen
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, 116044, China
| | - Renwu Liu
- Central Hospital of Dalian, University of Technology, Dalian, Liaoning, 116044, China.
| | - Aiguo Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, 116044, China.
| |
Collapse
|
|
2
|
Dong L, Zhang N, Chen J, Dong P, Mao N, Li H, Wang A. Triiodothyronine (T3) suppresses hepatic tumorigenesis and development by inhibiting the phosphorylation of ERK. Mol Carcinog 2024; 63:1988-2000. [PMID: 39031486 DOI: 10.1002/mc.23788] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2023] [Revised: 06/04/2024] [Accepted: 06/25/2024] [Indexed: 07/22/2024]
Abstract
The effect of triiodothyronine (T3) on the phosphorylation of ERK and the occurrence and development of hepatocellular carcinoma (HCC) is controversial and remains to be clarified. In the present study, both in vitro (hepatoma cell lines) and in vivo (wild-type mice [WT] and mouse models of HCC [HrasG12Vand KrasG12Dtransgenic mice (Hras-Tg and Kras-Tg)]) systems were used to investigate the effect of T3 on p-ERK and hepatocarcinogenesis. The results showed that, in vitro, T3 treatment elevated the levels of p-ERK in hepatoma cells within 30 min. However, p-ERK levels returned to normal after 1 h with no significant effects on cellular proliferation or apoptosis. Interestingly, in vivo, T3 induced early rapid and transient activation of ERK and later persistent downregulation of p-ERK in liver tissues of WT. In Hras-Tg, liver weight, liver/body weight ratio, hepatic tumor numbers and sizes were significantly reduced withT3treatment compared with the untreated group. Furthermore, the levels of albumin, HrasG12V, and p-ERK in hepatic precancerous and tumor tissues were all significantly downregulated with T3 treatment; however, the levels of endogenous Hras were not affected. In WT, T3 also induced downregulation of Albumin in liver tissues, but without influence on the expression of endogenous Hras and p-MEK. Especially, the inhibitory effect of T3 on p-ERK and hepatic tumorigenesis and development without influence on the levels of KrasG12D and p-MEK was further confirmed in Kras-Tg. In conclusion, T3 suppresses hepatic tumorigenesis and development by independently and substantially inhibiting the phosphorylation of ERK in vivo.
Collapse
Affiliation(s)
- Lili Dong
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Nan Zhang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Jun Chen
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Penghui Dong
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Nan Mao
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Huiling Li
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Aiguo Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| |
Collapse
|
|
3
|
Smiriglia A, Lorito N, Serra M, Perra A, Morandi A, Kowalik MA. Sex difference in liver diseases: How preclinical models help to dissect the sex-related mechanisms sustaining NAFLD and hepatocellular carcinoma. iScience 2023; 26:108363. [PMID: 38034347 PMCID: PMC10682354 DOI: 10.1016/j.isci.2023.108363] [Citation(s) in RCA: 11] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/02/2023] Open
Abstract
Only a few preclinical findings are confirmed in the clinic, posing a critical issue for clinical development. Therefore, identifying the best preclinical models can help to dissect molecular and mechanistic insights into liver disease pathogenesis while being clinically relevant. In this context, the sex relevance of most preclinical models has been only partially considered. This is particularly significant in NAFLD and HCC, which have a higher prevalence in men when compared to pre-menopause women but not to those in post-menopausal status, suggesting a role for sex hormones in the pathogenesis of the diseases. This review gathers the sex-relevant findings and the available preclinical models focusing on both in vitro and in vivo studies and discusses the potential implications and perspectives of introducing the sex effect in the selection of the best preclinical model. This is a critical aspect that would help to tailor personalized therapies based on sex.
Collapse
Affiliation(s)
- Alfredo Smiriglia
- Department of Experimental and Clinical Biomedical Sciences, University of Florence, 50134 Florence, Italy
| | - Nicla Lorito
- Department of Experimental and Clinical Biomedical Sciences, University of Florence, 50134 Florence, Italy
| | - Marina Serra
- Department of Biomedical Sciences, University of Cagliari, 09042 Monserrato, Italy
| | - Andrea Perra
- Department of Biomedical Sciences, University of Cagliari, 09042 Monserrato, Italy
| | - Andrea Morandi
- Department of Experimental and Clinical Biomedical Sciences, University of Florence, 50134 Florence, Italy
| | - Marta Anna Kowalik
- Department of Biomedical Sciences, University of Cagliari, 09042 Monserrato, Italy
| |
Collapse
|
|
4
|
Kim HS, Na MJ, Son KH, Yang HD, Kim SY, Shin E, Ha JW, Jeon S, Kang K, Moon K, Park WS, Nam SW. ADAR1-dependent miR-3144-3p editing simultaneously induces MSI2 expression and suppresses SLC38A4 expression in liver cancer. Exp Mol Med 2023; 55:95-107. [PMID: 36599932 PMCID: PMC9898302 DOI: 10.1038/s12276-022-00916-8] [Citation(s) in RCA: 13] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2022] [Revised: 10/26/2022] [Accepted: 11/07/2022] [Indexed: 01/06/2023] Open
Abstract
Aberrant adenosine-to-inosine (A-to-I) RNA editing, catalyzed by adenosine deaminase acting on double-stranded RNA (ADAR), has been implicated in various cancers, but the mechanisms by which microRNA (miRNA) editing contributes to cancer development are largely unknown. Our multistage hepatocellular carcinogenesis transcriptome data analyses, together with publicly available data, indicated that ADAR1 was the most profoundly dysregulated gene among RNA-editing enzyme family members in liver cancer. Targeted inactivation of ADAR1 inhibited the in vitro tumorigenesis of liver cancer cells. An integrative computational analyses of RNA-edited hotspots and the known editing frequency of miRNAs suggested that the miRNA miR-3144-3p was edited by ADAR1 during liver cancer progression. Specifically, ADAR1 promoted A-to-I editing of canonical miR-3144-3p to replace the adenosine at Position 3 in the seed region with a guanine (ED_miR-3144-3p(3_A < G)) in liver cancer cells. We then demonstrated that Musashi RNA-binding protein 2 (MSI2) was a specific target of miR-3144-3p and that MSI2 overexpression was due to excessive ADAR1-dependent over-editing of canonical miR-3144-3p in liver cancer. In addition, target prediction analyses and validation experiments identified solute carrier family 38 member 4 (SLC38A4) as a specific gene target of ED_miR-3144-3p(3_A < G). The ectopic expression of both ADAR1 and the ED_miR-3144-3p(3_A < G) mimic enhanced mitotic activities, and ADAR1 suppressed SLC38A4 expression in liver cancer cells. Treatments with mouse-specific ADAR1-, MSI2-siRNA-, or SLC38A4-expressing plasmids suppressed tumorigenesis and tumor growth in a mouse model of spontaneous liver cancer. Our findings suggest that the aberrant regulation of ADAR1 augments oncogenic MSI2 effects by excessively editing canonical miR-3144-3p and that the resultant ED_miR-3144-3p(3_A < G) simultaneously suppresses tumor suppressor SLC38A4 expression, contributing to hepatocellular carcinogenesis.
Collapse
Affiliation(s)
- Hyung Seok Kim
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Min Jeong Na
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School, The Catholic University of Korea, Seoul, 06591, Korea
| | - Keun Hong Son
- Department of Microbiology, Dankook University, Cheonan, 31116, Republic of Korea
| | - Hee Doo Yang
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- NEORNAT Inc., Rm. #5104 Bldg. A, Omnibus Park, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Sang Yean Kim
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- NEORNAT Inc., Rm. #5104 Bldg. A, Omnibus Park, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Eunbi Shin
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School, The Catholic University of Korea, Seoul, 06591, Korea
| | - Jin Woong Ha
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School, The Catholic University of Korea, Seoul, 06591, Korea
| | - Soyoung Jeon
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School, The Catholic University of Korea, Seoul, 06591, Korea
| | - Keunsoo Kang
- Department of Microbiology, Dankook University, Cheonan, 31116, Republic of Korea
| | - Kiho Moon
- NEORNAT Inc., Rm. #5104 Bldg. A, Omnibus Park, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Won Sang Park
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Suk Woo Nam
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea.
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea.
- Department of Biomedicine & Health Sciences, Graduate School, The Catholic University of Korea, Seoul, 06591, Korea.
- NEORNAT Inc., Rm. #5104 Bldg. A, Omnibus Park, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea.
| |
Collapse
|
|
5
|
hnRNPC induces isoform shifts in miR-21-5p leading to cancer development. Exp Mol Med 2022; 54:812-824. [PMID: 35729324 PMCID: PMC9256715 DOI: 10.1038/s12276-022-00792-2] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2021] [Revised: 03/28/2022] [Accepted: 04/20/2022] [Indexed: 02/07/2023] Open
Abstract
MicroRNA (miRNA) processing is a critical step in mature miRNA production. Its dysregulation leads to an increase in miRNA isoforms with heterogenous 5'-ends (isomiRs), which can recognize distinct target sites because of their shifted seed sequence. Although some miRNA genes display productive expression of their 5'-isomiRs in cancers, how their production is controlled and how 5'-isomiRs affect tumor progression have yet to be explored. In this study, based on integrative analyses of high-throughput sequencing data produced by our group and publicly available data, we demonstrate that primary miR-21 (pri-miR-21) is processed into the cancer-specific isomiR isomiR-21-5p | ±1, which suppresses growth hormone receptor (GHR) in liver cancer. Treatment with antagomirs against isomiR-21-5p | ±1 inhibited the in vitro tumorigenesis of liver cancer cells and allowed the recovery of GHR, whereas the introduction of isomiR-21-5p | ±1 mimics attenuated these effects. These effects were validated in a mouse model of spontaneous liver cancer. Heterogeneous nuclear ribonucleoprotein C and U2 small nuclear RNA auxiliary factor 2 were predicted to bind upstream of pre-miR-21 via a poly-(U) motif and influence Drosha processing to induce the production of isomiR-21-5p | ±1. Our findings suggest an oncogenic function for the non-canonical isomiR-21-5p | ±1 in liver cancer, and its production was shown to be regulated by hnRNPC.
Collapse
|
|
6
|
Kim H, Jeong M, Na DH, Ryu SH, Jeong EI, Jung K, Kang J, Lee HJ, Sim T, Yu DY, Yu HC, Cho BH, Jung YK. AK2 is an AMP-sensing negative regulator of BRAF in tumorigenesis. Cell Death Dis 2022; 13:469. [PMID: 35585049 PMCID: PMC9117275 DOI: 10.1038/s41419-022-04921-7] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/01/2022] [Revised: 05/03/2022] [Accepted: 05/05/2022] [Indexed: 12/14/2022]
Abstract
The RAS-BRAF signaling is a major pathway of cell proliferation and their mutations are frequently found in human cancers. Adenylate kinase 2 (AK2), which modulates balance of adenine nucleotide pool, has been implicated in cell death and cell proliferation independently of its enzyme activity. Recently, the role of AK2 in tumorigenesis was in part elucidated in some cancer types including lung adenocarcinoma and breast cancer, but the underlying mechanism is not clear. Here, we show that AK2 is a BRAF-suppressor. In in vitro assays and cell model, AK2 interacted with BRAF and inhibited BRAF activity and downstream ERK phosphorylation. Energy-deprived conditions in cell model and the addition of AMP to cell lysates strengthened the AK2-BRAF interaction, suggesting that AK2 is involved in the regulation of BRAF activity in response to cell metabolic state. AMP facilitated the AK2-BRAF complex formation through binding to AK2. In a panel of HCC cell lines, AK2 expression was inversely correlated with ERK/MAPK activation, and AK2-knockdown or -knockout increased BRAF activity and promoted cell proliferation. Tumors from HCC patients showed low-AK2 protein expression and increased ERK activation compared to non-tumor tissues and the downregulation of AK2 was also verified by two microarray datasets (TCGA-LIHC and GSE14520). Moreover, AK2/BRAF interaction was abrogated by RAS activation in in vitro assay and cell model and in a mouse model of HRASG12V-driven HCC, and AK2 ablation promoted tumor growth and BRAF activity. AK2 also bound to BRAF inhibitor-insensitive BRAF mutants and attenuated their activities. These findings indicate that AK2 monitoring cellular AMP levels is indeed a negative regulator of BRAF, linking the metabolic status to tumor growth.
Collapse
Affiliation(s)
- Hyunjoo Kim
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| | - Muhah Jeong
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| | - Do-Hyeong Na
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| | - Shin-Hyeon Ryu
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| | - Eun Il Jeong
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| | - Kwangmin Jung
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| | - Jaemin Kang
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| | - Ho-June Lee
- grid.418158.10000 0004 0534 4718Departments of Discovery Oncology, Genentech, Inc., South San Francisco, CA 94080 USA
| | - Taebo Sim
- grid.35541.360000000121053345Chemical Kinomics Research Center, Korea Institute of Science and Technology, Seoul, 02792 Korea
| | - Dae-Yeul Yu
- grid.249967.70000 0004 0636 3099Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea
| | - Hee Chul Yu
- grid.411545.00000 0004 0470 4320Department of Surgery, Chonbuk National University Medical School, Jeonju, 561-180 Korea
| | - Baik-Hwan Cho
- grid.411545.00000 0004 0470 4320Department of Surgery, Chonbuk National University Medical School, Jeonju, 561-180 Korea
| | - Yong-Keun Jung
- grid.31501.360000 0004 0470 5905School of Biological Science, Seoul National University, Gwanak-gu, Seoul, 08826 Korea
| |
Collapse
|
|
7
|
Kim SY, Shen Q, Son K, Kim HS, Yang HD, Na MJ, Shin E, Yu S, Kang K, You JS, Yu KR, Jeong SM, Lee EK, Ahn YM, Park WS, Nam SW. SMARCA4 oncogenic potential via IRAK1 enhancer to activate Gankyrin and AKR1B10 in liver cancer. Oncogene 2021; 40:4652-4662. [PMID: 34140644 DOI: 10.1038/s41388-021-01875-6] [Citation(s) in RCA: 24] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2020] [Accepted: 05/28/2021] [Indexed: 01/04/2023]
Abstract
SWItch/Sucrose Non-Fermentable (SWI/SNF) is a multiprotein complex essential for the regulation of eukaryotic gene expression. SWI/SNF complex genes are genetically altered in over 20% of human malignancies, but the aberrant regulation of the SWI/SNF subunit genes and subsequent dysfunction caused by abnormal expression of subunit gene in cancer, remain poorly understood. Among the SWI/SNF subunit genes, SMARCA4, SMARCC1, and SMARCA2 were identified to be overexpressed in human hepatocellular carcinoma (HCC). Modulation of SMARCA4, SMARCC1, and SMARCA2 inhibited in vitro tumorigenesis of HCC cells. However, SMARCA4-targeting elicited remarkable inhibition in an in vivo Ras-transgenic mouse HCC model (Ras-Tg), and high expression levels of SMARCA4 significantly associated with poor prognosis in HCC patients. Furthermore, most HCC patients (72-86%) showed SMARCA4 overexpression compared to healthy controls. To identify SMARCA4-specific active enhancers, mapping, and analysis of chromatin state in liver cancer cells were performed. Integrative analysis of SMARCA4-regulated genes and active chromatin enhancers suggested 37 genes that are strongly activated by SMARCA4 in HCC. Through chromatin immunoprecipitation-qPCR and luciferase assays, we demonstrated that SMARCA4 activates Interleukin-1 receptor-associated kinase 1 (IRAK1) expression through IRAK1 active enhancer in HCC. We then showed that transcriptional activation of IRAK1 induces oncoprotein Gankyrin and aldo-keto reductase family 1 member B10 (AKR1B10) in HCC. The regulatory mechanism of the SMARCA4-IRAK1-Gankyrin, AKR1B10 axis was further demonstrated in HCC cells and in vivo Ras-Tg mice. Our results suggest that aberrant overexpression of SMARCA4 causes SWI/SNF to promote IRAK1 enhancer to activate oncoprotein Gankyrin and AKR1B10, thereby contributing to hepatocarcinogenesis.
Collapse
Affiliation(s)
- Sang Yean Kim
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Qingyu Shen
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Keunhong Son
- Department of Microbiology, Dankook University, Cheonan, 31116, Republic of Korea
| | - Hyung Seok Kim
- NEORNAT Inc, Rm# 305 Medical School Building, 222 Banpo-daero, Seocho-gu, 06591, Seoul, Republic of Korea
| | - Hee Doo Yang
- NEORNAT Inc, Rm# 305 Medical School Building, 222 Banpo-daero, Seocho-gu, 06591, Seoul, Republic of Korea
| | - Min Jeong Na
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Eunbi Shin
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Suji Yu
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Keunsoo Kang
- Department of Microbiology, Dankook University, Cheonan, 31116, Republic of Korea
| | - Jueng Soo You
- Department of Biochemistry, Research Institute of Medical Science, School of Medicine, Konkuk University, Seoul, 05029, Republic of Korea
| | - Kyung-Rok Yu
- Department of Agricultural Biotechnology, Seoul National University, Seoul, 08826, Republic of Korea
| | - Seung Min Jeong
- Department of Biochemistry, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Eun Kyung Lee
- Department of Biochemistry, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Young Min Ahn
- Department of Kidney System, College of Oriental Medicine, Kyung Hee University, Seoul, 02447, Republic of Korea
| | - Won Sang Park
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea
| | - Suk Woo Nam
- Department of Pathology, College of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea.
- Functional RNomics Research Center, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea.
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, The Catholic University of Korea, 222 Banpo-daero, Seocho-gu, Seoul, 06591, Republic of Korea.
- NEORNAT Inc, Rm# 305 Medical School Building, 222 Banpo-daero, Seocho-gu, 06591, Seoul, Republic of Korea.
| |
Collapse
|
|
8
|
Kim H, Choi J, Yu DY, Choi HJ. Expression of Organic Anion Transporting Polypeptides in an H-Ras 12V Transgenic Mouse Model of Spontaneous Hepatocellular Carcinoma. Yonsei Med J 2021; 62:622-630. [PMID: 34164960 PMCID: PMC8236347 DOI: 10.3349/ymj.2021.62.7.622] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/14/2021] [Revised: 03/29/2021] [Accepted: 04/06/2021] [Indexed: 11/27/2022] Open
Abstract
PURPOSE Expression of organic anion transporting polypeptides (OATPs) 1B1/1B3 in hepatocellular carcinoma (HCC) induces a paradoxical enhancement of gadoxetic acid on liver magnetic resonance imaging (MRI). We examined the expression profile of OATPs with regard to tumor differentiation in a genetically modified H-Ras 12V mouse model of spontaneous HCC that undergoes multistep hepatocarcinogenesis with minimal inter-individual variation. MATERIALS AND METHODS Tumor nodules were harvested from transgenic H-Ras 12V mice. Hematoxylin and eosin-stained slides were examined for tumor differentiation and high-grade pathological components (tumor necrosis, thickened trabeculae, or vascular invasion). Immunohistochemistry of OATP 1B1/1B3 was performed, and OATP expression was assessed. RESULTS We examined well-differentiated HCCs (n=59) in which high-grade pathological components were absent (n=49) or present (n=10). Among the well-differentiated HCCs without high-grade pathological components (n=49), OATP expression was negative, weak positive, and moderate positive in 23, 17, and nine cases, respectively. Among the well-differentiated HCCs with high-grade pathological components (n=10), OATP expression was negative, weak positive, and moderate positive in one, two, and seven cases, respectively. The ratio of positive OATP 1B1/1B3 expressing tumors was higher in HCCs with high-grade pathological components than in those without high-grade pathological components (p=0.004). CONCLUSION Our findings support those of previous clinical studies that have reported the frequent appearance of gadoxetic acid-enhanced MRI in moderately differentiated HCC.
Collapse
Affiliation(s)
- Honsoul Kim
- Department of Radiology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea
- Department of Health Science and Technology, SAIHST, Sungkyunkwan University, Seoul, Korea
| | - Junjeong Choi
- Department of Pharmacy, College of Pharmacy, Yonsei Institute of Pharmaceutical Science, Yonsei University, Incheon, Korea
| | - Dae Yeul Yu
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
| | - Hye Jin Choi
- Division of Oncology, Department of Internal Medicine, Severance Hospital, Yonsei University College of Medicine, Seoul, Korea.
| |
Collapse
|
|
9
|
Zhang J, Li H, Dong J, Zhang N, Liu Y, Luo X, Chen J, Wang J, Wang A. Omics-Based Identification of Shared and Gender Disparity Routes in Hras12V-Induced Hepatocarcinogenesis: An Important Role for Dlk1-Dio3 Genomic Imprinting Region. Front Genet 2021; 12:620594. [PMID: 34135934 PMCID: PMC8202007 DOI: 10.3389/fgene.2021.620594] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2020] [Accepted: 03/02/2021] [Indexed: 12/13/2022] Open
Abstract
The phenomenon of gender disparity is very profound in hepatocellular carcinoma (HCC). Although previous research has revealed important roles of microRNA (miRNA) in HCC, there are no studies investigating the role of miRNAs in gender disparity observed hepatocarcinogenesis. In the present study, we investigated the global miRNAomics changes related to Ras-induced male-prevalent hepatocarcinogenesis in a Hras12V-transgenic mouse model (Ras-Tg) by next-generation sequencing (NGS). We identified shared by also unique changes in miRNA expression profiles in gender-dependent hepatocarcinogenesis. Two hundred sixty-four differentially expressed miRNAs (DEMIRs) with q value ≤0.05 and fold change ≥2 were identified. A vertical comparison revealed that the lower numbers of DEMIRs in the hepatic tumor (T) compared with the peri-tumor precancerous tissue (P) of Ras-Tg and normal liver tissue of wild-type C57BL/6J mice (W) in males indicated that males are more susceptible to develop HCC. The expression pattern analysis revealed 43 common HCC-related miRNAs and 4 Ras-positive-related miRNAs between males and females. By integrating the mRNA transcriptomic data and using 3-node FFL analysis, a group of significant components commonly contributing to HCC between sexes were filtered out. A horizontal comparison showed that the majority of DEMIRs are located in the Dlk1-Dio3 genomic imprinting region (GIR) and that they are closely related to not only hepatic tumorigenesis but also to gender disparity in hepatocarcinogenesis. This is achieved by regulating multiple metabolic pathways, including retinol, bile acid, and steroid hormones. In conclusion, the identification of shared and gender-dependent DEMIRs in hepatocarcinogenesis provides valuable insights into the mechanisms that contribute to male-biased Ras-induced hepatic carcinogenesis.
Collapse
Affiliation(s)
- Jing Zhang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Huiling Li
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Jianyi Dong
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Nan Zhang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Yang Liu
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Xiaoqin Luo
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Jun Chen
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Jingyu Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| | - Aiguo Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, China
| |
Collapse
|
|
10
|
Cyclic Peptide Mimotopes for the Detection of Serum Anti-ATIC Autoantibody Biomarker in Hepato-Cellular Carcinoma. Int J Mol Sci 2020; 21:ijms21249718. [PMID: 33352757 PMCID: PMC7766137 DOI: 10.3390/ijms21249718] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/12/2020] [Revised: 12/14/2020] [Accepted: 12/16/2020] [Indexed: 12/31/2022] Open
Abstract
Tumor-associated (TA) autoantibodies have been identified at the early tumor stage before developing clinical symptoms, which holds hope for early cancer diagnosis. We identified a TA autoantibody from HBx-transgenic (HBx-tg) hepatocellular carcinoma (HCC) model mouse, characterized its target antigen, and examined its relationship to human HCC. The mimotopes corresponding to the antigenic epitope of TA autoantibody were screened from a random cyclic peptide library and used for the detection of serum TA autoantibody. The target antigen of the TA autoantibody was identified as an oncogenic bi-functional purine biosynthesis protein, ATIC. It was upregulated in liver cancer tissues of HBx-tg mouse as well as human HCC tissues. Over-expressed ATIC was also secreted extracellularly via the cancer-derived exosomes, which might cause auto-immune responses. The cyclic peptide mimotope with a high affinity to anti-ATIC autoantibody, CLPSWFHRC, distinguishes between serum samples from HCC patients and healthy subjects with 70.83% sensitivity, 90.68% specificity (AUC = 0.87). However, the recombinant human ATIC protein showed a low affinity to anti-ATIC autoantibody, which may be incompatible as a capture antigen for serum TA autoantibody. This study indicates that anti-ATIC autoantibody can be a potential HCC-associated serum biomarker and suggests that autoantibody biomarker's efficiency can be improved by using antigenic mimicry to native antigens present in vivo.
Collapse
|
|
11
|
Li H, Rong Z, Wang H, Zhang N, Pu C, Zhao Y, Zheng X, Lei C, Liu Y, Luo X, Chen J, Wang F, Wang A, Wang J. Proteomic analysis revealed common, unique and systemic signatures in gender-dependent hepatocarcinogenesis. Biol Sex Differ 2020; 11:46. [PMID: 32792008 PMCID: PMC7427087 DOI: 10.1186/s13293-020-00316-5] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/27/2019] [Accepted: 07/02/2020] [Indexed: 02/08/2023] Open
Abstract
Hepatocellular carcinoma (HCC) is the most common liver cancer and is highly malignant. Male prevalence and frequent activation of the Ras signaling pathway are distinct characteristics of HCC. However, the underlying mechanisms remain to be elucidated. By exploring Hras12V transgenic mice showing male-biased hepatocarcinogenesis, we performed a high-throughput comparative proteomic analysis based on tandem-mass-tag (TMT) labeling combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) on the tissue samples obtained from HCC (T) and their paired adjacent precancerous (P) of Hras12V transgenic male and female mice (Ras-Tg) and normal liver (W) of wild-type male and female mice (Non-Tg). The further validation and investigation were performed using quantitative real-time PCR and western blot. Totally, 5193 proteins were quantified, originating from 5733 identified proteins. Finally, 1344 differentially expressed proteins (DEPs) (quantified in all examined samples; |ratios| ≥ 1.5, p < 0.05) were selected for further analysis. Comparison within W, P, and T of males and females indicated that the number of DEPs in males was much higher than that in females. Bioinformatics analyses showed the common and unique cluster-enriched items between sexes, indicating the common and gender-disparate pathways towards HCC. Expression change pattern analysis revealed HCC positive/negative-correlated and ras oncogene positive/negative-correlated DEPs and pathways. In addition, it showed that the ras oncogene gradually and significantly reduced the responses to sex hormones from hepatocytes to hepatoma cells and therefore shrunk the gender disparity between males and females, which may contribute to the cause of the loss of HCC clinical responses to the therapeutic approaches targeting sex hormone pathways. Additionally, gender disparity in the expression levels of key enzymes involved in retinol metabolism and terpenoid backbone/steroid biosynthesis pathways may contribute to male prevalence in hepatocarcinogenesis. Further, the biomarkers, SAA2, Orm2, and Serpina1e, may be sex differences. In conclusion, common and unique DEPs and pathways toward HCC initiated by ras oncogene from sexually dimorphic hepatocytes provide valuable and novel insights into clinical investigation and practice.
Collapse
Affiliation(s)
- Huiling Li
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Zhuona Rong
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China.,Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Biochemistry and Molecular Biology, Peking University Cancer Hospital & Institute, No. 52 Fucheng Road, Beijing, 100142, China
| | - Hong Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Nan Zhang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Chunwen Pu
- Department of Biobank, The Affiliated Sixth People's Hospital of Dalian Medical University, Dalian, 116031, China
| | - Yi Zhao
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Xu Zheng
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Chuanyi Lei
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Yang Liu
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Xiaoqin Luo
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Jun Chen
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China
| | - Fujin Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China.
| | - Aiguo Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China.
| | - Jingyu Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, 116044, Liaoning, China.
| |
Collapse
|
|
12
|
Lei C, Chen J, Li H, Fan T, Zheng X, Wang H, Zhang N, Liu Y, Luo X, Wang J, Wang A. Role of the Mitochondrial Citrate-malate Shuttle in Hras12V-Induced Hepatocarcinogenesis: A Metabolomics-Based Analysis. Metabolites 2020; 10:metabo10050193. [PMID: 32414018 PMCID: PMC7281175 DOI: 10.3390/metabo10050193] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2020] [Accepted: 05/07/2020] [Indexed: 12/15/2022] Open
Abstract
The activation of the Ras signaling pathway is a crucial process in hepatocarcinogenesis. Till now, no reports have scrutinized the role of dynamic metabolic changes in Ras oncogene-induced transition of the normal and precancerous liver cells to hepatocellular carcinoma in vivo. In the current study, we attempted a comprehensive investigation of Hras12V transgenic mice (Ras-Tg) by concatenating nontargeted metabolomics, transcriptomics analysis, and targeted-metabolomics incorporating [U-13C] glucose. A total of 631 peaks were detected, out of which 555 metabolites were screened. Besides, a total of 122 differently expressed metabolites (DEMs) were identified, and they were categorized and subtyped with the help of variation tendency analysis of the normal (W), precancerous (P), and hepatocellular carcinoma (T) liver tissues. Thus, the positive or negative association between metabolites and the hepatocellular carcinoma and Ras oncogene were identified. The bioinformatics analysis elucidated the hepatocarcinogenesis-associated significant metabolic pathways: glycolysis, mitochondrial citrate-malate shuttle, lipid biosynthesis, pentose phosphate pathway (PPP), cholesterol and bile acid biosynthesis, and glutathione metabolism. The key metabolites and enzymes identified in this analysis were further validated. Moreover, we confirmed the PPP, glycolysis, and conversion of pyruvate to cytosol acetyl-CoA by mitochondrial citrate-malate shuttle, in vivo, by incorporating [U-13C] glucose. In summary, the current study presented the comprehensive bioinformatics analysis, depicting the Ras oncogene-induced dynamic metabolite variations in hepatocarcinogenesis. A significant finding of our study was that the mitochondrial citrate-malate shuttle plays a crucial role in detoxification of lactic acid, maintenance of mitochondrial integrity, and enhancement of lipid biosynthesis, which, in turn, promotes hepatocarcinogenesis.
Collapse
Affiliation(s)
- Chuanyi Lei
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Jun Chen
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Huiling Li
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Tingting Fan
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Xu Zheng
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Hong Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Nan Zhang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Yang Liu
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Xiaoqin Luo
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Jingyu Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| | - Aiguo Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian 116044, Liaoning, China
| |
Collapse
|
|
13
|
Zhen H, Qian X, Fu X, Chen Z, Zhang A, Shi L. Regulation of Shaoyao Ruangan Mixture on Intestinal Flora in Mice With Primary Liver Cancer. Integr Cancer Ther 2019; 18:1534735419843178. [PMID: 31006277 PMCID: PMC6477757 DOI: 10.1177/1534735419843178] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Background: Shaoyao Ruangan mixture (SRM) has been applied
clinically for more than 20 years in Zhejiang Cancer Hospital to treat patients
with primary liver cancer (PLC). Intestinal microecology plays an important role
in the emergence of liver diseases. This study aimed to reveal connections among
SRM, intestinal microbiota and PLC, and the potential targets of SRM for liver
cancer. Methods: We established a control group, a PLC model group,
and a treatment group of mice to analyze the inhibitory effect of SRM on PLC and
its intestinal flora target. We also evaluated drug efficacy of SRM and analyzed
specific changes in intestinal flora by 16S rDNA sequencing of stools. As the
serum interleukin (IL)-10 level could be an independent prognostic factor for
unresectable liver cancer, we detected IL-10 levels and analyzed their
association with the abundance of specific bacteria. Results: Liver
tumors in the treatment group were smaller and fewer than those in the model
group (P = .046). The abundance of Bacteroides
was significantly higher in the model group than that in the control group,
while SRM significantly reduced the increasing abundance of
Bacteroides in mice with PLC. We found that the IL-10 level
was positively correlated with the abundance of Bacteroides.
Conclusion: SRM can effectively inhibit the progression of PLC
and increase Bacteroides abundance. In view of the association
between Bacteroides and liver cancer and the significant
positive correlation between Bacteroides and IL-10 levels,
Bacteroides may be the target intestinal flora of SRM to
inhibit PLC.
Collapse
Affiliation(s)
- Hongde Zhen
- Second Clinical Medical College,
Zhejiang Chinese Medical University, Hangzhou, People’s Republic of China
- Zhejiang Cancer Hospital, Hangzhou,
People’s Republic of China
| | - Xiang Qian
- Zhejiang Cancer Hospital, Hangzhou,
People’s Republic of China
| | - Xiaoxuan Fu
- Second Clinical Medical College,
Zhejiang Chinese Medical University, Hangzhou, People’s Republic of China
| | - Zhuo Chen
- Second Clinical Medical College,
Zhejiang Chinese Medical University, Hangzhou, People’s Republic of China
| | - Aiqin Zhang
- Zhejiang Cancer Hospital, Hangzhou,
People’s Republic of China
| | - Lei Shi
- Zhejiang Cancer Hospital, Hangzhou,
People’s Republic of China
- Lei Shi, Department of Mammary Oncology,
Zhejiang Cancer Hospital, No. 1 East Banshan Road, Gongshu District, Hangzhou
310022, People’s Republic of China.
| |
Collapse
|
|
14
|
Yang HD, Kim HS, Kim SY, Na MJ, Yang G, Eun JW, Wang HJ, Cheong JY, Park WS, Nam SW. HDAC6 Suppresses Let-7i-5p to Elicit TSP1/CD47-Mediated Anti-Tumorigenesis and Phagocytosis of Hepatocellular Carcinoma. Hepatology 2019; 70:1262-1279. [PMID: 30991448 DOI: 10.1002/hep.30657] [Citation(s) in RCA: 63] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/22/2018] [Accepted: 03/30/2019] [Indexed: 12/24/2022]
Abstract
Histone deacetylase 6 (HDAC6) uniquely serves as a tumor suppressor in hepatocellular carcinogenesis, but the underlying mechanisms leading to tumor suppression are not fully understood. To identify comprehensive microRNAs (miRNAs) regulated by HDAC6 in hepatocellular carcinogenesis, differential miRNA expression analysis of HDAC6-transfected Hep3B cells was performed. Using integrative analyses of publicly available transcriptome data and miRNA target prediction, we selected five candidate miRNAs and, through in vitro functional validation, showed that let-7i-5p specifically suppressed thrombospondin-1 (TSP1) in hepatocellular carcinoma (HCC). Ectopic expression of antisense let-7i-5p (AS-let-7i-5p) inhibited in vitro tumorigenesis of HCC cells. In addition, treatments of partially purified TSP1 from culture cell media (ppTSP1) and recombinant TSP1 (rTSP1) exhibited similar effects with AS-let-7i-5p treatment on the same HCC cells, whereas TSP1 neutralizing antibody treatment significantly attenuated these effects. Notably, treatments of HDAC6 plasmid, AS-let-7i-5p, ppTSP1, and rTSP1 significantly suppressed in vitro angiogenesis and metastatic potential of HCC cells, but the co-treatment of TSP1 antibody specific to cluster of differentiation 47 (CD47) binding domain successfully blocked these effects in the same cells. Furthermore, we demonstrated that recovery of HDAC6 elicited let-7i-5p suppression to de-repress TSP1 expression; therefore, it occupied the CD47 receptor to block CD47-SIRPα-mediated anti-phagocytosis of macrophage in HCC. We also observed that HCC-derived exosomal let-7i-5p suppressed TSP1 of recipient hepatocyte cells. Treatments of HDAC6 plasmid, AS-let-7i-5p, and rTSP1 suppressed tumor incidence as well as tumor growth rates in a spontaneous mouse HCC model. Conclusion: Our findings suggest that the HDAC6-let-7i-5p-TSP1 regulatory pathway suppresses neoplastic and antiphagocytic behaviors of HCC by interacting with cell surface receptor CD47 in HCC and neighboring cells of tumor microenvironment, providing a therapeutic target for the treatment of liver malignancy and metastasis.
Collapse
Affiliation(s)
- Hee Doo Yang
- Department of Pathology, College of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
- Functional RNomics Research Center, the Catholic University of Korea, Seoul, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
| | - Hyung Seok Kim
- Department of Pathology, College of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
- Functional RNomics Research Center, the Catholic University of Korea, Seoul, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
| | - Sang Yean Kim
- Department of Pathology, College of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
- Functional RNomics Research Center, the Catholic University of Korea, Seoul, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
| | - Min Jeong Na
- Department of Pathology, College of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
- Functional RNomics Research Center, the Catholic University of Korea, Seoul, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
| | - Gyeongdeok Yang
- Department of Pathology, College of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
- Functional RNomics Research Center, the Catholic University of Korea, Seoul, Republic of Korea
| | - Jung Woo Eun
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, Republic of Korea
| | - Hee Jung Wang
- Department of Surgery, Ajou University School of Medicine, Suwon, Republic of Korea
| | - Jae Youn Cheong
- Department of Gastroenterology, Ajou University School of Medicine, Suwon, Republic of Korea
| | - Won Sang Park
- Department of Pathology, College of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
- Functional RNomics Research Center, the Catholic University of Korea, Seoul, Republic of Korea
| | - Suk Woo Nam
- Department of Pathology, College of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
- Functional RNomics Research Center, the Catholic University of Korea, Seoul, Republic of Korea
- Department of Biomedicine & Health Sciences, Graduate School of Medicine, the Catholic University of Korea, Seoul, Republic of Korea
| |
Collapse
|
|
15
|
Heo CK, Hwang HM, Lee HJ, Kwak SS, Yoo JS, Yu DY, Lim KJ, Lee S, Cho EW. Serum anti-EIF3A autoantibody as a potential diagnostic marker for hepatocellular carcinoma. Sci Rep 2019; 9:11059. [PMID: 31363116 PMCID: PMC6667438 DOI: 10.1038/s41598-019-47365-4] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2018] [Accepted: 07/11/2019] [Indexed: 02/06/2023] Open
Abstract
Tumor-associated autoantibodies are promising diagnostic biomarkers for early detection of tumors. We have screened a novel tumor-associated autoantibody in hepatocellular carcinoma (HCC) model mice. Its target antigen was identified as eukaryotic translation initiation factor 3 subunit A (EIF3A) by proteomic analysis, and the elevated expression of EIF3A in HCC tissues of tumor model mice as well as human patients was shown. Also, its existence in tumor-derived exosomes was revealed, which seem to be the cause of tumor-associated autoantibody production. To use serum anti-EIF3A autoantibody as biomarker, ELISA detecting anti-EIF3A autoantibody in human serum was performed using autoantibody-specific epitope. For the sensitive detection of serum autoantibodies its specific conformational epitopes were screened from the random cyclic peptide library, and a streptavidin antigen displaying anti-EIF3A autoantibody-specific epitope, XC90p2(-CPVRSGFPC-), was used as capture antigen. It distinguished patients with HCC (n = 102) from healthy controls (n = 0285) with a sensitivity of 79.4% and specificity of 83.5% (AUC = 0.87). Also, by simultaneously detecting with other HCC biomarkers, including alpha-fetoprotein, HCC diagnostic sensitivity improved from 79.4% to 85%. Collectively, we suggest that serum anti-EIF3A autoantibody is a useful biomarker for the diagnosis of HCC and the combinational detection of related biomarkers can enhance the accuracy of the cancer diagnosis.
Collapse
Affiliation(s)
- Chang-Kyu Heo
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141, South Korea.,College of Bioscience and Biotechnology, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon, 34134, South Korea
| | - Hai-Min Hwang
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141, South Korea.,College of Bioscience and Biotechnology, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon, 34134, South Korea
| | - Hye-Jung Lee
- Proteometech Inc., 1101 Wooree Venture Town, 466 Gangseo-ro, Gangseo-gu, Seoul, 03722, South Korea.,Graduate Program for Nanomedical Science, Yonsei University, 50 Yonsei-ro Seodaemun-gu, Seoul, 03722, South Korea
| | - Sang-Seob Kwak
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141, South Korea.,Department of Functional Genomics, University of Science and Technology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141, South Korea
| | - Jong-Shin Yoo
- Biomedical Omics Group, Korea Basic Science Institute, 162 YeonGuDanji-Ro, Ochang-eup, Cheongju, Chungbuk, 28119, South Korea
| | - Dae-Yeul Yu
- Disease Model Research Laboratory, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141, South Korea
| | - Kook-Jin Lim
- Proteometech Inc., 1101 Wooree Venture Town, 466 Gangseo-ro, Gangseo-gu, Seoul, 03722, South Korea
| | - Soojin Lee
- College of Bioscience and Biotechnology, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon, 34134, South Korea.
| | - Eun-Wie Cho
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141, South Korea. .,Department of Functional Genomics, University of Science and Technology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141, South Korea.
| |
Collapse
|
|
16
|
mTOR and ERK regulate VKORC1 expression in both hepatoma cells and hepatocytes which influence blood coagulation. Clin Exp Med 2018; 19:121-132. [PMID: 30306378 DOI: 10.1007/s10238-018-0528-z] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/06/2018] [Accepted: 09/22/2018] [Indexed: 02/08/2023]
Abstract
Deficiency of γ-glutamyl carboxylation of coagulation factors, as evidenced by the elevated level of Des-γ-carboxyl prothrombin (DCP), is a common feature in hepatocellular carcinoma patients. Additionally, treatment of cancer patients with mTOR inhibitors significantly increases hemorrhagic events. However, the underlying mechanisms remain unknown. In the present study, Vitamin K epoxide reductase complex subunit 1 (VKORC1) was found to be significantly down-regulated in clinical hepatoma tissues and most tested hepatoma cell lines. In vitro investigations showed that VKORC1 expression was promoted by p-mTOR at the translational level and repressed by p-ERK at the transcriptional level. By exploring Hras12V transgenic mice, a hepatic tumor model, VKROC1 was significantly down-regulated in hepatic tumors and showed prolonged activated partial prothrombin time (APTT). In vivo investigations further showed that VKORC1 expression was promoted by p-mTOR and repressed by p-ERK in both hepatoma and hepatocytes. Consistently, APTT and prothrombin time were significantly prolonged under the mTOR inhibitor treatment and significantly shortened under the ERK inhibitor treatment. Conclusively, these findings indicate that mTOR and ERK play crucial roles in controlling VKORC1 expression in both hepatoma and hepatocytes, which provides a valuable molecular basis for preventing hemorrhage in clinical therapies.
Collapse
|
|
17
|
Hwang HM, Heo CK, Lee HJ, Kwak SS, Lim WH, Yoo JS, Yu DY, Lim KJ, Kim JY, Cho EW. Identification of anti-SF3B1 autoantibody as a diagnostic marker in patients with hepatocellular carcinoma. J Transl Med 2018; 16:177. [PMID: 29954402 PMCID: PMC6025833 DOI: 10.1186/s12967-018-1546-z] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/05/2018] [Accepted: 06/12/2018] [Indexed: 12/31/2022] Open
Abstract
BACKGROUND Tumor-associated (TA) autoantibodies, which are generated by the immune system upon the recognition of abnormal TA antigens, are promising biomarkers for the early detection of tumors. In order to detect autoantibody biomarkers effectively, antibody-specific epitopes in the diagnostic test should maintain the specific conformations that are as close as possible to those presenting in the body. However, when using patients' serum as a source of TA autoantibodies the characterization of the autoantibody-specific epitope is not easy due to the limited amount of patient-derived serum. METHODS To overcome these limits, we constructed a B cell hybridoma pool derived from a hepatocellular carcinoma (HCC) model HBx-transgenic mouse and characterized autoantibodies derived from them as tumor biomarkers. Their target antigens were identified by mass spectrometry and the correlations with HCC were examined. With the assumption that TA autoantibodies generated in the tumor mouse model are induced in human cancer patients, the enzyme-linked immunosorbent assays (ELISA) based on the characteristics of mouse TA autoantibodies were developed for the detection of autoantibody biomarkers in human serum. To mimic natural antigenic structures, the specific epitopes against autoantibodies were screened from the phage display cyclic random heptapeptide library, and the streptavidin antigens fused with the specific epitopes were used as coating antigens. RESULTS In this study, one of HCC-associated autoantibodies derived from HBx-transgenic mouse, XC24, was characterized. Its target antigen was identified as splicing factor 3b subunit 1 (SF3B1) and the high expression of SF3B1 was confirmed in HCC tissues. The specific peptide epitopes against XC24 were selected and, among them, XC24p11 cyclic peptide (-CDATPPRLC-) was used as an epitope of anti-SF3B1 autoantibody ELISA. With this epitope, we could effectively distinguish between serum samples from HCC patients (n = 102) and healthy subjects (n = 85) with 73.53% sensitivity and 91.76% specificity (AUC = 0.8731). Moreover, the simultaneous detection of anti-XC24p11 epitope autoantibody and AFP enhanced the efficiency of HCC diagnosis with 87.25% sensitivity and 90.59% specificity (AUC = 0.9081). CONCLUSIONS ELISA using XC24p11 peptide epitope that reacts against anti-SF3B1 autoantibody can be used as a novel test to enhance the diagnostic efficiency of HCC.
Collapse
Affiliation(s)
- Hai-Min Hwang
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
- Department of Microbiology and Molecular Biology, College of Bioscience and Biotechnology, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon, 34134 South Korea
| | - Chang-Kyu Heo
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
- Department of Microbiology and Molecular Biology, College of Bioscience and Biotechnology, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon, 34134 South Korea
| | - Hye Jung Lee
- Proteometech Inc., 1101 Wooree Venture Town, 466 Gangseo-ro, Gangseo-gu, Seoul, 07573 South Korea
- Graduate Program for Nanomedical Science, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul, 03722 South Korea
| | - Sang-Seob Kwak
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
- Department of Functional Genomics, University of Science and Technology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
| | - Won-Hee Lim
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
- Department of Functional Genomics, University of Science and Technology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
| | - Jong-Shin Yoo
- Biomedical Omics Group, Korea Basic Science Institute, 162 YeonGuDanji-ro, Ochang-eup, Cheongju, Chungbuk 28119 South Korea
| | - Dae-Yuel Yu
- Disease Model Research Laboratory, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
| | - Kook Jin Lim
- Proteometech Inc., 1101 Wooree Venture Town, 466 Gangseo-ro, Gangseo-gu, Seoul, 07573 South Korea
- Graduate Program for Nanomedical Science, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul, 03722 South Korea
| | - Jeong-Yoon Kim
- Department of Microbiology and Molecular Biology, College of Bioscience and Biotechnology, Chungnam National University, 99 Daehak-ro, Yuseong-gu, Daejeon, 34134 South Korea
| | - Eun-Wie Cho
- Rare Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
- Department of Functional Genomics, University of Science and Technology, 125 Gwahak-ro, Yuseong-gu, Daejeon, 34141 South Korea
| |
Collapse
|
|
18
|
Kopanja D, Pandey A, Huang S, Al Raheed MRH, Guzman G, Raychaudhuri P. p19Arf inhibits aggressive progression of H-ras-driven hepatocellular carcinoma. Carcinogenesis 2018; 39:318-326. [PMID: 29228217 PMCID: PMC5862269 DOI: 10.1093/carcin/bgx140] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2017] [Accepted: 11/15/2017] [Indexed: 11/14/2022] Open
Abstract
Arf, a well-established tumor suppressor, is either mutated or downregulated in a wide array of cancers. However, its role in hepatocellular carcinoma (HCC) progression is controversial. Conflicting observations have been published regarding its expression in HCC. In this study, we provide clear genetic evidence demonstrating a protective role of p19Arf in hepatocarcinogenesis. Using Ras-induced mouse model, we show that p19Arf deficiency accelerates progression of aggressive HCC in vivo. To investigate the role of p14ARF in human liver cancers, we analyzed its expression in human HCC using immunohistochemistry (IHC). We observe lack of nucleolar p14ARF in 43.02% of human HCC samples and that low expression of p14ARF strongly correlates with the early onset of HCC. Importantly, cirrhotic livers that did not progress to HCC harbor higher expression of the p14ARF protein in hepatocytes compared with that in cirrhotic livers with HCC. These results are significant because they suggest that nucleolar p14ARF can be used as early prognostic marker in chronic liver disease to reliably identify patients with high risk for developing liver cancer. Currently, there is no effective systemic therapy for advanced liver cancer; hence, more efficient patient screening and early detection of HCC would significantly contribute to the eradication of this devastating disease.
Collapse
Affiliation(s)
- Dragana Kopanja
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, USA
| | - Akshay Pandey
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, USA
| | - Shuo Huang
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, USA
| | | | - Grace Guzman
- Department of Pathology, University of Illinois, College of Medicine, USA
| | - Pradip Raychaudhuri
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, USA
- Department of Research, Jesse Brown VA Medical Center, USA
| |
Collapse
|
|
19
|
Han B, Shin HJ, Bak IS, Bak Y, Jeong YL, Kwon T, Park YH, Sun HN, Kim CH, Yu DY. Peroxiredoxin I is important for cancer-cell survival in Ras-induced hepatic tumorigenesis. Oncotarget 2018; 7:68044-68056. [PMID: 27517622 PMCID: PMC5356538 DOI: 10.18632/oncotarget.11172] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/11/2016] [Accepted: 07/27/2016] [Indexed: 02/06/2023] Open
Abstract
Peroxiredoxin I (Prx I), an antioxidant enzyme, has multiple functions in human cancer. However, the role of Prx I in hepatic tumorigenesis has not been characterized. Here we investigated the relevance and underlying mechanism of Prx I in hepatic tumorigenesis. Prx I increased in tumors of hepatocellular carcinoma (HCC) patients that aligned with overexpression of oncogenic H-ras. Prx I also increased in H-rasG12V transfected HCC cells and liver tumors of H-rasG12V transgenic (Tg) mice, indicating that Prx I may be involved in Ras-induced hepatic tumorigenesis. When Prx I was knocked down or deleted in HCC-H-rasG12V cells or H-rasG12V Tg mice, cell colony or tumor formation was significantly reduced that was associated with downregulation of pERK pathway as well as increased intracellular reactive oxygen species (ROS) induced DNA damage and cell death. Overexpressing Prx I markedly increased Ras downstream pERK/FoxM1/Nrf2 signaling pathway and inhibited oxidative damage in HCC cells and H-rasG12V Tg mice. In this study, we found Nrf2 was transcriptionally activated by FoxM1, and Prx I was activated by the H-rasG12V/pERK/FoxM1/Nrf2 pathway and suppressed ROS-induced hepatic cancer-cell death along with formation of a positive feedback loop with Ras/ERK/FoxM1/Nrf2 to promote hepatic tumorigenesis.
Collapse
Affiliation(s)
- Bing Han
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea.,Department of Biology, Chungnam National University, Daejeon, 305-764, Korea
| | - Hye-Jun Shin
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea
| | - In Seon Bak
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea.,Department of Toxicology Evaluation, Graduate School of Preclinical Laboratory Science, Konyang University, Daejeon, 363-700, Korea
| | - Yesol Bak
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea.,Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, Seoul, 143-701, Korea
| | - Ye-Lin Jeong
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea.,Department of Animal Biosystem Sciences, Chungnam National University, Daejeon, 305-764, Korea
| | - Taeho Kwon
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea
| | - Young-Ho Park
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea
| | - Hu-Nan Sun
- College of Life Science and Biotechnology, Heilongjiang Bayi Agricultural University, Daqing, 163319, China
| | - Cheol-Hee Kim
- Department of Biology, Chungnam National University, Daejeon, 305-764, Korea
| | - Dae-Yeul Yu
- Genome Editing Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 305-806, Korea
| |
Collapse
|
|
20
|
IGF-II induced by hepatitis B virus X protein regulates EMT via SUMO mediated loss of E-cadherin in mice. Oncotarget 2018; 7:56944-56957. [PMID: 27486970 PMCID: PMC5302964 DOI: 10.18632/oncotarget.10922] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2016] [Accepted: 07/18/2016] [Indexed: 12/31/2022] Open
Abstract
Hepatocellular carcinoma (HCC) is one of the most common cancers and a leading cause of cancer mortality. Prognosis of this disease largely depends on its stage. An Enlarged liver, due to dysplasia, may be a critical point in the multi-step progression to HCC. The mechanism underlying hepatomegaly in human and mouse models are poorly understood. We previously reported we observed enlarged liver in hepatitis B virus X protein (HBx) expressing mice (HBx mice). Here we identify the critical role of HBx induced IGF-II in hepatomegaly in mice and abnormal cell growth in human hepatoma cells. We found that HBx induced IGF-II is essential to induce epithelial-mesenchymal transition (EMT) through loss of E-cadherin. In mouse liver, loss of E-cadherin was mediated by post-translational regulation, at least in part, by protease and SUMOylation not by transcriptional regulation. In contrast, in hepatoma cell line (HepG2 cells) Akt signal pathway controls the mRNA expression level of EMT-related transcription factors, especially Twist, in addition to post- translational modification through SUMOylation. Thus, IGF-II-mediated loss of E-cadherin is central in developing hepatomegaly in mice and abnormal cell growth in the hepatoma cell line. HBx induced IGF-II represents a potential biomarker, which is also a therapeutic target in HCC.
Collapse
|
|
21
|
Fan T, Rong Z, Dong J, Li J, Wang K, Wang X, Li H, Chen J, Wang F, Wang J, Wang A. Metabolomic and transcriptomic profiling of hepatocellular carcinomas in Hras12V transgenic mice. Cancer Med 2017; 6:2370-2384. [PMID: 28941178 PMCID: PMC5633588 DOI: 10.1002/cam4.1177] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/16/2017] [Revised: 07/31/2017] [Accepted: 08/07/2017] [Indexed: 12/19/2022] Open
Abstract
Activation of the Ras/MAPK pathway is prevalently involved in the occurrence and development of hepatocellular carcinoma (HCC). However, its effects on the deregulated cellular metabolic processes involved in HCC in vivo remain unknown. In this study, a mouse model of HCC induced by hepatocyte-specific expression of the Hras12V oncogene was investigated using an integrative analysis of metabolomics and transcriptomics data. Consistent with the phenotype of abundant lipid droplets in HCC, the lipid biosynthesis in HCC was significantly enhanced by (1) a sufficient supply of acetyl-CoA from enhanced glycolysis and citrate shuttle activity; (2) a sufficient supply of NADPH from enhanced pentose phosphate pathway (PPP) activity; (3) upregulation of key enzymes associated with lipid biosynthesis; and (4) downregulation of key enzymes associated with bile acid biosynthesis. In addition, glutathione (GSH) was significantly elevated, which may result from a sufficient supply of 5-oxoproline and L-glutamate as well as an enhanced reduction in the process of GSSG being turned into GSH by NADPH. The high level of GSH along with elevated Bcl2 and Ucp2 expression may contribute to a normal level of reactive oxygen species (ROS) in HCC. In conclusion, our results suggest that the lipid metabolism, glycolysis, PPP, tricarboxylic acid (TCA) cycle, citrate shuttle activity, bile acid synthesis, and redox homeostasis in the HCC induced by ras oncogene are significantly perturbed, and these altered metabolic processes may play crucial roles in the carcinogenesis, development, and pathological characteristics of HCC.
Collapse
Affiliation(s)
- Tingting Fan
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Zhuona Rong
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Jianyi Dong
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Juan Li
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Kangwei Wang
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Xinxin Wang
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Huiling Li
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Jun Chen
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Fujin Wang
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Jingyu Wang
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| | - Aiguo Wang
- Laboratory animal center, Dalian medical University, Dalian, Liaoning, 116044, China
| |
Collapse
|
|
22
|
Wang K, Nie X, Rong Z, Fan T, Li J, Wang X, Li H, Dong J, Chen J, Wang F, Wang J, Wang A. B lymphocytes repress hepatic tumorigenesis but not development in Hras12V transgenic mice. Int J Cancer 2017; 141:1201-1214. [PMID: 28580661 DOI: 10.1002/ijc.30823] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2016] [Revised: 04/20/2017] [Accepted: 05/23/2017] [Indexed: 01/10/2023]
Abstract
Increasing reports show noninflammation underlying HCC, challenging our understanding of the roles of the immune system in hepatocarcinogenesis. By exploring a mouse model of hepatic tumor induced by hepatocyte-specific expression of the Hras12V oncogene without obvious inflammation, we found that the proportion of B cells, but not T cells, progressively and significantly decreased in 3, 5-month-old transgenic mice (Tg) compared with non-transgenic mice. Notably, the proportions of total and activated B and T cells all significantly decreased in 9-month-old Tg with multiple massive hepatic tumors. Together with the decreased B cell proportion, serum IgG1/2 also significantly decreased in 5, 9-month-old Tg. Interestingly, homozygous Tg showed significantly higher B cell proportion and IgG2 levels, accompanied by significantly lower incidences of liver nodules but not adenomas and carcinomas compared with heterozygous Tg. Treatment of Tg with PCI-32765, a potent Bruton's tyrosine kinase (BTK) inhibitor for suppressing B cell proliferation and activation, significantly decreased the B cell proportion and IgG2 levels, accompanied by a significantly higher incidence of liver nodules, but had no effects on adenoma and carcinoma. Treatment of Tg with insulin-like growth factor 1 (IGF-1) significantly increased the B cell proportion and IgG2 levels, accompanied by a significantly lower incidence of liver nodules and carcinoma, but had no effects on adenoma. Conclusively, B cells and IgG2 may play important roles in suppressing hepatic tumorigenesis, but not development. In addition, hepatocyte-specific expression of the ras oncogene may play roles in suppressing B cells, while developed hepatic tumors suppress both B and T cells.
Collapse
Affiliation(s)
- Kangwei Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Xin Nie
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Zhuona Rong
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Tingting Fan
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Juan Li
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Xinxin Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Huiling Li
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Jianyi Dong
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Jun Chen
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Fujin Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Jingyu Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| | - Aiguo Wang
- Department of Comparative Medicine, Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning, People's Republic of China
| |
Collapse
|
|
23
|
Rong Z, Fan T, Li H, Li J, Wang K, Wang X, Dong J, Chen J, Wang F, Wang J, Wang A. Differential Proteomic Analysis of Gender-dependent Hepatic Tumorigenesis in Hras12V Transgenic Mice. Mol Cell Proteomics 2017; 16:1475-1490. [PMID: 28512230 DOI: 10.1074/mcp.m116.065474] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2016] [Revised: 04/29/2017] [Indexed: 12/18/2022] Open
Abstract
Male prevalence is an outstanding characteristic of hepatocellular carcinoma (HCC), and the underlying mechanisms for this have remained largely unknown. In the present study, Hras12V transgenic mice, in which hepatocyte-specific expression of the ras oncogene induces male-biased hepatic tumorigenesis, were studied, and altered proteins were detected by two-dimensional fluorescence difference gel electrophoresis (2D-DIGE). Protein samples from hepatic tumor tissues (T) and peritumor tissues (P) of transgenic males and females and the corresponding normal liver tissues (Wt) of nontransgenic males and females were subjected to pairwise comparisons based on proteomic analysis. Among 2381 autodetected protein spots, more than 1600 were differentially expressed based on a pairwise comparison (|ratio| > = 1.5, p < = 0.05). Of these, 180 spots were randomly selected for matrix-assisted laser desorption ionization tandem time-of-flight mass spectrometry (MALDI-TOF/TOF MS) identification; finally, 89 distinct proteins were obtained. Among these 89 proteins, 7 and 50 proteins were further validated by Western blotting and literature investigation, respectively. Intriguingly, compared with Wt, the altered proteins were relatively concentrated in T in transgenic females but in P in transgenic males. Consistently, the levels of p-ERK and p-mTOR were significantly higher in the T of females compared with that of males. The pathway enrichment assay showed that 5 pathways in males but only 1 in females were significantly altered in terms of the upregulated proteins in T compared with Wt. These data indicate that female hepatocytes are disturbed by oncogenes with great difficulty, whereas male hepatocytes readily do so. In addition, 33 proteins were gender-dependently altered in hepatic tumorigenesis. Moreover, 4% DNA packaging and 4% homeostasis-related functional proteins were found in females but not in males, and more nucleus proteins were found in females (8%) than in males (3%). In conclusion, the proteomic data and comparative analysis presented here offer crucial clues for elucidating the mechanisms that underlie the male prevalence in HCC.
Collapse
Affiliation(s)
- Zhuona Rong
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Tingting Fan
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Huiling Li
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Juan Li
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Kangwei Wang
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Xinxin Wang
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Jianyi Dong
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Jun Chen
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Fujin Wang
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China
| | - Jingyu Wang
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China.
| | - Aiguo Wang
- From the ‡Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116000, China.
| |
Collapse
|
|
24
|
Park YH, Kim SU, Kwon TH, Kim JM, Song IS, Shin HJ, Lee BK, Bang DH, Lee SJ, Lee DS, Chang KT, Kim BY, Yu DY. Peroxiredoxin II promotes hepatic tumorigenesis through cooperation with Ras/Forkhead box M1 signaling pathway. Oncogene 2015; 35:3503-13. [PMID: 26500057 DOI: 10.1038/onc.2015.411] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2015] [Revised: 09/09/2015] [Accepted: 09/18/2015] [Indexed: 12/14/2022]
Abstract
The current study was carried out to define the involvement of Peroxiredoxin (Prx) II in progression of hepatocellular carcinoma (HCC) and the underlying molecular mechanism(s). Expression and function of Prx II in HCC was determined using H-ras(G12V)-transformed HCC cells (H-ras(G12V)-HCC cells) and the tumor livers from H-ras(G12V)-transgenic (Tg) mice and HCC patients. Prx II was upregulated in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg mouse tumor livers, the expression pattern of which highly similar to that of forkhead Box M1 (FoxM1). Moreover, either knockdown of FoxM1 or site-directed mutagenesis of FoxM1-binding site of Prx II promoter significantly reduced Prx II levels in H-ras(G12V)-HCC cells, indicating FoxM1 as a direct transcription factor of Prx II in HCC. Interestingly, the null mutation of Prx II markedly decreased the number and size of tumors in H-ras(G12V)-Tg livers. Consistent with this, knockdown of Prx II in H-ras(G12V)-HCC cells reduced the expression of cyclin D1, cell proliferation, anchorage-independent growth and tumor formation in athymic nude mice, whereas overexpression of Prx II increased or aggravated the tumor phenotypes. Importantly, the expression of Prx II was correlated with that of FoxM1 in HCC patients. The activation of extracellular signal-related kinase (ERK) pathway and the expression of FoxM1 and cyclin D1 were highly dependent on Prx II in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg livers. Prx II is FoxM1-dependently-expressed antioxidant in HCC and function as an enhancer of Ras(G12V) oncogenic potential in hepatic tumorigenesis through activation of ERK/FoxM1/cyclin D1 cascade.
Collapse
Affiliation(s)
- Y-H Park
- Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.,National Primate Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.,Department of Functional Genomics, University of Science and Technology, Daejeon, Korea
| | - S-U Kim
- Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.,National Primate Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.,Department of Functional Genomics, University of Science and Technology, Daejeon, Korea
| | - T-H Kwon
- Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea
| | - J-M Kim
- School of Medicine, Chungnam National University, Daejeon, Korea
| | - I-S Song
- Cardiovascular and Metabolic Disease Center, Inje University, Busan, Korea
| | - H-J Shin
- Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea
| | - B-K Lee
- Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea
| | - D-H Bang
- School of Medicine, Wonkwang University, Iksan, Korea
| | - S-J Lee
- Research Institute for Natural Sciences, Hanyang University, Seoul, Korea
| | - D-S Lee
- College of Natural Sciences, Kyungpook National University, Daegu, Korea
| | - K-T Chang
- National Primate Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea
| | - B-Y Kim
- World Class Institute, Korea Research Institute of Bioscience and Biotechnology, Ochang, Korea
| | - D-Y Yu
- Aging Intervention Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Korea.,Department of Functional Genomics, University of Science and Technology, Daejeon, Korea
| |
Collapse
|
|
25
|
Kopanja D, Pandey A, Kiefer M, Wang Z, Chandan N, Carr JR, Franks R, Yu DY, Guzman G, Maker A, Raychaudhuri P. Essential roles of FoxM1 in Ras-induced liver cancer progression and in cancer cells with stem cell features. J Hepatol 2015; 63:429-436. [PMID: 25828473 PMCID: PMC4508215 DOI: 10.1016/j.jhep.2015.03.023] [Citation(s) in RCA: 71] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/20/2014] [Revised: 03/13/2015] [Accepted: 03/19/2015] [Indexed: 12/13/2022]
Abstract
BACKGROUND & AIMS Overexpression of FoxM1 correlates with poor prognosis in hepatocellular carcinoma (HCC). Moreover, the Ras-signaling pathway is found to be ubiquitously activated in HCC through epigenetic silencing of the Ras-regulators. We investigated the roles of FoxM1 in Ras-driven HCC, and on HCC cells with stem-like features. METHODS We employed a transgenic mouse model that expresses the oncogenic Ras in the liver. That strain was crossed with a strain that harbor floxed alleles of FoxM1 and the MxCre gene that allows conditional deletion of FoxM1. FoxM1 alleles were deleted after development of HCC, and the effects on the tumors were analyzed. Also, FoxM1 siRNA was used in human HCC cell lines to determine its role in the survival of the HCC cells with stem cell features. RESULTS Ras-driven tumors overexpress FoxM1. Deletion of FoxM1 inhibits HCC progression. There was increased accumulation of reactive oxygen species (ROS) in the FoxM1 deleted HCC cells. Moreover, FoxM1 deletion caused a disproportionate loss of the CD44+ and EpCAM+ HCC cells in the tumors. We show that FoxM1 directly activates expression of CD44 in human HCC cells. Moreover, the human HCC cells with stem cell features are addicted to FoxM1 for ROS-regulation and survival. CONCLUSION Our results provide genetic evidence for an essential role of FoxM1 in the progression of Ras-driven HCC. In addition, FoxM1 is required for the expression of CD44 in HCC cells. Moreover, FoxM1 plays a critical role in the survival of the HCC cells with stem cell features by regulating ROS.
Collapse
Affiliation(s)
- Dragana Kopanja
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607, United States
| | - Akshay Pandey
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607, United States
| | - Megan Kiefer
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607, United States
| | - Zebin Wang
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607, United States
| | - Neha Chandan
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607, United States
| | - Janai R Carr
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607, United States; Department of Medicine, University of California, San Francisco, CA, United States
| | - Roberta Franks
- Transgenic Production Facility, University of Illinois, College of Medicine, 909 S. Wolcott Ave, Chicago, IL 60612, United States
| | - Dae-Yeul Yu
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 305-806, Republic of Korea
| | - Grace Guzman
- Department of Pathology, University of Illinois, College of Medicine, 840 S. Wood St, Chicago, IL 60612, United States
| | - Ajay Maker
- Department of Medicine, University of Illinois, College of Medicine, 909 S. Wolcott Ave, Chicago, IL 60612, United States
| | - Pradip Raychaudhuri
- Department of Biochemistry and Molecular Genetics (M/C 669), University of Illinois, College of Medicine, 900 S. Ashland Ave., Chicago, IL 60607, United States; Jesse Brown VA Medical Center, 820 S. Damen Ave., Chicago, IL 60612, United States.
| |
Collapse
|
|
26
|
Kim SK, Kim H, Koh GY, Lim DS, Yu DY, Kim MD, Park MS, Lim JS. Mouse Hepatic Tumor Vascular Imaging by Experimental Selective Angiography. PLoS One 2015; 10:e0131687. [PMID: 26131558 PMCID: PMC4489182 DOI: 10.1371/journal.pone.0131687] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2015] [Accepted: 06/05/2015] [Indexed: 12/18/2022] Open
Abstract
PURPOSE Human hepatocellular carcinoma (HCC) has unique vascular features, which require selective imaging of hepatic arterial perfusion and portal venous perfusion with vascular catheterization for sufficient evaluation. Unlike in humans, vessels in mice are too small to catheterize, and the importance of separately imaging the feeding vessels of tumors is frequently overlooked in hepatic tumor models. The purpose of this study was to perform selective latex angiography in several mouse liver tumor models and assess their suitability. MATERIALS AND METHODS In several ectopic (Lewis lung carcinoma, B16/F10 melanoma cell lines) and spontaneous liver tumor (Albumin-Cre/MST1fl/fl/MST2fl/fl, Albumin-Cre/WW45fl/fl, and H-ras12V genetically modified mouse) models, the heart left ventricle and/or main portal vein of mice was punctured, and latex dye was infused to achieve selective latex arteriography and/or portography. RESULTS H-ras12V transgenic mice (a HCC and hepatic adenoma model) developed multiple liver nodules that displayed three different perfusion patterns (portal venous or hepatic artery perfusion predominant, mixed perfusion), indicating intra-tumoral vascular heterogeneity. Selective latex angiography revealed that the Lewis lung carcinoma implant model and the Albumin-Cre/WW45fl/fl model reproduced conventional angiography findings of human HCC. Specifically, these mice developed tumors with abundant feeding arteries but no portal venous perfusion. CONCLUSION Different hepatic tumor models showed different tumor vessel characteristics that influence the suitability of the model and that should be considered when designing translational experiments. Selective latex angiography applied to certain mouse tumor models (both ectopic and spontaneous) closely simulated typical characteristics of human HCC vascular imaging.
Collapse
Affiliation(s)
- Sang Kyum Kim
- Department of Pathology, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Honsoul Kim
- Department of Radiology, Research Institute of Radiological Science, Yonsei University College of Medicine, Seoul, Republic of Korea
- * E-mail:
| | - Gou Young Koh
- National Research Laboratory of Vascular Biology and Stem Cell, Graduate School of Medical Science and Engineering, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Republic of Korea
| | - Dae-Sik Lim
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, Republic of Korea
| | - Dae-Yeul Yu
- Aging Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea
| | - Man Deuk Kim
- Department of Radiology, Research Institute of Radiological Science, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Mi-Suk Park
- Department of Radiology, Research Institute of Radiological Science, Yonsei University College of Medicine, Seoul, Republic of Korea
| | - Joon Seok Lim
- Department of Radiology, Research Institute of Radiological Science, Yonsei University College of Medicine, Seoul, Republic of Korea
| |
Collapse
|
|
27
|
Wang AG, Song YN, Chen J, Li HL, Dong JY, Cui HP, Yao L, Li XF, Gao WT, Qiu ZW, Wang FJ, Wang JY. Activation of RAS/ERK alone is insufficient to inhibit RXRα function and deplete retinoic acid in hepatocytes. Biochem Biophys Res Commun 2014; 452:801-7. [PMID: 25218146 DOI: 10.1016/j.bbrc.2014.09.007] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2014] [Accepted: 09/01/2014] [Indexed: 12/19/2022]
Abstract
Activation of RAS/ERK signaling pathway, depletion of retinoid, and phosphorylation of retinoid X receptor alpha (RXRα) are frequent events found in liver tumors and thought to play important roles in hepatic tumorigenesis. However, the relationships among them still remained to be elucidated. By exploring the transgenic mouse model of hepatic tumorigenesis induced by liver-specific expression of H-ras12V oncogene, the activation of RAS/ERK, the mRNA expression levels of retinoid metabolism-related genes, the contents of retinoid metabolites, and phosphorylation of RXRα were determined. RAS/ERK signaling pathway was gradually and significantly activated in hepatic tumor adjacent normal liver tissues (P) and hepatic tumor tissues (T) of H-ras12V transgenic mice compared with normal liver tissues (Wt) of wild type mice. On the contrary, the mRNA expression levels of retinoid metabolism-related genes were significantly reduced in P and T compared with Wt. Interestingly, the retinoid metabolites 9-cis-retinoic acid (9cRA) and all-trans-retinoic acid (atRA), the well known ligands for nuclear transcription factor RXR and retinoic acid receptor (RAR), were significantly decreased only in T compared with Wt and P, although the oxidized polar metabolite of atRA, 4-keto-all-trans-retinoic-acid (4-keto-RA) was significantly decreased in both P and T compared with Wt. To our surprise, the functions of RXRα were significantly blocked only in T compared with Wt and P. Namely, the total protein levels of RXRα were significantly reduced and the phosphorylation levels of RXRα were significantly increased only in T compared with Wt and P. Treatment of H-ras12V transgenic mice at 5-week-old or 5-month-old with atRA had no effect on the prevention of tumorigenesis or cure of developed nodules in liver. These events imply that the depletion of 9cRA and atRA and the inhibition of RXRα function in hepatic tumors involve more complex mechanisms besides the activation of RAS/ERK pathway.
Collapse
Affiliation(s)
- Ai-Guo Wang
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China.
| | - Ya-Nan Song
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Jun Chen
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Hui-Ling Li
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Jian-Yi Dong
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Hai-Peng Cui
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Liang Yao
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Xue-Feng Li
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Wen-Ting Gao
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Ze-Wen Qiu
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Fu-Jin Wang
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China
| | - Jing-Yu Wang
- Laboratory Animal Center, Dalian Medical University, Dalian, Liaoning 116044, PR China.
| |
Collapse
|
|
28
|
Kim JH, Shin HJ, Ha HL, Park YH, Kwon TH, Jung MR, Moon HB, Cho ES, Son HY, Yu DY. Methylsulfonylmethane suppresses hepatic tumor development through activation of apoptosis. World J Hepatol 2014; 6:98-106. [PMID: 24575169 PMCID: PMC3934636 DOI: 10.4254/wjh.v6.i2.98] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/29/2013] [Revised: 11/06/2013] [Accepted: 12/13/2013] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the effect of methylsulfonylmethane (MSM), recently reported to have anti-cancer effects, in liver cancer cells and transgenic mice.
METHODS: Three liver cancer cell lines, HepG2, Huh7-Mock and Huh7-H-rasG12V, were used. Cell growth was measured by Cell Counting Kit-8 and soft agar assay. Western blot analysis was used to detect caspases, poly (ADP-ribose) polymerase (PARP), and B-cell lymphoma 2 (Bcl-2) expressions. For in vivo study, we administered MSM to H-ras12V transgenic mice for 3 mo.
RESULTS: MSM decreased the growth of HepG2, Huh7-Mock and Huh7-H-rasG12V cells in a dose-dependent manner. That was correlated with significantly increased apoptosis and reduced cell numbers in MSM treated cells. Cleaved caspase-8, cleaved caspase-3 and cleaved PARP were remarkably increased in the liver cancer cells treated with 500 mmol/L of MSM; however, Bcl-2 was slightly decreased in 500 mmol/L. Liver tumor development was greatly inhibited in the H-ras12V transgenic mice treated with MSM, compared to control, by showing reduced tumor size and number. Cleaved PARP was significantly increased in non-tumor treated with MSM compared to control.
CONCLUSION: Liver injury was also significantly attenuated in the mice treated with MSM. Taken together, all the results suggest that MSM has anti-cancer effects through inducing apoptosis in liver cancer.
Collapse
|
|
29
|
Kim YK, Minai-Tehrani A, Lee JH, Cho CS, Cho MH, Jiang HL. Therapeutic efficiency of folated poly(ethylene glycol)-chitosan-graft-polyethylenimine-Pdcd4 complexes in H-ras12V mice with liver cancer. Int J Nanomedicine 2013; 8:1489-98. [PMID: 23620665 PMCID: PMC3630991 DOI: 10.2147/ijn.s42949] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/23/2023] Open
Abstract
Background Chitosan and chitosan derivatives have been proposed as alternative and biocompatible cationic polymers for nonviral gene delivery. However, the low transfection efficiency and low specificity of chitosan is an aspect of this approach that must be addressed prior to any clinical application. In the present study, folated poly(ethylene glycol)-chitosan-graft-polyethylenimine (FPCP) was investigated as a potential folate receptor-overexpressed cancer cell targeting gene carrier. Methods The FPCP copolymer was synthesized in two steps. In the first step, folate-PEG was synthesized by an amide formation reaction between the activated carboxyl groups of folic acid and the amine groups of bifunctional poly(ethylene glycol) (PEG). In the second step, FPCP was synthesized by an amide formation reaction between the activated carboxyl groups of folate-PEG and amine groups of CHI-g-polyethyleneimine (PEI). The composition of FPCP was characterized by 1H nuclear magnetic resonance. Results: FPCP showed low cytotoxicity in various cell lines, and FPCP-DNA complexes showed good cancer cell specificity as well as good transfection efficiency in the presence of serum. Further, FPCP-Pdcd4 complexes reduced tumor numbers and progression more effectively than PEI 25 kDa in H-ras12V liver cancer mice after intravenous administration. Conclusion Our data suggest that FPCP, which has improved transfection efficiency and cancer cell specificity, may be useful in gene therapy for liver cancer.
Collapse
Affiliation(s)
- You-Kyoung Kim
- State Key Laboratory of Natural Medicines, Department of Pharmaceutics, China Pharmaceutical University, Nanjing, People's Republic of China
| | | | | | | | | | | |
Collapse
|
|
30
|
Shin JY, Chung YS, Kang B, Jiang HL, Yu DY, Han K, Chae C, Moon JH, Jang G, Cho MH. Co-delivery of LETM1 and CTMP synergistically inhibits tumor growth in H-ras12V liver cancer model mice. Cancer Gene Ther 2013; 20:186-94. [PMID: 23392203 DOI: 10.1038/cgt.2013.6] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
As hepatocellular carcinoma (HCC) is one of the most common tumors worldwide, development of novel therapeutic approaches for HCC is urgently needed. Two different genes, LETM1 and CTMP, which target mitochondrial functions, were chosen and linked using 2A-peptide sequence. Successful self-cleavage of 2A-peptide induced synergistic antitumor effect in the liver of H-ras12V, the HCC model mice, by simultaneous activation of LETM1 (Leucine zipper/EF hand-containing transmembrane-1) and CTMP (carboxyl-terminal modulator protein). Overexpression of LETM1 and CTMP significantly reduced the incidence of tumorigenesis, which were confirmed by gross and microscopic observations. Morphological changes in mitochondria, such as swelling and loss of cristae, were significant, and the prolonged activation of defects in mitochondrial function led to mitochondria-mediated apoptosis. Furthermore, with CTMP as a direct binding partner of Akt1, and LETM1 as a binding partner of CTMP, LETM1-2A-CTMP downregulated the Akt1 pathway at both Ser473 and Thr308 sites of phosphorylation. Proliferation and angiogenesis, which are important in cancer prognosis, were reduced in tumor sites after introduction of LETM1-2A-CTMP. Taken together, the results indicate that introduction of the mitochondria-targeting genes, LETM1 and CTMP, and self-processing capacity of 2A-peptide sequence exerts an antitumor effect in liver of H-ras12V mice, suggesting its potential as a tool for gene therapy.
Collapse
Affiliation(s)
- J-Y Shin
- Laboratory of Toxicology, College of Veterinary Medicine, Seoul National University, Seoul, Korea
| | | | | | | | | | | | | | | | | | | |
Collapse
|
|
31
|
Heo CK, Hwang HM, Ruem A, Yu DY, Lee JY, Yoo JS, Kim IG, Yoo HS, Oh S, Ko JH, Cho EW. Identification of a mimotope for circulating anti-cytokeratin 8/18 antibody and its usage for the diagnosis of breast cancer. Int J Oncol 2012; 42:65-74. [PMID: 23128437 PMCID: PMC3583721 DOI: 10.3892/ijo.2012.1679] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2012] [Accepted: 09/14/2012] [Indexed: 12/22/2022] Open
Abstract
A novel circulating tumor-associated autoantibody, K94, obtained from a hepatocellular carcinoma (HCC) mouse model was characterized. The target antigen of K94 autoanti-body was expressed in various tumor cell lines including liver cancer, and its secretion was detectable using MCF-7 breast carcinoma cells. Proteomic analysis revealed that the protein bands reactive to K94 included cytokeratin (CK) 8 and 18, which are known to be related to tumorigenesis and form a heterotypic complex with each other. However, K94 showed no activity toward CK8 or CK18 separately. The epitope of the K94 antibody was only presented by a complex between CK8 and CK18, which was confirmed by analysis using recombinant CK8 and CK18 proteins. To formulate an assay for anti-CK8/18 complex autoantibody, a mimotope peptide reactive to K94 was selected from loop-constrained heptapeptide (-CX7C-) display phage library, of which sequence was CISPDAHSC (K94p1). A mimotope enzyme-linked immunosorbent assay (ELISA) using phage-displayed K94p1 peptide as a coating antigen was able to discriminate breast cancer (n=30) patients from normal subjects (n=30) with a sensitivity of 50% and a specificity of 82.61%. CA15.3 was detected at very low levels in the same breast cancer subjects and did not discriminate breast cancer patients from normal subjects, although it is a conventional biomarker of breast cancer. These results suggest that a mimotope ELISA composed of K94p1 peptide may be useful for the diagnosis of breast cancer.
Collapse
Affiliation(s)
- Chang-Kyu Heo
- Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea
| | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
32
|
Figueiredo ML, Stein TJ, Jochem A, Sandgren EP. Mutant Hras(G12V) and Kras(G12D) have overlapping, but non-identical effects on hepatocyte growth and transformation frequency in transgenic mice. Liver Int 2012; 32:582-91. [PMID: 22221894 PMCID: PMC4319543 DOI: 10.1111/j.1478-3231.2011.02732.x] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/04/2011] [Accepted: 11/28/2011] [Indexed: 12/24/2022]
Abstract
BACKGROUND Mouse hepatocarcinogenesis is associated with mutations in Hras, but infrequently in Kras. The effect on carcinogenesis of developmental age at the time of ras mutation remains unknown. AIM We sought to compare quantitatively the effects of expressing mutant H- or Kras genes in fetal vs. adult mouse liver. METHODS We established an inducible system of gene expression in mouse liver to define disease pathogenesis associated with activation of oncogene expression. RESULTS Diffuse expression of either oncogene in fetal or adult hepatocytes caused hepatomegaly. For mutant Hras(G12V), this phenotype was almost fully reversible and accompanied by apoptosis, indicating that maintenance of hepatomegaly requires continuous Hras(G12V) expression. We also examined the effect of ras expression on growth of transplanted hepatocytes in an in vivo system that allows us to quantify hepatocyte growth effects in both permissive and restrictive hepatic growth environments. Mutant Kras(G12D) had no effect on hepatocyte growth in this system. In contrast, Hras(G12V) induced increased hepatocyte focus growth in quiescent liver, the hallmark of a cell autonomous growth stimulus. Hras(G12V) also increased the fraction of donor hepatocyte foci that displayed extreme growth, a characteristic of preneoplastic lesions. CONCLUSIONS The primary effect of diffuse, whole-liver expression of either mutant ras gene in fetal or adult mouse liver is diffuse and progressive hepatic growth. Hras(G12V) mutation influences hepatocarcinogenesis by conferring cell autonomous growth potential upon foci of expressing cells and by increasing the risk of neoplastic progression. Kras(G12D) does not share these latter carcinogenic effects in mouse liver.
Collapse
Affiliation(s)
- Marxa L. Figueiredo
- Department of Pathobiological Sciences; School of Veterinary Medicine; University of Wisconsin-Madison; Madison; WI; USA
| | - Timothy J. Stein
- Department of Pathobiological Sciences; School of Veterinary Medicine; University of Wisconsin-Madison; Madison; WI; USA
| | - Adam Jochem
- Department of Pathobiological Sciences; School of Veterinary Medicine; University of Wisconsin-Madison; Madison; WI; USA
| | - Eric P. Sandgren
- Department of Pathobiological Sciences; School of Veterinary Medicine; University of Wisconsin-Madison; Madison; WI; USA
| |
Collapse
|
|
33
|
Kim JH, Minai-Tehrani A, Kim YK, Shin JY, Hong SH, Kim HJ, Lee HD, Chang SH, Yu KN, Bang YB, Cho CS, Yoon TJ, Yu DY, Jiang HL, Cho MH. Suppression of tumor growth in H-ras12V liver cancer mice by delivery of programmed cell death protein 4 using galactosylated poly(ethylene glycol)-chitosan-graft-spermine. Biomaterials 2011; 33:1894-902. [PMID: 22153867 DOI: 10.1016/j.biomaterials.2011.11.024] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2011] [Accepted: 11/13/2011] [Indexed: 02/04/2023]
Abstract
Non-viral gene delivery systems based on polyethyleneimine (PEI) are efficient due to their proton-sponge effect within endosomes, but they have poor physical characteristics such as slow dissociation, cytotoxicity, and non targeted gene delivery. To overcome many of the problems associated with PEI, we synthesized a galactosylated poly(ethylene glycol)-chitosan-graft-spermine (GPCS) copolymer with low cytotoxicity and optimal gene delivery to hepatocytes using an amide bond between galactosylated poly(ethylene glycol) and chitosan-graft-spermine. The GPCS copolymer formed complexes with plasmid DNA, and the GPCS/DNA complexes had well-formed spherical shapes. The GPCS/DNA complexes were nanoscale size with homogenous size distribution and a positive zeta potential by dynamic light scattering (DLS). The GPCS copolymer had lower cytotoxicity than that of PEI 25K in HepG2, HeLa, and A549 cell lines at various concentrations and showed good hepatocyte-targeting ability. Furthermore, GPCS/DNA complexes showed higher levels of GFP expression in the liver in model mice after intravenous injection than naked DNA and metoxy-poly(ethylene glycol)-chitosan-graft-spermine as controls without remarkable fibrosis, inflammation, lipidosis, or necrosis. In a tumor suppression study, an intravenous injection of the GPCS/Pdcd4 complexes significantly suppressed tumor growth, activated apoptosis, and suppressed proliferation and angiogenesis in liver tumor-bearing H-ras12V mice. Our results indicate that the GPCS copolymer has potential as a hepatocyte-targeting gene carrier.
Collapse
Affiliation(s)
- Ji-Hye Kim
- Laboratory of Toxicology, College of Veterinary Medicine, Seoul National University, Seoul 151-742, Republic of Korea
| | | | | | | | | | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
34
|
Steatosis induced by the accumulation of apolipoprotein A-I and elevated ROS levels in H-ras12V transgenic mice contributes to hepatic lesions. Biochem Biophys Res Commun 2011; 409:532-8. [PMID: 21600874 DOI: 10.1016/j.bbrc.2011.05.039] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2011] [Accepted: 05/06/2011] [Indexed: 11/22/2022]
Abstract
Hepatic steatosis is considered to have an important impact on liver tumorigenesis, despite a lack of clear experimental evidence. Histopathological analysis of H-ras12V transgenic mice showed liver lesions on a steatosis background had significantly higher incidence than on a non-steatosis background. Further investigation showed that apolipoprotein A-I was elevated and accumulated around fatty vacuoles. This elevated level of apolipoprotein A-I was coupled with an elevated level of H-ras12V protein and ROS. In conclusion, our results suggest that the expression of H-ras12V oncogene leads to elevated levels of ROS and apolipoprotein A-I that contribute to steatosis. The steatosis, in turn, promotes the development of hepatic lesions induced by H-ras12V oncogene.
Collapse
|
|
35
|
Sugamori KS, Brenneman D, Grant DM. Liver-selective expression of human arylamine N-acetyltransferase NAT2 in transgenic mice. Drug Metab Dispos 2011; 39:882-90. [PMID: 21317369 DOI: 10.1124/dmd.111.038216] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022] Open
Abstract
Human arylamine N-acetyltransferase 2 (NAT2) mediates the biotransformation of arylamine drugs and procarcinogens into either innocuous or reactive DNA-damaging metabolites and is expressed predominantly in liver. Interspecies differences and incongruous results between in vitro, in vivo, and epidemiological studies make it difficult to extrapolate animal results to human risk. We have generated human NAT2 transgenic mice on both C57BL/6 (hNAT2(tg)) and Nat1/2 null backgrounds [hNAT2(tg)Nat1/2(-/-)], in which liver-selective expression of human NAT2 is driven by the mouse albumin promoter. We detected expression of the human NAT2 transcript and protein in mouse liver by real-time PCR and Western blot analysis. NAT2 enzyme activity, measured using the human NAT2-selective substrate sulfamethazine (SMZ), was 40- to 80-fold higher in liver cytosols from hNAT2(tg)Nat1/2(-/-) mice than in wild-type mice. An unexpected gender difference was observed, with males displaying 2-fold higher activity than females. Transgenic mice also had an increased in vivo plasma clearance of SMZ and higher levels of N-acetylated SMZ than wild-type mice. Liver expression of human NAT2 did not affect the disposition of the human NAT1-selective substrate p-aminosalicylic acid (PAS), because hNAT2(tg)Nat1/2(-/-) mice displayed in vivo PAS pharmacokinetic profiles similar to those of Nat1/2(-/-) mice. The metabolism of 4-aminobiphenyl was similar between hNAT2(tg)Nat1/2(-/-) and wild-type mice with the exception of a more liver-restricted pattern in hNAT2(tg)Nat1/2(-/-) mice and lower activity in females. Overall, the hNAT2(tg)Nat1/2(-/-) mouse mimics human expression of NAT2 and may thus be of value in clarifying the role of human NAT2 in arylamine clearance, detoxification, and bioactivation.
Collapse
Affiliation(s)
- Kim S Sugamori
- Department of Pharmacology and Toxicology, University of Toronto, Toronto, Canada
| | | | | |
Collapse
|
|
36
|
Abstract
Several studies have shown physiological functions of interleukin (IL)-32, a novel cytokine. However, the role of IL-32 in cancer development has not been reported. In this study, we showed that IL-32γ inhibited tumor growth in IL-32γ-overexpressing transgenic mice inoculated with melanoma as well as colon tumor growth in xenograft nude mice inoculated with IL-32γ-transfected colon cancer cells (SW620). The inhibitory effect of IL-32γ on tumor growth was associated with the inhibition of constitutive activated nuclear transcription factor-κB (NF-κB) and of signal transducer and activator of transcription 3 (STAT3). The expression of antiapoptotic, cell proliferation and tumor-promoting genes (bcl-2, X-chromosome inhibitor of apoptosis protein (IAP), cellular IAP and cellular FADD-like IL-1β-converting enzyme-inhibitory protein, cyclin D), cyclin-dependent kinase 4, cycolooxygenase-2 and inducible nitric oxide synthase was decreased, whereas the expression of apoptotic target genes (caspase-3 and -9, bax) increased. In tumor, spleen and blood, the number of cytotoxic CD8+ T cells and CD57+ natural killer cells and the levels of IL-10 increased, but that of tumor necrosis factor-α (TNF-α), IL-1β and IL-6 decreased. We also found that forced overexpression of IL-32γ inhibited colon cancer cell (SW620 and HCT116) growth accompanied with the inhibition of activated NF-κB and STAT3 in vitro. In addition, when IL-32γ was knocked down by small interfering RNA (siRNA) or neutralized with an anti-IL-32γ antibody, IL-32γ-induced colon cancer cell growth inhibition, the IL-32γ-induced decrease of TNF-α, IL-1 and IL-6 production, and the increase of IL-10 production were abolished. However, siRNA of NF-κB and STAT3 augmented IL-32γ-induced colon cancer cell growth inhibition. These findings indicate significant pathophysiological roles of IL-32γ in cancer development.
Collapse
|
|
37
|
Bang DH, Park SH, Jun HY, Moon HB, Kim SU, Yu DY, Yoon KH. Gd-EOB-DTPA enhanced micro-MR imaging of hepatic tumors in H-ras 12V transgenic mice. Acad Radiol 2011; 18:13-9. [PMID: 20926317 DOI: 10.1016/j.acra.2010.08.007] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/02/2010] [Revised: 08/13/2010] [Accepted: 08/13/2010] [Indexed: 01/11/2023]
Abstract
RATIONALE AND OBJECTIVES The aims of this study were to evaluate the morphologic characteristics and growth pattern of hepatic tumors in H-ras 12V transgenic (TG) mice using a micro-magnetic resonance (MR) system and to assess the usefulness of gadolinium ethoxybenzyl diethylenetriamine penta-acetic acid (Gd-EOB-DTPA) enhancement for the detection of hepatic tumors in these mice. MATERIALS AND METHODS Hepatocellular carcinoma lines were established to allow insertion of the H-ras 12V transgene under the control of the albumin enhancer/promoter. Seven H-ras 12V TG mice and four wild-type mice were included in this study. The mice underwent various MR imaging examinations, including T1-weighted imaging (repetition time, 300 ms; echo time, 11 ms), Gd-EOB-DTPA-enhanced T1-weighted imaging (dose, 0.025 mmol/kg), and T2-weighted imaging (repetition time, 3500 ms; echo time, 36 ms), with a 4.7-T MR scanner, at 4, 6, 8, and 9 months of age. All mice were euthanized after the final MR imaging procedure, except for one TG mouse and two wild-type mice that were euthanized after MR imaging procedures at 4 months of age. For imaging analysis, the tumor characteristics in each MR sequence, including tumor size, number, and signal intensity (SI), were recorded, and the contrast-to-noise ratio and contrast enhancement ratio were calculated to quantify the SI of the tumor. The MR images were correlated with the findings of histopathologic examinations. RESULTS No tumors were detected in the four wild-type mice. In the six TG mice, a total of 67 tumors were found in histopathologic specimens obtained at 9 months of age. Of the 67 tumors, 62 were detected on Gd-EOB-DTPA-enhanced T1-weighted images with fat saturation. The majority of hepatic tumors showed high SI on T1-weighted images without fat saturation. The SI diminished on T1-weighted images with fat saturation. The tumor contrast-to-noise ratio for Gd-EOB-DTPA-enhanced T1-weighted imaging was significantly better than that for the other sequences. The tumors were histopathologically confirmed as hepatocellular adenomas (n = 32) and well-differentiated hepatocellular carcinomas (n = 35). CONCLUSIONS Micro-MR imaging can reveal the characteristics of hepatic tumors in a live murine model. Gd-EOB-DTPA-enhanced T1-weighted imaging is helpful in the detection of hepatic tumors in H-ras 12V TG mice.
Collapse
Affiliation(s)
- Dong-Ho Bang
- Department of Radiology, Institute for Radiological Imaging Science, Wonkwang University School of Medicine, Iksan, Jeonbuk, Korea
| | | | | | | | | | | | | |
Collapse
|
|
38
|
Ha HL, Yu DY. HBx-induced reactive oxygen species activates hepatocellular carcinogenesis via dysregulation of PTEN/Akt pathway. World J Gastroenterol 2010; 16:4932-7. [PMID: 20954279 PMCID: PMC2957601 DOI: 10.3748/wjg.v16.i39.4932] [Citation(s) in RCA: 52] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the role of hepatitis B virus X-protein (HBx)-induced reactive oxygen species (ROS) on liver carcinogenesis in HBx transgenic mice and HepG2-HBx cells.
METHODS: Cell growth rate was analyzed, and through western blotting, mitogenic signaling was observed. Endogenous ROS from wild and HBx transgenic mice and HepG2-Mock and HBx cells were assayed by FACScalibur. Identification of oxidized and reduced phosphatase and tensin homolog (PTEN) was analyzed through N-ethylmaleimide alkylation, nonreducing electrophoresis.
RESULTS: We observed that the cell-proliferation-related phosphoinositide 3-kinase/Akt pathway is activated by HBx in vivo and in vitro. Increased ROS were detected by HBx. Tumor suppressor PTEN, via dephosphorylation of Akt, was oxidized and inactivated by increased ROS. Increased oxidized PTEN activated the mitogenic pathway through over-activated Akt. However, treatment with ROS scavenger N-acetyl cysteine can reverse PTEN to a reduced form. Endogenously produced ROS also stimulated HBx expression.
CONCLUSION: HBx induced ROS promoted Akt pathways via oxidized inactive PTEN. HBx and ROS maintained a positive regulatory loop, which aggravated carcinogenesis.
Collapse
|
|
39
|
Lee HS, Lee SY, Ha HL, Han DC, Han JM, Jeong TS, Yu DY, Yeom YI, Kwon BM, Moon EY. 2'-Benzoyloxycinnamaldehyde inhibits tumor growth in H-ras12V transgenic mice via downregulation of metallothionein. Nutr Cancer 2010; 61:723-34. [PMID: 19838947 DOI: 10.1080/01635580902825613] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/20/2022]
Abstract
Cinnamaldehydes have been reported to induce apoptosis in human carcinomas through the generation of reactive oxygen species (ROS). 2'-benzoyloxycinnamaldehyde (BCA) has been reported to inhibit tumor formation in H-ras12V transgenic mice. To see the antitumor effects of BCA, BCA was administrated intraperitoneally (50 mg/kg) to H-ras12V transgenic mice for 3 wk, and it was found that the hepatic tumor volume and the total number of tumors were decreased in BCA-treated mice as compared to control H-ras12V transgenic mice. To identify possible target genes responsible for BCA antitumor effects in H-ras12V transgenic mice, cDNA microarray analyses were performed comparing gene expression between BCA treated and control transgenic mice. We found that 42 genes were downregulated, and 40 genes were upregulated in the BCA-treated transgenic mice. The downregulated genes included several genes involved in ROS regulation and immune response (aconitase, metallothionein-1, metallothionein-2, and purine nucleoside phosphorylase). The expression of ROS-related genes, metallothionein 1 and metallothionein 2, was decreased more than twofold with BCA treatment (P < 0.001). It was confirmed by RT-PCR and immunohistochemical analyses. The inhibition of tumor formation and growth in H-ras12V transgenic mice by BCA was mediated through inhibition of the expression of the ROS scavengers metallothionein 1 and metallothionein 2.
Collapse
Affiliation(s)
- Heun-Sik Lee
- Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea
| | | | | | | | | | | | | | | | | | | |
Collapse
|
|
40
|
Wang AG, Lee DS, Moon HB, Kim JM, Cho KH, Choi SH, Ha HL, Han YH, Kim DG, Hwang SB, Yu DY. Non-structural 5A protein of hepatitis C virus induces a range of liver pathology in transgenic mice. J Pathol 2009; 219:253-62. [PMID: 19621337 DOI: 10.1002/path.2592] [Citation(s) in RCA: 50] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
Hepatitis C virus (HCV) is a major cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma (HCC). However, the mechanism of HCV pathogenesis is not well understood. Our previous in vitro studies suggested that non-structural 5A (NS5A) protein may play an important role in liver pathogenesis. To elucidate the mechanism of HCV-induced liver pathogenesis, we investigated the histopathological changes of liver in transgenic mice harbouring the NS5A gene. We generated transgenic mice harbouring HCV NS5A gene under the control of hepatitis B virus (HBV) enhancer. Pathological changes were analysed by immunohistochemical staining and western blot analysis. Lipid composition and reactive oxygen species (ROS) production in NS5A transgenic mice were analysed. HCV NS5A transgenic mice developed extraordinary steatosis over 6 months old and induced HCC in some mice. NS5A was co-localized with apolipoprotein A-I in fatty hepatocytes. In addition, the extraordinarily high levels of ROS, NF-kappaB and STAT3 were detected in hepatocytes of NS5A transgenic mice. These data suggest that NS5A, independent of other HCV viral proteins, may play an important role in the development of hepatic pathologies, including steatosis and hepatoceullular carcinoma in transgenic mice.
Collapse
Affiliation(s)
- Ai-Guo Wang
- Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea
| | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
41
|
Extracellular signal-regulated kinase 2-dependent phosphorylation induces cytoplasmic localization and degradation of p21Cip1. Mol Cell Biol 2009; 29:3379-89. [PMID: 19364816 DOI: 10.1128/mcb.01758-08] [Citation(s) in RCA: 66] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023] Open
Abstract
p21(Cip1) is an inhibitor of cell cycle progression that promotes G(1)-phase arrest by direct binding to cyclin-dependent kinase and proliferating cell nuclear antigen. Here we demonstrate that mitogenic stimuli, such as epidermal growth factor treatment and oncogenic Ras transformation, induce p21(Cip1) downregulation at the posttranslational level. This downregulation requires the sustained activation of extracellular signal-regulated kinase 2 (ERK2), which directly interacts with and phosphorylates p21(Cip1), promoting p21(Cip1) nucleocytoplasmic translocation and ubiquitin-dependent degradation, thereby resulting in cell cycle progression. ERK1 is not likely involved in this process. Phosphopeptide analysis of in vitro ERK2-phosphorylated p21(Cip1) revealed two phosphorylation sites, Thr57 and Ser130. Double mutation of these sites abolished ERK2-mediated p21(Cip1) translocation and degradation, thereby impairing ERK2-dependent cell cycle progression at the G(1)/S transition. These results indicate that ERK2 activation transduces mitogenic signals, at least in part, by downregulating the cell cycle inhibitory protein p21(Cip1).
Collapse
|
|
42
|
Park JA, Lee JJ, Jung JC, Yu DY, Oh C, Ha S, Kim TJ, Chang Y. Gd-DOTA Conjugate of RGD as a Potential Tumor-Targeting MRI Contrast Agent. Chembiochem 2008; 9:2811-3. [DOI: 10.1002/cbic.200800529] [Citation(s) in RCA: 62] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
|
|
43
|
Zender L, Kubicka S. Androgen receptor and hepatocarcinogenesis: what do we learn from HCC mouse models? Gastroenterology 2008; 135:738-40. [PMID: 18692055 DOI: 10.1053/j.gastro.2008.07.034] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
|
|
44
|
Wang AG, Fang W, Han YH, Cho SM, Choi JY, Lee KH, Kim WH, Kim JM, Park MG, Yu DY, Kim NS, Lee DS. Expression of the RERG gene is gender-dependent in hepatocellular carcinoma and regulated by histone deacetyltransferases. J Korean Med Sci 2006; 21:891-6. [PMID: 17043425 PMCID: PMC2722001 DOI: 10.3346/jkms.2006.21.5.891] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/04/2023] Open
Abstract
Ras-related, estrogen-regulated, and growth-inhibitory gene (RERG) is a novel gene that was first reported in breast cancer. However, the functions of RERG are largely unknown in other tumor types. In this study, RERG expression was analyzed in hepatocellular carcinomas of human patients using reverse transcriptase PCR analysis. In addition, the possible regulation of RERG expression by histone deacetyltransferases (HDACs) was studied in several cell lines. Interestingly, the expression of RERG gene was increased in hepatocellular carcinoma (HCC) of male patients (57.9%) but decreased in HCC of females (87.5%) comparison with paired peri-tumoral tissues. Moreover, RERG gene expression was increased in murine hepatoma Hepa1-6 cells, human breast tumor MDA-MB-231 cells, and mouse normal fibroblast NIH3T3 cells after treated by HDAC inhibitor, trichostatin A. Our results suggest that RERG may function in a gender-dependent manner in hepatic tumorigenesis and that the expression of this gene may be regulated by an HDAC-related signaling pathway.
Collapse
Affiliation(s)
- Ai-Guo Wang
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
| | - Wan Fang
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
| | - Ying-Hao Han
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
| | - Sang-Mi Cho
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
| | - Jong Young Choi
- Department of Internal Medicine, Kangnam St. Mary's Hospital, The Catholic University of Korea, Seoul, Korea
| | - Kee Ho Lee
- Laboratory of Molecular Oncology, Korea Institute of Radiological and Medical Sciences, Seoul, Korea
| | - Wook Hwan Kim
- Department of Surgery, School of Medicine Ajou University, Suwon, Korea
| | - Jin Man Kim
- Department of Pathology, College of Medicine, Chungnam National University, Daejeon, Korea
| | - Moon Gi Park
- Department of Surgery, SUN General Hospital, Daejeon, Korea
| | - Dae-Yeul Yu
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
| | - Nam-Soon Kim
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
| | - Dong-Seok Lee
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon, Korea
- Animal Resources Science, Kangwon National University, Chuncheon, Korea
| |
Collapse
|
|
45
|
Wang AG, Moon HB, Chun SY, Lee TH, Yu DY, Lee DS. Orchiectomy reduces hepatotumorigenesis of H-ras12V transgenic mice via the MAPK pathway. Life Sci 2006; 79:1974-80. [PMID: 16846616 DOI: 10.1016/j.lfs.2006.06.032] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2006] [Revised: 04/22/2006] [Accepted: 06/20/2006] [Indexed: 10/24/2022]
Abstract
A common characteristic of hepatocellular carcinoma in humans and animals is a striking male prevalence. However, the underlying mechanisms remain largely unknown. We have reported that hepatotumorigenesis is prevalent in male H-ras12V transgenic mice (Wang et al., 2005). Orchiectomy and ovariectomy were performed to determine if the presence of sex organ-related factors contributes to hepatotumorigenesis. After orchiectomy and ovariectomy, the mice were sampled at 6 months of age. The pathological diagnosis, sex hormone levels, expression of the H-ras12V transgene, and ras related signal pathways were determined. Orchiectomy significantly reduced the incidence of hepatotumorigenesis in males. However, no significant difference was detected in the incidence of tumorigenesis between ovariectomized and non-ovariectomized females. Molecular biochemical experiments showed that the sex organ-related factors significantly influenced transgene expression, which contributed to activation of the MAPK signaling pathway. The present study demonstrates that testis-related factors play important roles in hepatotumorigenesis in H-ras12V transgenic mice.
Collapse
Affiliation(s)
- Ai-Guo Wang
- Laboratory of Human Genomics, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 305-806, South Korea
| | | | | | | | | | | |
Collapse
|
|
46
|
Moon EY, Lee MR, Wang AG, Lee JH, Kim HC, Kim HM, Kim JM, Kwon BM, Yu DY. Delayed occurrence of H-ras12V-induced hepatocellular carcinoma with long-term treatment with cinnamaldehydes. Eur J Pharmacol 2006; 530:270-5. [PMID: 16405947 DOI: 10.1016/j.ejphar.2005.11.053] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2005] [Accepted: 11/15/2005] [Indexed: 10/25/2022]
Abstract
Cinnamaldehyde from the bark of Cinnamomum cassia has been reported to have antitumor activity mediated by the inhibition of farnesyl transferase. We assessed in vivo the chemo-preventive effect of cinnamaldehydes on H-ras12V-induced hepatocellular carcinoma formation. A mouse model of hepatocellular carcinoma was established by using the transgene of mutated H-ras12V under the regulation of albumin enhancer/promoter. When treated with cinnamaldehyde for 10 weeks, hepatic tumor development was delayed with 2'-benzoyloxycinnamaldehyde (BCA) compared with control hepatocellular carcinoma formation. The effect of 2'-hydroxycinnamaldehyde (HCA) was comparable. The number of lesions and the size of each lesion were significantly reduced by BCA. Cell proliferation in the lesion was detected by incorporation of 5-bromo-2'-deoxyuridine (BrdU). BCA increased the number of splenocytes, concanavalin A-stimulated splenocyte proliferation and the infiltration of lymphocytes into liver. Data suggest that the delayed hepatic tumor development observed with BCA could be mediated by a long-term immunostimulating effect on T cells.
Collapse
MESH Headings
- Acrolein/analogs & derivatives
- Acrolein/pharmacology
- Animals
- Antineoplastic Agents/pharmacology
- Antineoplastic Agents, Phytogenic/pharmacology
- Benzoates/pharmacology
- Carcinoma, Hepatocellular/drug therapy
- Carcinoma, Hepatocellular/genetics
- Carcinoma, Hepatocellular/pathology
- Cell Proliferation/drug effects
- Female
- Genes, ras/genetics
- Hepatocytes/drug effects
- Hepatocytes/metabolism
- Liver Neoplasms, Experimental/drug therapy
- Liver Neoplasms, Experimental/genetics
- Liver Neoplasms, Experimental/pathology
- Male
- Mice
- Mice, Inbred C57BL
- Mice, Transgenic
- NIH 3T3 Cells
- Spleen/cytology
- Tumor Burden/drug effects
Collapse
Affiliation(s)
- Eun-Yi Moon
- Department of Human Genomics, Korea Research Institute of Bioscience and Biotechnology, 52 Oun-Dong, Yusong-Gu, Taejeon 305-806, Korea.
| | | | | | | | | | | | | | | | | |
Collapse
|