Brief Article Open Access
Copyright ©2012 Baishideng Publishing Group Co., Limited. All rights reserved.
World J Gastroenterol. Mar 21, 2012; 18(11): 1249-1256
Published online Mar 21, 2012. doi: 10.3748/wjg.v18.i11.1249
CCL7 and CCL21 overexpression in gastric cancer is associated with lymph node metastasis and poor prognosis
Tsann-Long Hwang, Li-Yu Lee, Chee-Chan Wang, Ying Liang, Shu-Fang Huang, Chi-Ming Wu
Tsann-Long Hwang, Shu-Fang Huang, Department of Surgery, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan
Tsann-Long Hwang, Department of Medicine, Chang Gung University, Taoyuan 333, Taiwan
Li-Yu Lee, Department of Pathology, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan
Chee-Chan Wang, Chi-Ming Wu, Department of Cosmetic Science, Vanung University, Chungli 320, Taiwan
Ying Liang, Molecular Medicine Research Center, Chang Gung University, Taoyuan 333, Taiwan
Author contributions: Hwang TL and Wu CM designed the research; Lee LY, Wang CC, Liang Y and Huang SF performed the research and analyzed the data; and Wu CM wrote the paper.
Supported by Grants NSC 98-2314-B-238-001 from the National Science Council; and VIT-98-CM-01 from Vanung University, Taiwan
Correspondence to: Chi-Ming Wu, Associate Professor, Department of Cosmetic Science, Vanung University, No.1 Van-Nung Rd, Chungli 320, Taiwan. chimingwu@mail.vnu.edu.tw
Telephone: +886-3-4515811 Fax: +886-3-4345846
Received: April 6, 2011
Revised: June 23, 2011
Accepted: June 30, 2011
Published online: March 21, 2012

Abstract

AIM: To investigate how a complex network of CC chemokine ligands (CCLs) and their receptors influence the progression of tumor and metastasis.

METHODS: In the present study, we used immunohistochemistry to examine the expression of CCL7, CCL8 and CCL21 in 194 gastric cancer samples and adjacent normal tissues. We analyzed their correlation with tumor metastasis, clinicopathologic parameters and clinical outcome.

RESULTS: We found that the higher expression of CCL7 and CCL21 in cancer tissues than in normal tissues was significantly correlated with advanced depth of wall invasion, lymph node metastasis and higher tumor node metastasis stage. Moreover, Kaplan-Meier survival analysis revealed that CCL7 and CCL21 overexpression in cancer tissues was correlated with poor prognosis.

CONCLUSION: These results suggest that overexpression of these two CC chemokine ligands is associated with tumor metastasis and serves as a prognostic factor in patients with gastric cancer.

Key Words: CC chemokine, Chemokine ligand 7, Chemokine ligand 21, Gastric cancer, Lymph node metastasis, Poor prognosis



INTRODUCTION

Chemokine ligands (CCLs) belong to the small molecule chemoattractive cytokine family and are grouped into CC and CXC chemokines ligands on the basis of the characteristic presence of four conserved cysteine residues[1-3]. Chemokines mediate their chemical effect on target cells through G-protein-coupled receptors, which are characterized structurally by seven transmembrane spanning domains and are involved in the attraction and activation of mononuclear and polymorphonuclear leukocytes. CCLs and their receptors play an important role in angiogenesis and tumor growth, however the role of CCLs in metastasis has only recently been explored[4,5]. CCL7 promoted the invasion and migration of oral squamous cell carcinoma[4]. CCL21 was significantly highly expressed in breast tumor cells with lymph node metastasis and prognosis[5].

Gastric cancer is one of the commonest malignant tumors of the alimentary tract and is characterized by late clinical presentation, rapid progression, and poor survival[6]. The reason for this poor prognosis is that, at the time of diagnosis, gastric cancer usually shows extensive local tumor invasion and frequent spread to metastatic sites, particularly lymph nodes. Spread of malignant tumors is a multistep process and many of the stages of tumor invasion require degradation or breakdown of the extracellular matrix and connective tissue surrounding tumor cells[7,8]. The matrix metalloproteinases (MMPs) are a family of zinc containing enzymes which are involved in the degradation of different components of the extracellular matrix, and there is considerable evidence to indicate that individual MMPs have important roles in tumor invasion and tumor spread[9-11]. A recent study showed that increased levels of CCL recruit immature myeloid cells that carry the chemokine ligand receptor (CCR) from the blood to the tumor invasion front. These immature myeloid cells produced MMP9 and MMP2 and help the tumor cells to migrate and invade[12].

In the present study, we used immunohistochemistry to examine the expression of CCL7, CCL8 and CCL21 in 194 gastric cancer samples and adjacent normal tissues. We analyzed their correlation with tumor metastasis, clinicopathologic parameters and clinical outcome.

MATERIALS AND METHODS
Patients and specimens

A consecutive series of 194 tissue specimens were collected from patients with gastric cancer who received subtotal or total gastroectomy resection in Chang Gung Memorial Hospital (CGMH) in Taiwan. All operations were performed between January 2001 and December 2002. Written informed consent was obtained before sample collection and this study was approved by the Institutional Review Board of CGMH. There were 114 males and 80 females with a mean age of 62 years (range, 24-90 years). The age and gender of patients, tumor location, tumor size, cell differentiation, depth of wall invasion, status of lymph node metastasis, vascular invasion, lymphatic invasion and desmoplastic reaction were obtained from histopathology records. Stage of gastric cancer was described according to the 1997 tumor node metastasis (TNM) classification of malignant tumors by the American Joint Committee on Cancer. All patients were followed until December 2007 with a minimum 5 years of follow-up. All tissue specimens were formalin-fixed and paraffin-embedded. Formalin fixed tissue sections were stained with haematoxylin and eosin and classified by a pathologist. These results were compared with the histopathology records from CGMH. Final pathology was determined by consensus and review if necessary.

Immunohistochemistry

The tissue blocks were constructed according to the me-thod of Schraml et al[13] and the best representative morphological areas of tumors were used in this study. The specimen sections were deparaffinized, treated with 3% hydrogen peroxide and microwaved after pretreatment in 10 mmol/L citric acid to retrieve antigenicity. The sections were incubated with blocking solution containing phosphate buffered saline and 1% bovine serum albumin for 20 min at room temperature, and then incubated overnight at 4 °C with an anti-CCL7 antibody (1:100, R and D), an anti-CCL8 monoclonal antibody (1:50, R and D), or an anti-CCL21 monoclonal antibody (1:50, R and D), respectively. After washing 4 times with Tris Buffered Saline, the sections were incubated with biotinylated secondary antibody (Santa Cruz Biotechnology). The immuno-complex was visualized by the immonoglobulin enzyme bridge technique using the DAKO LSAB 2 System, HRP kit (DAKO corp. Carpinteria, CA) with 3,3’ diaminobenzidine tetrachloride as a substrate. The sections were counterstained with hematoxylin, dehydrated with graded alcohols, cleared with xylene and mounted with a coverslip.

Scoring of the immunohistochemical staining

The immunostaining results were scored as follows, according to a previous report[14]. The immunostaining reaction was evaluated by subjective assessments of the median staining intensity (0, no stain; 1, weak; 2, moderate; and 3, strong stain) and by the fraction of stained cells in percentage categories (0, 0%-9%; 1, 10%-49%; 2, 50%-89%; and 3, ≥ 90%). This scoring system was previously shown to be reproducible[15]. The scores of 0 to 3 were obtained as follows: percentage categories and staining were each ranked as indicated above. The ranks for percentage and staining intensity were multiplied by each other, divided by 3, and rounded up to the nearest whole number[15]. The results of immunostaining in tumor and normal tissues were divided into three groups, higher (rank of tumor tissue > rank of normal tissue), equal (rank of tumor tissue = rank of normal tissue), and lower (rank of tumor tissue < rank of normal tissue) (Figures 1-3).

Figure 1
Figure 1 Immunohistochemistry of chemokine ligand 7 in gastric cancer and adjacent normal tissues. Chemokine ligand 7 (CCL7) staining is higher in the cytoplasm of gastric cancer cells (A) than in the cytoplasm of adjacent normal cells (B); CCL7 staining is equal in the cytoplasm of gastric cancer cells (C) and in the cytoplasm of adjacent normal cells (D); CCL7 staining is lower in the cytoplasm of gastric cancer cells (E) than in the cytoplasm of adjacent normal cells (F). (magnification, × 400).
Figure 2
Figure 2 Immunohistochemistry of chemokine ligand 8 in gastric cancer and adjacent normal tissues. Chemokine ligand 8 (CCL8) staining is higher in the cytoplasm of gastric cancer cells (A) than in the cytoplasm of adjacent normal cells (B); CCL8 staining is equal in the cytoplasm of gastric cancer cells (C) and in the cytoplasm of adjacent normal cells (D); CCL8 staining is lower in the cytoplasm of gastric cancer cells (E) than in the cytoplasm of adjacent normal cells (F). (magnification, × 400).
Figure 3
Figure 3 Immunohistochemistry of chemokine ligand 21 in gastric cancer and adjacent normal tissues. Chemokine ligand 21 (CCL21) staining is higher in the cytoplasm of gastric cancer cells (A) than in the cytoplasm of adjacent normal cells (B); CCL21 staining is equal in the cytoplasm of gastric cancer cells (C) and in the cytoplasm of adjacent normal cells (D); CCL21 staining is lower in the cytoplasm of gastric cancer cells (E) than in the cytoplasm of adjacent normal cells (F). (magnification, × 400).
Statistical analysis

χ2 or Fisher’s exact test was used to test for an association between CCL7, CCL8 and CCL21 expression and patient clinicopathologic parameters. Disease-free survival was defined as the time from surgery to the first relapse of cancer, occurrence of a second primary tumor, or death from any cause. Univariate survival analysis was assessed by the Kaplan-Meier method and significance of difference between groups was analysed using log rank test or log rank test for trend. Stepwise multivariate survival analysis was performed by the Cox proportional hazards model. All reported P values were two-sided and a P value < 0.05 was considered significant.

RESULTS
CCL7, CCL8 and CCL21 expression in gastric cancer and adjacent normal tissues

The percentages of the higher expression of CCL7, CCL8 and CCL21 in cancer tissues than in normal tissues were 42.3% (82 of 194), 29.9% (58 of 194) and 44.8% (87 of 194), respectively (Figures 1-3 and Table 1). The percentages of the equal expression of CCL7, CCL8 and CCL21 in cancer tissues and in normal tissues were 35.6% (69 of 194), 33% (64 of 194) and 32.5% (63 of 194), respectively (Figures 1-3 and Table 1). The percentages of the lower expression of CCL7, CCL8 and CCL21 in cancer tissues than in normal tissues were 22.2% (43 of 194), 37.1% (72 of 194) and 22.7% (44 of 194), respectively (Figures 1-3 and Table 1).

Table 1 Association of chemokine ligand 7, chemokine ligand 8 and chemokine ligand 21 expression with the clinicopathologic parameters.
FactorsCasesCCL-7 expressionCCL-8 expressionCCL-21 expression
HigherEqualLowerP valueHigherEqualLowerP valueHigherEqualLowerP value
n = 82n = 69n = 43n = 58n = 64n = 72n = 87n = 63n = 44
Age (yr)
≤ 60833131210.4491728380.0263424250.101
> 60111513822413634533919
Gender
Male1145240220.4123640380.4294745220.041
Female80302921222434401822
Tumor location
Upper2210660.02546120.0048950.026
Middle4010171310624131116
Lower124584620424834604321
Whole8404242602
Tumor size (cm)
≤ 3952936300.0012428430.0683435260.043
> 399533313343629532818
Differentiation
Well186840.1775670.4227740.130
Moderate552722617231530187
Poor52251512191320271213
Signet ring cell69242421172230232620
Depth of wall invasion
T1471219160.001815240.07091820< 0.0001
T2378181113121291810
T3101582914353135642413
T49432261531
Lymph node metastasis
N0883036220.0202129380.4882833270.003
N147161615161714201710
N223137376101355
N336231031412102682
Vascular invasion
No1687060380.8005058600.5617157400.163
Yes26129586121664
Lymphatic invasion
No1043739280.0862734430.3253638300.006
Yes90453015313029512514
Desmoplastic reaction
None2861480.00656170.0717912< 0.0001
Mild64222022182125172621
Moderate754325723302244247
Marked2711106127819443
TNM stage
I641627210.0081420300.427132625< 0.0001
II4417171013141718188
III38201261314112486
IV482913618161432115
CCL7, CCL8 and CCL21 overexpression in relation to clinicopathologic parameters

The overexpression of CCL7 in cancer tissues compared with normal tissues was significantly correlated with tumor location (P = 0.025) and tumor size (P = 0.001). The overexpression of CCL7 was significantly higher in gastric cancer with advanced depth of wall invasion (P = 0.001), lymph node metastasis (P = 0.020), desmoplastic reaction (P = 0.006) and higher TNM stage (P = 0.008), but was not correlated with age, gender, differentiation, vascular invasion or lymphatic invasion (Table 1).

The overexpression of CCL8 was significantly correlated with age (P = 0.026) and tumor location (P = 0.004), but not with gender, tumor size, differentiation, depth of wall invasion, lymph node metastasis, vascular invasion, lymphatic invasion, desmoplastic reaction or TNM stage.

The overexpression of CCL21 was significantly higher in females than in males (P = 0.041) and was correlated with tumor location (P = 0.026), tumor size (P = 0.043) and lymphatic invasion (P = 0.006). As with CCL7, the overexpression of CCL21 was significantly higher in gastric cancer with an advanced depth of wall invasion (P < 0.0001), lymph node metastasis (P = 0.003), desmoplastic reaction (P < 0.0001) and higher TNM stage (P < 0.0001), but was not correlated with age, differentiation or vascular invasion (Table 1).

Prognostic implications of CCL7, CCL8 and CCL21 overexpression in gastric cancer

CCL7 and CCL21 overexpression was correlated with a poor prognosis (P = 0.002 and 0.001, Table 2 and Figure 4A and C). CCL8 overexpression was not correlated with survival (Table 2, Figure 4B). Other significant prognostic factors were tumor location, tumor size, differentiation, depth of invasion, lymph node metastases, vascular invasion, lymphatic invasion, marked desmoplastic reaction and higher TNM stage. In multivariate analysis, depth of invasion, lymph node metastasis and desmoplastic reaction were independent prognostic factors (Table 3).

Table 2 Univariate analysis of the clinicopathologic parameters influencing the disease-free survival of 194 gastric cancer patients undergoing gastrectomy.
FactorsCasesMean survival (mo)95% CI of mean5-year survival (%)P value
Age (yr)
≤ 608357.3350.22-64.4460.2 0.516
> 6011153.3446.93-59.7656.1
Gender
Male11457.9051.74-64.0562.2 0.174
Female8051.0943.61-58.5651.7
Type of gastrectomy
Total4131.8523.28-40.4333.5< 0.0001
Subtotal15360.2155.14-65.4864.5
Tumor location
Upper2226.0115.81-36.2127.3< 0.0001
Middle4069.2261.15-71.2977.0
Lower12456.5650.70-62.4359.5
Diffuse823.417.46-39.3625.0
Margin
Negative17357.7152.73-62.6862.30.0001
Positive2127.4117.26-37.5612.6
Tumor size (cm)
≤ 39570.9465.97-75.9280.7< 0.0001
> 39939.0232.23-45.8134.6
Differentiation
Well1865.5859.43-71.7394.4< 0.0001
Moderate5543.1135.16-51.0654.4
Poor5237.9130.28-45.5433.6
Signet ring cell6962.1654.89-69.4367.9
Depth of invasion
T14779.5375.46-83.6095.7< 0.0001
T23771.9264.56-79.2880.6
T310140.0433.53-46.5435.4
T4910.575.94-15.210.0
Lymph node metastasis
N08873.6469.17-78.1183.8< 0.0001
N14757.6747.75-67.5964.9
N22331.6121.57-41.6523.9
N33615.3910.83-19.950.0
Vascular invasion
No16860.3655.48-65.2365.4< 0.0001
Yes2617.7411.39-24.084.4
Lymphatic invasion
No10470.5865.84-75.3379.2< 0.0001
Yes9036.4829.43-43.5332.1
Perineural invasion
No10967.5762.27-72.8876.3< 0.0001
Yes8437.3630.70-44.0234.1
Desmoplastic reaction
None2867.5556.87-78.2478.6< 0.0001
Mild6470.2064.03-76.3778.7
Moderate7541.9934.35-49.6238.8
Marked2736.6724.49-48.8536.7
TNM stage
I6478.8675.66-82.0792.1< 0.0001
II4466.5558.17-74.9274.2
III3846.5236.06-56.9845.7
IV4814.7110.94-18.480.0
CCL-7
Higher8245.6237.97-53.2843.6 0.002
Equal6960.0952.62-67.5764.7
Lower4363.2854.24-72.3271.8
CCL-8
Higher5851.0542.41-59.7050.0 0.269
Equal6451.8943.39-60.3957.4
Lower7259.9752.60-67.3464.6
CCL-21
Higher8744.9637.83-52.1045.8 0.001
Equal6356.6048.30-64.9059.5
Lower4470.5863.24-77.9379.2
Table 3 Cox’s proportional hazards analysis.
FactorsHazard ratio95% CIP value
upper-lower
Depth of invasion
T2/T17.8501.454-42.3900.017
T3/T123.2004.733-113.7160.000
T4/T165.05210.830-390.730< 0.0001
Lymph node metastasis
N1/N01.8560.865-3.9820.112
N2/N03.5201.597-7.7580.002
N3/N07.2273.349-15.596< 0.0001
Desmoplastic reaction
Mild/none3.6631.272-10.6380.016
Moderate/none3.6231.304-10.1010.014
Marked/none4.9261.590-15.1520.006
CCL-70.801
CCL-80.620
CCL-210.084
Figure 4
Figure 4 Kaplan-Meier survival curves for disease-free survival of 194 patients with gastric cancer. A: Categorized by chemokine ligand 7 (CCL7) expression, survival was significantly worse for patients with higher CCL7 expression than those with equal or lower CCL7 expression (P = 0.002); B: Categorized by CCL8 expression, no significant difference was observed among the three groups (P = 0.269); C: Categorized by CCL21 expression, survival was significantly worse for patients with higher CCL21 expression than those with equal or lower CCL21 expression (P = 0.001).
DISCUSSION

In this study, CCL7, CCL8 and CCL21 expression levels were examined in 194 cases of gastric cancer for correlation with patient clinicopathologic factors. We found that the higher expression of CCL7 and CCL21 in cancer tissues than in normal tissues was significantly correlated with advanced depth of wall invasion, lymph node metastasis and higher TNM stage. The mechanism for chemokine ligand promotion of tumor invasion and metastasis is not clear. Using a model of colorectal tumor progression, Kitamura et al[12] showed that tumor-stromal interaction could promote tumor invasion. The colonic tumor can promote the production of CCL9. Increased levels of CCL9 recruited immature myeloid cells that carry the CCL9 receptor CCR1 from the blood to the tumor invasion front. The immature myeloid cells produce MMP2 and MMP9 and help the tumor epithelium to migrate and invade into the stroma. Jung et al[4] also showed the importance of tumor-stromal crosstalk in invasion of oral squamous cell carcinoma (OSCC) via CCL7[4]. To identify key molecular regulators expressed by carcinoma-associated fibroblasts (CAF) that promote cancer cell invasion, Jung et al[4] used microarrays to compare cocultured OSCC and CAF with monoculture controls. Microarray and real-time polymerase chain reaction analysis identified marked upregulation of CCL7 in cocultured CAF. Enzyme-linked immunosorbent assay showed an elevated level of CCL7 secretion from CAF stimulated by coculture with OSCC cells. CCL7 promoted the invasion and migration of OSCC cells, and the invasiveness was inhibited by treatment with CCL7 neutralizing antibody.

However, other studies have shown that CC chemokine ligands promote T cells to kill the tumor cells. Wu et al[16] investigated the effect of exogenous CCL21 expressed in breast cancer MCF-7 cells on human monocyte-derived dendritic cells (DCs). Stimulation of CCL21-transfected MCF-7 cells prompted DC function: migration, antigen uptake and presentation. The stimulated DCs facilitated Th 1 type cytokine production, perforin-forming CD8+ T cell transformation and final T cell-associated clearance of MCF-7 cells. Wetzel et al[17] showed that human CCL7 can reduce tumorigenicity and augment infiltration of dendritic cells and neutrophils toward mouse mastocytoma; it also inhibits mouse melanoma growth through activation of T lymphocytes and natural killer cells.

The differences between the expression of CCL7 and CCL21 correlated with clinicopathologic parameters were gender and lymphatic invasion. The overexpression of CCL7 in gastric cancer was not correlated with gender and lymphatic invasion, but that of CCL21 was correlated with these two parameters. The overexpression of CCL21 was significantly higher in females than in males. The reason for the significance is not clear and more studies are necessary to clarify the significance. The overexpression of CCL21 was also correlated with lymphatic invasion. Recently, metastatic gastric carcinoma cells have been shown to express the receptor for chemokine CCL21, chemokine receptors CCR7, a property that may allow them to access the lymphatic system and spread to regional lymph nodes[18]. Thus the “chemoattraction” theory of metastasis may be reflected by malignant cells expressing functional chemokine receptors that can respond to organ-specific chemoattractant molecules and migrate directionally along chemokine gradients to set up site-specific metastases in the target organs. Such chemotactic migration of tumors would mirror the physiologic mechanisms of lymphocyte homing into lymphoid organs.

Kaplan-Meier survival analysis revealed that CCL7 and CCL21 overexpression in cancer tissues was correlated with poor prognosis. If tumor-infiltrating leukocytes are able, in some instances, to promote cancer, then the local production of chemokines that attract leukocytes could be a poor prognostic sign. This is the case in human breast cancer, where levels of CCL5 and CCL2 correlate with tumor progression and there is a positive correlation between the extent of the macrophage infiltrate, lymph-node metastasis and clinical aggressiveness[19-21]. In esophageal squamous cell carcinoma, CCL2 expression has been associated with the extent of macrophage infiltration, tumor cell invasion and tumor vascularity[22].

In conclusion, the higher expression of CCL7 and CCL21 in gastric cancer tissues than in normal tissues was significantly correlated with advanced depth of wall invasion, lymph node metastasis and higher TNM stage. Moreover, Kaplan-Meier survival analysis revealed that CCL7 and CCL21 overexpression in cancer tissues was correlated with poor prognosis. These results suggest that overexpression of these two CC chemokine ligands is associated with tumor metastasis and serves as a prognostic factor in patients with gastric cancer.

COMMENTS
Background

Gastric cancer is one of the commonest malignant tumors of the alimentary tract and is characterized by late clinical presentation, rapid progression, and poor survival. The reason for this poor prognosis is that, at the time of diagnosis, gastric cancer usually shows extensive local tumor invasion and frequent spread to metastatic sites, particularly lymph nodes. Spread of malignant tumors is a multistep process and many of the stages of tumor invasion require degradation or breakdown of the extracellular matrix and connective tissue surrounding tumor cells.

Research frontiers

The matrix metalloproteinases (MMPs) are a family of zinc containing enzymes which are involved in the degradation of different components of the extracellular matrix, and there is considerable evidence to indicate that individual MMPs have important roles in tumor invasion and tumor spread. A recent study showed that increased levels of chemokine ligand (CCL) recruit immature myeloid cells that carry chemokine ligand receptor from the blood to the tumor invasion front. These immature myeloid cells produced MMP9 and MMP2 and help the tumor cells to migrate and invade.

Innovations and breakthroughs

In the present study, the authors used immunohistochemistry to examine the expression of CCL7, CCL8 and CCL21 in 194 gastric cancer samples and adjacent normal tissues. The authors analyzed their correlation with tumor metastasis, clinicopathologic parameters and clinical outcome. They found that the higher expression of CCL7 and CCL21 in cancer tissues than in normal tissues was significantly correlated with advanced depth of wall invasion, lymph node metastasis and higher tumor node metastasis (TNM) stage. Moreover, Kaplan-Meier survival analysis revealed that CCL7 and CCL21 overexpression in cancer tissues was correlated with poor prognosis.

Applications

These results suggest that overexpression of CCL7 and CCL21 is associated with tumor metastasis and serves as a prognostic factor in patients with gastric cancer.

Peer review

The authors used immunohistochemistry to examine the expression of CCL7, CCL8 and CCL21 in 194 gastric cancer samples and adjacent normal tissues. They found that the higher expression of CCL7 and CCL21 in cancer tissues than in normal tissues was significantly correlated with advanced depth of wall invasion, lymph node metastasis and higher TNM stage. Moreover, Kaplan-Meier survival analysis revealed that CCL7 and CCL21 overexpression in cancer tissues was correlated with poor prognosis.

Footnotes

Peer reviewer: Richard Hu, MD, MSc, Division of Gastroenterology, Department of Medicine, Olive view-UCLA Medical Center, 14445 Olive View Drive, Los Angeles, CA 91342, United States

S- Editor Tian L L- Editor O’Neill M E- Editor Li JY

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