Most of the available preclinical Metabolic dysfunction-associated steatotic liver disease (MASLD) and steatohepatitis (MASH) models fail to resemble metabolic comorbidities and liver fibrosis. To establish a standard MASLD/MASH model, we characterized some morphological, biochemical, and transcriptomic features in a Western diet-induced MASLD model in mice, depicting its similarities to the corresponding human disease. Male C57BL/6J mice received a hypercaloric diet containing sucrose, saturated fat, and cholesterol-rich chow, and high sugar solution for 24 weeks. This model featured a distinct MASH phenotype with obesity, impaired glucose metabolism, hypercholesterolemia, extensive macro and microvesicular, liver steatosis, and slight-to-moderate pericellular/perisinusoidal fibrosis, which was in keeping with the increased hepatic levels of IL-6 and TNF-α, and upregulation of 18 collagen subunit genes (as Col1a1, Col1a2, Col3a1, Col5a2, Col4a1, Col6a3, Col14a1, Col6a2, Col5a1), 34 cytokines or chemokines or related receptors-coding genes (as Il15, Cxcl9, Ccl22), 18 TNF-related genes (as Tnfaip8l3, Tnfrsf21, Tnfaip8, Tnfrfs12a) and 12 metalloproteinase/tissue inhibitors of metalloproteinases-related genes (as Mmp2, Mmp7). The downregulated genes were negative regulators of gluconeogenesis, insulin secretion, and lipid biosynthesis, most belonging to the major urinary protein (MUP) family. The computational analysis of human samples revealed a similarity between our bioassay and human steatohepatitis, with the upregulation of fibrosis- and inflammation-associated orthologs (COL1A1, COL1A2, COL3A1, COL5A2, COL4A1, COL6A3, COL14A1, COL6A2, COL5A1, TNFAIP8L3, TNFRSF21, TNFAIP8, TNFRFS12A, IL15, CXCL9, CCL22, MMP2, MMP7). Our mouse model may be applied as a standard MASH translational bioassay, providing valuable insights into the inflammatory/fibrosis axis of this chronic disease, from the pathogenesis to therapeutic intervention.

There are few effective pharmacological interventions for intracerebral hemorrhage (ICH). Atorvastatin (Ato) has been shown to exert a substantial protective effect on ischemic stroke and is effective in alleviating neuroinflammation. Lipocalin-2 (LCN2), an important inflammation-regulating protein, has been demonstrated to play pivotal roles in post-ICH neuroinflammation. However, the exact role of Ato and whether LCN2 is involved after ICH remain largely unknown. In the current study, the BV2 (microglia) cell line, which was transfected with or without LCN2 for overexpression/interference, was co-cultured with primary cultured neurons and received blood infusion from C57BL/6 mice in vitro. For the in vivo study, atorvastatin was injected peritoneally into an ICH mouse model, and LCN2 specific knockout using the flox/cre system was performed in mice for mechanism study. Behavioral tests were conducted before ICH and on days 1, 3, and 7 post-ICH, and the brains and cultured cells were collected for protein, histological, and morphological studies. Our results showed that atorvastatin treatment alleviates neural damage and promotes neurological outcomes after ICH. Moreover, M1 activation and pro-inflammatory polarization are inhibited by atorvastatin. In both in vivo and in vitro models, the upregulation of LCN2 after ICH is substantially inhibited by atorvastatin. Studies on LCN2 transgenic mice and LCN2 overexpression/interference cells demonstrated that the suppression of macrophage/microglia (M/M) LCN2 participates in atorvastatin-mediated anti-neuroinflammation and neural protection effects. Therefore, our study suggests that atorvastatin treatment attenuates M/M-related neuroinflammation and protects neural recovery by down-regulating LCN2 after ICH. This study identified a potential novel therapeutic target for ICH treatment.

Lipocalin 2 (LCN2) and the TWEAK/Fn14 signaling pathways are pivotal in psoriasis, influencing epidermal development, inflammatory cell chemotaxis, and inflammatory factor release. Despite their significant roles, the intricate relationship between LCN2 and TWEAK/Fn14 pathways remains unclear. Our study revealed the correlation between the expression of TWEAK, LCN2, and Fn14 in psoriatic lesions. We found that TWEAK is expressed by keratinocytes and macrophages, while LCN2 is expressed by keratinocytes and neutrophils. Surface plasmon resonance experiments demonstrated binding between LCN2 and Fn14, which was further validated by co-immunoprecipitation and cellular co-localization via immunofluorescence. In vitro, LCN2 promoted macrophage differentiation and TWEAK secretion, enhanced TWEAK and Fn14 expression in keratinocytes, and activated the MAPK signaling pathway. TWEAK upregulated LCN2 expression in neutrophils but not in keratinocytes. Bulk RNA-seq analysis revealed a synergistic effect of LCN2 and TWEAK in promoting inflammatory cytokine expression in keratinocytes, with enhanced MAPK pathway activation in the presence of M5 cytokines. Lcn2 knockout reduced Fn14 expression in skin lesions and serum TWEAK levels of imiquimod-induced murine psoriasis model, while Fn14 knockout attenuated the epidermal hyperplasia-promoting effects of TWEAK and LCN2. Overexpression of Fn14 in keratinocytes led to higher TWEAK expression upon LCN2 stimulation, suggesting a self-reinforcing loop among TWEAK, LCN2, and Fn14. We propose that LCN2 synergizes with TWEAK through Fn14 to drive psoriasis pathogenesis.

Chitinase-3-like protein 1 (Chi3l1) is a member of the mammalian Chitinase-like protein family, and several studies reported that Chi3l1 is associated with various inflammatory diseases as well as liver diseases. Acetaminophen (APAP) is usually used for antipyretic drug, but its overdose induces acute liver injury (ALI). Several studies reported that subsequent inflammatory responses of the immune system play a critical role in the severity and outcome of APAP-induced ALI. In the present study, we investigated the role of Chi3l1 and its mechanism during APAP-induced ALI using Chi3l1 knock-out (KO) mice. We explored the function of Chi3l1 using APAP-injected KO mice and sought proteins associated with Chi3l1 through biological research data program for investigating mechanism. Liver histological analysis revealed that APAP-induced ALI was attenuated in KO mice compared to wild-type (WT) mice. We observed that APAP-induced neutrophil infiltration was decreased in the liver of KO mice compared to WT mice. To investigate this mechanism, we sought proteins potentially associated with Chi3l1 by mRNA sequencing and protein correlation analysis data. We found lipocalin-2 (Lcn2) and examined Chi3l1, Lcn2, and their relationship in the APAP-induced ALI model using recombinant proteins and antibodies. Our results suggest that Chi3l1 deficiency ameliorates APAP-induced liver injury through abrogating Lcn2-mediated neutrophil infiltration in the liver.

Treating metabolic dysfunction-associated fatty liver disease (MAFLD) and reducing the occurrence of MAFLD-associated liver cancer remain challenging. Two-dimensional (2D) tantalum carbide (Ta4C3) MXene nanozymes (MXenzymes) exhibit antioxidant and anti-inflammatory activities and have thus attracted considerable attention in the fields of oncology and engineering. However, the potential mechanism of action and bioactive properties of Ta4C3 in MAFLD remain uncertain. In our study, Ta4C3 not only inhibited lipid accumulation and disrupted lipid metabolism in hepatocytes but also reduced cell death caused by fatty acids by decreasing intracellular reactive oxygen species (ROS) levels, which significantly promoted the polarization of M1 macrophages to M2 macrophages by alleviating oxidative stress and further suppressing inflammatory factor expression. In mice fed a methionine-choline-deficient (MCD) diet, Ta4C3 reduced lipid accumulation, the infiltration of inflammatory cells, and liver cell apoptosis by modulating the cellular microenvironment through its anti-inflammatory and antioxidant properties. Therefore, Ta4C3 can be used as a multifunctional bioactive material to alleviate hepatic steatosis and inflammation in individuals with MAFLD/metabolic dysfunction-associated steatohepatitis (MASH) because of its robust antioxidant and anti-inflammatory effects.

Tumor immune evasion extends beyond T cells, affecting innate immune elements like natural killer cells (NK) and macrophages within the tumor-immune microenvironment (TIME). Nevertheless, translational strategies to trigger collaboration of NK cells and macrophages to initiate sufficient anti-tumor cytoxicity remain scarce and are urgently needed.

In this study, TCGA datasets was used to confirm the prognosis value of the expression level of NLR family pyrin domain containing 4 (NLRP4) in NSCLC and the tumor tissues microarray was used to further check its clinical-relevance at protein-level. Subsequently, a tumor cell line with stable NLRP4 overexpression was established and subcutaneous tumor models in C57BL/6J mice were used to validate the anti-tumor characteristics of NLRP4. After analyzing the tumor microenvironment using flow cytometry and multiplex immunofluorescence, we further validated our findings through co-culture transwell assays and TCGA analysis. Utilizing bulk-RNA sequencing, proteomics, and mass spectrometry of mouse tumor tissues, we innovatively identified the downstream pathways of NLRP4 and verified them through co-immunoprecipitation (co-IP) and Western blot (WB) experiments.

NLRP4 could trigger a distinct anti-tumor ecosystem organized by TIGIT+TNFA+ NK and iNOS+ M1 in lung cancer, discovered in TCGA analysis and verified in murine model. NLRP4-eco exerted tumor-suppression capacity through chemokine reprogramming including CCL5 and CXCL2. Meanwhile, the cytoxicity of NK could be facilitated by iNOS+M1. Mechanistically, NLRP4 stimulated PI3K/Akt-NF-kB axis through suppression of the activity of PP2A. Besides, knockdown of CCL5 and blockade of CXCL2-CXCR2 axis abolished chemotaxis of TIGIT+TNFA+ NK and iNOS+ M1 respectively, as well as for LB-100, a PP2A inhibitor.

Altogether, we delineated NLRP4's unexplored facets and discovered an NLRP4-driven anti-tumor ecosystem composed of TIGIT+TNFA+ NK and iNOS+ M1. Finally, targeting PP2A by its inhibitor successfully mimicked the anti-tumor capacity of the overexpression of NLRP4.

Metabolic Dysfunction Associated Steatotic Liver Disease (MASLD) and its progressive form, Metabolic Dysfunction Associated Steatohepatitis (MASH), represent significant health concerns associated with the metabolic syndrome. These conditions are characterized by excessive hepatic fat accumulation, inflammation, and potential progression to cirrhosis and hepatocellular carcinoma. Neutrophils are innate immune cells that play a pivotal role in the development of MASLD and MASH. They can infiltrate the hepatic microenvironment in response to inflammatory cytokines and damage associated molecular patterns (DAMPs) derived from the liver and exacerbate tissue damage by releasing of reactive oxygen species (ROS), cytokines, and neutrophil extracellular traps (NETs). Moreover, neutrophils can disrupt the metabolism of hepatocytes through key factors such as neutrophil elastase (NE) and human neutrophil peptides-1 (HNP-1), leading to inflammation and fibrosis, while myeloperoxidase (MPO) and lipocalin (LCN2) are involved in inflammatory and fibrotic processes. In contrast, neutrophils contribute to liver protection and repair through mechanisms involving microRNA-223 and matrix metalloproteinase 9 (MMP9). This dual role of neutrophils highlights their significance in the pathogenesis of MASLD and MASH. This review summarizes current understanding from recent studies on the involvement of neutrophils in MASLD and MASH. Understanding complex roles of neutrophils within the liver's unique microenvironment offers insights into novel therapeutic strategies, emphasizing the need for further research to explore neutrophil-targeted interventions for managing MASLD and MASH. [BMB Reports 2025; 58(3): 116-123].

Diabetic cardiomyopathy (DbCM) is one of the common complications in diabetic patients, but there is no effective treatment for it up to now. Ketone bodies such as β-OHB have been widely reported to be beneficial for metabolic diseases including various diabetic complications. However, the role of ketone metabolism, especially the relevant enzymes, in the pathogenesis of DbCM is poorly understood.

In this study, we firstly observed BDH1, the rate-limiting enzyme of ketone metabolism, was markedly diminished in cardiac tissues from db/db mice and diabetic patients, as well as in H9C2 cells treated with palmitic acid. Genetic deletion of BDH1 aggravated, whereas AAV-mediated BDH1 overexpression attenuated, the diastolic dysfunction and pathogenic progression including apoptosis, fibrosis and inflammation of hearts from db/db mice. Likewise, BDH1 knockdown promoted, whereas BDH1 overexpression reversed, the palmitic acid-induced lipotoxicity in H9C2 cells. Transcriptome analysis revealed that BDH1 negatively regulated LCN2 expression and LCN2 overexpression largely abrogated BDH1 overexpression-mediated myocardial protection in vitro and in vivo. Mechanistically, BDH1 overexpression reprogrammed ketone metabolism with increased AcAc and decreased β-OHB, thereby resulting in decreased β-hydroxybutyrylation of H3K9 on promoter region of LCN2, which repressed transcription of LCN2 and ultimately inhibited NF-κB activity through weakening interaction between NF-κB and RPS3. Furthermore, oral administration of β-hydroxybutyrylation inhibitor A485 to diabetic mice mitigated the cardiac injury concurrently with decreased expression of LCN2.

Our results uncovered a novel mechanism whereby myocardial BDH1 ameliorates DbCM via epigenetic regulation of LCN2, which highlights the potential of BDH1/LCN2-based therapeutics in DbCM.

Alcoholic liver disease (ALD) and non-alcoholic fatty liver disease (NAFLD) are two major chronic liver diseases worldwide. The triggers for fatty liver can be derived from external sources such as adipose tissue, the gut, personal diet, and genetics, or internal sources, including immune cell responses, lipotoxicity, hepatocyte death, mitochondrial dysfunction, and extracellular vesicles. However, their pathogenesis varies to some extent. This review summarizes various immune mechanisms and therapeutic targets associated with these two types of fatty liver disease. It describes the gut-liver axis and adipose tissue-liver crosstalk, as well as the roles of different immune cells (both innate and adaptive immune cells) in fatty liver disease. Additionally, mitochondrial dysfunction, extracellular vesicles, microRNAs (miRNAs), and gastrointestinal hormones are also related to the pathogenesis of fatty liver. Understanding the pathogenesis of fatty liver and corresponding therapeutic strategies provides a new perspective for developing novel treatments for fatty liver disease.

Intervertebral disc degeneration (IVDD) stands as a primary pathophysiological driver of low back pain, yet no therapeutic intervention effectively arrests its progression. Evidence shows that certain Sirt1 agonists may confer protective effects on intervertebral discs, but the underlying mechanisms remain unclear. This study aims to delineate the interaction between Sirt1 and the inflammatory microenvironment, offering potential novel avenues for IVDD prevention and treatment.

In vitro IL-1β-induced nucleus pulposus cells (NPCs) degenerative model and in vivo a mouse annulus fibrosus needle puncture model in Sirt1 transgenic (Sirt1TG) and the same litter WT mice were used to investigate the role of Sirt1 in homeostasis and inflammation. Mechanistic insights were obtained through RNA sequencing, co-immunoprecipitation (Co-IP), luciferase assays, and chromatin immunoprecipitation-(ChIP)-PCR. A co-culture system of Raw264.7 and NPCs was employed to assess the involvement of Lipocalin 2.

Our study demonstrated reduced Sirt1 expression in degenerating human nucleus pulposus (NP) tissue. Both in vitro and in vivo data revealed that NP-specific overexpression of Sirt1 inhibited extracellular matrix degradation and inflammation. Mechanistically, Sirt1 suppressed the acetylation of RelA/p65 at lysine 310 and phosphorylation at serine 536, with the C-terminus of Sirt1 and the RHD-NLS domain of RelA mediating to their interaction. Furthermore, NPCs-derived Lipocalin 2 was identified as a cytokine involved in macrophage chemotaxis and M1 polarization to exacerbate inflammation.

Our work revealed that Sirt1 negatively regulates Lipocalin 2, thereby ameliorating the inflammatory milieu and blocking NPCs and macrophages crosstalk.

This study illuminates the crucial role and molecular mechanisms of Sirt1 in regulating the NP microenvironment. These insights shed light on strategies for the prevention and treatment of IVDD-related herniation and low back pain. By pinpointing specific biological targets, the screening of smallmolecule compounds with significant clinical implications can be facilitated. This translational innovation promises to optimize cells communication within intervertebral disc microenvironment via localized drug delivery, potentially improving patient outcomes and satisfaction following spinal fusion or discectomy surgeries.

Dysfunction of the intestinal barrier is a prevalent phenomenon observed across a spectrum of diseases, encompassing conditions such as mesenteric artery dissection, inflammatory bowel disease, cirrhosis, and sepsis. In these pathological states, the integrity of the intestinal barrier, which normally serves to regulate the selective passage of substances between the gut lumen and the bloodstream, becomes compromised. This compromised barrier function can lead to a range of adverse consequences, including increased permeability to harmful substances, the translocation of bacteria and their products into systemic circulation, and heightened inflammatory responses within the gut and beyond. Understanding the mechanisms underlying intestinal barrier dysfunction in these diverse disease contexts is crucial for the development of targeted therapeutic interventions aimed at restoring barrier integrity and ameliorating disease progression. Lipocalin-2 (LCN2) expression is significantly upregulated during episodes of intestinal inflammation, making it a pivotal indicator for gauging the extent of such inflammatory processes. Notably, however, LCN2 derived from distinct cellular sources, whether intestinal epithelial cells or immune cells, exhibits notably divergent functional characteristics. Furthermore, the multifaceted nature of LCN2 is underscored by its varying roles across different diseases, sometimes even demonstrating contradictory effects.

Previous studies have reported an association between telomere length and non-alcoholic fatty liver disease (NAFLD). This study aimed to explore the involvement of alternative lengthening of telomere-related genes (ALTRGs) in the pathology of NAFLD, construct a risk signature, and evaluate both treatment and prognosis. Three NAFLD datasets (GSE48452, GSE89632, and GSE63067) were collected from the GEO database and merged into combined GEO datasets. ALTRGs were collected from GeneCards and PubMed databases. Differentially expressed genes (DEGs) were identified, and functional enrichment analysis was performed. This study employed a support vector machine algorithm and least absolute shrinkage and selection operator regression analysis to identify key genes for constructing a diagnostic model. High- and low-risk groups were identified from the combined GEO datasets using the diagnostic model. Gene set enrichment analysis, regulatory network analysis, and intergroup immune infiltration analysis were performed. This study identified the key genes using receiver operating characteristic and Friends analysis. Expression of these genes was validated in a mouse model of NAFLD. Twenty-five genes were differentially expressed, with a positive correlation between FOS and EGR1 and a negative correlation between MYC and CEBPA. A diagnostic model was constructed using 12 genes, and high- and low-risk groups were identified. CAMK2G, ERBB2, FOSB, WT1, and CEBPA showed certain accuracy, and their expression levels were significantly different in the model. Immune infiltration analysis between the risk groups revealed that six immune cells were statistically significant. This includes a strong negative interaction between type 2 T helper cells and SPHK2 in the high-risk group. These findings suggest that ALTRDEGs are potential therapeutic targets and prognostic indicators for NAFLD. However, further investigations are required to elucidate the specific underlying mechanisms.

The mammalian immune system is constantly surveying our tissues to clear pathogens and maintain tissue homeostasis. In order to fulfill these tasks, immune cells take up nutrients to supply energy for survival and for directly regulating effector functions via their cellular metabolism, a process now known as immunometabolism. Neutrophilic granulocytes, the most abundant leukocytes in the human body, have a short half-life and are permanently needed in the defense against pathogens. According to a long-standing view, neutrophils were thought to primarily fuel their metabolic demands via glycolysis. Yet, this view has been challenged, as other metabolic pathways recently emerged to contribute to neutrophil homeostasis and effector functions. In particular during neutrophilic development, the pentose phosphate pathway, glycogen synthesis, oxidative phosphorylation, and fatty acid oxidation crucially promote neutrophil maturation. At steady state, both glucose and lipid metabolism sustain neutrophil survival and maintain the intracellular redox balance. This review aims to comprehensively discuss how neutrophilic metabolism adapts during development, which metabolic pathways fuel their functionality, and how these processes are reconfigured in case of various diseases. We provide several examples of hereditary diseases, in which mutations in metabolic enzymes validate their critical role for neutrophil function.

This review explores the biological aspects of neutrophils, their contributions to the development of steatotic liver disease, and their potential as therapeutic targets for the disease. Although alcohol-associated and metabolic dysfunction-associated liver diseases originate from distinct etiological factors, the two diseases frequently share excessive lipid accumulation as a common contributor to their pathogenesis, thereby classifying them as types of steatotic liver disease. Dysregulated lipid deposition in the liver induces hepatic injury, triggering the activation of the innate immunity, partially through neutrophil recruitment. Traditionally recognized for their role in microbial clearance, neutrophils have recently garnered attention for their involvement in sterile inflammation, a pivotal component of steatotic liver disease pathogenesis. In conclusion, technological innovations, including single-cell RNA sequencing, have gradually disclosed the existence of various neutrophil subsets; however, how the distinct subsets of neutrophil population contribute differentially to the development of steatotic liver disease remains unclear.

Lipocalin-2, a neutrophil gelatinase-associated lipocalin, is a 25-kDa secreted protein implicated in a broad range of inflammatory diseases affecting the brain and periphery. It is a pleotropic protein expressed by various immune and nonimmune cells throughout the body. Importantly, the surge in lipocalin-2 levels in disease states has been associated with a myriad of undesirable effects, further exacerbating the ongoing pathological processes. In the brain, glial cells are the principal source of lipocalin-2, which plays a definitive role in determining their functional phenotypes. In different central nervous system pathologies, an increased expression of glial lipocalin-2 has been linked to neurotoxicity. Lipocalin-2 mediates a crosstalk between central and peripheral immune cells under neuroinflammatory conditions. One intriguing aspect is that elevated lipocalin-2 levels in peripheral disorders, such as cancer, metabolic conditions, and liver diseases, potentially incite an inflammatory activation of glial cells while disrupting neuronal functions. This review comprehensively summarizes the influence of lipocalin-2 on the exacerbation of neuroinflammation by regulating various cellular processes. Additionally, this review explores lipocalin-2 as a mediator of neuroimmune crosstalk in various central nervous system pathologies and highlights the role of lipocalin-2 in carrying inflammatory signals along the neuroimmune axis.

The transitory emergence of myeloid-derived suppressor cells (MDSCs) in infants is important for the homeostasis of the immune system in early life. The composition and functional heterogeneity of MDSCs in newborns remain elusive, hampering the understanding of the importance of MDSCs in neonates. In this study, we unraveled the maturation trajectory of polymorphonuclear (PMN)-MDSCs from the peripheral blood of human newborns by performing single-cell RNA sequencing. Results indicated that neonatal PMN-MDSCs differentiated from self-renewal progenitors, antimicrobial PMN-MDSCs, and immunosuppressive PMN-MDSCs to late PMN-MDSCs with reduced antimicrobial capacity. We also established a simple framework to distinguish these distinct stages by CD177 and CXCR2. Importantly, preterm newborns displayed a reduced abundance of classical PMN-MDSCs but increased late PMN-MDSCs, consistent with their higher susceptibility to infections and inflammation. Furthermore, newborn PMN-MDSCs were distinct from those from cancer patients, which displayed minimum expression of genes about antimicrobial capacity. This study indicates that the heterogeneity of PMN-MDSCs is associated with the maturity of human newborns.

Metabolic dysfunction-associated steatohepatitis (MASH) is the severe form of non-alcoholic fatty liver disease which has a high potential to progress to cirrhosis and hepatocellular carcinoma, yet adequate effective therapies are lacking. Hypoadiponectinemia is causally involved in the pathogenesis of MASH. This study investigated the pharmacological effects of adiponectin replacement therapy with the adiponectin-derived peptide ALY688 (ALY688-SR) in a mouse model of MASH. Human induced pluripotent stem (iPS) cell-derived hepatocytes were used to test cytotoxicity and signaling of unmodified ALY688 in vitro. High-fat diet with low methionine and no added choline (CDAHF) was used to induce MASH and test the effects of ALY688-SR in vivo. Histological MASH activity score (NAS) and fibrosis score were determined to assess the effect of ALY688-SR. Transcriptional characterization of mice through RNA sequencing was performed to indicate potential molecular mechanisms involved. In cultured hepatocytes, ALY688 efficiently induced adiponectin-like signaling, including the AMP-activated protein kinase and p38 mitogen-activated protein kinase pathways, and did not elicit cytotoxicity. Administration of ALY688-SR in mice did not influence body weight but significantly ameliorated CDAHF-induced hepatic steatosis, inflammation, and fibrosis, therefore effectively preventing the development and progression of MASH. Mechanistically, ALY688-SR treatment markedly induced hepatic expression of genes involved in fatty acid oxidation, whereas it significantly suppressed the expression of pro-inflammatory and pro-fibrotic genes as demonstrated by transcriptomic analysis. ALY688-SR may represent an effective approach in MASH treatment. Its mode of action involves inhibition of hepatic steatosis, inflammation, and fibrosis, possibly via canonical adiponectin-mediated signaling.

The regenerative capability of the liver is remarkable, but further research is required to understand the role that neutrophils play in this process. In the present study, we reanalyzed single-cell RNA sequencing data from a mouse partial hepatectomy (PH) model to track the transcriptional changes in hepatocytes and non-parenchymal cells. Notably, we unraveled the regenerative capacity of hepatocytes at diverse temporal points after PH, unveiling the contributions of three distinct zones in the liver regeneration process. In addition, we observed that the depletion of neutrophils reduced the survival and liver volume after PH, confirming the important role of neutrophils in liver regeneration. CellChat analysis revealed an intricate crosstalk between neutrophils and macrophages promoting liver regeneration and, using weighted gene correlation network analysis, we identified the most significant genetic module associated with liver regeneration. Our study found that hepatocytes in the periportal zone of the liver are more active than in other zones, suggesting that the crosstalk between neutrophils and macrophages might be a potential target for liver regeneration treatment.

FICZ (6-formylindolo[3,2-b]carbazole) is a potent aryl hydrocarbon receptor agonist that has a poorly understood function in the regulation of inflammation. In this study, we investigated the effect of aryl hydrocarbon receptor activation by FICZ in a murine model of autoimmune hepatitis induced by concanavalin A. High-throughput sequencing techniques such as single-cell RNA sequencing and assay for transposase accessible chromatin sequencing were used to explore the mechanisms through which FICZ induces its effects. FICZ treatment attenuated concanavalin A-induced hepatitis, evidenced by decreased T-cell infiltration, decreased circulating alanine transaminase levels, and suppression of proinflammatory cytokines. Concanavalin A revealed an increase in natural killer T cells, T cells, and mature B cells upon concanavalin A injection while FICZ treatment reversed the presence of these subsets. Surprisingly, concanavalin A depleted a subset of CD55+ B cells, while FICZ partially protected this subset. The immune cells showed significant dysregulation in the gene expression profiles, including diverse expression of migratory markers such as CCL4, CCL5, and CXCL2 and critical regulatory markers such as Junb. Assay for transposase accessible chromatin sequencing showed more accessible chromatin in the CD3e promoter in the concanavalin A-only group as compared to the naive and concanavalin A-exposed, FICZ-treated group. While there was overall more accessible chromatin of the Adgre1 (F4/80) promoter in the FICZ-treated group, we observed less open chromatin in the Itgam (CD11b) promoter in Kupffer cells, supporting the ability of FICZ to reduce the infiltration of proinflammatory cytokine producing CD11b+ Kupffer cells. Taken together, these data demonstrate that aryl hydrocarbon receptor activation by FICZ suppresses liver injury through the limitation of CD3+ T-cell activation and CD11b+ Kupffer cell infiltration.

Metabolic dysfunction-associated steatohepatitis (MASH) is the progressive form of metabolic dysfunction-associated steatotic liver disease (MASLD), and closely associated with a high risk of liver-related morbidity and mortality. Although enhanced neutrophil infiltration of the liver is a histological hallmark of MASH, the morphological pattern of hepatic neutrophils and their relevance to the definition of MASH remain unknown. This clinicopathological study aimed to determine the association of neutrophilic crown-like structures (CLSs) in liver biopsies and evaluate their relevance to the histological diagnosis of MASH. A total of 483 morbidly obese adults who underwent bariatric surgery were recruited. Neutrophilic CLSs in liver biopsies were detected by immunohistochemistry for neutrophil elastase and proteinase 3. All participants were classified into 4 histological subgroups: no MASLD (118, 24.4%), MASLD (76, 15.7%), borderline MASH (185, 38.3%), and definite MASH (104, 21.5%). In the discovery cohort (n = 379), the frequency of neutrophilic CLSs increased in line with the severity of liver disease. The number of neutrophilic CLSs was positively correlated with established histological characteristics of MASH. At a cutoff value of <0.3 per 20× microscopic field, the number of neutrophilic CLSs yielded a robust diagnostic accuracy to discriminate no MASLD and MASLD from borderline MASH and definite MASH; a cutoff at >1.3 per 20× microscopic field exhibited a statistically significant accuracy to distinguish definite MASH from other groups (no MASLD, MASLD, and borderline MASH). The significance of neutrophilic CLSs in identifying borderline MASH and definite MASH was confirmed in an external validation cohort (n = 104). The frequency of neutrophilic CLSs was significantly higher than that of macrophagic CLSs. In conclusion, neutrophilic CLSs in the liver represent a typical histological characteristic of MASH and may serve as a promising indicator to improve the diagnostic accuracy of MASH during histological assessment of liver biopsies.

Neutrophils play a crucial role in host defense against infection. Aberrant neutrophil activation may induce tissue damage via sterile inflammation. Neutrophil accumulation has been identified as a feature of the inflammatory response observed in metabolic dysfunction-associated steatohepatitis (MASH) and has been associated with liver fibrosis and cirrhosis. Here, we performed the transcriptomic analysis of circulating neutrophils from mild and advanced MASH patients to identify the potential mechanism behind neutrophil contribution to MASH progression. Our findings demonstrated that circulating neutrophils from mild and advanced MASH display an increased activated transcriptional program, with the expression of pro-inflammatory factors and an amplified lifespan compared to cells from non-diseased controls. Our results also suggest that MASH progression is associated with a dynamic shift in the profile of circulating neutrophils. In the early stages of MASH, mature neutrophils predominate in the bloodstream. As hepatic inflammation and fibrosis progress, the premature release of immature neutrophils into the circulation occurs. These immature neutrophils exhibit a pro-inflammatory profile that may exacerbate inflammation and promote fibrosis in MASH.

Obeticholic acid (OCA), a farnesoid X receptor (FXR) agonist with favorable effects on fatty and glucose metabolism, has been considered the leading candidate drug for nonalcoholic steatohepatitis (NASH) treatment. However, its limited effectiveness in resolving liver fibrosis and lipotoxicity-induced cell death remains a major drawback. Ferroptosis, a newly recognized form of cell death characterized by uncontrolled lipid peroxidation, is involved in the progression of NASH. Nitric oxide (NO) is a versatile biological molecule that can degrade extracellular matrix. In this study, we developed a PEGylated thiolated hollow mesoporous silica nanoparticles (MSN) loaded with OCA, as well as a ferroptosis inhibitor liproxsatin-1 and a NO donor S-nitrosothiol (ONL@MSN). Biochemical analyses, histology, multiplexed flow cytometry, bulk-tissue RNA sequencing, and fecal 16S ribosomal RNA sequencing were utilized to evaluate the effects of the combined nanoparticle (ONL@MSN) in a mouse NASH model. Compared with the OCA-loaded nanoparticles (O@MSN), ONL@MSN not only protected against hepatic steatosis but also greatly ameliorated fibrosis and ferroptosis. ONL@MSN also displayed enhanced therapeutic actions on the maintenance of intrahepatic macrophages/monocytes homeostasis, inhibition of immune response/lipid peroxidation, and correction of microbiota dysbiosis. These findings present a promising synergistic nanotherapeutic strategy for the treatment of NASH by simultaneously targeting FXR, ferroptosis, and fibrosis.

Metabolic dysfunction-associated steatotic liver disease (MASLD) has emerged as the most common liver disease worldwide in recent years. MASLD commonly presents as simple hepatic steatosis, but ~25% of patients develop liver inflammation, progressive fibrosis, liver cirrhosis and related hepatocellular carcinoma. Liver inflammation and the degree of fibrosis are key determinants of the prognosis. The pathophysiology of liver inflammation is incompletely understood and involves diverse factors and specifically innate and adaptive immune responses. More specifically, diverse mediators of innate immunity such as proinflammatory cytokines, adipokines, inflammasomes and various cell types like mononuclear cells, macrophages and natural killer cells are involved in directing the inflammatory process in MASLD. The activation of innate immunity is driven by various factors including excess lipids and lipotoxicity, insulin resistance and molecular patterns derived from gut commensals. Targeting pathways of innate immunity might therefore appear as an attractive therapeutic strategy in the future management of MASLD and possibly its complications.

Patients with inflammatory liver diseases, particularly alcohol-associated liver disease and metabolic dysfunction-associated fatty liver disease (MAFLD), have higher incidence of infections and mortality rate due to sepsis. The current focus in the development of drugs for MAFLD is the resolution of non-alcoholic steatohepatitis and prevention of progression to cirrhosis. In patients with cirrhosis or alcoholic hepatitis, sepsis is a major cause of death. As the metabolic center and a key immune tissue, liver is the guardian, modifier, and target of sepsis. Septic patients with liver dysfunction have the highest mortality rate compared with other organ dysfunctions. In addition to maintaining metabolic homeostasis, the liver produces and secretes hepatokines and acute phase proteins (APPs) essential in tissue protection, immunomodulation, and coagulation. Inflammatory liver diseases cause profound metabolic disorder and impairment of energy metabolism, liver regeneration, and production/secretion of APPs and hepatokines. Herein, the author reviews the roles of (1) disorders in the metabolism of glucose, fatty acids, ketone bodies, and amino acids as well as the clearance of ammonia and lactate in the pathogenesis of inflammatory liver diseases and sepsis; (2) cytokines/chemokines in inflammatory liver diseases and sepsis; (3) APPs and hepatokines in the protection against tissue injury and infections; and (4) major nuclear receptors/signaling pathways underlying the metabolic disorders and tissue injuries as well as the major drug targets for inflammatory liver diseases and sepsis. Approaches that focus on the liver dysfunction and regeneration will not only treat inflammatory liver diseases but also prevent the development of severe infections and sepsis.

Adipose tissues (AT) are an important endocrine organ that secretes various functional adipokines, peptides, non-coding RNAs, and acts on AT themselves or other distant tissues or organs through autocrine, paracrine, or endocrine manners. An accumulating body of evidence has suggested that many adipokines play an important role in liver metabolism. Besides the traditional adipokines such as adiponectin and leptin, many novel adipokines have recently been identified to have regulatory effects on the liver. Additionally, AT can produce extracellular vesicles (EVs) that act on peripheral tissues. However, under pathological conditions, such as obesity and diabetes, dysregulation of adipokines is associated with functional changes in AT, which may cause liver diseases. In this review, we focus on the newly discovered adipokines and EVs secreted by AT and highlight their actions on the liver under the context of obesity, nonalcoholic fatty liver diseases (NAFLD), and some other liver diseases. Clarifying the action of adipokines and adipose tissue-derived EVs on the liver would help to identify novel therapeutic targets or biomarkers for metabolic diseases.

Sterol regulatory element-binding protein (SREBP)-1c is involved in cellular lipid homeostasis and cholesterol biosynthesis and is highly increased in nonalcoholic steatohepatitis (NASH). However, the molecular mechanism by which SREBP-1c regulates hepatic stellate cells (HSCs) activation in NASH animal models and patients have not been fully elucidated. In this study, we examined the role of SREBP-1c in NASH and the regulation of LCN2 gene expression. Wild-type and SREBP-1c knockout (1cKO) mice were fed a high-fat/high-sucrose diet, treated with carbon tetrachloride (CCl4), and subjected to lipocalin-2 (LCN2) overexpression. The role of LCN2 in NASH progression was assessed using mouse primary hepatocytes, Kupffer cells, and HSCs. LCN2 expression was examined in samples from normal patients and those with NASH. LCN2 gene expression and secretion increased in CCl4-induced liver fibrosis mice model, and SREBP-1c regulated LCN2 gene transcription. Moreover, treatment with holo-LCN2 stimulated intracellular iron accumulation and fibrosis-related gene expression in mouse primary HSCs, but these effects were not observed in 1cKO HSCs, indicating that SREBP-1c-induced LCN2 expression and secretion could stimulate HSCs activation through iron accumulation. Furthermore, LCN2 expression was strongly correlated with inflammation and fibrosis in patients with NASH. Our findings indicate that SREBP-1c regulates Lcn2 gene expression, contributing to diet-induced NASH. Reduced Lcn2 expression in 1cKO mice protects against NASH development. Therefore, the activation of Lcn2 by SREBP-1c establishes a new connection between iron and lipid metabolism, affecting inflammation and HSCs activation. These findings may lead to new therapeutic strategies for NASH.

Introduction. Staphylococcus aureus is the leading cause of acute medical implant infections, representing a significant modern medical concern. The success of S. aureus as a pathogen in these cases resides in its arsenal of virulence factors, resistance to multiple antimicrobials, mechanisms of immune modulation, and ability to rapidly form biofilms associated with implant surfaces. S. aureus device-associated, biofilm-mediated infections are often persistent and notoriously difficult to treat, skewing innate immune responses to promote chronic reoccurring infections. While relatively little is known of the role neutrophils play in response to acute S. aureus biofilm infections, these effector cells must be efficiently recruited to sites of infection via directed chemotaxis. Here we investigate the effects of modulating CXC chemokine receptor 2 (CXCR2) activity, predominantly expressed on neutrophils, during S. aureus implant-associated infection.Hypothesis. We hypothesize that modulation of CXCR2 expression and/or signalling activities during S. aureus infection, and thus neutrophil recruitment, extravasation and antimicrobial activity, will affect infection control and bacterial burdens in a mouse model of implant-associated infection.Aim. This investigation aims to elucidate the impact of altered CXCR2 activity during S. aureus biofilm-mediated infection that may help develop a framework for an effective novel strategy to prevent morbidity and mortality associated with implant infections.Methodology. To examine the role of CXCR2 during S. aureus implant infection, we employed a mouse model of indwelling subcutaneous catheter infection using a community-associated methicillin-resistant S. aureus (MRSA) strain. To assess the role of CXCR2 induction or inhibition during infection, treatment groups received daily intraperitoneal doses of either Lipocalin-2 (Lcn2) or AZD5069, respectively. At the end of the study, catheters and surrounding soft tissues were analysed for bacterial burdens and dissemination, and Cxcr2 transcription within the implant-associated tissues was quantified.Results. Mice treated with Lcn2 developed higher bacterial burdens within the soft tissue surrounding the implant site, which was associated with increased Cxcr2 expression. AZD5069 treatment also resulted in increased implant- and tissues-associated bacterial titres, as well as enhanced Cxcr2 expression.Conclusion. Our results demonstrate that CXCR2 plays an essential role in regulating the severity of S. aureus implant-associated infections. Interestingly, however, perturbation of CXCR2 expression or signalling both resulted in enhanced Cxcr2 transcription and elevated implant-associated bacterial burdens. Thus, CXCR2 appears finely tuned to efficiently recruit effector cells and mediate control of S. aureus biofilm-mediated infection.

The skeleton is traditionally known for its structural support, organ protection, movement, and maintenance of mineral homeostasis. Over the last 10 years, bone has emerged as an endocrine organ with diverse physiological functions. The two key molecules in this context are fibroblast growth factor 23 (FGF23), secreted by osteocytes, and osteocalcin, a hormone produced by osteoblasts. FGF23 affects mineral homeostasis through its actions on the kidneys, and osteocalcin has beneficial effects in improving glucose homeostasis, muscle function, brain development, cognition, and male fertility. In addition, another osteoblast-derived hormone, lipocalin 2 (LCN2) has emerged into the researchers' field of vision. In this review, we mainly focus on LCN2's role in appetite regulation and glucose metabolism and also briefly introduce its effects in other pathophysiological conditions, such as nonalcoholic fatty liver disease, sarcopenic obesity, and cancer-induced cachexia.

Non-alcoholic steatohepatitis (NASH) is one of the fastest growing liver diseases worldwide, and oxidative stress is one of NASH main key drivers. Nicotinamide adenine dinucleotide phosphate (NADPH) is the ultimate donor of reductive power to a number of antioxidant defences. Here, we explored the potential of increasing NADPH levels to prevent NASH progression. We used nicotinamide riboside (NR) supplementation or a G6PD-tg mouse line harbouring an additional copy of the human G6PD gene. In a NASH mouse model induced by feeding mice a methionine-choline deficient (MCD) diet for three weeks, both tools increased the hepatic levels of NADPH and ameliorated the NASH phenotype induced by the MCD intervention, but only in female mice. Boosting NADPH levels in females increased the liver expression of the antioxidant genes Gsta3, Sod1 and Txnrd1 in NR-treated mice, or of Gsr for G6PD-tg mice. Both strategies significantly reduced hepatic lipid peroxidation. NR-treated female mice showed a reduction of steatosis accompanied by a drop of the hepatic triglyceride levels, that was not observed in G6PD-tg mice. NR-treated mice tended to reduce their lobular inflammation, showed a reduction of the NK cell population and diminished transcription of the damage marker Lcn2. G6PD-tg female mice exhibited a reduction of their lobular inflammation and hepatocyte ballooning induced by the MCD diet, that was related to a reduction of the monocyte-derived macrophage population and the Tnfa, Ccl2 and Lcn2 gene expression. As conclusion, boosting hepatic NADPH levels attenuated the oxidative lipid damage and the exhausted antioxidant gene expression specifically in female mice in two different models of NASH, preventing the progression of the inflammatory process and hepatic injury.

Neutrophils and Lipocalin-2 (LCN2) play pivotal roles in cerebral ischemiareperfusion (I/R) injury. However, their contribution is not fully clarified.

This study aimed to explore the role of LCN2 and its association with neutrophil polarization in I/R injury.

A mouse model of middle cerebral artery occlusion (MCAO) was used to induce cerebral ischemia. LCN2mAb was administered 1 h and Anti-Ly6G was administered for 3d before MCAO. The role of LCN2 in the polarity transition of neutrophils was explored using an in vitro HL-60 cell model.

LCN2mAb pretreatment had neuroprotective effects in mice. The expression of Ly6G was not significantly different, but the expression of N2 neutrophils was increased. In the in vitro study, LCN2mAb-treated N1-HL-60 cells induced N2-HL-60 polarization.

LCN2 may affect the prognosis of ischemic stroke by mediating neutrophil polarization.

Peripheral arterial disease (PAD) is a leading cause of morbimortality worldwide. Lipocalin-2 (LCN2) has been associated with higher risk of amputation or mortality in PAD and might be involved in muscle regeneration. Our aim is to unravel the role of LCN2 in skeletal muscle repair and PAD.

WT and Lcn2-/- mice underwent hindlimb ischemia. Blood and crural muscles were analyzed at the inflammatory and regenerative phases. At day 2, Lcn2-/- male mice, but not females, showed increased blood and soleus muscle neutrophils, and elevated circulating pro-inflammatory monocytes (p < 0.05), while locally, total infiltrating macrophages were reduced (p < 0.05). Moreover, Lcn2-/- soleus displayed an elevation of Cxcl1 (p < 0.001), and Cxcr2 (p < 0.01 in males), and a decrease in Ccl5 (p < 0.05). At day 15, Lcn2 deficiency delayed muscle recovery, with higher density of regenerating myocytes (p < 0.04) and arterioles (αSMA+, p < 0.025). Reverse target prediction analysis identified miR-138-5p as a potential regulator of LCN2, showing an inverse correlation with Lcn2 mRNA in skeletal muscles (rho = -0.58, p < 0.01). In vitro, miR-138-5p mimic reduced Lcn2 expression and luciferase activity in murine macrophages (p < 0.05). Finally, in human serum miR-138-5p was inversely correlated with LCN2 (p ≤ 0.001 adjusted, n = 318), and associated with PAD (Odds ratio 0.634, p = 0.02, adjusted, PAD n = 264, control n = 54).

This study suggests a possible dual role of LCN2 in acute and chronic conditions, with a probable role in restraining inflammation early after skeletal muscle ischemia, while being associated with vascular damage in PAD, and identifies miR-138-5p as one potential post-transcriptional regulator of LCN2.

Lipedema is a chronic disorder that mainly affects women. It is often misdiagnosed, and its etiology remains unknown. Recent research indicates an accumulation of macrophages and a shift in macrophage polarization in lipedema. One known protein superfamily that contributes to macrophage accumulation and polarization is the macrophage migration inhibitory factor (MIF) family. MIF-1 and MIF-2 are ubiquitously expressed and also regulate inflammatory processes in adipose tissue. In this study, the expression of MIF-1, MIF-2 and CD74-a common receptor for both cytokines-was analyzed in tissue samples of 11 lipedema and 11 BMI-matched, age-matched and anatomically matched control patients using qPCR and immunohistochemistry (IHC). The mRNA expression of MIF-1 (mean 1.256; SD 0.303; p = 0.0485) and CD74 (mean 1.514; SD 0.397; p = 0.0097) were significantly elevated in lipedema patients, while MIF-2 expression was unaffected (mean 1.004; SD 0.358; p = 0.9718). The IHC analysis corroborated the results for CD74 expression on a cellular level. In conclusion, our results provide first evidence for a potential involvement of the MIF family, presumably via the MIF-1-CD74 axis, in lipedema.

CD24+CK19+/CD24+SOX9+ resident liver cells are activated and expanded after chronic liver injury in a ductular reaction. However, the sources and functions of these cells in liver damage remain disputed.

The current study combined genetic lineage tracing with in vitro small-molecule-based reprogramming to define liver progenitor cells (LPCs) derived from hepatic parenchymal and non-parenchymal tissues. tdTom+ hepatocytes were isolated from ROSA26tdTomato mice following AAV8-Tbg-Cre-mediated recombination, EpCAM+ biliary epithelial cells (BECs) from wild-type intrahepatic bile ducts and ALB/GFP-EpCAM- cells were isolated from AlbCreERT/R26GFP mice. A cocktail of small molecules was used to convert the isolated cells into LPCs. These in vitro cultured LPCs with CD24 and SOX9 expression regained the ability to proliferate. Transcriptional profiling showed that the in-vitro cultured LPCs derived from the resident LPCs in non-parenchymal tissues expressed Lipocalin-2 (Lcn2) at high levels. Accordingly, endogenous Cd24a+Lcn2+ LPCs were identified by integration of sc-RNA-sequencing and pathological datasets of liver dysfunction which indicates that LPCs produced by ductular reactions might also originate from the resident LPCs. Transplantation of in-vitro cultured Cd24a+Lcn2+ LPCs into CCl4-induced fibrotic livers exacerbated liver damage and dysfunction, possibly due to LCN2-dependent macrophage inflammatory response.

CD24+LCN2+ LPCs constituted the expanding ductular reaction and contributed to macrophage-mediated inflammation in chronic liver damage. The current findings highlight the roles of LPCs from distinct origins and expose the possibility of targeting LPCs in the treatment of chronic hepatic diseases.

Cancer metastatic organotropism is still a mystery. The liver is known to be susceptible to cancer metastasis and alcoholic injury. However, it is unclear whether and how alcohol facilitates liver metastasis and how to intervene. Here, we show that alcohol preferentially promotes liver metastasis in colon-cancer-bearing mice and post-surgery pancreatic cancer patients. The mechanism is that alcohol triggers an extra- and intrahepatic crosstalk to reshape an immunosuppressive liver microenvironment. In detail, alcohol upregulates extrahepatic IL-6 and hepatocellular IL-6 receptor expression, resulting in hepatocyte STAT3 signaling activation and downstream lipocalin-2 (Lcn2) upregulation. Furthermore, LCN2 promotes T cell-exhaustion neutrophil recruitment and cancer cell epithelial plasticity. In contrast, knocking out hepatocellular Stat3 or systemic Il6 in alcohol-treated mice preserves the liver microenvironment and suppresses liver metastasis. This mechanism is reflected in hepatocellular carcinoma patients, in that alcohol-associated signaling elevation in noncancerous liver tissue indicates adverse prognosis. Accordingly, we discover a novel application for BBI608, a small molecular STAT3 inhibitor that can prevent liver metastasis. BBI608 pretreatment protects the liver and suppresses alcohol-triggered premetastatic niche formation. In conclusion, under extra- and intrahepatic crosstalk, the alcoholic injured liver forms a favorable niche for cancer cell metastasis, while BBI608 is a promising anti-metastatic agent targeting such microenvironments.

Neutrophils are the most abundant circulating granulocytes, linking innate and adaptive immunity. Neutrophils can regulate inflammatory and immune responses through degranulation, reactive oxygen species generation, the production of cytokines and chemokines, and NETosis. Emerging evidence has indicated that neutrophils contribute to the pathogenesis of various noncancer liver diseases, including nonalcoholic fatty liver disease, alcohol-associated liver disease, hepatic ischemia-reperfusion injury, and liver fibrosis. Cellular interactions among neutrophils, other immune cells, and nonimmune cells constitute a complex network that regulates the immune microenvironment of the liver. This review summarizes novel neutrophil subtypes, including CD177+ neutrophils and low-density neutrophils. Moreover, we provide an overview of the cellular cros stalk of neutrophils in noncancer liver diseases, aiming to shed new light on mechanistic studies of novel neutrophil subtypes. In addition, we discuss the potential of neutrophils as therapeutic targets in noncancer liver diseases, including inhibitors targeting NETosis, granule proteins, and chemokines.