Pub Date : 2025-01-01DOI: 10.1016/j.jcmgh.2025.101460
Alexandra M. Vázquez Salgado , Chunmiao Cai , Markcus Lee II , Dingzi Yin , Marie-Lise Chrystostome , Adrienne F. Gefre , Shirui He , Julia E. Kieckhaefer , Kirk J. Wangensteen
Background & Aims
Hepatocellular carcinoma (HCC) frequently undergoes regional chromosomal amplification, resulting in elevated gene expression levels. We aimed to elucidate the role of these poorly understood genetic changes by using CRISPR activation (CRISPRa) screening in mouse livers to identify which genes within these amplified loci are cancer driver genes.
Methods
We used data from The Cancer Genome Atlas to identify that frequently copy number-amplified and up-regulated genes all reside on human chromosomes 1q and 8q. We generated CRISPRa screening transposons that contain oncogenic Myc to drive tumor formation. We conducted CRISPRa screens in vivo in the liver to identify tumor driver genes. We extensively validated the findings in separate mice and performed RNA sequencing analysis to explore mechanisms driving tumorigenesis.
Results
We targeted genes that frequently undergo amplification in human HCC using an in vivo CRISPRa screening system in mice, which induced extensive liver tumorigenesis. Human chromosome 1q genes Zbtb7b, Vps72, Gba1, and Mrpl9 emerged as drivers of liver tumorigenesis. In human HCC there is a trend in correlation between levels of MRPL9, VPS72, or GBA1 and poor survival. In validation assays, activation of Vps72, Gba1, or Mrpl9 resulted in extensive liver tumorigenesis and decreased survival in mice. RNA sequencing revealed different mechanisms driving HCC, with Mrpl9 activation altering genes functionally related to mitochondrial function, Vps72 levels altering phospholipid metabolism, and Gba1 activation enhancing endosomal-lysosomal activity, all leading to promotion of cellular proliferation. Analysis of human tumor tissues with high levels of MRPL9, VPS72, or GBA1 revealed congruent results, indicating conserved mechanisms driving HCC.
Conclusions
This study reveals chromosome 1q genes Vps72, Gba1, and Mrpl9 as drivers of HCC. Future efforts to prevent or treat HCC can focus on these new driver genes.
{"title":"In Vivo CRISPR Activation Screening Reveals Chromosome 1q Genes VPS72, GBA1, and MRPL9 Drive Hepatocellular Carcinoma","authors":"Alexandra M. Vázquez Salgado , Chunmiao Cai , Markcus Lee II , Dingzi Yin , Marie-Lise Chrystostome , Adrienne F. Gefre , Shirui He , Julia E. Kieckhaefer , Kirk J. Wangensteen","doi":"10.1016/j.jcmgh.2025.101460","DOIUrl":"10.1016/j.jcmgh.2025.101460","url":null,"abstract":"<div><h3>Background & Aims</h3><div>Hepatocellular carcinoma (HCC) frequently undergoes regional chromosomal amplification, resulting in elevated gene expression levels. We aimed to elucidate the role of these poorly understood genetic changes by using CRISPR activation (CRISPRa) screening in mouse livers to identify which genes within these amplified loci are cancer driver genes.</div></div><div><h3>Methods</h3><div>We used data from The Cancer Genome Atlas to identify that frequently copy number-amplified and up-regulated genes all reside on human chromosomes 1q and 8q. We generated CRISPRa screening transposons that contain oncogenic <em>Myc</em> to drive tumor formation. We conducted CRISPRa screens in vivo in the liver to identify tumor driver genes. We extensively validated the findings in separate mice and performed RNA sequencing analysis to explore mechanisms driving tumorigenesis.</div></div><div><h3>Results</h3><div>We targeted genes that frequently undergo amplification in human HCC using an in vivo CRISPRa screening system in mice, which induced extensive liver tumorigenesis. Human chromosome 1q genes <em>Zbtb7b</em>, <em>Vps72</em>, <em>Gba1</em>, and <em>Mrpl9</em> emerged as drivers of liver tumorigenesis. In human HCC there is a trend in correlation between levels of <em>MRPL9</em>, <em>VPS72,</em> or <em>GBA1</em> and poor survival. In validation assays, activation of <em>Vps72</em>, <em>Gba1</em>, or <em>Mrpl9</em> resulted in extensive liver tumorigenesis and decreased survival in mice. RNA sequencing revealed different mechanisms driving HCC, with <em>Mrpl9</em> activation altering genes functionally related to mitochondrial function, <em>Vps72</em> levels altering phospholipid metabolism, and <em>Gba1</em> activation enhancing endosomal-lysosomal activity, all leading to promotion of cellular proliferation. Analysis of human tumor tissues with high levels of <em>MRPL9</em>, <em>VPS72,</em> or <em>GBA1</em> revealed congruent results, indicating conserved mechanisms driving HCC.</div></div><div><h3>Conclusions</h3><div>This study reveals chromosome 1q genes <em>Vps72</em>, <em>Gba1</em>, and <em>Mrpl9</em> as drivers of HCC. Future efforts to prevent or treat HCC can focus on these new driver genes.</div></div>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 5","pages":"Article 101460"},"PeriodicalIF":7.1,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143048930","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Retinoblastoma-binding protein 9 (RBBP9) was initially reported as cell cycle regulator via RB/E2F. Accumulating evidence has revealed the importance of RBBP9 in physiological and pathological states including inflammatory disease. However, the functional role of RBBP9 in ulcerative colitis (UC) and colitis-associated cancer (CAC) remains elusive.
Methods
Human samples of UC and CAC were examined by immunohistochemical and bioinformatics analyses. We established dextran sodium sulfate (DSS)-induced colitis, azoxymethane (AOM)/DSS-induced CAC model, and ApcMin/+ sporadic tumor model using wild-type and Rbbp9-/- mice. RNA sequencing was analyzed to identify the phenotype alternation upon Rbbp9 deletion. In addition, genetic and pharmacological inhibition of the Janus kinase (JAK)/signal transducer and activator of transcription 1 (STAT1) pathway was performed.
Results
The expression of RBBP9 was reduced in human UC and CAC samples. The loss of RBBP9 enhanced the activation of interferon (IFN)/JAK/STAT1 signaling, resulting in susceptibility to DSS-induced colitis and AOM/DSS-induced CAC tumors by increasing epithelial cell apoptosis and immune activation. An in vitro kinase assay revealed that RBBP9 directly regulated JAK/STAT1 signaling by suppressing STAT1 phosphorylation. A positive feedback loop involving epithelial cell apoptosis, commensal microbiome invasion, and activation of submucosal immune activity was identified in Rbbp9-/- mouse intestines through enhanced JAK/STAT1 signaling in RBBP9-deficient epithelial cells and macrophages. The genetic inhibition of STAT1 or treatment with the JAK/STAT inhibitor reversed epithelial cell apoptosis and mitigated the enhanced susceptibility to DSS-induced colitis in Rbbp9-/- mice.
Conclusions
RBBP9 suppresses the intestinal inflammation by negatively regulating JAK/STAT1 signaling pathway.
{"title":"Retinoblastoma-binding Protein 9 Suppresses Intestinal Inflammation and Inflammation-induced Tumorigenesis in Mice","authors":"Kensuke Hamada , Yuki Nakanishi , Yu Muta , Mayuki Omatsu , Kosuke Iwane , Munehiro Ikeda , Jiayu Chen , Yoko Masui , Naoki Aoyama , Nobukazu Agatsuma , Go Yamakawa , Takahiro Utsumi , Hiroki Kitamoto , Makoto Okabe , Yoshiro Itatani , Takumi Adachi , Koubun Yasuda , Shuji Yamamoto , Akihisa Fukuda , Etsushi Kuroda , Hiroshi Seno","doi":"10.1016/j.jcmgh.2024.101435","DOIUrl":"10.1016/j.jcmgh.2024.101435","url":null,"abstract":"<div><h3>Background & Aims</h3><div>Retinoblastoma-binding protein 9 (RBBP9) was initially reported as cell cycle regulator via RB/E2F. Accumulating evidence has revealed the importance of RBBP9 in physiological and pathological states including inflammatory disease. However, the functional role of RBBP9 in ulcerative colitis (UC) and colitis-associated cancer (CAC) remains elusive.</div></div><div><h3>Methods</h3><div>Human samples of UC and CAC were examined by immunohistochemical and bioinformatics analyses. We established dextran sodium sulfate (DSS)-induced colitis, azoxymethane (AOM)/DSS-induced CAC model, and <em>Apc</em><sup><em>Min/+</em></sup> sporadic tumor model using wild-type and <em>Rbbp9</em><sup><em>-/-</em></sup> mice. RNA sequencing was analyzed to identify the phenotype alternation upon <em>Rbbp9</em> deletion. In addition, genetic and pharmacological inhibition of the Janus kinase (JAK)/signal transducer and activator of transcription 1 (STAT1) pathway was performed.</div></div><div><h3>Results</h3><div>The expression of RBBP9 was reduced in human UC and CAC samples. The loss of RBBP9 enhanced the activation of interferon (IFN)/JAK/STAT1 signaling, resulting in susceptibility to DSS-induced colitis and AOM/DSS-induced CAC tumors by increasing epithelial cell apoptosis and immune activation. An <em>in vitro</em> kinase assay revealed that RBBP9 directly regulated JAK/STAT1 signaling by suppressing STAT1 phosphorylation. A positive feedback loop involving epithelial cell apoptosis, commensal microbiome invasion, and activation of submucosal immune activity was identified in <em>Rbbp9</em><sup><em>-/-</em></sup> mouse intestines through enhanced JAK/STAT1 signaling in RBBP9-deficient epithelial cells and macrophages. The genetic inhibition of STAT1 or treatment with the JAK/STAT inhibitor reversed epithelial cell apoptosis and mitigated the enhanced susceptibility to DSS-induced colitis in <em>Rbbp9</em><sup><em>-/-</em></sup> mice.</div></div><div><h3>Conclusions</h3><div>RBBP9 suppresses the intestinal inflammation by negatively regulating JAK/STAT1 signaling pathway.</div></div>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 3","pages":"Article 101435"},"PeriodicalIF":7.1,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11786897/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142781960","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-01DOI: 10.1016/j.jcmgh.2024.101429
Xianghu Wang , Yuanguo Wang , Bing Bai , Aurpita Shaha , Wenming Bao , Lianping He , Tian Wang , Gaspar J. Kitange , Ningling Kang
Background & Aims
Transforming growth factor (TGF)β1 induces plasma membrane (PM) accumulation of glucose transporter 1 (Glut1) required for glycolysis of hepatic stellate cells (HSCs) and HSC activation. This study aimed to understand how Glut1 is anchored/docked onto the PM of HSCs.
Methods
HSC expression of protein kinase M zeta isoform (PKMζ) was detected by reverse transcription polymerase chain reaction (RT-PCR), Western blotting, and immunofluorescence. PKMζ level was manipulated by short hairpin RNA (shRNA) or overexpression; HSC activation was assessed by cell expression of activation markers; PM Glut1, glucose uptake, and glycolysis of HSCs were analyzed by biotinylation, 2-NBDG-based assay, and Seahorse Glycolysis Stress Test. Phospho-mutants of vasodilator-stimulated phosphorylated protein (VASP) were created by site-directed mutagenesis. TGFβ transcriptome was obtained by RNA sequencing. Single-cell RNA sequencing datasets and immunofluorescence were leveraged to analyze PKMζ expression in cancer-associated fibroblasts (CAFs) of colorectal liver metastases. Function of HSC PKMζ was determined by tumor/HSC co-implantation study.
Results
Primary human and murine HSCs express PKMζ, but not full-length PKCζ. PKMζ knockdown suppresses, whereas PKMζ overexpression potentiates PM accumulation of Glut1, glycolysis, and HSC activation induced by TGFβ1. Mechanistically, PKMζ binds to and induces VASP phosphorylation at serines 157 and 239 facilitating anchoring/docking of Glut1 onto the PM of HSCs. PKMζ expression is increased in the CAFs of murine and patient colorectal liver metastases compared with quiescent HSCs. Targeting PKMζ suppresses transcriptome, CAF activation of HSCs, and colorectal tumor growth in mice.
Conclusions
Because HSCs are also a major contributor of liver fibrosis, our data highlight PKMζ and VASP as targets to inhibit metabolic reprogramming, HSC activation, liver fibrosis, and the pro-metastatic microenvironment of the liver.
{"title":"PKMζ, a Brain-specific PKCζ Isoform, is Required for Glycolysis and Myofibroblastic Activation of Hepatic Stellate Cells","authors":"Xianghu Wang , Yuanguo Wang , Bing Bai , Aurpita Shaha , Wenming Bao , Lianping He , Tian Wang , Gaspar J. Kitange , Ningling Kang","doi":"10.1016/j.jcmgh.2024.101429","DOIUrl":"10.1016/j.jcmgh.2024.101429","url":null,"abstract":"<div><h3>Background & Aims</h3><div>Transforming growth factor <strong>(</strong>TGF)β1 induces plasma membrane (PM) accumulation of glucose transporter 1 (Glut1) required for glycolysis of hepatic stellate cells (HSCs) and HSC activation. This study aimed to understand how Glut1 is anchored/docked onto the PM of HSCs.</div></div><div><h3>Methods</h3><div>HSC expression of protein kinase M zeta isoform (PKMζ) was detected by reverse transcription polymerase chain reaction (RT-PCR), Western blotting, and immunofluorescence. PKMζ level was manipulated by short hairpin RNA (shRNA) or overexpression; HSC activation was assessed by cell expression of activation markers; PM Glut1, glucose uptake, and glycolysis of HSCs were analyzed by biotinylation, 2-NBDG-based assay, and Seahorse Glycolysis Stress Test. Phospho-mutants of vasodilator-stimulated phosphorylated protein (VASP) were created by site-directed mutagenesis. TGFβ transcriptome was obtained by RNA sequencing. Single-cell RNA sequencing datasets and immunofluorescence were leveraged to analyze PKMζ expression in cancer-associated fibroblasts (CAFs) of colorectal liver metastases. Function of HSC PKMζ was determined by tumor/HSC co-implantation study.</div></div><div><h3>Results</h3><div>Primary human and murine HSCs express PKMζ, but not full-length PKCζ. PKMζ knockdown suppresses, whereas PKMζ overexpression potentiates PM accumulation of Glut1, glycolysis, and HSC activation induced by TGFβ1. Mechanistically, PKMζ binds to and induces VASP phosphorylation at serines 157 and 239 facilitating anchoring/docking of Glut1 onto the PM of HSCs. PKMζ expression is increased in the CAFs of murine and patient colorectal liver metastases compared with quiescent HSCs. Targeting PKMζ suppresses transcriptome, CAF activation of HSCs, and colorectal tumor growth in mice.</div></div><div><h3>Conclusions</h3><div>Because HSCs are also a major contributor of liver fibrosis, our data highlight PKMζ and VASP as targets to inhibit metabolic reprogramming, HSC activation, liver fibrosis, and the pro-metastatic microenvironment of the liver.</div></div>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 3","pages":"Article 101429"},"PeriodicalIF":7.1,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11750446/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142633578","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-01DOI: 10.1016/j.jcmgh.2024.101413
Linxi Xie , Hejiao Zhang , Long Xu
Previously, eosinophils were primarily regarded as effector toxic cells involved in allergic diseases and parasitic infections. Nevertheless, new research has shown that eosinophils are diverse and essential for immune regulation and tissue homeostasis. Their functional plasticity has been observed in patients with inflammatory diseases, cancer, infections, and other disorders. Although eosinophils are infrequently observed within the liver during periods of homeostasis, they are recruited to the liver in various liver diseases, including liver parasitosis, acute liver injury, autoimmune liver disease, and hepatocellular carcinoma. Furthermore, eosinophils have demonstrated the capacity to promote liver regeneration. This article explores the multifaceted roles of eosinophils in liver diseases, aiming to provide insights that could lead to more effective clinical therapies for these conditions.
{"title":"The Role of Eosinophils in Liver Disease","authors":"Linxi Xie , Hejiao Zhang , Long Xu","doi":"10.1016/j.jcmgh.2024.101413","DOIUrl":"10.1016/j.jcmgh.2024.101413","url":null,"abstract":"<div><div>Previously, eosinophils were primarily regarded as effector toxic cells involved in allergic diseases and parasitic infections. Nevertheless, new research has shown that eosinophils are diverse and essential for immune regulation and tissue homeostasis. Their functional plasticity has been observed in patients with inflammatory diseases, cancer, infections, and other disorders. Although eosinophils are infrequently observed within the liver during periods of homeostasis, they are recruited to the liver in various liver diseases, including liver parasitosis, acute liver injury, autoimmune liver disease, and hepatocellular carcinoma. Furthermore, eosinophils have demonstrated the capacity to promote liver regeneration. This article explores the multifaceted roles of eosinophils in liver diseases, aiming to provide insights that could lead to more effective clinical therapies for these conditions.</div></div>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 2","pages":"Article 101413"},"PeriodicalIF":7.1,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11719855/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142333077","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-01DOI: 10.1016/j.jcmgh.2024.101421
Elias Broeckhoven, Kai Dallmeier
{"title":"Mouse Models for Chronic Hepatitis B: Old Challenges, Novel Approaches","authors":"Elias Broeckhoven, Kai Dallmeier","doi":"10.1016/j.jcmgh.2024.101421","DOIUrl":"10.1016/j.jcmgh.2024.101421","url":null,"abstract":"","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 1","pages":"Article 101421"},"PeriodicalIF":7.1,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142565402","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-01DOI: 10.1016/j.jcmgh.2024.101440
Hui Leng, Theo Thijs, Louis Desmet, Guillaume Vanotti, Mona Farhadipour, Inge Depoortere
Background & Aims
Circadian disturbances result in adverse health effects, including gastrointestinal symptoms. We investigated which physiological pathways in jejunal mucosa were disrupted during chronic jetlag and prevented during time-restricted feeding (TRF). Enteroids from Bmal1+/+ and Bmal1-/- mice were used to replicate the processes that were affected by chronic jetlag and rescued by TRF.
Methods
C57BL/6J male mice were subjected to chronic jetlag or night-TRF for 4 weeks. An around-the-clock bulk-RNA sequencing study was performed on the jejunal mucosa. Bmal1+/+ and Bmal1-/- mouse enteroids were generated to study the jejunal epithelial clock dependency of rhythmic jejunal processes.
Results
Chronic jetlag disrupted the rhythmicity of jejunal clock genes and the jejunal transcriptome, which was partially rescued by TRF. Genes whose rhythm was altered by chronic jetlag but prevented by TRF were primarily associated with nutrient transport, lipid metabolism, ketogenesis, and cellular organization. In vivo, chronic jetlag caused a phase shift in the rhythmic accumulation of neutral lipids and induced a diurnal rhythm in the number of crypt epithelial cells, both of which were prevented by TRF. In vitro, enteroids replicated the in vivo rhythmic accumulation of neutral lipids in a clock-dependent manner, whereas the rhythm of S phase proliferation was ultradian in both genotypes of enteroids.
Conclusions
This pioneering transcriptomic study demonstrates that TRF acts as a robust entrainer during chronic jetlag, realigning disturbances in the circadian clock and the transcriptome involved in metabolic functions in the jejunal mucosa. Enteroids can replicate the rhythmic accumulation of neutral lipids dependent on the jejunal epithelial clock, enabling these functions to be studied in vitro.
{"title":"Time-Restricted Feeding Reinforces Gut Rhythmicity by Restoring Rhythms in Intestinal Metabolism in a Jetlag Mouse Model","authors":"Hui Leng, Theo Thijs, Louis Desmet, Guillaume Vanotti, Mona Farhadipour, Inge Depoortere","doi":"10.1016/j.jcmgh.2024.101440","DOIUrl":"10.1016/j.jcmgh.2024.101440","url":null,"abstract":"<div><h3>Background & Aims</h3><div>Circadian disturbances result in adverse health effects, including gastrointestinal symptoms. We investigated which physiological pathways in jejunal mucosa were disrupted during chronic jetlag and prevented during time-restricted feeding (TRF). Enteroids from <em>Bmal1</em><sup>+/+</sup> and <em>Bmal1</em><sup>-/-</sup> mice were used to replicate the processes that were affected by chronic jetlag and rescued by TRF.</div></div><div><h3>Methods</h3><div>C57BL/6J male mice were subjected to chronic jetlag or night-TRF for 4 weeks. An around-the-clock bulk-RNA sequencing study was performed on the jejunal mucosa. <em>Bmal1</em><sup>+/+</sup> and <em>Bmal1</em><sup>-/-</sup> mouse enteroids were generated to study the jejunal epithelial clock dependency of rhythmic jejunal processes.</div></div><div><h3>Results</h3><div>Chronic jetlag disrupted the rhythmicity of jejunal clock genes and the jejunal transcriptome, which was partially rescued by TRF. Genes whose rhythm was altered by chronic jetlag but prevented by TRF were primarily associated with nutrient transport, lipid metabolism, ketogenesis, and cellular organization. In vivo, chronic jetlag caused a phase shift in the rhythmic accumulation of neutral lipids and induced a diurnal rhythm in the number of crypt epithelial cells, both of which were prevented by TRF. In vitro, enteroids replicated the in vivo rhythmic accumulation of neutral lipids in a clock-dependent manner, whereas the rhythm of S phase proliferation was ultradian in both genotypes of enteroids.</div></div><div><h3>Conclusions</h3><div>This pioneering transcriptomic study demonstrates that TRF acts as a robust entrainer during chronic jetlag, realigning disturbances in the circadian clock and the transcriptome involved in metabolic functions in the jejunal mucosa. Enteroids can replicate the rhythmic accumulation of neutral lipids dependent on the jejunal epithelial clock, enabling these functions to be studied in vitro.</div></div>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 4","pages":"Article 101440"},"PeriodicalIF":7.1,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142820351","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Hepatitis B virus (HBV)-DNA integration into the host genome contributes to hepatocellular carcinoma (HCC) development. KMT2B is the second most frequent locus of HBV-DNA integration in HCC; however, its role and function remain unclear. We aimed to clarify the impact of HBV-KMT2B integration in HCC development using a human genome-edited induced pluripotent stem cell (iPSCs) model.
Methods
Based on the genetic information on HBV-KMT2B integration in HCC, we determined its complete DNA sequence and transcript variants. To exclude the effect of other oncogenic mutations, we reproduced HBV integration in healthy donor iPSCs with an intact genome and analyzed its effects using iPSC-derived hepatic progenitor cells (HPCs) and hepatocytes (iPS-Heps).
Results
The reproduced HBV-KMT2B integration significantly upregulated the proliferation of hepatic cells. Comprehensive transcriptional and epigenetic analyses revealed enhanced expression of cell cycle-related genes in hepatic cells with HBV-KMT2B integration based on perturbation of histone 3 lysine 4 tri-methylation (H3K4me3), mimicking that in the original HCC sample. Long-read RNA-sequence detected the common KMT2B transcript variants in the HCC sample and HPCs. Overexpression of the truncated variant significantly enhanced proliferation of hepatic cells, whereas HBV-KMT2B fusion transcripts did not enhance proliferation. HBV-KMT2B-integrated HPCs exhibited replication stress and DNA damage, indicating that our model initiated the process of hepatocarcinogenesis due to abnormally promoted KMT2B function.
Conclusions
Our disease model using genetically engineered iPSCs provides the first insight into both the KMT2B function in HCC development and the oncogenic processes by HBV-KMT2B integration. We clarified the novel oncogenic mechanism in HBV-related HCC due to aberrant KMT2B function.
{"title":"Hepatitis B Virus-KMT2B Integration Drives Hepatic Oncogenic Processes in a Human Gene-edited Induced Pluripotent Stem Cells-derived Model","authors":"Jun Tsuchiya , Masato Miyoshi , Sei Kakinuma , Fukiko Kawai-Kitahata , Akihide Kamiya , Taro Shimizu , Ayako Sato , Keiya Watakabe , Tomohiro Mochida , Kento Inada , Rion Kamimae , Shun Kaneko , Miyako Murakawa , Sayuri Nitta , Mina Nakagawa , Mamoru Watanabe , Yasuhiro Asahina , Ryuichi Okamoto","doi":"10.1016/j.jcmgh.2024.101422","DOIUrl":"10.1016/j.jcmgh.2024.101422","url":null,"abstract":"<div><h3>Background & Aims</h3><div>Hepatitis B virus (HBV)-DNA integration into the host genome contributes to hepatocellular carcinoma (HCC) development. KMT2B is the second most frequent locus of HBV-DNA integration in HCC; however, its role and function remain unclear. We aimed to clarify the impact of HBV-<em>KMT2B</em> integration in HCC development using a human genome-edited induced pluripotent stem cell (iPSCs) model.</div></div><div><h3>Methods</h3><div>Based on the genetic information on HBV-<em>KMT2B</em> integration in HCC, we determined its complete DNA sequence and transcript variants. To exclude the effect of other oncogenic mutations, we reproduced HBV integration in healthy donor iPSCs with an intact genome and analyzed its effects using iPSC-derived hepatic progenitor cells (HPCs) and hepatocytes (iPS-Heps).</div></div><div><h3>Results</h3><div>The reproduced HBV-<em>KMT2B</em> integration significantly upregulated the proliferation of hepatic cells. Comprehensive transcriptional and epigenetic analyses revealed enhanced expression of cell cycle-related genes in hepatic cells with HBV-<em>KMT2B</em> integration based on perturbation of histone 3 lysine 4 tri-methylation (H3K4me3), mimicking that in the original HCC sample. Long-read RNA-sequence detected the common <em>KMT2B</em> transcript variants in the HCC sample and HPCs. Overexpression of the truncated variant significantly enhanced proliferation of hepatic cells, whereas HBV-<em>KMT2B</em> fusion transcripts did not enhance proliferation. HBV-<em>KMT2B</em>-integrated HPCs exhibited replication stress and DNA damage, indicating that our model initiated the process of hepatocarcinogenesis due to abnormally promoted KMT2B function.</div></div><div><h3>Conclusions</h3><div>Our disease model using genetically engineered iPSCs provides the first insight into both the KMT2B function in HCC development and the oncogenic processes by HBV-<em>KMT2B</em> integration. We clarified the novel oncogenic mechanism in HBV-related HCC due to aberrant KMT2B function.</div></div>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":"19 2","pages":"Article 101422"},"PeriodicalIF":7.1,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11664014/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142481781","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Background & aims: Sora is the first-line drug for advanced hepatocellular carcinoma (HCC). However, acquired resistance to Sora treatment largely hinders its therapeutic efficacy, and the mechanisms underlying Sora resistance remain poorly understood. Here, we revealed a new mechanism by which Sora promotes the differentiation of regulatory T (Treg) cells to suppress the immune response in the HCC tumor microenvironment (TME) and induce Sora resistance.
Methods: Human liver tissues were obtained from HCC patients. Female C57BL/6J, OT-II, and Foxp3GFP mice were also used. Flow cytometry was used to analyze immune cells in TME. Flow cytometry, real-time polymerase chain reaction, and enzyme-linked immunosorbent assay were performed to evaluate Treg cell differentiation. Immunoblotting was conducted to identify relevant proteins. Mouse and human tumor tissues were evaluated via multiplex immunofluorescence staining. Sora-treated HCC tissues and Sora-treated Treg cells were subjected to RNA sequencing analysis. Tumor models were generated and treated with Sora, Sora combined with an anti-CD25 antibody, or Sora combined with the Foxo1 inhibitor AS1842856.
Results: First, we found through bioinformatic analysis that Sora suppresses the immune response in HCC. Furthermore, Sora increased the Treg cell population to promote the formation of an immunosuppressive TME in HCC. In vitro, Sora promoted Treg cell differentiation and increased the immunosuppressive activity of Treg cells. Activating VEGF and AKT abolished the effect of Sora on Treg cell differentiation, whereas inhibiting Foxo1 compromised Sora-induced Treg cell differentiation, indicating that the induction of Treg cells by Sora is dependent on the VEGFR/AKT/Foxo1 pathway. Finally, Treg inactivation by an anti-CD25 antibody or the Foxo1 inhibitor AS1842856 in combination with Sora showed greater efficacy in the treatment of HCC.
Conclusions: Sora induced Treg cell differentiation by inhibiting VEGFR/AKT signaling and activating Foxo1, thus suppressing the immune response and reducing Sora efficacy. Treg inactivation might be a promising strategy to alleviate the immunosuppressive TME and overcome Sora resistance.
{"title":"Sorafenib Promotes Treg Cell Differentiation To Compromise Its Efficacy via VEGFR/AKT/Foxo1 Signaling in Hepatocellular Carcinoma.","authors":"Yingying Shen, Hanliang Wang, Zeyu Ma, Minyan Hao, Shuowang Wang, Junwei Li, Yue Fang, Lei Yu, Yingying Huang, Changrong Wang, Jingjing Xiang, Zhijian Cai, Jianli Wang, Hongchuan Jin, Jia Zhou, Jufeng Guo, Pingting Ying, Xian Wang","doi":"10.1016/j.jcmgh.2024.101454","DOIUrl":"10.1016/j.jcmgh.2024.101454","url":null,"abstract":"<p><strong>Background & aims: </strong>Sora is the first-line drug for advanced hepatocellular carcinoma (HCC). However, acquired resistance to Sora treatment largely hinders its therapeutic efficacy, and the mechanisms underlying Sora resistance remain poorly understood. Here, we revealed a new mechanism by which Sora promotes the differentiation of regulatory T (Treg) cells to suppress the immune response in the HCC tumor microenvironment (TME) and induce Sora resistance.</p><p><strong>Methods: </strong>Human liver tissues were obtained from HCC patients. Female C57BL/6J, OT-II, and Foxp3<sup>GFP</sup> mice were also used. Flow cytometry was used to analyze immune cells in TME. Flow cytometry, real-time polymerase chain reaction, and enzyme-linked immunosorbent assay were performed to evaluate Treg cell differentiation. Immunoblotting was conducted to identify relevant proteins. Mouse and human tumor tissues were evaluated via multiplex immunofluorescence staining. Sora-treated HCC tissues and Sora-treated Treg cells were subjected to RNA sequencing analysis. Tumor models were generated and treated with Sora, Sora combined with an anti-CD25 antibody, or Sora combined with the Foxo1 inhibitor AS1842856.</p><p><strong>Results: </strong>First, we found through bioinformatic analysis that Sora suppresses the immune response in HCC. Furthermore, Sora increased the Treg cell population to promote the formation of an immunosuppressive TME in HCC. In vitro, Sora promoted Treg cell differentiation and increased the immunosuppressive activity of Treg cells. Activating VEGF and AKT abolished the effect of Sora on Treg cell differentiation, whereas inhibiting Foxo1 compromised Sora-induced Treg cell differentiation, indicating that the induction of Treg cells by Sora is dependent on the VEGFR/AKT/Foxo1 pathway. Finally, Treg inactivation by an anti-CD25 antibody or the Foxo1 inhibitor AS1842856 in combination with Sora showed greater efficacy in the treatment of HCC.</p><p><strong>Conclusions: </strong>Sora induced Treg cell differentiation by inhibiting VEGFR/AKT signaling and activating Foxo1, thus suppressing the immune response and reducing Sora efficacy. Treg inactivation might be a promising strategy to alleviate the immunosuppressive TME and overcome Sora resistance.</p>","PeriodicalId":55974,"journal":{"name":"Cellular and Molecular Gastroenterology and Hepatology","volume":" ","pages":"101454"},"PeriodicalIF":7.1,"publicationDate":"2024-12-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142916336","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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