Biology subjects

Sanyal, A. J.

Publications and source records attributed to Sanyal, A. J..

4 recordsLinked to original sources

bioRxiv2026

Cholangiocyte RUNX1 Orchestrates Fibrogenic and Inflammatory Signaling to Drive Biliary Fibrosis

IntroductionBiliary fibrosis and inflammation are central to the pathogenesis of cholangiopathies such as primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC). Inflammatory and fibrogenic stimuli, such as transforming growth factor-{beta} (TGF{beta}) and lipopolysaccharide (LPS) signaling, drive these processes, but their underlying transcriptional mechanisms in cholangiocytes remain incompletely defined. We investigated the role of Runt-related transcription factor 1 (RUNX1) as a transcriptional co-regulator of fibroinflammatory signaling in cholangiocytes. MethodsHuman PSC-derived cholangiocytes (PSC-Cs) and mouse large biliary epithelial cells (MLEs) were subjected to RUNX1 knockdown or pharmacologic inhibition (Ro5-3335 or AI-10-104). Cytokine secretion was profiled by Luminex multiplexing; RUNX1 genomic binding and protein interactome were assessed by ChIP-qPCR, ChIP-seq, and LC-MS/MS. In vivo, Mdr2-/- mice received Ro5-3335, and cholangiocyte-selective Runx1 knockout mice (Krt19-CreERT) were challenged with a DDC diet, followed by evaluation of fibrosis and inflammation. ResultsRUNX1 expression was significantly increased in cholangiocytes from PSC and PBC patients, and Mdr2-/- mice. RUNX1 knockdown or inhibition reduced IL6, TNF, and other proinflammatory cytokines in PSC-Cs and attenuated TGF{beta}-, LPS-, and TNF-induced Il6 and Ccl2 expression in MLEs. ChIP-qPCR and ChIP-seq revealed TGF{beta}-induced RUNX1 binding to the Il6 promoter and 727 additional genomic sites enriched for fibrosis and inflammatory pathways; predicted upstream regulators included TGF{beta}, TNF, and NF{kappa}B signaling. Proteomic analysis identified TGF{beta}-induced RUNX1 interactions with SMAD2 and NF{kappa}B2. In vivo, Ro5-3335 treatment in Mdr2-/- mice reduced hepatic collagen, ECM gene expression, immune cell infiltration, and serum liver injury markers and bile acids. Similarly, cholangiocyte-specific Runx1 deletion mitigated fibrosis, inflammation, and liver injury in DDC-fed mice. ConclusionRUNX1 is a central transcriptional hub integrating TGF{beta} and inflammatory signals in cholangiocytes. Its inhibition attenuates biliary fibrosis and inflammation in cholestatic models, supporting RUNX1 as a potential therapeutic target in fibroinflammatory cholangiopathies.

physiology↗

bioRxiv2025

Hepatic isomiR landscaping reveals new biological insights into metabolic dysfunction in steatotic liver disease

Post-transcriptionally modified microRNA (miRNA), called isomiRs, expand the repertoire of transcripts that can leveraged for therapeutic targets and biological insights. However, the expression of isomiRs has not been characterized in metabolic dysfunction-associated steatotic liver disease (MASLD). Therefore, we assessed the isomiR expression profile in liver biopsies from 79 patients with MASLD and modeled their potential role in disease biology. MiRNAs represented 75% of the sequencing reads and over 65% of them were attributed to isomiRs, demonstrating their higher expression and diversity compared to canonically annotated miRNAs. Differential expression and machine-learning analyses were used to identify 173 isomiRs associated to MASLD severity and 58 isomiRs associated to fibrosis score. Candidate target mRNAs were identified for each isomiR based on sequence complementarity. Using matched mRNA sequencing data, and supported by data from an independent study, we proposed key dysregulated mRNA targets involved in a selection of 33 disease-associated pathways. Importantly, isomiRs offered novel and unique mRNA targets compared to the canonical miRNA, e.g. isomiR-122 targeting INSIG1 (insulin and cholesterol metabolism), and isomiR-21 targeting HMGCS2 and PPARA (PARR and TGF-beta signaling). Our work advances knowledge regarding the role of isomiRs in MASLD and lays a foundation for therapeutic targets identification. HighlightsOur results provide a comprehensive analysis of microRNA (isomiRs) in liver tissue. Machine learning identified isomiRs whose expression is associated with MASLD. Multi-omic analysis uncovered novel isomiR regulatory mechanisms involved in MASLD.

bioinformatics↗

bioRxiv2024

Aramchol attenuates fibrosis in mouse models of biliary fibrosis and blocks the TGFβ-induced fibroinflammatory mediators in cholangiocytes

BackgroundFibroinflammatory cholangiopathies, such as primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC), are characterized by inflammation and biliary fibrosis, driving disease-related complications. In biliary fibrosis, cholangiocytes activated by transforming growth factor-{beta} (TGF{beta}) release signals that recruit immune cells to drive inflammation and activate hepatic myofibroblasts to deposit the extracellular matrix (ECM). TGF{beta} regulates stearoyl-CoA desaturase (SCD), an enzyme that catalyzes the synthesis of monounsaturated fatty acids, in stimulating fibroinflammatory lipid signaling. However, the role of SCD or its inhibitor, Aramchol, has not been investigated in biliary fibrosis or TGF{beta}-mediated cholangiocyte activation. Method10-16-week-old multi-drug resistance 2 knockout (Mdr2-/-) and 3,5-diethoxycarboncyl-1,4-dihydrocollidine (DDC) diet-fed mice were orally gavaged daily with Aramchol at 12.5 mg/kg/day for 4 and 3 weeks, respectively. Liver and serum were harvested for the assessment of fibrosis and inflammation. Transformed human cholangiocyte cells (H69) and mouse large biliary epithelial cells (MLEs) were used to test the effects of the SCD inhibitor, Aramchol, at varying doses on TGF{beta}-mediated expression of fibroinflammatory signals and were confirmed in PSC-derived cholangiocytes (PSC-Cs) using ELISA, qPCR, and Western blot analyses. ResultsAramchol treatment of Mdr2-/- mice with established biliary fibrosis (treatment) and DDC diet-induced (prevention) models of cholestatic injury and fibrosis demonstrated significant reductions in both measures of ECM synthesis (mRNA expression of ECM components in the liver), collagen content of the liver (picrosirius red staining and hydroxyproline content) and myofibroblast activation (SMA staining). Il6 and Tnfa were also reduced with Aramchol in the liver. RNA-seq analysis of H69 cells showed that Aramchol co-treatment led to significant inhibition of TGF{beta}-induced hepatic fibrosis pathways while upregulating peroxisome proliferator-activated receptor (PPAR) signaling. SCD expression was significantly increased in TGF{beta}-treated H69 cells (2-fold, p<0.05). Aramchol in a dose-dependent manner significantly attenuated the increased expression of the fibrotic marker, plasminogen activator inhibitor-1 (PAI-1/SERPINE1), and hepatic stellate cell-activating genes (VEGFA and PDGFB) in TGF{beta}-activated H69 and MLEs. Aramchol also markedly reduced the expression of the inflammatory cytokine, interleukin 6 (IL6). SCD siRNA knockdown produced similar results in H69 cells. Furthermore, in PSC-Cs, the expressions of SCD, VEGFA and IL6 were significantly reduced with Aramchol. The expression of the anti-fibroinflammatory factors PPAR and -{gamma} were modestly increased in cholangiocyte cell lines with increased expression of PPAR-responsive genes and increased nuclear binding of DNA PPAR response elements with Aramchol co-treatment compared to TGF{beta} only. ConclusionAramchol, an SCD inhibitor, both attenuates and prevents biliary fibrosis in mouse models of cholestatic injury and fibrosis. This effect is partially due to Aramchol inhibiting TGF{beta}-induced fibroinflammatory mediators in cholangiocytes by upregulating PPAR and -{gamma} expression and activity. These findings, along with Aramchols excellent safety profile in clinical trials, provide the rationale for assessing Aramchol in further clinical studies in patients with biliary fibrosis, particularly PSC, where a treatment is desperately needed.

cell biology↗

bioRxiv2024

Effective anti-tumor immune response against HCC is orchestrated by immune cell partnership network that functions through hepatic homeostatic pathways, not direct cytotoxicity

BackgroundMetabolic dysfunction-associated fatty liver disease (MAFLD) can progress to hepatocellular carcinoma (HCC), yet the immune mechanisms driving this transition remain unclear. MethodsIn a chronic Western diet (WD) mouse model, we performed single-nuclei RNA sequencing to track MAFLD progression into HCC and subsequent tumor inhibition upon dietary correction. ResultsCarcinogenesis begins during MAFLD, with tumor cells entering dormancy when HCC is mitigated. Rather than purely tolerogenic, the liver actively engages immune responses targeting myofibroblasts, fibroblasts and hepatocytes to maintain tissue homeostasis. Cytotoxic cells contribute to turnover of liver cells but do not primarily target the tumor. NKT cells predominate under chronic WD, while monocytes join them in HCC progression on a WD. Upon dietary correction, monocyte-driven immunity confers protection against HCC through targeting tissue homeostatic pathways and antioxidant mechanisms. Crucially, liver tissue response--not merely immune activation--dictates whether tumors grow or regress, emphasizing the importance of restoring liver tissue integrity. Also, protection against HCC is linked to a distinct immunological pattern, differing from healthy controls, underscoring the need for immune reprogramming. ConclusionThese findings reveal the dual roles of similar pathways, where immune patterns targeting different cells shape distinct outcomes. Restoring tissue homeostasis and regeneration creates a tumor-hostile microenvironment, whereas tumor-directed approaches fail to remodel the TME. This underscores the need for tissue remodeling strategies in cancer prevention and treatment. Lay summaryOur study challenges the traditional view that the liver is purely tolerant to immune responses, revealing that it actively regulates immunity to maintain tissue health. We found that liver cancer (HCC) begins during fatty liver disease (MAFLD) but can be halted if immune cells--especially monocytes--restore tissue integrity. Instead of focusing solely on killing tumors, effective immunotherapy should harness the bodys natural ability to repair the liver, creating an environment where cancer cannot thrive. This discovery paves the way for innovative treatments that promote immune-driven tissue regeneration as a strategy for cancer prevention and therapy.

immunology↗