Compound C(Shahid Beheshti University)

Previous studies highlight the critical role of AMP-activated protein kinase (AMPK) in hepatic lipid metabolism, yet natural activators for metabolic dysfunction-associated steatohepatitis (MASH) remain underexplored. This study investigated whether cichoriin from Cichorium intybus mitigates MASH fibrosis. C57BL/6J mice fed a Gubra-Amylin NASH diet for 20 weeks received cichoriin (100 or 200 mg/kg/day, oral gavage) for 6 weeks. Primary murine hepatocytes and hepatic stellate cells (HSCs) were co-cultured under fatty acid stimulation with cichoriin intervention to examine cellular mechanisms. AMPK dependency was validated using the pharmacological inhibitor Compound C. Cichoriin treatment significantly reduced hepatic steatosis, serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, and fibrotic markers while improving histological features. Mechanistically, molecular dynamics simulations indicated stable binding between cichoriin and AMPK. Cichoriin enhanced AMPK and acetyl-CoA carboxylase (ACC) phosphorylation, promoted fatty acid β-oxidation, suppressed lipogenesis, and reduced oxidative stress and pro-inflammatory cytokine production. Proteomic analysis revealed significant enrichment in AMPK-related and lipid metabolic pathways. Importantly, cichoriin decreased platelet-derived growth factor-AA (PDGF-AA) secretion by hepatocytes, thereby inhibiting HSCs activation and extracellular matrix deposition indirectly. Inhibition of AMPK via Compound C or siRNA knockdown reversed these protective effects, confirming AMPK dependency. This study demonstrates that cichoriin protects against MASH-associated liver injury through AMPK pathway activation and disruption of profibrotic hepatocyte-stellate cell signaling, supporting its development as a MASH fibrosis therapeutic.

AMP-activated protein kinase (AMPK) is a key regulator of cellular energy homeostasis that critically regulates oocyte maturation. A previous study indicates that 25 μM docosahexaenoic acid (DHA) improves energy metabolism and developmental competence during porcine oocyte in vitro maturation (IVM), demonstrating that its effects could be mediated by AMPK activation since AMPK plays a pivotal role in energy metabolism and homeostasis maintenance. This study aimed to investigate whether the effects of DHA during porcine oocyte IVM are mediated by AMPK activation. To this end, the effect of AMPK activity on oocyte maturation and developmental competence was evaluated in four experimental groups: control, DHA-treated, AMPK inhibitor (10 μM compound C; CC)-treated, and co-treatment with DHA and the inhibitor. Immunofluorescence assays showed that phosphorylated AMPK increased following DHA treatment and decreased with CC treatment during porcine oocyte IVM. Further, expression analyses of AMPK downstream target genes revealed that genes positively regulated by AMPK activity were upregulated, whereas that of the gene suppressed by AMPK activity was downregulated; CC treatment exhibited the opposite pattern. Importantly, co-treatment with DHA and CC partially restored these changes. In addition, AMPK inhibition impaired oocyte maturation, mitochondrial function, ATP content, and embryonic development, while increasing apoptosis in blastocysts. DHA mitigated these detrimental effects. Collectively, these results demonstrate that DHA effectively modulates AMPK signaling during porcine oocyte IVM, thereby improving mitochondrial function and energy metabolism and ultimately enhancing oocyte quality and developmental potential.