The roles of nuclear receptor subfamily 1 group d member 1 (NR1D1) and the circadian clock in liver fibrosis remain unclear. Here, we showed that liver clock genes, especially NR1D1, were dysregulated in mice with carbon tetrachloride (CCl₄)-induced liver fibrosis. In turn, disruption of the circadian clock exacerbated experimental liver fibrosis. NR1D1-deficient mice were more sensitive to CCl₄-induced liver fibrosis, supporting a critical role of NR1D1 in liver fibrosis development. Validation at the tissue and cellular levels showed that NR1D1 was primarily degraded by N6-methyladenosine (m⁶A) methylation in a CCl₄-induced liver fibrosis model, and this result was also validated in rhythm-disordered mouse models. In addition, the degradation of NR1D1 further inhibited the phosphorylation of dynein-related protein 1-serine site 616 (DRP1^S616), resulting in weakened mitochondrial fission function and increased mitochondrial DNA (mtDNA) release in hepatic stellate cell (HSC), which in turn activated the cGMP-AMP synthase (cGAS) pathway. Activation of the cGAS pathway induced a local inflammatory microenvironment that further stimulated liver fibrosis progression. Interestingly, in the NR1D1 overexpression model, we observed that DRP1^S616 phosphorylation was restored, and cGAS pathway was also inhibited in HSCs, resulting in improved liver fibrosis. Taken together, our results suggest that targeting NR1D1 may be an effective approach to liver fibrosis prevention and management.

Zbtb7a in alveolar macrophages influences early inflammation and autoimmunity in antibody-mediated rejection of transplanted lungs.

We investigated whether blockade of the CD47 signaling pathway could reduce ischemia‐reperfusion injury (IRI) of renal allografts donated after cardiac death (DCD) in a porcine animal model of transplantation. Renal allografts were subjected to 30 minutes of warm ischemia, 3.5 hours of cold ischemia, and then perfused with a humanized anti‐CD47 monoclonal antibody (CD47mAb) in the treatment group or HTK solution in the control group (n = 4/group). The animals were euthanized five days after transplantation. At the time of reperfusion, indocyanine green‐based in vivo imaging showed that CD47mAb‐treated organs had greater and more uniform reperfusion. On post‐transplant days 3‐5, the treatment group had lower values compared to the control for creatinine and blood urea nitrogen. Histological examination of allograft tissues showed a significant decrease of acute tubular injury in the CD47mAb‐treated group compared to control. Compared to the control group, CD47mAb treatment significantly decreased genes expression related to oxidative stress (sod‐1, gpx‐1, and txn), the inflammatory response (il‐2, il‐6, inf‐g, and tgf‐b), as well as reduced protein levels of BAX, Caspase‐3, MMP2, and MMP9. These data demonstrate that CD47mAb blockade decreases IRI and subsequent tissue injury in DCD renal allografts in a large animal transplant model.

Hepatitis B virus (HBV)/hepatitis C virus (HCV) coinfection accelerates liver fibrosis progression compared with HBV or HCV monoinfection. Octamer binding transcription factor 4 (OCT4) and Nanog are direct targets of the profibrogenic TGF-β1 signaling cascade. We leveraged a coculture model to monitor the effects of HBV and HCV coinfection on fibrogenesis in both sodium taurocholate cotransporting polypeptide–transfected Huh7.5.1 hepatoma cells and LX2 hepatic stellate cells (HSCs). We used CRISPR-Cas9 to knock out OCT4 and Nanog to evaluate their effects on HBV-, HCV-, or TGF-β1–induced liver fibrogenesis. HBV/HCV coinfection and HBx, HBV preS2, HCV Core, and HCV NS2/3 overexpression increased TGF-β1 mRNA levels in sodium taurocholate cotransporting polypeptide–Huh7.5.1 cells compared with controls. HBV/HCV coinfection further enhanced profibrogenic gene expression relative to HBV or HCV monoinfection. Coculture of HBV and HCV monoinfected or HBV/HCV coinfected hepatocytes with LX2 cells significantly increased profibrotic gene expression and LX2 cell invasion and migration. OCT4 and Nanog guide RNA independently suppressed HBV-, HCV-, HBV/HCV-, and TGF-β1–induced α-SMA, TIMP-1, and Col1A1 expression and reduced Huh7.5.1, LX2, primary hepatocyte, and primary human HSC migratory capacity. OCT4/Nanog protein expression also correlated positively with fibrosis stage in liver biopsies from patients with chronic HBV or HCV infection. In conclusion, HBV and HCV independently and cooperatively promote liver fibrogenesis through a TGF-β1–induced OCT4/Nanog-dependent pathway.

Antioxidant defence mechanisms, such as the nuclear factor-erythroid 2-related-factor-2 (NRF2) axis, are integral to oxidative stress responses and ischemic injury. Hepatic antioxidant capacity is contingent on parenchymal quality, and there is a need to develop new insights into key molecular mechanisms in marginal liver allografts that might provide therapeutic targets. This study examines the clinical relevance of NRF2 in donor livers and its response to normothermic machine perfusion (NMP). Discarded donor livers (n = 40) were stratified into a high NRF2 and low NRF2 group by quantifying NRF2 expression. High NRF2 livers had significantly lower transaminase levels, hepatic vascular inflammation and peri-portal CD3⁺ T cell infiltration. Human liver allografts (n = 8) were then exposed to 6-h of NMP and high NRF2 livers had significantly reduced liver enzyme alterations and improved lactate clearance. To investigate these findings further, we used a rat fatty-liver model, treating livers with an NRF2 agonist during NMP. Treated livers had increased NRF2 expression and reduced transaminase derangements following NMP compared to vehicle control. These results support the association of elevated NRF2 expression with improved liver function. Targeting this axis could have a rationale in future studies and NRF2 agonists may represent a supplemental treatment strategy for rescuing marginal donor livers.

It remains controversial whether renal allografts from donation after circulatory death (DCD) have a higher risk of acute rejection (AR). In the porcine large animal kidney transplant model, we investigated the AR and function of DCD renal allografts compared to the non‐DCD renal allografts and the effects of increased immunosuppression. We found that the AR was significantly increased along with elevated MHC‐I expression in the DCD transplants receiving low‐dose immunosuppression; however, AR and renal function were significantly improved when given high‐dose immunosuppressive therapy postoperatively. Also, high‐dose immunosuppression remarkably decreased the mRNA levels of ifn‐g, il‐6, tgf‐b, il‐4, and tnf‐a in the allograft at day 5 and decreased serum cytokines levels of IFN‐g and IL‐17 at day 4 and day 5 after operation. Furthermore, Western blot analysis showed that higher immunosuppression decreased phosphorylation of signal transducer and activator of transcription 3 and nuclear factor kappa‐light‐chain‐enhancer of activated B cells‐p65, increased phosphorylation of extracellular‐signal‐regulated kinase, and reduced the expression of Bcl‐2‐associated X protein and caspase‐3 in the renal allografts. These results suggest that the DCD renal allograft seems to be more vulnerable to AR; enhanced immunosuppression reduces DCD‐associated AR and improves early allograft function in a preclinical large animal model.