Interaction with YAP underlies the species differences between humans and rodents in CAR-dependent hepatocyte proliferation.

Constitutive androstane receptor (CAR), a nuclear receptor predominantly expressed in the liver, is activated by diverse chemicals and induces hepatocyte proliferation and hepatocarcinogenesis in rodents. However, the underlying mechanism responsible for CAR-dependent hepatocyte proliferation remains unclear. Importantly, this phenomenon has not been observed in the human liver. This study aimed to investigate the molecular mechanism underlying CAR-induced hepatocyte proliferation and to explore the species differences in hepatocyte proliferation between humans and rodents. Treatment of mice with the CAR activator TCPOBOP induced hepatocyte proliferation and nuclear accumulation of yes-associated protein (YAP), a known liver cancer inducer. This induction was abolished in CAR-knockout mice. Exogenously expressed YAP in cultured cells was accumulated in the nucleus by the coexpression with mouse CAR but not human CAR. Pull-down analysis of recombinant proteins revealed that mouse CAR interacted with YAP, whereas human CAR did not. Further investigations using YAP deletion mutants identified the WW domain of YAP as essential for interacting with CAR and showed that the PY motif (PPAY) in mouse CAR was crucial for binding to the WW domain, whereas human CAR with its mutated motif (PPAH) failed to interact with YAP. A mouse model harboring the Y150H mutation (PPAY to PPAH) in CAR displayed drastically attenuated TCPOBOP-induced hepatocyte proliferation and nuclear accumulation of YAP. CAR induces the nuclear accumulation of YAP through the PY motif-WW domain interaction to promote hepatocyte proliferation. The absence of this interaction in human CAR contributes to the lack of CAR-dependent hepatocyte proliferation in human livers.

Vancomycin induces reactive oxygen species-dependent apoptosis via mitochondrial cardiolipin peroxidation in renal tubular epithelial cells.

Vancomycin (VCM) is a first-line antibiotic for serious infections caused by methicillin-resistant Staphylococcus aureus. However, nephrotoxicity is one of the most complaint in VCM therapy. We previously reported that VCM induced apoptosis in a porcine proximal tubular epithelial cell line (LLC-PK1), in which mitochondrial complex I may generate superoxide, leading to cell death. In the present study, VCM caused production of mitochondrial reactive oxygen species and peroxidation of the mitochondrial phospholipid cardiolipin that was reversed by administration of the mitochondrial uncoupler carbonyl cyanide-4-(trifluoromethoxy) phenylhydrazone (FCCP). FCCP also significantly suppressed VCM-induced depolarization of the mitochondrial membrane and apoptosis. Moreover, the lipophilic antioxidant vitamin E and a mitochondria-targeted antioxidant, mitoTEMPO, also significantly suppressed VCM-induced depolarization of mitochondrial membrane and apoptosis, whereas vitamin C, n-acetyl cysteine, or glutathione did not provide significant protection. These findings suggest that peroxidation of the mitochondrial membrane cardiolipin mediated the VCM-induced production of intracellular reactive oxygen species and initiation of apoptosis in LLC-PK1 cells. Furthermore, regulation of mitochondrial function using a mitochondria-targeted antioxidant, such as mitoTEMPO, may constitute a potential strategy for mitigation of VCM-induced proximal tubular epithelial cell injury.