Inhibition of cysteine-serine-rich nuclear protein 1 ameliorates ischemia-reperfusion injury during liver transplantation in an MAPK-dependent manner.

Hepatic ischemia-reperfusion injury (HIRI) is a critical pathophysiological process during liver transplantation (LT). Multiple genes and signal pathways are dysregulated during HIRI. This study aims to identify genes as potential therapeutic targets for ameliorating HIRI. Datasets containing samples from the human donor liver (GSE151648) and mouse HIRI model (GSE117066) were analyzed to determine differentially expressed genes (DEGs). The selected DEGs were confirmed by real-time PCR and western blot in the hepatocyte hypoxia-reoxygenation (HR) model, mouse HIRI model, and human liver samples after transplantation. Genetic inhibition was used to further clarify the underlying mechanism of the gene in vitro and in vivo. Among the DEGs, CSRNP1 was significantly upregulated (|log FC|= 2.08, P < 0.001), and was positively correlated with the MAPK signal pathway (R = 0.67, P < 0.001). CSRNP1 inhibition by siRNA significantly suppressed apoptosis in the AML-12 cell line after HR (mean Annexin+ ratio = 60.62% vs 42.47%, P = 0.0019), but the protective effect was eliminated with an additional MAPK activator. Knocking down CSRNP1 gene expression by intravenous injection of AAV-shRNA markedly reduced liver injury in mouse HIRI model (ALT: AAV-NC vs AAV-shCsrnp1 = 26,673.5 ± 2761.2 vs 3839.7 ± 1432.8, P < 0.001; AST: AAV-NC vs AAV-shCsrnp1 = 8640.5 ± 1450.3 vs 1786.8 ± 518.3, P < 0.001). Liver-targeted delivery of siRNA by nanoparticles effectively inhibited intra-hepatic genetic expression of Csrnp1 and alleviated IRI by reducing tissue inflammation and hepatocyte apoptosis. Furthermore, CSRNP1 inhibition was associated with reduced activation of the MAPK pathway both in vitro and in vivo. In conclusion, our results demonstrated that CSRNP1 could be a potential therapeutic target to ameliorate HIRI in an MAPK-dependent manner.

PAK1 inhibition increases TRIM21-induced PD-L1 degradation and enhances responses to anti-PD-1 therapy in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDA) is a common malignancy with a 5-year survival <10 %. Immunosuppressive tumor microenvironment (TME) plays a critical role in the progression of PDA. In recent years, programmed death-ligand 1 (PD-L1)/programmed cell death protein-1 (PD-1) blockade has emerged as a potent anti-tumor immunotherapy, while is yet to achieve significant clinical benefits for PDA patients. P21-Activated kinase 1 (PAK1) is highly upregulated in PDA and has been reported to be involved in the regulation of anti-tumor immunity. This study aims to investigate the combined effect of PAK1 inhibition and anti-PD-1 therapy on PDA and the underlying mechanisms. We have shown that PAK1 expression positively correlated with PD-L1 in PDA patients, and that inhibition of PAK1 downregulated PD-L1 expression of PDA cells. More importantly, we have demonstrated that PAK1 competed with PD-L1 in binding to tripartite motif-containing protein 21 (TRIM21), a ubiquitin E3 ligase, resulting in less ubiquitination and degradation of PD-L1. Moreover, PAK1 inhibition promoted CD8+ T cells activation and infiltration. In a murine PDA model, the combination of PAK1 inhibition and anti-PD-1 therapy showed significant anti-tumor effects compared with the control or monotherapy. Our results indicated that the combination of PAK1 inhibition and anti-PD-1 therapy would be a more effective treatment for PDA patients.

Incorporation of protein induced by vitamin K absence or antagonist-II into transplant criteria expands beneficiaries of liver transplantation for hepatocellular carcinoma: a multicenter retrospective cohort study in China.

INTRODUCTION: In order to maximize the utilization of precious donor liver, precisely determining potential hepatocellular carcinoma (HCC) candidates who will benefit from liver transplantation (LT) is essential. As a crucial diagnostic biomarker for HCC, protein induced by vitamin K absence or antagonist-II (PIVKA-II) has become one of the key indicators for assessing tumor recurrence risk after LT. This study aims to investigate the role of PIVKA-II in recipient selection and prognostic stratification. METHODS: The clinicopathologic data of HCC patients undergoing LT from 2015 to 2020 in six Chinese transplant centers were collected. Univariate and multivariate analyses were performed to determine risk factors for disease free survival (DFS). Based on these risk factors, survival analysis was made by Kaplan-Meier method and their value in prognostic stratification was assessed. RESULTS: A total of 522 eligible HCC patients with pre-LT PIVKA-II records were finally included in this study. Tumor burden>8 cm, α-fetoprotein>400 ng/ml, histopathologic grade III and PIVKA-II>240 mAU/ml were identified as independent risk factors for DFS. DFS of patients with PIVKA-II≤240 mAU/ml ( N =288) were significantly higher than those with PIVKA-II>240 mAU/ml ( N =234) (1-year, 3-year, and 5-year DFS: 83.2, 77.3, and 75.9% vs. 75.1, 58.5, and 50.5%; P <0.001). Compared with Hangzhou criteria ( N =305), incorporating PIVKA-II into Hangzhou criteria (including tumor burden, α-fetoprotein, and histopathologic grade) increased the number of patients with eligibility for LT by 21.6% but achieved comparable DFS and overall survival. CONCLUSIONS: Incorporating PIVKA-II into existing LT criteria could increase the number of eligible HCC patients without compromising post-LT outcomes.

Inhibition of the mitochondrial complex-1 protects against carbon tetrachloride-induced acute liver injury.

Mitochondrial dysfunction has been documented to play a crucial role in the pathogenesis of liver injury. In the present study, we investigated the role of rotenone, a mitochondrial complex-1 inhibitor, in carbon tetrachloride (CCl4) -induced acute liver injury, as well as the underlying mechanisms. Before CCl4 administration, the mice were pretreated with rotenone at a dose of 250 ppm in food for three days. Then CCl4 was administered to the mice for 16 h by intraperitoneal injection. The liver injury, mitochondrial status, oxidative stress, and inflammation were examined. Strikingly, CCl4 treatment markedly induced liver injury as shown by enhanced serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) and morphological lesions (HE stating), which was significantly attenuated by rotenone treatment in line with the reduced activity of mitochondrial complex-1. Meanwhile, oxidative stress markers of malondialdehyde (MDA), 4-hydroxynonenal (HNE), and dihydroethidium (DHE) and the inflammatory markers of IL-1ß, MCP-1, TNF-α, TLR-4, and IL-6 were also significantly suppressed by rotenone. More importantly, the mitochondrial abnormalities shown by the reduction of SOD2, mitochondrial transcription factor A (TFAM), mitochondrial NADH dehydrogenase subunit 1 (mtND1), and Cytb were significantly restored, indicating that rotenone protected against mitochondrial damage induced by CCl4 in liver. Moreover, rotenone treatment alone did not significantly alter liver morphology and liver enzymes ALT and AST. CYP2E1, a metabolic enzyme of CCl4, was also not significantly affected by rotenone. In conclusion, rotenone protected the liver from CCl4-induced damage possibly by inhibiting the mitochondrial oxidative stress and inflammation.