Abnormal expression of serum miR-4746-5p in liver cancer patients after interventional chemotherapy and its possible mechanism.

Interventional chemotherapy is a common operation in the clinical treatment of liver cancer. The aim of this study was to investigate the expression and molecular mechanism of serum miR-4746-5p in liver cancer patients before and after interventional chemotherapy. The levels of miR-4746-5p and CDKN1C in serum samples from liver cancer patients were detected using real-time fluorescence quantitative polymerase chain reaction. Receiver operating characteristic curves revealed the diagnostic value of miR-4746-5p in tumors. Differences in clinical indicators between liver cancer patients and healthy controls were assessed using Pearson correlation analysis. Luciferase reporter gene assays confirmed the targeted interaction between miR-4746-5p and CDKN1C. In vitro cellular assays were validated by Cell Counting Kit-8, Transwell assay, and chemoresistance assay. Serum miR-4746-5p levels were increased in liver cancer patients but were downregulated after chemotherapy intervention. CDKN1C expression showed the opposite trend. Low levels of miR-4746-5p mediated cell growth and metastasis by targeting and negatively regulating CDKN1C expression, while silencing CDKN1C restored cell activity. Inhibition of miR-4746-5p reduced chemoresistance, while downregulation of CDKN1C affected cell sensitivity. miR-4746-5p may be a potential therapeutic factor for liver cancer diagnosis and interventional chemotherapy.

Upregulation of lncRNA PTOV1-AS1 in hepatocellular carcinoma contributes to disease progression and sorafenib resistance through regulating miR-505.

Increasing evidence has displayed the vital influence of lncRNA in tumorigenesis and chemoresistance of cancer treatment. This study investigated the function of lncRNA PTOV1-AS1 in hepatocellular carcinoma (HCC) and its role in sorafenib resistance. The relative expression of lncRNA and miRNA was measured by RT-qPCR. The cellular activities including cell proliferation and invasion were explored by CCK-8 and Transwell assays. Bioinformatics analysis and dual-luciferase reporter assay were used to predict the targeting miRNA of PTOV1-AS1. The expression levels of PTOV1-AS1 were higher in HCC tissues than that in the normal tissues and associated with patients' overall survival. Knockdown of PTOV1-AS1 decreased cell proliferation rate and invasion number. After treatment with different concentrations of sorafenib, the sorafenib-resistant hepatoma cells were conducted. PTOV1-AS1 expression levels were increased in HepG2-SR and Huh7-SR cells. PTOV1-AS1 knockdown repressed the proliferation, invasion, and drug resistance of sorafenib-resistant HCC cells by targeting the expression of miR-505. In conclusion, the expression of PTOV1-AS1 is increased in HCC and sorafenib-resistance HCC cells, as well as is associated with patients' prognosis. Inhibition of PTOV1-AS1 expression can reduce the resistance of sorafenib-resistant HCC cells, which may play a role by targeting the negative regulation of miR-505 expression.

Toripalimab in combination with Anlotinib for unresectable hepatocellular carcinoma after SBRT: A prospective, single-arm, single-center clinical study.

Objective: Exposing tumor antigens to the immune system is the key to ensuring the efficacy of immunotherapy. SBRT is the main way to reveal the specifical antigens of tumors which can enhance the immune response. We aimed to explore the clinical efficacy and safety of Toripalimab combined with Anlotinib for uHCC after SBRT. Methods: This is a prospective, single-arm, explorative clinical study. uHCC patients with an ECOG PS score of 0-1, Child-Pugh class A or B, and BCLC stage B or C were included and treated with SBRT(8Gy*3) followed by 6-cycle combinational therapy with Toripalimab and Anlotinib. The primary endpoint was progression-free survival (PFS) and the secondary endpoints were objective response rate (ORR), disease control rate (DCR), overall survival (OS), and incidence of treatment-related adverse events (TRAEs). Continuous variables were presented as medians and ranges. Survivals were studied with the Kaplan-Meier method. Categorical data were expressed as n (percentage). Results: Between June 2020 and October 2022, a total of 20 patients with intermediate-advanced uHCC were enrolled. All cases had multiple intrahepatic metastases, or macrovascular invasion, or both, among whom 5 cases with lymph node or distant metastases. Until September 2022, the median follow-up time was 7.2 months (range, 1.1-27.7 months). Median survival time could not be assessed at the moment, based on iRecist, median PFS was 7.4 months (range, 1.1-27.7 months), ORR 15.0%, and DCR 50.0%. 14 patients experienced treatment-related adverse events with an incidence of 70%. The overall survival rates at 18 months and 24 months were 61.1% and 50.9%, respectively. And the progression-free survival rates were 39.3% and 19.7%. Conclusion: Exposure of specific antigens of HCC via SBRT may improve the efficacy of combinational therapy with Toripalimab and Anlotinib for uHCC with manageable adverse effects, which deserves further exploration. Clinical trial registration: www.clinicaltrials.gov, identifier ChiCTR2000032533.

Upregulation of microRNA miR-652-3p is a prognostic risk factor for hepatocellular carcinoma and regulates cell proliferation, migration, and invasion.

As powerful regulatory factors, microRNAs (miRNAs) are involved in tumor progression. The current research aimed to excavate the prognostic significance and potential regulatory mechanisms of miR-652-3p in hepatocellular carcinoma (HCC). Expression of miR-652-3p in HCC tissues and cells was exposed by Quantitative real-time polymerase chain reaction (RT-qPCR) assay, and we found that miR-652-3p was elevated in HCC tissues and cells than in the control group (P < 0.05). Then, the relationship between miR-652-3p levels and clinical characteristics was obtained from the Chi-square test. Kaplan-Meier survival analysis and Cox regression model to explore the outcome of miR-652-3p on the prognosis of HCC. The results investigated that overexpression of miR-652-3p was related to clinical tumor-node-metastasis (TNM) stage (P = 0.020) and differentiation (P = 0.031). HCC patients with elevated miR-652-3p levels were correlated with poor overall survival (log-rank, P = 0.007), and maybe a possible prognostic marker for HCC. Finally, CCK-8, colony formation, wound healing and Transwell assay was detected after transfection of HCC cells with miR-652-3p mimic or inhibitor. And the results confirmed that elevation miR-652-3p promoted the proliferation, migration, and invasion of tumor cells (P < 0.05). All data indicated that elevated miR-652-3p is a prognostic marker and would be able to participate in tumor progression of HCC by regulating cell proliferation, migration, and invasion.

microR-505/heterogeneous nuclear ribonucleoprotein M inhibits hepatocellular carcinoma cell proliferation and induces cell apoptosis through the Wnt/ß-catenin signaling pathway.

Aim: This study aimed to investigate the expression of microRNA-505 (miR-505) and explore its clinical significance, biological function and mechanisms in hepatocellular carcinoma (HCC). Methods: Expression of miR-505 was measured in 128 paired HCC tissues and five cell lines by quantitative real-time polymerase chain reaction (qRT-PCR). MTT assay, Transwell migration, invasion assays and apoptosis assay were performed to explore the functional role of miR-505. The target gene of miR-505 was assessed using the bioinformatics assay and the related signaling pathway was confirmed using western blot. Results: Expression of miR-505 in HCC serum and tissues were downregulated. The overexpression of miR-505 in HCC cells inhibited cell proliferation and metastasis, as well as enhanced cell apoptosis by directly downregulating heterogeneous nuclear ribonucleoprotein M (HNRNPM). The activity of the Wnt/ß-catenin signaling pathway was suppressed by the overexpression of miR-505 but was promoted by the upregulation of HNRNPM. Conclusion: The results suggest that the regulation of miR-505/HNRNPM may be a novel strategy to improve the targeted therapy of HCC.

Glycochenodeoxycholic acid impairs transcription factor E3 -dependent autophagy-lysosome machinery by disrupting reactive oxygen species homeostasis in L02 cells.

Cholestasis represents pathophysiologic syndromes defined as impaired bile flow from the liver. As an outcome, bile acids accumulate and promote hepatocyte injury, followed by liver cirrhosis and liver failure. Glycochenodeoxycholic acid (GCDCA) is relatively toxic and highly concentrated in bile and serum after cholestasis. However, the mechanism underlying GCDCA-induced hepatotoxicity remains unclear. In this study, we found that GCDCA inhibits autophagosome formation and impairs lysosomal function by inhibiting lysosomal proteolysis and increasing lysosomal pH, contributing to defects in autophagic clearance and subsequently leading to the death of L02 human hepatocyte cells. Notably, through tandem mass tag (TMT)-based quantitative proteomic analysis and database searches, 313 differentially expressed proteins were identified, of which 71 were increased and 242 were decreased in the GCDCA group compared with those in the control group. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that GCDCA suppressed the signaling pathway of transcription factor E3 (TFE3), which was the most closely associated with autophagic flux impairment. In contrast, GCDCA-inhibited lysosomal function and autophagic flux were efficiently attenuated by TFE3 overexpression. Specifically, the decreased expression of TFE3 was closely related to the disruption of reactive oxygen species (ROS) homeostasis, which could be prevented by inhibiting intracellular ROS with N-acetyl cysteine (NAC). In summary, our study is the first to demonstrate that manipulation of ROS/TFE3 signaling may be a therapeutic approach for antagonizing GCDCA-induced hepatotoxicity.

Suppression of microRNA­27a protects against liver ischemia/reperfusion injury by targeting PPARγ and inhibiting endoplasmic reticulum stress.

Liver ischemia­reperfusion (I/R) injury is an important clinical issue related to liver transplantation. Recent studies suggest that microRNAs are implicated in various biological and pathological processes, including liver I/R injury. This study aimed to investigate the role and potential mechanism of miR­27a during liver I/R injury. A liver I/R model was induced via 60 min of ischemia and reperfusion for 6 h in rats. Cells were transfected with miR­27a mimics or the miR­27a inhibitor to examine the effect of miR­27a on liver I/R. Apoptotic cells were detected by flow cytometry and TUNEL staining. The expression of miR­27a was measured by real­time PCR. The expression of peroxisome proliferator­activated receptor γ (PPARγ); gastrin­releasing peptide 78 (GRP78) and C/EBP homologous protein (CHOP) were detected by western blot analysis. The results showed that miR­27a was significantly upregulated during I/R injury in vivo and in vitro. In addition, miR­27a inhibitors attenuated hypoxia/reoxygenation (H/R)­induced oxidative stress, endoplasmic reticulum stress (ERS) and apoptosis in AML12 cells. By contrast, miR­27a mimics promoted hypoxia/reoxygenation­induced ERS, and apoptosis. Furthermore, PPARγ was identified as a target gene of miR­27a using bioinformatic analysis and a dual­luciferase reporter assay. Knockdown of PPARγ significantly abrogated the inhibitory effect of miR­27a inhibitors on the ERS pathway. Moreover, the miR­27a antagomir attenuated liver I/R injury in rats, a finding manifested by reduced ALT/AST, hepatocyte apoptosis, oxidative stress and inhibition of the ERS pathway. Taken together, these findings demonstrate that suppression of miR­27a protects against liver I/R injury by targeting PPARγ and by inhibiting the ERS pathway.