ABSTRACT
To explore the effects of ß-Sitosterol upon hepatocellular carcinoma cell proliferation, apoptosis, migration, invasion, and epithelial-mesenchymal transition (EMT), and to investigate the underlying mechanism using network pharmacology. Human hepatocellular carcinoma cell lines (Huh-7 and HCCLM3) were expose to gradient concentrations of ß-Sitosterol (5 µg/mL, 10 µg/mL, and 20 µg/mL). Cell viability and proliferation were assessed using MTT, CCK-8, colony formation, and EdU assays.Flow cytometry was employed to evaluate cell cycle and apoptosis. Scratch and Transwell assays were performed, respectively, to detect cell migration and invasion. The levels of apoptosis-associated proteins (BAX, BCL2, and cleaved caspase3) as well as EMT-associated proteins (E-cadherin, N-cadherin, Snail, and Vimentin) were detected in Huh-7 and HCCLM3 cell lines using Western blot analysis. The drug target gene for ß-Sitosterol was screened via PubChem and subsequently evaluated for expression in the GSE112790 dataset. In addition, the expression level of glycogen synthase kinase 3 beta (GSK3B) within the Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) database was analyzed, along with its correlation to the survival outcomes of patients with hepatocellular carcinoma. The diagnostic efficiency of GSK3B was assessed by analyzing the ROC curve. Subsequently, Huh-7 and HCCLM3 cell lines were transfected with the overexpression vector of GSK3B and then treated with ß-Sitosterol to further validate the association between GSK3B and ß-Sitosterol. GSK3B demonstrated a significantly elevated expression in patients with hepatocellular carcinoma, which could predict hepatocellular carcinoma patients' impaired prognosis based on GEO dataset and TCGA database. GSK3B inhibitor (CHIR-98014) notably inhibited cell proliferation and invasion, promoted cell apoptosis and cell cycle arrest at G0/G1 phase in hepatocellular carcinoma cells. ß-Sitosterol treatment further promoted the efffects of GSK3B inhibitor on hepatocellular carcinoma cells. GSK3B overexpression has been found to enhance the proliferative and invasive capabilities of hepatocellular carcinoma cells. Furthermore it has been observed that GSK3B overexpression, it has been obsear can partially reverse the inhibitory effect of ß-Sitosterol upon hepatocellular. ß-Sitosterol suppressed hepatocellular carcinoma cell proliferation and invasion, and enhanced apoptosis via inhibiting GSK3B expression.
Subject(s)
Apoptosis , Carcinoma, Hepatocellular , Cell Proliferation , Epithelial-Mesenchymal Transition , Glycogen Synthase Kinase 3 beta , Liver Neoplasms , Sitosterols , Humans , Sitosterols/pharmacology , Glycogen Synthase Kinase 3 beta/metabolism , Glycogen Synthase Kinase 3 beta/genetics , Carcinoma, Hepatocellular/pathology , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/drug therapy , Carcinoma, Hepatocellular/metabolism , Liver Neoplasms/pathology , Liver Neoplasms/genetics , Liver Neoplasms/drug therapy , Liver Neoplasms/metabolism , Cell Proliferation/drug effects , Cell Proliferation/genetics , Apoptosis/drug effects , Apoptosis/genetics , Cell Line, Tumor , Epithelial-Mesenchymal Transition/drug effects , Epithelial-Mesenchymal Transition/genetics , Cell Movement/drug effects , Cell Movement/genetics , Gene Expression/genetics , Gene Expression/drug effects , Phenotype , Neoplasm Invasiveness/genetics , Cell Survival/drug effects , Cell Survival/genetics , Network Pharmacology , Gene Expression Regulation, Neoplastic/drug effectsABSTRACT
Plant-derived exosome-like nanoparticles (PELNs) are natural nanocarriers and effective delivery systems for plant microRNAs (miRNAs). These PELN-carrying plant miRNAs can regulate mammalian genes across species, thereby increasing the diversity of miRNAs in mammals and exerting multi-target effects that play a crucial role in diseases, particularly cancer. PELNs demonstrate exceptional stability, biocompatibility, and targeting capabilities that protect and facilitate the up-take and cross-kingdom communication of plant miRNAs in mammals. Primarily ingested and absorbed within the gastrointestinal tract of mammals, PELNs preferentially act on the intestine to regulate intestinal homeostasis through functional miRNA activity. The oncogenesis and progression of cancer are closely associated with disruptions in intestinal barriers, ecological imbalances, as well as secondary changes, such as abnormal inflammatory reactions caused by them. Therefore, it is imperative to investigate whether PELNs exert their anticancer effects by regulating mammalian intestinal homeostasis and inflammation. This review aims to elucidate the intrinsic crosstalk relationships and mechanisms of PELNs-mediated miRNAs in maintaining intestinal homeostasis, regulating inflammation and cancer treatment. Furthermore, serving as exceptional drug delivery systems for miRNAs molecules, PELNs offer broad prospects for future applications, including new drug research and development along with drug carrier selection within targeted drug delivery approaches for cancer therapy.
