Regulates -Mediated Autophagy in High Glucose-Induced Vascular Smooth Muscle Cells.

Background: Vascular smooth muscle cells(VSMCs) phenotypic switching under hyperglycemic conditions accelerates atherosclerotic progression. Notch receptor 3(NOTCH3), a critical stabilizer of VSMC homeostasis implicated in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) pathogenesis, ensures vascular integrity; however, its interplay with RAN Binding Protein 1() during pathological hyperglycemia remains uncharacterized. We hypothesize that hyperglycemia-induced autophagic dysregulation is mechanistically governed by theNotch receptor 3 ()/ axis, proliferative capacity, and apoptotic signaling in high glucose (HG)-stimulated VSMCs. The aim of this study was to elucidate the regulatory mechanisms of autophagy in VSMCs under HG conditions, with a focus on the / axis and its implications for vascular health.

Methods: Bioinformatics analysis was performed on sequencing data, including weighted gene co-expression network analysis (WGCNA), screening of differentially expressed genes (DEGs), and construction of a protein-protein interaction (PPI) network, to identify the key gene, RANBP1. In vitro experiments, including cell counting kit-8 (CCK-8) assays, quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting (WB), and flow cytometry, were conducted to examine the effects of knockdown combined with overexpression on glucose-induced autophagy marker expression and cell viability in VSMCs.

Results: knockdown suppressed VSMC proliferation and induced apoptosis, and the cell cycle was stopped at the S phase. Analysis of VSMC sequencing data revealed 38 overlapping genes between the turquoise module and DEGs, 11 (, , , , , , , , , , and ) of which were significantly underexpressed in VSMC samples with si-. In a dose-dependent manner, HG therapy altered the expression of autophagy-related markers, upregulated , and downregulated phosphorylated mammalian target of rapamycin (p-mTOR). Downregulation of aggravated the effects of HG on cell viability and autophagy, whereas overexpression of reversed these effects, suggesting an offsetting effect on HG-induced autophagy.

Conclusion: On the basis of sequencing technology, bioinformatics analysis and cell experiments, we conclude that the / axis is essential for the control of autophagy and survival of VSMCs under hyperglycemic stress and could provide new insight for the clinical treatment of VSMC-related diseases.