O-GlcNAcylation promotes astroglial-mesenchymal transition via the connexin43 pathway under high-glucose conditions.

Abstract

This study aimed to investigate the effects of O-linked N-acetylglucosamine modification (O-GlcNAcylation) on astroglial-mesenchymal transition through connexin43 (Cx43) pathway under high-glucose conditions. The primary rat astrocytes were cultured under normal and high-glucose conditions, and level of GFAP, α-SMA and Cx43 was investigated. To explore the influence of O-GlcNAcylation on astroglial-mesenchymal transition, Thiamet G treatment was employed to enhance O-GlcNAcylation, while Alloxan was used to decrease it. Cx43 knockdown was acquired through lentivirus constructs to explore its role in astrocyte transition. The levels of GFAP and α-SMA expressions were examined, while astrocyte proliferation was evaluated using the CCK-8 assay, and migration was assessed through wound healing assays. The results showed that primary rat astrocytes were identified by GFAP antibody staining. Under high-glucose conditions, the levels of GFAP, α-SMA, and Cx43 increased, as confirmed by Western blot and immunofluorescence. O-GlcNAcylation augmentation induced by Thiamet G treatment significantly increased the expression of GFAP, α-SMA, and Cx43 compared to both normal and high-glucose conditions. Conversely, the inhibition of O-GlcNAcylation reversed the high-glucose-induced increase in GFAP and α-SMA. Cx43 knockout led to the downregulation of GFAP and α-SMA compared to high-glucose and O-GlcNAcylation-augmented conditions. Additionally, levels of O-GlcNAcylation and VEGF were reduced in Cx43 knockout group. Consistently, CCK8 and wound healing assays demonstrated that Cx43 knockout could inhibit astrocyte proliferation and migration compared to the high-glucose and O-GlcNAcylation augmented groups. These findings demonstrate that astroglial-mesenchymal transition occurs under high-glucose conditions, and can be promoted by O-GlcNAcylation augmentation, but suppressed by Cx43 knockout. The study underscores the significant role of Cx43 in this transition and its potential involvement in diabetic complications.