Long non-coding RNA VIM Antisense RNA 1 (VIM-AS1) sponges microRNA-29 to participate in diabetic retinopathy

Abstract

Aims: Long non-coding RNA (lncRNA) VIM Antisense RNA 1 (VIM-AS1) has been reported to be correlated with type 2 diabetes (T2D) susceptibility, while the roles of this lncRNA in T2D and its complications remain unclear. This study aimed to explore the role of VIM-AS1 in diabetic retinopathy (DR).

Methods: Gene expression levels in both human specimens and in vitro cultivated cells were determined by qPCR and western blot. Overexpression experiments were performed to analyze gene interactions. Cell apoptosis after transfections was detected by cell apoptosis assay.

Results: We found that VIM-AS1 was significantly downregulated in T2D patients in comparison with that in healthy controls. Specifically, the expression levels of VIM-AS1 were lowest among T2D patients complicated with DR. Bioinformatics analysis showed that VIM-AS1 can interact with microRNA 29 (miR-29), which is a critical player in high glucose-induced apoptosis of human retinal pigment epithelial cells (RPEs). Dual-luciferase assay also revealed the direct interaction between them. High glucose treatment led to upregulated miR-29 and downregulated VIM-AS1. However, overexpression of VIM-AS1 and miR-29 did not affect the expression of each other. Cell apoptosis analysis showed that overexpression of VIM-AS1 reduced the enhancing effects of miR-29 overexpression on RPEs cell proliferation.

Conclusions: Therefore, VIM-AS1 may sponge miR-29 to participate in DR.

Keywords: Apoptosis; Diabetic retinopathy; Retinal pigment epithelial cell; T2D; VIM-AS1; miR-29.

Fig. 1

VIM-AS1 was downregulated in DR. Expression levels of in plasma samples from DR group (n = 60), DN group (n = 60), DG group (n = 60), DU group (n = 60), T2D group (n = 60) and control (n = 60) group were measured by qPCR. *p < 0.05

Fig. 2

VIM-AS1 can interact with miR-29 but they were not significantly correlated in DR patients. The interaction between VIM-AS1 and miR-29 was analyzed by IntaRNA (https://rna.informatik.uni-freiburg.de/IntaRNA/Input.jsp). It was observed that miR-29 can form strong base pairing with VIM-AS1 (a). Dual luciferase assay was performed to further confirm the interaction between VIM-AS1 and miR-29 (b). Levels of miR-29 in plasma from DR patients were measured by qPCR, and the correlation between miR-29 and VIM-AS1 was analyzed by linear regression (c). *p < 0.05

Fig. 3

VIM-AS1 and miR-29 did not affect the expression of each other. H1RPE7 cells were transfected with VIM-AS1 expression vector and miR-29 mimic to further analyze the interactions between them. Overexpression of VIM-AS1 and miR-29 was confirmed by qPCR as 24 h post-transfection (a). The effects of overexpression of VIM-AS1 on miR-29 (b) and the effects of overexpression of miR-29 on VIM-AS1 (c) were analyzed by qPCR. Experiments were repeated 3 times and mean values were presented. *p < 0.05

Fig. 4

Overexpression of VIM-AS1 attenuated the effects of miR-29 on glucose-induced h1RPE7 cell apoptosis. H1RPE7 cells were cultivated in medium containing 5, 10, 20 and 30 mM d-glucose for 24 h, followed by the measurement of the levels of VIM-AS1 (a) and miR-29 (b) expression. Cell apoptosis analysis was performed to analyze the effects of overexpressing VIM-AS1 and miR-29 on h1RPE7 cell apoptosis during the treatment of 30 mM d-glucose (c). Experiments were repeated 3 times and mean values were presented. *p < 0.05