miR-142-3p overexpression targets CLTA and impairs clathrin-mediated endocytosis in HLSECs induced by high glucose.

OBJECTIVE

This study aims to explore the effect of miR-142-3p on the endocytosis in human liver sinusoidal endothelial cells (HLSECs) induced by high glucose and to investigate the underlying mechanisms.

METHODS

Multi-database target prediction was performed to identify potential targets of hsa-miR-142-3p. HLSECs were cultured under 5.5 mmol/L glucose (normal) or 25 mmol/L glucose (high) conditions. qRT-PCR assessed the RNA expression levels of miR-142-3p and CLTA. Western blot assessed the protein expression levels of clathrin-mediated endocytosis(CME)-related genes. Immunofluorescence assay was used to observe the cellular localization of CLTA. A dual-luciferase assay investigated the targeting relationship between miR-142-3p and CLTA. DiI-LDL endocytosis assay was used to study the endocytosis in HLSECs.

RESULTS

Bioinformatics analysis predicted CLTA as a potential target of miR-142-3p. The expression level of miR-142-3p was increased and the expression levels of CLTA, α-adaptin, AP2M1 and VPS35 were reduced in high glucose-induced HLSECs (P < 0.05). DiI-LDL endocytosis assay demonstrated that the endocytosis was weakend in HLSECs induced by high glucose. Dual-luciferase reporter assay confirmed that miR-142-3p directly targets CLTA. Compared with the negative control group, inhibition of miR-142-3p significantly increased the expression levels of CLTA, AP2M1, α-adaptin and VPS35, and improved the endocytosis in HLSECs (P < 0.05). In contrast, overexpression of miR-142-3p showed the opposite effects.

CONCLUSIONS

miR-142-3p overexpression targets CLTA and impairs CME in HLSECs induced by high glucose. The concurrent changes in other CME-related genes further suggest a broader regulatory role of miR-142-3p on the endocytosis.