Inhibition of the Rho/ROCK pathway promotes the expression of developmental and migration-related genes in astrocytes exposed to alcohol

Abstract

Astrocytes are an important regulator of alcohol dependence. Furthermore, the downregulation of Rho-associated coiled coil-containing protein kinase 2 (ROCK2) attenuates alcohol-induced inflammation and oxidative stress in astrocytes. On the basis of these findings, we examined the effects of alcohol and a Rho/RACK kinases inhibitor on astrocyte function and investigated their effects on mRNA expression to further explore the protective mechanisms of a Rho/RACK kinases inhibitor in astrocytes after alcohol exposure. CTX TNA2 astrocytes were cultured with alcohol and Rho/RACK kinases inhibitor intervention before undergoing transcriptome sequencing, quantitative reverse transcription polymerase chain reaction (qRT-PCR), and wound healing assays. Alcohol exposure modulated cell morphology and inhibited astrocyte migration, whereas Fasudil improved cell morphology and promoted astrocyte migration after alcohol exposure. Transcriptome sequencing results indicated that alcohol exposure modulates the expression of genes involved in astrocyte development. Fasudil reversed the effects of alcohol exposure on the astrocyte developmental process. Four genes related to the developmental process and migration – Ccl2, Postn, Itga8, and Serpine1 – with the highest protein–protein interaction correlations (node degree >7) were selected for verification by qRT-PCR, and the results were consistent with those of the sequencing and wound healing assays. Our results suggest that the Rho/ROCK pathway is essential for alcohol to be able to interfere with astrocyte development and migration gene expression. The Rho/ROCK pathway inhibitor Fasudil reversed the adverse effects of alcohol exposure on astrocytes and may have clinical applications.