miR-32-3p Regulates the Vitality in Oxygen and Glucose Deprivation/ Reperfusion Injury by Targeting Smad3

ZHANG Aoqi^1,2, ZHAO Haile^2,3, BADE Rengui^4,5, ZHU Runxiu^2,3, YAO Yuan^1,2,3*

( ¹College of Life Science, Inner Mongolia University, Hohhot 010070, China; ²Department of Neurology, Inner Mongolia University People’s Hospital, Hohhot 010017, China; ³Department of Neurology, Inner Mongolia People’s Hospital, Hohhot 010017, China; ⁴ Inner Mongolia Key Laboratory of Hypoxic Translational Medicine, Baotou Medical College, Baotou 014060, China; ⁵ School of Medical Technology and Anesthesiology, Baotou Medical College, Baotou 014060, China)

Abstract:

This study aimed to investigate miR-32-3p regulates the vitality in OGD/R (oxygen-glucose deprivation and reperfusion) injury by targeting Smad3. For studying CIRI (cerebral ischemia reperfusion injury), SK-N-SH cells were used to construct an OGD/R model. The target genes of miR-32-3p were predicted and then used for function annotation through bioinformatic methods. The effect of miR-32-3p on cell viability by targeting Smad3 was verified by dual-luciferase reporter system, real-time quantitative PCR, Western blot, CCK-8 and live/ dead cell assay. The results showed that Smad3 is a novel target of miR-32-3p, and miR-32-3p is upregulated in SK-N-SH OGD/R model. CCK-8 assay demonstrated that cell viability was significantly decreased in sh-Smad3 and miR-32-3p-inhibitor+sh-Smad3 groups, compared with miR-32-3p inhibitor control and shNC groups. Subsequently, these results were further supported by the live/dead cell vitality assay. The more dead cells represented by red fluorescence were observed in sh-Smad3 and miR-32-3p-inhibitor+sh-Smad3 groups, compared with those in inhibitor NC and shNC groups. Compared with the SK-N-SH cells, the Lats2, Yap/Taz, and Smad3 levels were observed to decrease in OGD/R cells. These results indicated that lacking Smad3 might inhibit cell viability. These results suggest that Smad3, Lats2 and Yap/Taz in Hippo signaling pathway may jointly regulate cell vitality.

CSTR: 32200.14.cjcb.2022.09.0002