DHA Attenuates Hypoxia/Reoxygenation Injury by Activating SSeCKS in Human Cerebrovascular Pericytes

Docosahexaenoic acid (DHA) can alleviate cerebral ischemia/reperfusion injury by reducing blood–brain barrier permeability and maintaining its integrity, accompanied by an increased Ang-1/Ang-2 ratio; however, the underlying mechanisms of these effects remain unclear. Src-suppressed C kinase substrates (SSeCKS), a substrate of protein kinase C, plays an important role in maintaining cell junctions and cell morphology and regulating cell permeability. However, whether DHA can increase SSeCKS expression and then mediate the Ang-1/Ang-2 ratio still needs to be studied. Human cerebrovascular pericytes (HBVPs) cultured in vitro were divided into groups, treated with or without DHA along with SSeCKS siRNA to knockdown SSeCKS expression, and then subjected to 24 h of hypoxia followed by 6 h of reoxygenation. Cell viability; lactate dehydrogenase (LDH) release; and Ang-1, Ang-2 and VEGF activity were detected by using ELISA kits. The apoptosis rate was assessed by TUNEL flow cytometry. Expression of the SSeCKS, Ang-1, Ang-2 and VEGF proteins was evaluated by western blotting. Pretreatment with 10 μM or 40 μM DHA efficiently attenuated hypoxia/reoxygenation (H/R) injury by activating SSeCKS to increase the Ang-1/Ang-2 ratio and downregulate VEGF expression in HBVPs, as evidenced by decreased LDH release and apoptotic rates and increased HBVPs viability. Meanwhile, after we used SSeCKS siRNA to knock down SSeCKS protein expression, the protective effect of DHA on HBVPs following H/R injury was reversed. In conclusion, DHA can activate SSeCKS to increase the Ang-1/Ang-2 ratio and downregulate VEGF expression in HBVPs, thus reducing H/R injury.