Mechanism of leech extract improving cerebral ischemia-reperfusion injury based on RhoA/ROCK Ⅱ pathway

AimTo investigate the effect of leech extract on the blood-brain barrier of MCAO/R (middle cerebrovascular occlusion/reperfusion)mice by regulating Rho-associated coiled-coil-forming kinase (ROCK)pathway and its mechanism in vitro.MethodsAfter MCAO/R model was established by middle cerebral artery occlusion, SPF C57BL/6 male mice were divided into the sham operation group, MCAO/R group, aspirin group (aspirin, 13 mg·kg-1)and leech extract group (leech, 0.8, 1.6 and 2.4 mL·kg-1). The volume of cerebral infarction was observed by TTC staining; MNSS score was used to evaluate nerve defect; HE staining and Nissl staining were used to observe the pathological changes of cerebral ischemic penumbra in each group. Immunofluorescence was used to detect the expression of Laminin, and PDGFRβ/Biotinylated LEL immunofluorescence double staining was used to detect the dissociation between peripenumbra cells and cerebral microvascular endothelial cells. Western blot was used to detect the expression of RhoA and ROCK Ⅱ proteins, tight junction (TJs)related proteins ZO-1, Occludin and Claudin-5, basement membrane (BM) related proteins laminin (LN), fibronectin (FN), matrix metalloproteinase MMP-2 and MMP-9, and p-MLC/MLC protein.ResultsLeech obviously improved the nerve defect and reduced the volume of cerebral infarction in mice (P P P P ConclusionsLeech can improve BBB injury in cerebral ischemic penumbra of MCAO/R mice, and its mechanism may be related to regulating RhoA/ROCK Ⅱ signaling pathway.