Activity Screening And Study Of Small Molecule RhoA Inhibitor On Vasodilation

Objective: To screen the compounds which have relaxant effect on cerebralvasospasm after subarachnoid hemorrhage on the basis of structure modifying of smallmolecule RhoA inhibitor HL07. It will be useful to study the pathophysiological effects ofRhoA-mediated pathway and also to discover new leading compound for cardiovasculardisease medication.Methods:1) We applied G-LISA method to evaluate the inhibitory activities of33new compound by measuring LPA-induced RhoA activation in the human cerebrovascularsmooth muscle cells (HBVSMCs);2)We used Isometric tension determination to monitorthe changes of vascular tension caused by HL47/47K、HL79/79K and HL82/82K;3) Weestablished the rat SAH-CVS model using the method of double-heorrhage by injectingautologous arterial blood into the prechiasmatic cistern, and observed the influence ofHL47on the morphology and the perimeter of inside diameter of basilar artery (BA). AllSD rats were divided into7groups randomly: Normal group, SAH group, SAH+DMSOgroup, SAH+Fasudil group, SAH+HL47(L) group, SAH+HL47(M) group, SAH+HL47(H)group,(n=9in each group).Results:1) In the RhoA activation assay, three of the33new compounds(HL47、HL79、HL82) show better inhibitory activities than HL07，with IC50=1.66±0.28μM、1.26±0.13μM、1.17±0.19μM，respectively；2) In isolated endothelium-denuded ratthoracic aorta (TA) rings, both HL47(IC50=70.90±1.33μM）and HL82（IC50=73.43±2.61μM） causeddose-dependent relaxation on PE-induced contraction, and more potenrt thanHL07（IC50=123.22±8.93μM）(P<0.05),but HL79shows weak relaxation activities. Aftersalifying, all of the three compounds display better relaxationwith IC50=59.92±5.40 μM(HL47K)、66.64±2.32μM(HL82K)、111.88±15.71μM(HL79K),respectively;3) Thecomparison of the perimeter of inside diameter of BA in each group: there weresignificantly statistical difference between SAH group (386.25±11.25μm) and normalgroup (472.50±53.47μm)(P<0.05), and SAH+Fasudil group (468.83±43.53μm)(P<0.05),and SAH+HL07（L、M、H）group (464.00±50.46μm、475.60±63.79μm、513.42±40.15μm)(P<0.05); There were no significant difference between SAH group and SAH+DMSOgroup (390.93±20.60μm)、also between SAH+HL47(L、M、H)group and SAH+Fasudilgroup, as well as among SAH+HL07(L)、SAH+HL07(M)、SAH+HL07(H)group.Conclusion:1)After structural modification,some of the new compounds(HL47、HL79、HL82) show stronger RhoA inhibitory activities than HL07;2) Comparing HL07,HL47/47K、 HL82/82Kcausedbetterrelaxation on PE-induced contraction in arteryrings.Furthermore, the solubility of the compound has a great impact on its inhibitoryactivity;3) HL47could relieve the CVS of the BA after SAH. In summation, the resultsindicate that HL47can cause stronger relaxanteffect on cerebral vasospasm aftersubarachnoid hemorrhageby inhibitionof RhoA activation.