Influences Of Caveolae On Proliferation Of Vascular Smooth Muscle Cells And The Possible Mechanism

BACKGROUND AND PURPOSES:Delayed cerebral vasospasm(DCVS) is a serious complication after subarachnoidhemorrhage(SAH), and the main reason of death and disability. The main performance isthe affected arterial stenosis and the thickening of the artery walls, include that apoptosis ofendothelial cells and proliferation of VSMCs, but the pathogenesis is not yet been fullyelucidated[1].caveolae is a specific invagination of the cytoplasmic membrane surface, the structuresbrought together a variety of receptor molecules, such as transforming growth factorreceptor, epidermal growth factor receptor, the platelet-derived growth factor receptor,angiotensin receptor, etc, and plays an important role in the signaling pathway[3].Caveolin-1is a surface marker protein of caveolae, and keep the structure and function ofcaveolae complete. The study shows, caveolin-1in regulating the contraction of vascularsmooth muscle cells(VSMCs) and proliferation reaction plays an important role[4、5]. Recentresearch suggests that the DCVS occurrence may be associated with vascular endothelialgrowth factor、transforming growth factor-beta1(TGF-β1) and so on, these cell factorsactively participate in the cerebrovascular wall proliferation process[2]. And TGF-βcanactivate intracellular Smads signal system regulation of the expression[6]. Therefore, Weassume through the experiment in vitro cultivation of VSMCs,in the case of destruction ofcaveolae structure, whether can enhance TGF-β/Smad pathway signal pathway, therebyinhibiting the proliferation of VSMCs. Research the possible mechanisms of thecerebrovascular wall proliferation after SAH.METHODS:1. Dissect the SD rats, separate and remove the common carotid artery, in vitro tissuepiece method to cultivate the original generation of vascular smooth muscle cells and Identification of vascular smooth muscle cells by immunofluorescence;2. through the CCK-8method detects the changes of vascular smooth muscleproliferation in normal control group and experimental group;3. detect caveolin-1, TGF-beta R1expression level in cells with immunohistochemicalmethod;4. extract cell protein, detect caveolin-1, pSmad2protein expression with westernblot method;5. detect caveolin-1, pSmad2mRNA transcription with RT-PCR;RESULTS:1. success to separate the common carotid artery, and culture the vascular smoothmuscle cells in vitro;2. the vascular smooth muscle cells in experimental group after MβCD interferencewas inhibited significantly through CCK-8method(P<0.01);3. immunohistochemical method to detect the experimental group expressed in thecell membrane and cytoplasm caveolin-1reduced, TGF-beta R1expressionenhancement;4. RT-PCR method to detect caveolin-1gene expression level decreased obviously,while western blot method to detect caveolin-1protein expression level decreasedobviously in the experimental group;5. RT-PCR method to detect Smad2gene expression level increased significantly,while western blot method to detect pSmad2protein expression levels rose significantly inthe experimental group;CONCLUSIONS:Caveolae that interferes with MβCD, the structure was destroyed, TGF-β/Smadsignaling pathways was enhanced, and the proliferation of vascular smooth muscle cellswere inhibited;...