Mechanism Of Nerve Regeneration After Cerebral Vascular Laminin And Its Integrin Receptor α6β1 Regulation

Backround and ObjectiveThe third National Death Survey shows, cerebrovascular disease is the major cause of death in China’s urban and rural residents, accounting for22.45%of total deaths, is a major cause of death and disability, which has become a serious public health problem. Cerebral infarction is the most common in cerebral vascular disease, which accounted for about70%in China and85%in Europe and America. So, attaching importance to the prevention and treatment of cerebral infarction has great significance for the whole society. So far, there is no effective method to cure cerebral infarction except for thrombolytic therapy within4.5h after onset. But only a few patients are able to receive treatment due to the limited therapeutic window, and they need to undertake the risk of intracranial hemorrhage after treatment. Neurological damage after cerebral infarction lasts a lifetime, which brings a heavy burden to patients, their families and the community. Therefore, how to improve the treatment of cerebral infarction is still a core issue that people engaged in basic and clinical research strive to solve.Recently, there are numerous studies about cerebral infarction and the two phenomena of angiogenesis and endogenous neurogenesis attract much attention. In our previous study, angiogenesis induced by treatment could significantly promote endogenous neurogenesis, which may cause functional recovery after cerebral ischemia. However, whether angiogenesis is the direct cause of endogenous neurogenesis is still unknown, and the molecular mechanism of neurovascular regeneration requires further research. In this study, we will investigate the relationship between angiogenesis and endogenous neurogenesis through increasing and decreasing angiogenesis after cerebral ischemia, and the changes in the anatomy, modified neurological severity scores, and micro-PET will be examined. Additionally, the effect of laminin and its receptor α6β1integrin in the process of regulating angiogenesis and endogenous neurogenesis will be further studied. This study will provide theoretical basis for the treatment strategy of cerebral ischemia. MethodsEstablish the model of MCAO with longa method.24h after models established, according to the mNSS, randomly divided the50rats which scored10-13into five groups.①A:none-intervention control group.②B:VEGF-injection group:3d after models established, implant osmotic micropump (alzet1007D/kit2,100ul/1w)besides the infraction area with totally VEGF4ug;③C:endostatin-injection group:3d after models established, implant osmotic micropump (alzet1007D/kit2,100ul/1w) besides the infraction area with totally endostatin20ug;④D:intraventricular injection of CXCR4antibody group:3d after model established, implant osmotic micropump (alzet1007D/kit2,100ul/1w) into the right lateral ventricular with totally4ug of anti-CXCR4antibody;⑤E:intraventricular injection of α6β1antagonist(GoH3antibody)group:3d after model established, implant osmotic micropump (alzet1007D/kit2,100ul/1w) into the right lateral ventricular with totally4ug of GoH3antibody。(1) mNSS as well as sticker removal experiment evaluate the nerve function recovery.(2) PET-CT analyze rats’ changes of metabolism around the cerebral infarction before and after VEGF/endostatin injection.(3) Double immunofluorescence staining of BrdU and nestin, DCX, GFAP, NeuN and vWF, marking NSPCs’proliferation, migration, differentiation in SVZ, infarction periphery and the striatum.(4) SVZ whole amount:anatomically isolate SVZ in infraction side and mark proliferated stem cells, vascular matrix, NSPCs by BrdU, laminin and α6β1antibody. Study the relationship of NSPCs, laminin and vascular, measure the distance between NSPCs and nearest vascular, elaborate the vascular microenvironment in SVZ. Results1.2w after MCAO, compare VEGF group with control group. Behavioral recovery is more obvious and discrepancy has statistical significance (P<0.05), which shows mNSS Score decreased and rats’ removal of sticker has no obvious improvement in the sticker removal experiment.2.2w after MCAO, micro-PET result shows that rats in VEGF group showed increased glucose metabolic activity and reduced infract volume.3. Compared with control group, in VEGF group,rats’ cell of BrdU+/vWF+increased around the cerebral infarction and VEGF expression increased around the cerebral infarction,and discrepancy has statistical significance (P<0.05). However, the number of cells in endostatin group, anti-CXCR4group and GoH3group are less and VEGF expression reduced, and discrepancy has statistical significance (P<0.05). Conclusion Promoting angiogenesis after cerebral infarction could provide the proliferation, migration and differentiation of NSPCs a good vascular microenvironment and then promote endogenous neurogenesis. It is the interaction with α6β1integrin and laminin that keeps NSPCs and vascular a close contact. With the support of vascular microenvironment as well as the SDF-1chemotaxis, NSPCs succeed in finishing healthy migration from SVZ to neighboring infarction.