| BACKGROUND:SAH (subarachnoid hemorrhage,SAH) is the third acute cerebral vascular disease , the incidence is only second to cerebral infarction and cerebral hemorrhage.The vast majority of SAH due to rupture of cerebral aneurysms, the mortality rate is as high as 25% . About 1/3 of patients who survived was result of neurological deficit and need to rely on others to live, imposed a heavy burden to the family and community. Cerebral vasospasm (CVS) is the most common complication of SAH which often causes serious brain ischemic brain damage and results in death or serious disability. CVS and the second bleeding are two major complications of SAH which is most closely related to the prognosis of patients. At present, the occlusion of the cerebral aneurysm surgery and endovascular embolization has made great progress.The second bleeding after SAH has been significantly reduced.However, the overall prognosis of SAH has not been a corresponding improvement . It was identified that the CVS was more closely related to the prognosis of patients of SAH. As a result, on the basis of in-depth understanding of the mechanism of CVS after SAH, to search for a safe, effective, easy preventive measures is the key that improve the prognosis of SAH. The calcitonin gene-related peptide (CGRP) is an endogenous vasodilator peptide which has strong relaxation effect on the brain blood vessels, known as the most powerful endogenous vasodilator substances. CGRP is closely related to the pathophysiological process of cerebrovascular disease (CVD) ,may have potential value of the treatment of a variety of ischemic cerebrovascular disease. Animal studies showed that, CGRP could reverse the CVS after experimental SAH. CGRP is difficult through the blood-brain barrier and into the cerebrospinal fluid and brain tissue from the blood circulation due to large molecule. As a result ,the conventional route of administration will not be effective.The purpose of this study is to find a drug delivery pathway target for CNS which is simple,effective and non-invasive.To explore the feasibility of CGRP can bypass the blood-brain-barrier targeting for the central nervous system effectively by nasal administration. To observe whether CGRP has the ability to prevent cerebral CVS following SAH by nasal administrationOBJECTIVE:(1). Explore the nasal route of administration CGRP is effectively into the central nervous system bypass the BBB and clarify its distribution in the CNS.(2).To investigate the effects of CGRP given intranasally on the cerebral blood flow and circulating endothelial cells in rats after SAH.(3). To observe whether CGRP has the ability to prevent cerebral vasospasm CVS following SAH.METHODS:(1). 48 SPF level healthy Wistar rats were randomly divided into six groups :①Intranasal (IN) 125I-CGRP normal group: normal rats were given intranasally 125I-CGRP50μl (1μg, 20μCi)②Intranasal 125I-CGRP SAH group: SAH models were given intranasally 125I-CGRP50μl (1μg, 20μCi)③Intranasal normal saline solution (NS) control group: normal rats were given intranasally NS 50μl④Intravenous (IV) 125I-CGRP normal groups: normal rats intravenous injection of 125I-CGRP300μl (1μg, 20μCi)⑤Intravenous 125I-CGRP SAH groups: SAH models, intravenous injection of 125I-CGRP 300μl (1μg, 20μCi)⑥Intravenous normal saline solution (NS) control group: normal rats intravenous injection of NS 300μl. Using radioimmunoassay technology, compared nasal olfactory pathway (IN) with intravenous injection (IV)after 30min, the radioactive labelled CGRP in the brain, spinal cord, cerebrospinal fluid, blood and tissue surrounding the quantitative distribution.(2).Sixty-four healthy audlt Wistar rats were randomly divided into four groups: group A :normal group,group B: model of SAH group, group C:IN NS SAH group, group D:IN CGRP SAH group. SAH was induced in rats via blood injection into the cisternamagna。Group C was given 50μl NS by intranasal pathway after the second SAH. Group D was given CGRP1μg/50μl by intranasal pathway after the second SAH. The behavior changes of rats after SAH were observed. The regional cerebral blood flow (rCBF) was recorded with the Laser Doppler. On the third day after SAH, take blood samples to detect apoptosis of circulating endothelial cells(CEC). (3).Thirty-two New Zealand white rabbits were divided into four groups randomly (n=8). group A :normal group,group B: model of SAH group, group C:IN NS SAH group, group D:IN CGRP SAH group. SAH was induced in rats via blood injection into the cisternamagna.Group C was given 0.4ml NS by intranasal pathway after the second SAH. Group D was given CGRP10μg/0.4ml by intranasal pathway after the second SAH. The behavior changes of rats after SAH were observed. On the third day after SAH, all rabbits were given digital substract angiography .To quick decapitated and to obtain brain after iduction of SAH 3 days,to separate cortex and hippocampus,the the brain water content was detected with wet-dry weight method.RESULT:(1) The concentrations of CGRP in the district of the brain, CSF and the spinal cord is much higher(6-161 times) in group IN CGRP than group IV CGRP. The highest regin is olfactory bulb , followed by the CSF and spinal cord. The drug distribution is no significant difference between normal rats and SAH rats by IN or IV drug administration.(2) The rCBF of Group B and Group C is significantly lower than that of group A 30 min after SAH, down to the lowest in one hour, there was no clear trend to resume in 12h. The rCBF of the rats which was dealed with CGRP after SAH decreased slowly. The extent of reduction was smaller obviously than group B and Group C. The CEC count of group B and C was significantly higher than the group A (P<0.01), The group D was significantly lower than the group B and C (P<0.01).(3) Compared with the normal group, SAH group and IN NS SAH group the Basilar Artery (BA) vasospasm obviously ,the diameter of BA narrowing (P <0.01); Compared with the SAH group and the IN NS SAH group, the diameter of BA of IN CGRP SAH group was improved Significantly (P <0.01).On the 3d after SAH-induced,the brain tissue water content SAH group and the IN NS SAH group was significantly higher than the normal control group (P<0.01);Compared with the SAH group and the IN NS SAH group, the brain tissue water content of IN CGRP SAH group was significantly lower (P <0.01).CONCLUSION:(1) .The results suggest intranasally delivered CGRP can bypass the blood-brain barrier via olfactory pathways to rapidly elicit biological effects in the CNS.(2). The rCBF decreased after SAH, It can be significantly improved rCBF by intranasal (IN) delivery CGRP. It was concluded that VEC injury may play an important role in the pathogenesis of CVS after SAH. CGRP has protective function on CVS by preventing VEC from being damaged. The number of CECs was increased in the peripheral blood,so CECs can be used as a early warning factor for CVS after SAH.(3). It was concluded that the CVS after SAH can be prevented by intranasal (IN) delivery CGRP in rabbits. |
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