| Objective: Intracranial aneurysm(IA) has a tendency of spontaneous rupture hemorrhage, its morbidity and mortality are very high. At present,the therapies of IA are confined to surgery treatment and interventional treatment in the clinical, there are very few researches about non-surgical treatments such as repair therapy for IA wall. We found the proliferation activity of vascular smooth muscle cell(VSMC) function in the research of the mechanism of IA in rats. As the development of biological and gene therapy, it would be showed us to bring the hope to repair therapy of the unruptured IA wall. VSMC is the major component of artery wall, we found that in the process of IA formation, it put up strong proliferation activity and plasticity. Thus it could be seen that the research on proliferation of VSMC of blood vessel wall in the process of IA formation would become the main directions to repair therapy for IA wall. Therefore, in this experiment, we will establish rat model of IA and investigate the proliferation activity of VSMC of IA wall in rats, and the aim is to enhance the proliferative activity of VSMC in the aneurysm wall and increase the smooth muscle layer, so that we can provide theoretical basis to repair therapy of human IA in the future.Methods: A total of 180 adult male spontaneously hypertensive rats(SHR) were randomly divided into 6 groups: Control group, 1, 2, 3, 4 and 5 months experimental group(each group n = 30). In experimental groups, the left common carotid artery and posterior branches of both renal arteries of rats were ligated and those were not ligated in control group. The rats of all groups were fed the drinking water containing 1% salt solution and 0.12% β-aminopropionitrile(BAPN) feed from one week later after operation. Before surgery and sacrifice, the arterial blood pressure was measured at room temperature in quiet state, andintracranial arterywas examined with 7.0MRA. The circles of cerebral artery were obtained, the pathological changes of IA were observed by haemotoxylin and eosin(H&E) staining. The expression of proliferating cell nuclear antigen(PCNA) of IA wall in each group was tested by immunohistochemistry. The protein levels of α-mooth muscle actin(α-SMA), β-tubulinand osteopontin(OPN) in IA wall were detected by immunofluorescence staining and Western blot. The mRNA relative levels of SM22α and hypertension-related gene(HRG-1) were tested by Real-time Rolymerase Chain Reaction(RT-PCR). Apoptosis of IA wall VSMC was detected by Terminal Deoxynucleotidyl Transferase Mediated dUTP Nick End Labeling(TUNEL).Results:1.Blood pressure changes: incontrol group, the average blood pressurewas increased slowly.In experimental groups, the average blood pressures weregradually increased from 1 month group to 5 months group, and wereobviously higher than control group and preoperative blood pressures(P < 0.05).2. 7.0MRA: 6 rats of each group were randomly selected to examined with 7.0MRA. In control group, the left and right cerebral arteries were symmetrical, and the thickness was uniform, and the cerebral arteries had no significant expansion. In experimental groups, the cerebral arteries of all rats examined with 7.0MRA were thickening in different extent, and the rate of vascular disease was 100%. There were 3/6 of rats of IA formation in the thickening of the cerebral arteries in 4 months group, and there were 5/6 of rats of IA formation in the thickening of the cerebral arteries in 5 months group. 3.Aneurysms pathological changes: in control group, intracranial artery intima layer, smooth muscle cell layer and the outer layer were not damaged, and each layermaintained the integrity of the structure.In experimental groups, endometrial layer of cerebral arteriesbegan todestroy and disappear from 1 month group, VSMCwas degenerated and the structure arrangement was disordered, the amount of VSMCwas reduced, the aneurysm wall was thinninggradually, to 4, 5 months groups, endometrial layer had disappeared, elastic fiber fracture was disrupted and the aneurysm wallinfiltratedwith inflammatory cells, which in 5 months group were the most obvious, and the arterial wall was the thinnest. 4.Immunohistochemistry results: incontrol group, the positive cell rate of PCNA was very low.Compared with control group, the positive cell rate of PCNAwas significantly increased from 1 month group to 5 months group in experimental groups, which was the highestin 4 months group, began to decreased in 5 months group,and there was significantdifferencecompared with 4 months group(P <0.05).5.Western blot results: incontrol group, the expressions of α-SMA and β-tubulin were high, but OPN expression was very low. α-SMA and β-tubulin expressionswere gradually decreased from 1 month group to 5 months group in experimental groups, and were significantly lower than those in control group(P <0.05); compared withcontrol group, OPN expression was significantly increased from 1 month group to 5 months group, which was the highestin 4 months group, began to decreased in 5 months group,and there was significantdifferencecompared with 4 months group(P <0.05).6.Immunofluorescence stainingresults: incontrol group,α-SMA and β-tubulin relative fluorescence intensitise were high, but OPN relative fluorescence intensity was low. In experimental groups, α-SMA and β-tubulin relative fluorescence intensitisewere gradually decreased from 1 month group to 5 months group, and were significantly lower than those in control group(P <0.05); compared withcontrol group, OPN relative fluorescence intensity was significantly increased from 1 month group to 5 months group, which was the highestin 4 months group, began to decreased in 5 months group,and there was significantdifferencecompared with 4 months group(P <0.05).7.RT-PCR results: in control group, SM22αand HRG-1 mRNA relative expressionswere high. In experimental groups, SM22αand HRG-1 mRNA relative expressionswere gradually decreased from 1 month group to 5 months group, and were significantly lower than those in control group(P <0.05). 8.TUNEL staining results: in control group, the percentage ofapoptosis of VSMC was low. In experimental groups, the percentage ofapoptosis of VSMCwere gradually increased from 1 month group to 5 months group, and were significantly higher than those in control group(P <0.05).Conclusions: In the process of experimental rat IA formation, the aneurysm wall VSMC has proliferation activity, and the activeperiod of proliferation is obviously; the formation of IA is closely related to the aneurysmal wall VSMC proliferation activity weaken relatively. |
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