摘要
目的探讨缺氧对脑动静脉畸形（BAVM）猪模型血管内膜平滑肌细胞（VSMC）中内皮生长因子（VEGF）表达及VSMC细胞功能的影响。方法建立稳定的BAVM实验用猪模型，分离脑底微血管网（RM）的VSMC后行原代培养。取鉴定成功的VSMC及源自正常实验用猪的VSMC分为A、B、C、D4组，其中A组：正常实验用猪VSMC，21％O2环境培养；B组：BAVM模型猪VSMC，21％O2环境培养；C组：正常实验用猪VSMC，1％O2环境培养；D组：BAVM模型猪VSMC，1％O2环境培养。细胞免疫荧光染色实验检测VSMC密度；实时定量多聚酶链反应（RT-PCR）和Westernblotting检测VSMC中VEGFmRNA及蛋白表达情况：TUNEL法检测VSMC的凋亡情况；Transwell实验检测VSMC的侵袭能力。结果（1）4组VSMC密度（个／高倍镜）分别为71．65±4．22B、158．24±9．87、95．33±7．21、299．80±13．23，差异有统计学意义（F=119.351，P=0．000）；其中B、D组明显高于A、C组，差异有统计学意义（P〈0．05）。（2）4组细胞VEGFmRNA表达量分别为1．93±0．77、4．51±1．25、2．87±1．94、8．03±1．74，差异有统计学意义（F=119．351，P=0．000）；其中B、C、D组明显高于A组，D组明显高于B、C组，差异有统计学意义（P〈0．05）。（3）4组细胞VEGF蛋白表达量分别为3．83±0．63、6．99±1．77、5．02±1．23、8．27±1．50，差异有统计学意义（F=117．420，P=0．000）；其中B、C、D组明显高于A组，D组明显高于B、C组，差异有统计学意义（P〈0．05）。（4）24h、48h时4组VSMC凋亡和侵袭数目差异无统计学意义（P〉0．05）：72h时B、C和D组细胞凋亡数目和侵袭数目较A组显著增多，差异有统计学意义（P〈0．05）。结论缺氧可上调BAVM模型中VEGF表达，加剧VSMC凋亡及侵袭能力的增加。进而加速畸形血管团的形成。
Objective To investigate the effect of hypoxia on endothelial growth factor （VEGF） expression in vascular intimal smooth muscle cells （VSMC） and function of VSMC of swine models of brain arteriovenous malformations （BAVM） . Methods The stable swine models of BAVM were established; separation of cerebral microvascular network （RM） was performed and VSMC was selected and primarily cultured. VSMC from the above models and normal swines were divided into four groups： group A, VSMC from normal swines cultured at 21% O2; group B, VSMC from models cultured at 21% O2; group C, VSMC from normal swines cultured at 1% O2; group D, VSMC from models cultured at 1% O2. Quantitative real-time polymerase chain reaction （RT-PCR） and Western blotting were used to validate the mRNA and protein VEGF expressions in VSMC; TUNNEL was used to detect the apoptosis of VSMC, and Transwell assay was used to determine the VSMC invasion. Results （1） VSMC density in the four groups was 71.65±4.22, 158.24±9.87, 95.33±7.21 and 299.80±13.23 cells/field ingroup A-D, with significant differences （F=119.351, P=-0.000）. （2） VEGF mRNA expression quantity in the four groups was 1.93±0.77, 4.51±1.25, 2.87±1.94 and 8.03±1.74 in group A-D, with significant differences （F=119.351, P=0.000）; that in group B-D was significantly higher than that in group A （P〈0.05）, and that in group D was significantly higher than that in group B and C （P〈0.05）. （3） The VEGF protein expression quantity of the four groups was 3.83±0.63, 6.99±1.77, 5.02±1.23 and 8.27± 1.50, with significant differences （F=117.420, P=0.000）; that in group B, C and D was obviously higher than that in group A, and that in group D was obviously higher than that in group B and C （P〈 0.05）. （4） At the 24 and 48 h of culture, VSMC apoptosis and invasion number showed no statistical difference among the four groups （P〉0.05）; at the 72 h, the number ofapoptosis and invasion cells in group A was significantly decreased as compared with those in group B, C and D （P〈0.05）. Conclusion Hypoxia can increase the VEGF expression, aggravate the apoptosis and invasion of VSMC, and accelerate the formation of vascular malformation in BAVM models.
出处
《中华神经医学杂志》
CAS
CSCD
北大核心
2015年第10期994-999,共6页
Chinese Journal of Neuromedicine
基金
广东省中医药局科技项目（20151061）
