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目的研究丝状肌动蛋白(filament actin,F-actin)和 RhoA 在未成熟大鼠缺氧缺血(hypoxic-ischemia,HI)性脑白质损伤(white matter damage,WMD)中的变化,探讨两者作用及可能存在的关系。方法 2日龄 SD 大鼠(n=184只)随机分为14组:7个时段 WMD 组(HI 后12、24、48、72 h,7、14、28 d)和7个相应时段对照组。采用 Back 方法制作 WMD 动物模型。HE 染色观察脑组织病理学变化,电镜观察超微结构改变。采用荧光免疫组织化学方法(n=80只)和实时荧光定量 PCR(n=80只)分别观察 HI 后12、24、48、72 h,7 d 脑组织中 F-actin 和 RhoA 的变化。结果 (1)光镜和透射电镜检查脑组织,符合 WMD 的病理及超微结构也改变。(2)WMD 组胞膜荧光染色不连续细胞百分比与对照组相比明显增高(P<0.05),WMD 组分别为0.32±0.04,0.43±0.04,0.56±0.03,0.65±0.04,0.87±0.03;对照组为0.02±0.01,0.02±0.01,0.01±0.01,0.02±0.01,0.02±0.01。(3)WMD 组 RhoAmRNA 表达在 HI 后12、24、48、72 h 均明显高于对照组(P<0.05),WMD 组分别为1.205,2.415,4.830,1.500;对照组为0.300,0.375,0.375,0.530。HI 后7 d WMD 组 RhoAmRNA表达接近对照组(P>0.05)。结论 (1)2日龄未成熟大鼠 WMD 模型建立成功。(2)HI 后,F-actin在细胞内分布发生变化:细胞膜上分布减少,胞浆中分布增高,该变化可能与神经细胞生长锥的塌陷和回缩有关。(3)WMD 中 RhoA 可能在一定程度上参与了 F-actin 分布表达的变化,但并不是影响F-actin 的惟一因素。
Objective To investigate the changes of F-actin and RhoA in immature rats with hypoxic-ischemic (HI) white matter damage (WMD) The role and possible relationship. Methods Two-day-old SD rats (n = 184) were randomly divided into 14 groups: WMD group (12,24,48,72 h, 7,14,28 d after HI) and 7 control groups . Using Back method to make WMD animal model. The pathological changes of brain were observed by HE staining and the ultrastructure was observed by electron microscope. The changes of F-actin and RhoA at 12, 24, 48, 72 and 7 days after HI were observed by fluorescence immunohistochemistry (n = 80) and real-time fluorescence quantitative PCR (n = 80) respectively. Results (1) Light microscopy and transmission electron microscopy of brain tissue, in line with WMD pathology and ultrastructure also changed. (2) The percentage of discontinuous cells in the membrane fluorescence staining of WMD group was significantly higher than that in the control group (P <0.05), and the WMD group was 0.32 ± 0.04,0.43 ± 0.04,0.56 ± 0.03,0.65 ± 0.04,0.87 ± 0.03 respectively; The control group was 0.02 ± 0.01, 0.02 ± 0.01, 0.01 ± 0.01, 0.02 ± 0.01, 0.02 ± 0.01. (3) The expression of RhoAmRNA in WMD group was significantly higher than that in control group at 12, 24, 48 and 72 h after HI (P <0.05), and the WMD group was 1.205, 2.415, 4.830 and 1.500 respectively; the control group was 0.300, 0.375 and 0.375 , 0.530. RhoA mRNA expression in WMD group was similar to that in control group on the 7th day after HI (P> 0.05). Conclusion (1) The 2-day-old immature rat WMD model was established successfully. (2) After HI, the distribution of F-actin in the cells changed: the distribution on the cell membrane decreased and the cytoplasm distribution increased, which may be related to the collapse and retraction of the growth cone of nerve cells. (3) RhoA may play a part in the changes of F-actin distribution in WMD, but it is not the only factor that affects F-actin.