CAJ | 학술논문

目的：探讨Wnt5a/Frizzled-2信号在创伤性颅脑损伤(TBI)后神经细胞钙超载过程中的作用。方法体内实验：成年Sprague-Dawley大鼠(96只)，随机分为正常组A(32只)，单纯损伤组B(32只)和RNAi抑制组C(32只)。B组和C组Fenny式法制作中型颅脑损伤模型。其中C组模型建立前48 h采用立体定向海马区注射RNAi以抑制Frizzled-2的表达。伤后24 h处死大鼠，取伤侧海马组织，用western blotting方法检测其中Frizzled-2和Wnt5a含量，用免疫荧光染色和激光共聚焦显微镜检测其中钙离子浓度。单因素方差分析比较各组间的差别。结果 A组大鼠海马组织细胞中稳定表达着Wnt5a/Frizzled-2信号。TBI后，与A组相比，B组海马组织中的Wnt5a和Frizzled-2的表达分别升高了2倍和5倍(P<0.01)，海马细胞中钙离子的含量显著增高(P<0.01)。与B组相比，随着Frizzled-2的抑制，C组海马细胞中的Wnt5a和Frizzled-2的表达分别降低1倍、3.5倍(P<0.01)，同时细胞中的钙离子含量显著降低1.5倍，接近A组(P<0.01)。结论在生理和病理条件下，Wnt5a/Frizzled-2信号在调节神经细胞中的钙离子浓度变化中起到重要的作用，同时我们的研究提示该信号的相关重要组成因子Wnt5a，Fzd2和P-CamKII以及针对frizzled-2设计的特异性RNAi可以作为日后TBI研究和治疗的相关靶点。
목적：탐토Wnt5a/Frizzled-2신호재창상성로뇌손상(TBI)후신경세포개초재과정중적작용。방법체내실험：성년Sprague-Dawley대서(96지)，수궤분위정상조A(32지)，단순손상조B(32지)화RNAi억제조C(32지)。B조화C조Fenny식법제작중형로뇌손상모형。기중C조모형건립전48 h채용입체정향해마구주사RNAi이억제Frizzled-2적표체。상후24 h처사대서，취상측해마조직，용western blotting방법검측기중Frizzled-2화Wnt5a함량，용면역형광염색화격광공취초현미경검측기중개리자농도。단인소방차분석비교각조간적차별。결과 A조대서해마조직세포중은정표체착Wnt5a/Frizzled-2신호。TBI후，여A조상비，B조해마조직중적Wnt5a화Frizzled-2적표체분별승고료2배화5배(P<0.01)，해마세포중개리자적함량현저증고(P<0.01)。여B조상비，수착Frizzled-2적억제，C조해마세포중적Wnt5a화Frizzled-2적표체분별강저1배、3.5배(P<0.01)，동시세포중적개리자함량현저강저1.5배，접근A조(P<0.01)。결론재생리화병리조건하，Wnt5a/Frizzled-2신호재조절신경세포중적개리자농도변화중기도중요적작용，동시아문적연구제시해신호적상관중요조성인자Wnt5a，Fzd2화P-CamKII이급침대frizzled-2설계적특이성RNAi가이작위일후TBI연구화치료적상관파점。
Objective To investigate the role of Wnt5a/Frizzled-2 signal in the process of nerve cell calcium overload after traumatic brain injury (TBI). Methods In vivo experience:adult Sprague-Dawley rats (n=96) were randomly divided into Sham group A (n=32), Pure injury group B (n=32) and RNAi inhibition group C (n=32). Moderate TBI modal was made in group B and C by Fenny method. RNAi was stereotactic hippocampal injected 48 hours before modal were made in group C to inhibit the expression of Frizzled-2. Rats were killed 24 h after injury, the levels of Frizzled-2 and Wnt5a in the injured side of hippocampal tissue were tested by western blotting method, the levels of calcium concentration were tested by immunofluorescent staining and laser confocal microscope. The data among groups was compared by single factor analysis of variance. Results In group A, the expression of Wnt5a/Frizzled-2 signal in the hippocampal tissue cells were stable. Compared with group A, the expression of Wnt5a and Frizzled-2 in the hippocampal tissue cells after TBI were increased by 2 times and 5 times (P<0.01) respectively in group B, and the level of calcium ion increased significantly (P<0.01). Compared with group B, with the inhibition of Frizzled-2, the expression of Wnt5a and Frizzled-2 in the hippocampal cells were decreased by 1 times and 3.55 times (P<0.01) respectively in group C, meanwhile the level of calcium ion were significantly decreased 1.5 times, close to the group A (P<0.01). Conclusion Under the physiological and pathological conditions, Wnt5a/Frizzled-2 signal plays an important role in the change of calcium concertration in the regulation of nerve cells. Our research suggests that the important factor related to the signal of Wnt5a, Frizzled-2, P-CamKII and RNAi which specific designed for Frizzled-2 RNAi could become potential therapeutic targets for further study in TBI.