miR-126基因敲减对小鼠胸腺淋巴细胞发育的影响  

Effect of miR-126 knocking down on the development of mouse thymocytes

作　　者：郭萌萌[1] 胡燕[1] 赵娟娟[1] 陶弋婧 秦娜琳[1] 郑静[1] 徐林[1] 

机构地区：[1]遵义医学院免疫学教研室、贵州省免疫学研究生教育创新基地,贵州遵义563000

出　　处：《基础医学与临床》2016年第3期289-294,共6页Basic and Clinical Medicine

基　　金：国家自然科学基金(31370918);教育部新世纪优秀人才计划(NCET-12-0661)

摘　　要：目的研究miR-126基因敲减对小鼠胸腺细胞发育的影响,并初步探讨其意义。方法观察miR-126基因敲减(KD)小鼠胸腺的体积、重量和细胞总数;Real-time PCR检测小鼠胸腺miR-126表达水平;HE染色观察小鼠胸腺形态学结构;FACS检测胸腺中淋巴细胞比例、核抗原Ki-67的表达以及细胞凋亡的变化;最后,Western blot检测胸腺中磷酸化AKT(p-AKT)和磷酸化ERK1/2(p-ERK)的表达。结果 WT和miR-126KD小鼠胸腺体积大小、重量以及细胞总数均无明显差异,但miR-126KD小鼠胸腺miRNA-126表达水平显著下调(P<0.05)。miR-126KD小鼠胸腺组织结构出现形态学异常。与WT小鼠相比,miR-126KD小鼠胸腺CD4^+SP细胞比例及细胞绝对数明显增加(P<0.05),而CD4^+CD8^+(DP)细胞比例及绝对数则显著减少(P<0.05),miR-126KD小鼠胸腺CD4^+SP细胞核抗原Ki-67表达显著增加(P<0.05),且细胞凋亡明显减少(P<0.05),而DP细胞Ki-67表达则明显减少(P<0.05)。最后,miR-126KD小鼠胸腺淋巴细胞中磷酸化AKT和磷酸化ERK1/2的表达水平明显下调(P<0.05)。结论 miR-126基因敲减可显著影响胸腺细胞特别是CD4^+SP细胞的发育,为后续进一步深入探讨miR-126对T淋巴细胞发育以及功能的影响提供重要的实验基础。Objective To investigate the influence of miR-126 knocking down on the development of mouse thymo- cytes and to explore its significance. Methods The volume, weight and total cell number of thymus in miR-126 knockdown （KD） mice were detected. The expression of miRNA-126 in thymus was detected by real-time PCR. The pathologic change of thymus was observed by microscopy with HE staining. The proportion and expression of nuclear antigen Ki-67 and the apoptosis of lymphocytes in thymus from miR-126KD mice were determined by flow cytometry analysis. Finally, the expression level of p-AKT and p-ERK1/2 in the thymus was detected by Western blot. Results Compared with those of WT mice, the volume, weight and total cells number of miR-126KD mice thymus were not obviously differenced. The expression level of miRNA-126 in miR-126KD mouse thymus was sig- nificantly lower than that in the WT mice （ P 〈 0. 05 ）. Thymus morphology of miR- 126KD mice was abnormal. Compared with WT mice, the percentage and absolute number of thymic CD4 ＋ SP cells in miR-126KD mouseincreased obviously （P 〈 0.05 ） , however, the proportion and number of DP ceils were significantly decreased （P 〈 0.05）. Furthermore, the expression of nuclear antigen Ki-67 in thymic CD4^＋ SP cells from miR-126KD mouse abnormally increased and the apoptosis of thymic CD4^＋ SP ceils decreased （P 〈 0.05 ） ; Conversely, the expression of nuclear antigen Ki- 67 in DP cells decreased （ P 〈 0.05 ）. Finally, the expression of p-AKT and p-ERK1/2 in the thymus decreased in miR-126 KD mice （P 〈 0.05）. Conclusions miR-126 knocking down affected development of thymocytes in the thymus, especially on the development of CD4^＋T cells, which could provide an important experimental basis for further exploring the roles of miR-126 in the development and function of thymocytes.

关 键 词：微小RNA-126 基因敲减 胸腺 CD4+T细胞 

分 类 号：R392[医药卫生—免疫学]