低氧暴露对小鼠脾淋巴细胞增殖和凋亡的影响  

Effects of hypoxia exposure on proliferation and apoptosis of mouse spleen lymphocytes

作　　者：郭玉静 胡英 龙启福 许玉珍 李积东 永胜 GUO Yujing;HU Ying;LONG Qifu;XU Yuzhen;LI Jidong;YONG Sheng(Department of Immunology,School of Basic Medicine,Qinghai University School of Medical,Xining 810016,China)

机构地区：[1]青海大学医学院基础医学部免疫学教研室,810016

出　　处：《天津医药》2022年第10期1014-1020,共7页Tianjin Medical Journal

基　　金：国家自然科学基金资助项目(82060295);青海省科技计划项目基金(2019-ZJ-7059)。

摘　　要：目的研究低氧暴露诱导淋巴细胞数量减少与细胞增殖和凋亡的关系。方法从C57BL/6小鼠脾中分离淋巴细胞,将淋巴细胞分别在低氧(1%O2)和常氧(21%O2)环境中培养12、24、48 h。用流式细胞术检测T、B淋巴细胞数量;羧基荧光素乙酰乙酸和AnnexinⅤ-FITC/PI方法分别检测淋巴细胞的增殖和凋亡;扫描电镜观察淋巴细胞形态;实时荧光定量逆转录PCR和Western blot检测凋亡相关因子鼠源B细胞淋巴瘤/白血病-2(bcl-2)、bcl-2同源拮抗剂-杀伤蛋白(Bak)和胱天蛋白酶-3(caspase-3)的mRNA和蛋白表达水平。结果低氧暴露12、24、48 h,可降低T、B淋巴细胞数量,抑制淋巴细胞增殖,促进细胞凋亡;扫描电镜观察显示,同一时间下,低氧组淋巴细胞更早出现凋亡特征形态学变化;低氧暴露12、24、48 h,淋巴细胞Bak和caspase-3的mRNA和蛋白表达水平均上调,而bcl-2的mRNA和蛋白表达水平均下调。结论低氧暴露通过抑制淋巴细胞增殖、促进凋亡,从而介导淋巴细胞数量下降。Objective To investigate the relationship between lymphocyte number reduction,cell proliferation and apoptosis induced by hypoxia exposure.Methods Lymphocytes were isolated from spleen of C57BL/6 mice and cultured in hypoxia(1%O_(2))and normoxia(21%O_(2))for 12,24 and 48 h,respectively.T and B lymphocytes were assessed by flow cytometry.Proliferation and apoptosis of lymphocytes were detected by CFSE and AnnexinⅤ-FITC/PI method,respectively.The morphology of lymphocytes was observed by scanning electron microscope.The mRNA and protein expression levels of apoptosis-related factor bcl-2 homologous antagonist killer protein(Bak),murine B-cell lymphoma/leukemia-2(bcl-2)and cysteine aspartate protease-3(caspase-3)were detected by real-time quantitative reverse transcription PCR(qPCR)and Western blot assay.Results Hypoxic exposure for 12,24 and 48 h can reduce the number of T and B lymphocytes,inhibit the proliferation of lymphocytes and promote cell apoptosis.Scanning electron microscopy showed that at the same time,the characteristic morphological changes of apoptosis appeared earlier in the hypoxic group.After 12,24 and 48 h hypoxia exposure,the mRNA and protein expression levels of Bak and caspase-3 in lymphocytes were up-regulated,while the mRNA and protein expression levels of bcl-2 were down-regulated.Conclusion Hypoxic exposure mediates the decline of lymphocyte numbers by inhibiting lymphocyte proliferation and promoting apoptosis.

关 键 词：缺氧 淋巴细胞减少 细胞增殖 细胞凋亡 bcl-2同源对抗杀伤蛋白 胱天蛋白酶-3 

分 类 号：R392.12[医药卫生—免疫学]