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间歇性低压低氧通过促进细胞凋亡抗大鼠胶原诱导性关节炎

史敏, 崔芳, 刘爱京, 李姣, 马慧娟, 程明, 杨晶, 张翼^*

河北医科大学1生理学教研室；第二医院检验科；第二医院免疫风湿科，石家庄 050017

摘要

本文旨在研究慢性间歇性低压低氧(chronic intermittent hypobaric hypoxia, CIHH)对大鼠胶原诱导性关节炎(collagen-induced arthritis, CIA)的影响。雄性成年Sprague-Dawley大鼠50只，随机分为5组：CIHH预处理组(Pre-T)、预处理对照组(Pre-C)、CIHH后处理组(Post-T)、后处理对照组(Post-C)及空白对照组(Con)。Pre-T和Post-T大鼠分别在CIA造模前和造模开始后第12天给予28 d模拟海拔3 000 m (pO2 = 108.8 mmHg，14% O2)、每天5 h的CIHH处理；Pre-C和Post-C不接受CIHH处理，CIA造模处理分别与Pre-T和Post-T相同；Con大鼠不给予任何处理。通过螺旋测微器测定大鼠双后足爪厚度，以关节炎指数(arthritis index, AI)评价关节炎程度；HE染色法观察踝关节组织形态学变化；原位末端标记法(TUNEL)检测膝关节滑膜组织细胞凋亡率；流式细胞术检测脾脏CD3+ T淋巴细胞凋亡率；免疫组化SP法检测滑膜细胞和脾脏淋巴细胞Bcl-2、Bax蛋白表达水平。结果显示：(1) Pre-T大鼠CIA发生率明显低于Pre-C组大鼠(P < 0.05)；Pre-T和Post-T组大鼠的AI值分别明显低于Pre-C和Post-C组大鼠(P < 0.05)。(2) Pre-C及Post-C大鼠踝关节滑膜细胞明显增生，形成血管翳，侵蚀软骨及骨，炎细胞浸润增加；Pre-T和Post-T大鼠关节组织病理改变明显减轻。(3) Pre-T和Post-T大鼠滑膜细胞及脾脏淋巴细胞的凋亡率分别较Pre-C及Post-C大鼠明显增加(P < 0.05)。(4) 与Pre-C及Post-C大鼠相比较，Pre-T和Post-T动物滑膜组织细胞及脾脏淋巴细胞Bcl-2蛋白表达明显降低(P < 0.05)，而Bax蛋白表达则明显升高(P < 0.05)。以上结果表明，CIHH可通过抑制Bcl-2蛋白和增强Bax蛋白表达，促进关节滑膜细胞和淋巴细胞凋亡，从而发挥抗大鼠CIA作用。

关键词： 胶原诱导性关节炎; 慢性间歇性低压低氧; 细胞凋亡; T淋巴细胞; 大鼠

分类号：R392.32

Protection of chronic intermittent hypobaric hypoxia against collagen-induced arthritis in rat through increasing apoptosis

SHI Min, CUI Fang, LIU Ai-Jing, LI Jiao, MA Hui-Juan, CHENG Ming, YANG Jing, ZHANG Yi^*

Department of Physiology, Hebei Medical University； Department of Clinical Laboratory, Second Hospital, Hebei Medical University； 3Department of Immune and Rheumatism, Second Hospital, Hebei Medical University, Shijiazhuang 050017, China

Abstract

The aim of present study was to investigate the effect of chronic intermittent hypobaric hypoxia (CIHH) on collagen-induced arthritis (CIA) in rat. Fifty male adult Sprague-Dawley rats were randomly divided into 5 groups: CIHH pre-treatment group (Pre-T), pre-control group (Pre-C), CIHH post-treatment group (Post-T), post-control group (Post-C) and blank control group (Con). The rats in Pre-T and Post-T groups were exposed to 28 d of hypobaric hypoxia (simulated 3 000 m altitude, 5 h per day, pO2 = 108.8 mmHg, 14% O2) in a hypobaric chamber before and 12 days after CIA induction, respectively. The rats in Pre-C and Post-C groups were only experienced CIA induction, being control groups for Pre-T and Post-T groups, respectively. The rats in Con group were not given any treatment. The thickness of two-hind paw of rat was measured with spiral micrometer and the degree of arthritis was evaluated by arthritis index (AI). Morphological changes of ankle joint were observed through HE staining. The apoptotic rate in synovial tissue was measured by terminal dUTP nick end labeling (TUNEL) and the apoptotic rate of CD3+ T lymphocyte in spleen was measured by flow cytometry technique. The protein expressions of Bcl-2 and Bax were measured using immunohistochemistry SP method. The results showed that incidence rate of CIA in Pre-T rats was lower than that in Pre-C rats (P < 0.05). AI in Pre-T and Post-T rats were smaller than those in Pre-C and Post-C, respectively (P < 0.05). In Pre-C and Post-C rats, there were hyperplasia of synovial cell, pannus forming, infiltration with inflammatory cell, and destroyed cartilage and bone in ankle joint. On the contrary, pathological changes of ankle joint were alleviated significantly in Pre-T and Post-T rats. Compared with Pre-C and Post-C rats, apoptotic rates of synovial cell and T lymphocyte in Pre-T and Post-T rats were increased (P < 0.05). As to the possible anti-apoptosis mechanism, CIHH, no matter before and after CIA induction, decreased Bcl-2 expression and increased Bax expression in joint synovial cells and spleen T lymphocytes (P < 0.05), respectively. In conclusion, CIHH possesses a protective effect against CIA in rat by enhancing apoptosis of synovial cells and T lymphocytes, which may be related to the inhibition of Bcl-2 protein expression and the promotion of Bax protein expression.

Key words: collagen-induced arthritis; chronic intermittent hypobaric hypoxia; cell apoptosis; T lymphocytes; rat

收稿日期：2010-10-26　　录用日期：2011-02-11

通讯作者：张翼　　E-mail: zhyhenry@hotmail.com

引用本文：

史敏, 崔芳, 刘爱京, 李姣, 马慧娟, 程明, 杨晶, 张翼. 间歇性低压低氧通过促进细胞凋亡抗大鼠胶原诱导性关节炎[J]. 生理学报 2011; 63 (2): 115-123.

SHI Min, CUI Fang, LIU Ai-Jing, LI Jiao, MA Hui-Juan, CHENG Ming, YANG Jing, ZHANG Yi. Protection of chronic intermittent hypobaric hypoxia against collagen-induced arthritis in rat through increasing apoptosis. Acta Physiol Sin 2011; 63 (2): 115-123 (in Chinese with English abstract).