低氧对血清剥夺后山羊颞下颌关节盘细胞凋亡和自噬的影响  

Effects of Hypoxia on the Apoptosis and Autophagy of the Goat Temporomandibular Joint Disc Cells after Serum Deprivation

作　　者：张芳[1] 包广洁[1,2,3] 唐玉尧 刘琳[2,3] 保善英 康宏[1] Zhang Fang;Bao Guangjie;Tang Yuyao;Liu Lin;Bao Shanying;Kang Hong(Institute of Stomatology,Lanzhou University,Lanzhou 730000,China;Key Laboratory of Stomatology of State Ethnic Affairs Commission,Northwest Minzu University,Lanzhou 730030,China;Key Laboratory of Oral Diseases of Gansu Province,Northwest Minzu University,Lanzhou 730030,China)

机构地区：[1]兰州大学口腔医学研究所,兰州730000 [2]西北民族大学口腔医学国家民委重点实验室,兰州730030 [3]西北民族大学甘肃省口腔疾病研究重点实验室,兰州730030

出　　处：《中国细胞生物学学报》2018年第8期1295-1302,共8页Chinese Journal of Cell Biology

基　　金：国家自然科学基金(批准号:81660189);西北民族大学中央高校基本科研业务费资助项目(批准号:2yp2015014;31920170167)资助的课题~~

摘　　要：该研究体外分离山羊颞下颌关节盘(temporomandibular joint disc,TMJ disc)细胞并培养至p2代。通过HE(Hematoxylin-Eosin)染色、Hoechst 33258和丹磺酰戊二胺(dansylcadaverine,MDC)荧光染色观察血清剥夺后细胞的形态学变化以及是否存在自噬和凋亡。随后,通过流式细胞术和实时荧光定量聚合酶链反应(Real-time PCR)分别检测血清剥夺0 h、12 h、24 h、36 h、48 h和60 h后细胞的凋亡率和自噬水平的变化以及加入自噬抑制剂3-MA(3-methyladenine)后的凋亡变化。检测给予血清(10%FBS)或血清剥夺(0%FBS)的细胞在常氧(21%O2)或低氧(2%O2)条件下的凋亡和自噬的变化。结果发现:血清剥夺后,凋亡率随时间的延长逐渐上升,自噬率先上升后下降;当血清剥夺诱导的自噬被3-MA抑制后,细胞的凋亡率明显上升。自噬能够抑制血清剥夺引起的细胞凋亡,说明自噬在细胞的存活中有很重要的作用。与常氧培养的细胞相比,低氧条件下细胞的凋亡率和自噬率均下降。低氧通过降低细胞的过度自噬减少长期血清剥夺引起的细胞凋亡,比常氧更有利于细胞的存活。In this work, they isolated and cultured temporomandibular joint disc（TMJ disc） cells of goat to p2 generation in vitro. The morphological changes of cells were observed by Hematoxylin-Eosin（HE） staining, Hoechst 33258 and dansylcadaverine（MDC） fluorescence staining were used to observe whether autophagy or apoptosis existed after the serum deprivation. After 0 h, 12 h, 24 h, 36 h, 48 h and 60 h of serum deprivation, apoptosis rate and autophagy rate of cells were detected by flow cytometry and Real-time PCR, respectively. Changes of apoptosis was observed after applying 3-methyladenine（3-MA）, an autophagy inhibitor. The changes of apoptosis and autophagy with serum（10% FBS） or serum deprivation（0% FBS） were detected under the conditionsof oxygen（21% O2） or hypoxia（2% O2）. The results showed that the rate of apoptosis increased gradually with the prolongation of the time, and the autophagy first increased and then decreased after the serum deprivation. When the autophagy induced by serum deprivation was inhibited by 3-MA, the rate of apoptosis of cells increased obviously. Autophagy could inhibit the apoptosis induced by serum deprivation, which showed that autophagy played an important role in cell survival. Compared with normoxia-cultured cells, the apoptosis rate and autophagy rate of the cells decreased under hypoxia condition. Hypoxia reduced the apoptosis of cells caused by long-term serum deprivation by reducing excessive autophagy, which was more conducive to cell survival than normoxia.

关 键 词：颞下颌关节盘 自噬 凋亡 血清剥夺 低氧 

分 类 号：R782.6[医药卫生—口腔医学]