These tools will no longer be maintained as of December 31, 2024. Archived website can be found here. PubMed4Hh GitHub repository can be found here. Contact NLM Customer Service if you have questions.


Pubmed for Handhelds

PUBMED FOR HANDHELDS

Search MEDLINE/PubMed

  • Title: [Expression and significance of hypoxia-inducible factor 1α in endplate chondrocytes of rats].
    Author: Huang Z, Wang Y, Ma K.
    Journal: Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi; 2017 Mar 15; 31(3):351-356. PubMed ID: 29806267.
    Abstract:
    OBJECTIVE: To explore the expression and significance of hypoxia-inducible factor 1α (HIF-1α) in endplate chondrocytes, and to study the relations between HIF-1α expression and endplate chondrocytes apoptosis. METHODS: Eight Sprague Dawley rats were selected to obtain the L 1-5 intervertebral disc endplate; the endplate chondrocytes were isolated by enzyme digestion method, and the endplate chondrocytes at passage 3 were cultured under 20% O 2 condition (group A), and under 0.5% O 2 condition (group B). Cell morphology was observed by inverted phase contrast microscope and cell apoptosis was detected using flow cytometry after cultured for 24 hours; the mRNA expression of HIF-1α was detected by real-time fluorescent quantitative PCR, the protein expressions of HIF-1α, Bax, and Bcl-2 by Western blot. Gene clone technology to design and synthesize two siRNAs based on the sequence of HIF-1α mRNA. HIF-1α specific RNAi sequence compound was constructed and transfected into cells. The transfected endplate chondrocytes at passage 3 were cultured under 0.5% O 2 condition in group C and group D (HIF-1α gene was silenced). After cultured for 24 hours, cells were observed via immunofluorescence staining of HIF-1α, and cell apoptosis was detected using flow cytometry. Meanwhile, the mRNA expressions of HIF-1α, collagen type II (COL II), Aggrecan, and SOX9 were detected by real-time fluorescent quantitative PCR, and the protein expressions of HIF-1α, Bax, and Bcl-2 by Western blot. RESULTS: At 24 hours after culture, small amount of vacuoles necrotic cells could be observed in group A and group B; there was no significant difference in apoptosis rate between groups A and B ( t=1.026, P=0.471), and HIF-1α mRNA and protein expressions in group B were significantly higher than those in group A ( t=22.672, P=0.015; t=18.396, P=0.013), but, there was no significant difference in protein expressions of Bax and Bcl-2 between groups A and B ( t=0.594, P=0.781; t=1.251, P=0.342). The number of vacuolar necrosis cells in group D was significantly higher than that in group C, and HIF-1α positive cells were observed in group D. The apoptosis rate of group D was significantly higher than that of group C ( t=27.143, P=0.002). The mRNA expressions of HIF-1α, COL II, Aggrecan, and SOX9 in group D were significantly lower than those in group C ( t=21.097, P=0.015; t=34.829, P=0.002; t=18.673, P=0.022; t=31.949, P=0.007). The protein expressions of HIF-1α and Bcl-2 in group D were significantly lower than those in group C ( t=37.648, P=0.006; t=16.729, P=0.036), but the protein expression of Bax in group D was significantly higher than that in group C ( t=25.583, P=0.011). CONCLUSION: HIF-1α mRNA expression is up-regulated under hypoxia condition, which will increase the hypoxia tolerance of endplate chondrocytes. Cell apoptosis is suppressed by the activation of HIF-1α in endplate chondrocytes under hypoxia condition. 目的: 探讨低氧诱导因子 1α（hypoxia-inducible factor 1α，HIF-1α）基因在大鼠终板软骨细胞中的表达情况及其与终板软骨细胞凋亡的关系。. 方法: 取 8 只 10 周龄 SPF 级雄性 SD 大鼠 L 1～5 腰椎终板软骨组织，采用酶消化法分离培养终板软骨细胞并传代，取第 3 代细胞进行实验。首先将终板软骨细胞分为两组，分别于 20% O 2 常氧条件下（A 组）以及 0.5% O 2 低氧下条件（B 组）培养；培养 24 h，倒置相差显微镜观察细胞形态，流式细胞仪检测细胞凋亡率，实时荧光定量 PCR 检测 HIF-1α 基因表达，Western blot 检测凋亡蛋白 HIF-1α、Bax、Bcl-2 表达。然后，构建 Lipofectamin TM2000 载体试剂和 HIF-1α siRNA/ Lipofectamin TM2000 载体混合剂分别转染终板软骨细胞后，于 0.5% O 2 低氧条件下培养（分别为 C、D 组）。培养 24 h 后，倒置相差显微镜下观察细胞形态，行 HIF-1α 免疫荧光染色观察，流式细胞仪检测细胞凋亡率，实时荧光定量 PCR 检测 HIF-1α、Ⅱ 型胶原、蛋白多糖、SOX9 基因表达，Western blot 检测凋亡蛋白 HIF-1α、Bax、Bcl-2 表达。. 结果: 培养 24 h 后，A、B 组均见少量空泡坏死细胞；细胞凋亡率比较差异无统计学意义（ t=1.026， P=0.471）；B 组 HIF-1α 基因及蛋白相对表达量显著高于 A 组（ t=22.672， P=0.015； t=18.396， P=0.013）；两组 Bax、Bcl-2 蛋白表达比较，差异均无统计学意义（ t=0.594， P=0.781； t=1.251， P=0.342）。低氧培养 24 h 后，D 组空泡坏死细胞较 C 组增多，且可见大量 HIF-1α 阳性终板软骨细胞；与 C 组相比，D 组细胞凋亡率显著增高（ t=27.143， P=0.002），D 组 HIF-1α、Ⅱ型胶原、蛋白多糖、SOX9 基因表达较 C 组显著降低（ t=21.097， P=0.015； t=34.829， P=0.002； t=18.673， P=0.022； t=31.949， P=0.007），HIF-1α、Bcl-2 蛋白相对表达量较 C 组均显著降低（ t=37.648， P=0.006； t=16.729， P=0.036），而 Bax 蛋白表达较 C 组提高（ t=25.583， P=0.011）。. 结论: 缺氧条件下，终板软骨细胞中 HIF-1α 表达上调，以提高细胞耐氧性；HIF-1α 可抑制终板软骨细胞在低氧环境中发生凋亡。.
    [Abstract] [Full Text] [Related] [New Search]