p53-Pathway activity and apoptosis in hydrogen sulfide-exposed stem cells separated from human gingival epithelium
p53-Pathway activity and apoptosis in hydrogen sulfide-exposed stem cells separated from human gingival epithelium
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DOI：
10.1111/jre.12011
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发表时间：
2013-06-01
影响因子：
3.5
通讯作者：
Tanaka, T.
中科院分区：
文献类型：
--
作者：
Calenic, B.;Yaegaki, K.;Tanaka, T.
黄春华，黄春华，黄春华，黄春华，等。人类牙龈上皮细胞凋亡的研究进展[j]。[J] journal of chengdu electromechanical college；48: 322 - 330。(C) 2012 John Wiley & Sons A/S背景和目的硫化氢（H2S）是一种挥发性硫化合物，导致生理性口臭。H2S也被报道有牙周病理活动。龈沟上皮是抵抗牙周病原体及其产物的第一道屏障；口腔角质细胞干细胞oksc在维持这一屏障中起关键作用。p53通路负责调节关键的生物事件。细胞凋亡的增加和DNA修复的细胞周期阻滞可以影响角质细胞干细胞，直接影响口腔上皮组织的结构。然而，H2S、p53活性和OKSCs之间的联系尚未得到充分探讨。本研究的主要目的是探讨暴露于H2S后oksc中p53通路的影响。材料与方法从人牙龈上皮中分离出OKSCs，与生理水平的H2S孵育24和48小时。流式细胞术检测细胞凋亡和线粒体膜电位。采用特异性elisa检测细胞色素c、总p53、磷酸化p53和caspase活性。采用定量RT-PCR检测p53通路基因活性。结果在H2S作用下，细胞凋亡水平显著升高，尤其是在48h后（36.95 +/- 1.91% vs. 4.77 +/- 0.74%）。caspase 9和3被激活，而caspase 8的活性仍然很低。与对照组相比，总p53活性，特别是丝氨酸46处磷酸化的p53活性显著增强（48小时分别为47.11 +/- 9.84单位/mL vs. 1.5 +/- 0单位/mL和32.22 +/- 10.23单位/mL vs. 0.15 +/- 0单位/mL）。在p53通路基因中，凋亡相关基因（即磷酸酶和紧张素同源物（PTEN）、b细胞CLL/淋巴瘤2 （BCL2）、sirtuin 3 （SIRT3）和caspases）与对照组相比显著升高。此外，细胞周期进展基因[即E2F转录因子（E2F）家族和组蛋白去乙酰化酶（HDAC）]和dna修复基因[即生长阻滞和dna损伤诱导，γ (GADD45G)家族和丝氨酸/苏氨酸蛋白激酶Chk2 (CHEK2)]也有所增加。结论经H2S孵育后，OKSCs表达多种p53相关基因，包括程序性细胞死亡、细胞周期控制和dna修复基因。
Calenic B, Yaegaki K, Ishkitiev N, Kumazawa Y, Imai T, Tanaka T. p53-Pathway activity and apoptosis in hydrogen sulfide-exposed stem cells separated from human gingival epithelium. J Periodont Res 2013; 48: 322-330. (C) 2012 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Background and Objective Hydrogen sulfide (H2S) is a volatile sulfur compound responsible for physiological halitosis. H2S was also reported as having periodontal pathologic activities. Gingival crevicular epithelium is the first barrier against periodontal pathogens and their products; oral keratinocyte stem cells OKSCs play key roles in maintaining this barrier. The p53 pathway is responsible for regulating key biological events. Increased apoptosis and cell-cycle arrest of DNA repair can affect keratinocyte stem cells, having a direct impact on the architecture of the oral epithelial tissue. However, the link between H2S , p53 activity and OKSCs has not yet been fully explored. The main objective of the present study was to explore the implications of the p53 pathway in OKSCs following exposure to H2S . Material and Methods OKSCs were isolated from human gingival epithelium and incubated with physiological levels of H2S for 24 and 48h. Apoptosis and the mitochondrial membrane potential were detected using flow cytometry. Cytochrome c, total p53, phosphorylated p53 and caspase activity were assessed using specific ELISAs. p53 Pathway gene activity was assayed using quantitative RT-PCR. Results The levels of apoptosis were significantly increased following incubation in the presence of H2S, especially after 48h (36.95 +/- 1.91% vs. 4.77 +/- 0.74%). Caspases 9 and 3 were activated, whereas caspase-8 activity remained low. Total p53 activity and particularly phosphorylated p53 at serine 46, were significantly enhanced compared with controls (47.11 +/- 9.84units/mL vs. 1.5 +/- 0units/mL and 32.22 +/- 10.23units/mL vs. 0.15 +/- 0units/mL, respectively, at 48h). Among p53 pathway genes, apoptosis-related genes [i.e. phosphatase and tensin homolog ( PTEN ), B-cell CLL/lymphoma 2 ( BCL2), sirtuin 3 ( SIRT3) and caspases]) were dramatically increased when compared with controls. Moreover, cell-cycle progression genes [i.e. E2F transcription factor (E2F) family and histone deacetylase ( HDAC )] and DNA-repair genes [i.e. growth arrest and DNA-damage-inducible, gamma ( GADD45G ) family and serine/threonine-protein kinase Chk2 ( CHEK2)] were also increased. Conclusion Following incubation with H2S , OKSCs express multiple p53-associated genes, including programmed cell death, cell-cycle control and DNA-repair genes.