Study for periodontal pathogenesis and apoptosis in periodontal tissues induced by volatile sulfur compounds

挥发性硫化合物诱导牙周组织凋亡及牙周发病机制的研究

基本信息

批准号：
17390568
负责人：
YAEGAKI Ken
金额：
$ 10.15万
依托单位：
The Nippon Dental University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2007
项目状态：
已结题

项目摘要

Hydrogen sulfide (H_2S) is one of the main reasons of halitosis and periodontally pathogenic. H_2S causes apoptotic cell death in aorta smooth muscle cells and other tissues, and apoptosis plays an important role in the onset and progress of periodontitis. Therefore, we investigated if H_2S causes apoptosis in human gingival fibroblasts (HGF). Materials and Methods : Necrotic cell were determined using lactate dehydrogenase assay. Apoptosis was ascertained with detecting histone-complexed DNA fragments and a flow cytometry. Caspase 3, a key enzyme in apoptotic signaling, was assessed. Reactive oxygen species (ROS) were also detected and inhibition of superoxide dismutase (SOD) by H_2S was verified. DNA damage was examined with single-cell gel electrophoresis (SCGE). Results : Necrosis was found less than 10% of HGF during 72 hours incubation with 100 ng/ml H_2S, whereas significant increment of DNA fragmentation in cytoplasm was found (p < 0.05). Apoptotic HGF cells were also increased in a flow cytometry. Caspase 3 activity was significantly increased at 72 hours incubation (p < 0.01). Tail length, %DNA in tail and Tail moment obtained by SCGE increased in HGF exposed to H_2S for 48 and 72 hours. Moreover, ROS was significantly increased at 48 and 72 hour incubation with H_2S (p < 0.005 and p < 0.001, respectively), and SOD of HGF was strongly inhibited. Conclusion : H_2S caused apoptosis in HGF, and an inhibition of SOD by H_2S increased ROS, then induced apoptosis and DNA strand breaks.

硫化氢（H_2S）是口腔症和牙周致病性的主要原因之一。 H_2S导致主动脉平滑肌细胞和其他组织中的凋亡细胞死亡，凋亡在牙周炎的发作和进展中起重要作用。因此，我们研究了H_2S是否在人牙龈成纤维细胞（HGF）中引起凋亡。材料和方法：使用乳酸脱氢酶测定法确定坏死细胞。通过检测组蛋白复合的DNA片段和流式细胞术来确定凋亡。评估了caspase 3是凋亡信号传导中的关键酶。还验证了H_2S对超氧化物歧化酶（SOD）抑制活性氧（ROS）。用单细胞凝胶电泳（SCGE）检查了DNA损伤。结果：在72小时与100 ng/ml H_2S孵育期间，坏死率少于HGF的10％，而发现细胞质中DNA片段的显着增加（P <0.05）。在流式细胞仪中，凋亡的HGF细胞也增加了。在72小时孵育时，caspase 3的活性显着增加（p <0.01）。 SCGE获得的尾巴长度，尾部和尾部矩的尾巴矩，在H_2s中增加了48和72小时。此外，在48和72小时与H_2s孵育时ROS显着增加（分别为P <0.005和P <0.001），HGF的SOD被强烈抑制。结论：H_2S引起HGF的凋亡，H_2S对SOD的抑制增加了ROS，然后诱导的凋亡和DNA链断裂。