發(fā)布時間：2019-06-03 02:25

【摘要】： 目的：通過提取白色念珠菌刺激成分與ECV304細(xì)胞相互作用,觀察其對細(xì)胞增殖的影響,以解析白色念珠菌的致病性,以期揭示口腔念珠菌性白斑的發(fā)病機(jī)理。方法：體外培養(yǎng)人臍靜脈內(nèi)皮細(xì)胞(ECV304)模擬人口腔粘膜上皮細(xì)胞。培養(yǎng)白色念珠菌并提取白色念珠菌的上清液和滅活菌液,無菌驗(yàn)證后白色念珠菌刺激成分實(shí)驗(yàn)分組(上清液組、滅活菌液組、上清液和滅活菌液混合組)與ECV304細(xì)胞共培養(yǎng)。本課題具體從以下四個實(shí)驗(yàn)進(jìn)行研究探索：1、通過倒置顯微鏡觀察白色念珠菌上清液和滅活菌液對ECV304細(xì)胞密度變化的影響；2、通過MTT比色實(shí)驗(yàn)測定白色念珠菌刺激成分對ECV304細(xì)胞增殖的影響；3、通過細(xì)胞生長曲線測定白色念珠菌刺激成分對ECV304細(xì)胞增殖的影響；4、通過流式細(xì)胞術(shù)測定白色念珠菌刺激成分對ECV304細(xì)胞周期的影響。 結(jié)果：倒置顯微鏡觀察發(fā)現(xiàn)4倍稀釋的白色念珠菌上清液實(shí)驗(yàn)組細(xì)胞密度明顯增高；而4倍稀釋的白色念珠菌滅活菌液實(shí)驗(yàn)組細(xì)胞密度與對照組相比則無顯著差異。MTT比色實(shí)驗(yàn)觀察發(fā)現(xiàn)4倍稀釋的白色念珠菌上清液實(shí)驗(yàn)組的細(xì)胞0D值與對照組相比差異最為顯著。細(xì)胞生長曲線揭示在相同濃度、不同時間的培養(yǎng)條件下,隨著時間的增長,不同組分對細(xì)胞促增殖影響基本呈正相關(guān),以上清液組在作用48h時促增殖作用最為明顯；流式細(xì)胞術(shù)研究發(fā)現(xiàn)上清液和滅活菌液分別作用細(xì)胞40h后,上清液組細(xì)胞S期、G2／M期所占百分比顯著升高,反映細(xì)胞增殖活力指數(shù)PI增高,與對照組相比,有顯著性差異(P＜O.05)。而滅活菌液組的PI值無顯著性差異(乃O.05)。 結(jié)論：1.白色念珠菌的上清液和白色念珠菌的滅活菌液在一定程度上均可引起ECV30細(xì)胞增殖。實(shí)驗(yàn)表明,上清液作用明顯大于菌體作用。 2.白色念珠菌的代謝產(chǎn)物可引起ECV304細(xì)胞周期的改變。 3.白色念珠菌的代謝產(chǎn)物可引起ECV304細(xì)胞的增殖。
[Abstract]:Aim: to investigate the pathogenicity of candida albicans by extracting the stimulating components of candida albicans and interacting with ECV304 cells in order to reveal the pathogenesis of oral candida leukoplakia. Methods: human umbilical vein endothelial cells (ECV304) were cultured in vitro to simulate human oral epithelial cells. The culture of candida albicans was cultured and the supernatant and inactivated liquid of candida albicans were extracted. After aseptic verification, the stimulating components of candida albicans were divided into three groups (culture fluid group, inactivated bacteria solution group, culture medium and inactivated liquid mixed group) and ECV304 cells were co-cultured. In this paper, the following four experiments were carried out: 1. The effects of culture medium and inactivated liquid of candida albicans on the density of ECV304 cells were observed by inverted microscope. 2. The effect of candida albicans stimulating components on the proliferation of ECV304 cells was determined by MTT colorimetric assay, and the effect of candida albicans stimulating components on the proliferation of ECV304 cells was determined by cell growth curve. 4. The effect of stimulating components of candida albicans on ECV304 cell cycle was determined by flow cytometry. Results: the cell density of the experimental group with 4 times dilution of candida albicans was significantly increased by inverted microscope. However, there was no significant difference in cell density between the experimental group and the control group. The cell density of the experimental group was not significantly different from that of the control group. MTT colorimetric assay showed that the cell 0D value of the experimental group was similar to that of the control group. The difference is the most significant. The cell growth curve showed that under the same concentration and different time culture conditions, with the increase of time, the effect of different components on the proliferation of cells was basically positively correlated, and the above liquid group had the most obvious effect on cell proliferation at 48 h. Flow cytometry showed that the percentage of S phase and G2 / M phase in the culture medium group was significantly higher than that in the control group, which reflected the increase of cell proliferation activity index (PI), which was significantly higher than that in the control group after 40 hours of treatment with the culture medium and the inactivated bacteria solution, respectively. the percentage of the cells in the S phase and G2 / M phase increased significantly. There was significant difference (P < 0.05). However, there was no significant difference in PI value in inactivated bacteria group (0.05). Conclusion: 1. To a certain extent, the culture fluid of candida albicans and the inactivated solution of candida albicans could induce the proliferation of ECV30 cells. The results showed that the effect of the culture medium was significantly greater than that of the bacteria. two銆，

本文編號：2491603