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目的:研究不同理化及生物因素对肺炎克雷伯菌粘附宿主细胞的影响,探讨其粘附宿主细胞的可能机制。方法:用肺炎克雷伯菌临床分离株K.pnueumoniae 03183(K.p03183)建立粘附宿主细胞模型,通过结晶紫染色及扫描电镜观察其粘附效率;以不同理化及生物因素预处理K.p03183,平板菌落计数法观察上述因素对K.p03183粘附细胞的影响;用基因芯片技术,检测HMGN2蛋白预处理细菌对于其粘附相关基因表达的影响。结果:K.p03183可粘附至A549、T24、HeLa等3种上皮细胞系,其粘附率与E.coli 25992接近;氯化锂和盐酸胍、高碘酸钠及热预处理可明显降低K.p03183对于A549和T24的粘附率(P<0.05);同时,胃蛋白酶及HMGN2蛋白预处理K.p03183,也可抑制其对A549和T24的粘附(P<0.01);但胰蛋白酶预处理却能增加细菌对于A549细胞的粘附(P<0.05);同时,HMGN2还可通过上调dksA基因表达,从而干扰细菌粘附细胞。结论:肺炎克雷伯菌可能借助细胞壁及其表面蛋白和碳水化合物等成分粘附至宿主上皮细胞,通过非共价键结合表面蛋白或消除碳水化合物成分,抑制细菌粘附至宿主细胞。
Objective: To study the effects of different physico-chemical and biological factors on the adhesion of Klebsiella pneumoniae to host cells and to explore the possible mechanism of its adhesion to host cells. Methods: The adherent host cell model was established by Klebsiella pneumoniae clinical isolates K.pnueumoniae 03183 (K.p03183). The adhesion efficiency was observed by crystal violet staining and scanning electron microscopy. Pretreatment of K. with different physicochemical and biological factors. p03183 and plate colony counting method were used to observe the effect of these factors on the adhesion of K.p03183 cells. The effect of pretreatment of HMGN2 protein on the expression of adhesion-related genes was detected by gene chip technique. Results: K.p03183 could adhere to three kinds of epithelial cell lines such as A549, T24 and HeLa, and its adhesion rate was close to that of E.coli 25992. Lithium chloride, guanidine hydrochloride, sodium periodate and heat pretreatment could be significantly reduced (P <0.05). At the same time, pepsin and HMGN2 protein pretreatment K.p03183 also inhibited the adhesion to A549 and T24 (P <0.01), but trypsin Pretreatment increased the adhesion of bacteria to A549 cells (P <0.05). Meanwhile, HMGN2 could also interfere with bacterial adhesion cells by up-regulating the expression of dksA gene. Conclusion: Klebsiella pneumoniae may adhere to the host epithelial cells through the cell wall and its surface proteins and carbohydrates, and inhibit bacterial adhesion to host cells by non-covalently binding surface proteins or eliminating carbohydrate components.