牙龈卟啉单胞菌唾液酸酶基因突变株黏附与侵入KB细胞能力研究

Study on the adhesive and invasive potential of Porphyromonas gingivalis PG0352 mutant strain interacting with KB cell

目的研究PG0352基因缺失对牙龈卟啉单胞菌(P.gingivalis)黏附和侵入人口腔上皮癌KB细胞能力的影响。方法本研究于2014年7—12月在中国医科大学口腔医学院中心实验室完成。实验组:将P.gingivalis W83菌株(W83菌株组)和PG0352基因突变株(PG0352突变株组)分别以100∶1的比例与KB细胞共培养2 h,制备P.gingivalis与KB细胞的黏附和侵入模型;对照组:单纯KB细胞培养。透射电镜观察KB细胞表面及细胞内是否有P.gingivalis存在;用PBS清除未黏附于KB细胞的细菌,裂解细胞后涂板检测P.gingivalis黏附和侵入KB细胞情况,抗生素保护法检测P.gingivalis侵入KB细胞情况。结果透射电镜结果发现,W83菌株组和PG0352突变株组KB细胞内均有细菌侵入;通过涂板后菌落计数发现P.gingivalis W83菌株和PG0352基因突变株对KB细胞的黏附率分别为(15.559±2.020)%和(9.309±1.750)%,侵入率分别为(0.651±0.287)%和(1.517±0.233)%,两者差异均有统计学意义(均P<0.05)。结论 P.gingivalis W83菌株和PG0352基因突变株均能够黏附和侵入KB细胞;与W83菌株相比,PG0352基因突变株黏附上皮细胞的能力较弱而侵入能力较强。

ObjectiveTo study the adhesive and invasive potential of PG0352 mutant strain interacting with KB cell.MethodsThis study was carried out at Center Laboratory of School and Hospital of Stomatology,China Medical University from July to December 2014.Porphyromonas gingivalis（P.gingivalis）W83 or PG0352 mutant strain and KB cells were cocultured by MOI 100∶1 for 2 h.Transmission electron microscopy was used to see whether P.gingivalis W83 and PG0352 mutant strain existed in KB cell.P.gingivalis in the coculture medium were removed,and P.gingivalis,which adhered to and invaded KB cell were released after lysis KB cells and then P.gingivalis were coated to the BHI plate.The invasive ability of P.gingivalis W83 and PG0352 mutant strain was measured by means of an antibiotic protection assay.ResultsBoth P.gingivalis W83 and PG0352 mutant strain adhered to and entered KB cells.（15.559 ±2.020）% P.gingivalis W83 and（9.309 ± 1.750）% PG0352 mutant strain adhered to KB cell,and（0.651 ± 0.287）% P.gingivalis W83 and（1.517 ± 0.233）% PG0352 mutant strain invaded into KB cell,both with statistical difference.ConBoth W83 and PG0352 mu-clusionBoth P.gingivalis W83 and PG0352 mutant strain can adhere to and enter KB cell.Compared to P.gingivalis W83,less PG0352 mutant strain can adhere to KB cell and more PG0352 mutant strain can invade into KB cell.