牙龈卟啉单胞菌感染内皮细胞导致白介素-8表达升高的胞内机制初探

Intracellular mechanisms of Porphyromonas gingivalis induced interleukin-8 upregulation in endothelial cells

目的:探讨在牙龈卟啉单胞菌(Porphyromonas gingivalis,Pg)感染血管内皮细胞过程中引起白细胞介素(interleukin,IL)-8表达升高的胞内机制。方法:厌氧培养Pg,原代培养人脐静脉血管内皮细胞,用RNA干扰技术分别制作核苷酸结合寡聚化结构域样受体1(nucleotide binding oligomerization domain-containing protein 1,NOD1)、核苷酸结合寡聚化结构域样受体2(NOD-containing protein 2,NOD2)基因沉默的内皮细胞模型,用Pg感染正常的和基因沉默的内皮细胞,ELISA法检测细胞培养上清液中的IL-8水平,RT-PCR检测IL-8、NOD1、NOD2的mRNA和蛋白表达水平,Western blot法检测NOD1、NOD2的蛋白表达水平。用NOD1和NOD2的激动剂DAP和MDP分别激活内皮细胞,并检测IL-8、NOD1、NOD2的mRNA和蛋白表达水平。结果:正常人脐静脉血管内皮细胞受Pg感染后IL-8的mRNA和蛋白表达高于未感染的细胞,NOD1、NOD2表达亦均上调;NOD1、NOD2基因分别沉默的内皮细胞模型在受Pg感染后,其IL-8表达的mRNA和蛋白水平明显低于Pg感染的正常内皮细胞(P<0.01);NOD1和NOD2激动剂(DAP/MDP)激活内皮细胞后,IL-8表达的mRNA和蛋白表达水平均上调(P<0.05)。结论:Pg可诱导正常人脐静脉内皮细胞IL-8表达上调,NOD1和NOD2作为胞内模式识别受体在其中发挥重要作用。

Objective：To find out the mechanisms by which Porphyromonas gingivalis （P. gingivalis） regulates interleukin （IL）-8 expression in endothelial cells. Methods： P. gingivalis was applied to in- fect human umbilical vein endothelial cells （HUVECs）, and the expressions of nucleotide binding oligomerization domain （NOD） 1, NOD2 and IL-8 were detected at mRNA and protein levels. Then the NOD1/NOD2 gene was silenced by RNA interference targeting NOD1 or NOD2 mRNA, followed by P. gingivalis treatment in the HUVECs, and the expression levels of NOD1/NOD2 and IL-8 were examined by real-time PCR, Western-blot or ELISA. In order to confirm the relationship between NOD1/NOD2 and IL-8 in the HUVECs, the agonists for NOD1 and NOD2, DAP and MDP were used in this study. Results： P. gingivalis was activated the expressions of NOD1 and NOD2 in the HUVECs. Meanwhile, IL-8 expression level was also upreguated after P. gingivalis treatment （ P 〈 0.01 ）. Knocking down of NODI or NOD2, the expression level of NOD1 or NOD2 was decreased, and P. gingivalis-induced IL-8 expression was attenuated in the HUVECs （P 〈 0.01 ）. Compared with normal cells, the NOD1 and NOD2 agonists, DAP and MDP, successfully increased IL-8 expression respectively （P 〈 0.05 ）. Conclusion： NOD1 and NOD2 play an important role in the inflammation of HUVECs caused by P. gingivalis in the expression of IL-8.