Role for fimbriae and lysine-specific cysteine proteinase gingipain K in expression of interleukin-8 and monocyte chemoattractant protein in Porphyromonas gingivalis-infected endothelial cells
Role for fimbriae and lysine-specific cysteine proteinase gingipain K in expression of interleukin-8 and monocyte chemoattractant protein in Porphyromonas gingivalis-infected endothelial cells
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DOI：
10.1128/iai.70.1.268-276.2002
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发表时间：
2002-01-01
影响因子：
3.1
通讯作者：
Genco, CA
中科院分区：
文献类型：
--
作者：
Nassar, H;Chou, HH;Genco, CA
最近的横断面和前瞻性流行病学研究表明，牙周病和动脉粥样硬化和人类冠心病之间的关联。以前，我们已经建立了牙周病原体牙龈卟啉单胞菌是能够入侵主动脉，心脏和人脐静脉内皮细胞（HUVEC）。由于动脉粥样硬化是一种慢性炎症反应在血管壁开始，牙龈卟啉单胞菌与内皮细胞的相互作用和随后的宿主细胞对感染的反应可能是重要的动脉粥样硬化的发病机制。在这项研究中，我们研究了牙龈卟啉单胞菌感染的HUVEC的趋化因子白细胞介素-8（IL-8）和单核细胞趋化蛋白1（MCP-1）的表达的后果。发现HUVEC组成性地产生低水平的IL-8和MCP-1。将牙龈卟啉单胞菌菌毛蛋白特异性肽、脂多糖（LPS）或热灭活的全细胞制剂添加至HUVEC刺激适度的IL-8和MCP-1应答。相反，HUVEC与活牙龈卟啉单胞菌菌株A7436、33277或381的共培养消除了IL-8和MCP-1应答。IL-8和MCP-1产生的抑制不依赖于细菌粘附，因为用非粘附性牙龈卟啉单胞菌fimA突变体DPG 3或当牙龈卟啉单胞菌在加入HUVEC之前与菌毛蛋白肽抗血清预孵育时获得了类似的结果。此外，治疗1。用细胞松弛素D感染的HUVEC，阻止牙龈卟啉单胞菌的侵袭，也消除了组成性IL-8和MCP-1的反应。用E. coliLPS刺激了强烈的IL-8和MCP-1反应，当刺激的细胞与活牙龈卟啉单胞菌共培养时，这些反应被消除。通过RNA酶保护试验分析牙龈卟啉单胞菌感染的HUVEC培养物，显示IL-8转录物相对于未感染的HUVEC增加。在加入HUVEC之前用蛋白酶抑制剂预处理牙龈卟啉单胞菌，防止了牙龈卟啉单胞菌感染的HUVEC中IL-8和MCP-1产生的抑制，表明抑制是蛋白水解介导的。与牙龈卟啉单胞菌突变体赖氨酸特异性半胱氨酸蛋白酶（牙龈卟啉单胞菌蛋白酶K [Kgp]）缺陷的HUVEC共培养导致IL-8转录和蛋白表达相对于与牙龈卟啉单胞菌野生型菌株共培养的HUVEC中观察到的增加。这些结果表明，牙龈卟啉单胞菌可以通过菌毛和牙龈蛋白酶介导的机制在时间上调节内皮细胞中的趋化因子反应。
Recent cross-sectional and prospective epidemiological studies have demonstrated an association between periodontal disease and atherosclerosis and human coronary heart disease. Previously, we have established that the periodontal pathogen Porphyromonas gingivalis is capable of invading aortic, heart, and human umbilical vein endothelial cells (HUVEC). Since atherosclerosis is a chronic inflammatory response initiated at the vascular wall, interactions of P. gingivalis with endothelial cells and the subsequent host cell response to infection may be important in the pathogenesis of atherosclerosis. In this study we examined the consequences of P. gingivalis infection of HUVEC on the expression of the chemokines interleukin-8 (IL-8) and monocyte chemotactic protein 1 (MCP-1). HUVEC were found to constitutively produce low levels of IL-8 and MCP-1. The addition of P. gingivalis fimbrillin-specific peptides, lipopolysaccharides (LPS), or heat-killed whole cell preparations to HUVEC stimulated modest IL-8 and MCP-1 responses. In contrast, coculture of HUVEC with live P. gingivalis strain A7436, 33277, or 381 abolished the IL-8 and MCP-1 responses. Inhibition of IL-8 and MCP-1 production was not dependent on bacterial adherence since similar results were obtained with the nonadherent A gingivalis fimA mutant DPG3 or when P. gingivalis was preincubated with fimbrillin peptide antisera prior to the addition to HUVEC. Furthermore, treatment of 1. gingivalis-infected HUVEC with cytochalsin D, which prevented A gingivalis invasion, also abolished the constitutive IL-8 and MCP-1 responses. Treatment of HUVEC with E. coli LPS stimulated robust IL-8 and MCP-1 responses that were abolished when stimulated cells were cocultured with live A gingivalis. Analysis of A gingivalis-infected HUVEC cultures by an RNase protection assay revealed an increase in the IL-8 transcript relative to uninfected HUVEC. Pretreatment of A gingivalis with protease inhibitors prior to the addition to HUVEC prevented the inhibition of IL-8 and MCP-1 production in P. gingivalis-infected HUVEC, indicating that the inhibition was proteolytically mediated. Coculture of HUVEC with a A gingivalis mutant deficient in lysine-specific cysteine proteinase (gingipain K [Kgp]) resulted in an increase in both IL-8 transcription and protein expression relative to that observed in HUVEC cocultured with the A gingivalis wild-type strain. These results indicate that P. gingivalis can temporally modulate the chemokine response in endothelial cells through both fimbriae and gingipain-mediated mechanisms.