Morozumi T, Kubota T, Sugita N, Ohsawa Y, Yamazaki K, Yoshie H
Department of Periodontology, Faculty of Dentistry, Niigata University, Japan.
J Periodontal Res. 2001 Jun;36(3):160-8. doi: 10.1034/j.1600-0765.2001.360304.x.
Differential gene expression was examined in human neutrophils stimulated with lipopolysaccharide from Porphyromonas gingivalis (P. gingivalis-LPS) using RNA fingerprinting by arbitrarily primed polymerase chain reaction (RAP-PCR). LPS from Escherichia coli (E. coli-LPS) was used as control. More than 200 differently expressed transcripts were found in 8 subjects showing differential regulation at the transcriptional level. Densitometric analyses revealed that 42-100 genes were upregulated, while 53-116 genes were downregulated by P. gingivalis-LPS compared with E. coli-LPS. The results of sequencing identification showed the presence of supervillin (SVIL) and vascular endothelial growth factor (VEGF) genes in the clones which were upregulated by P. gingivalis-LPS. Consequently, semiquantitative analyses proved that the level of mRNA for SVIL and VEGF were significantly higher in P. gingivalis-LPS-stimulated neutrophils than in other bacterial (E. coli, Actinobacillus actinomycetemcomitans, Prevotella intermedia) LPS- or synthetic lipid A-stimulated neutrophils. Our findings suggest that an elevated mRNA expression for SVIL could be associated with impaired function of neutrophils when stimulated by P. gingivalis-LPS. Further, the VEGF mRNA over-expression might be related to the pathogenesis of P. gingivalis-associated periodontitis. The RAP-PCR technique used in this study enabled us to identify a number of P. gingivalis-LPS regulated genes which hitherto have not been reported.
利用任意引物聚合酶链反应(RAP-PCR)进行RNA指纹分析,检测牙龈卟啉单胞菌脂多糖(P. gingivalis-LPS)刺激的人中性粒细胞中的差异基因表达。以大肠杆菌脂多糖(E. coli-LPS)作为对照。在8名受试者中发现了200多个差异表达的转录本,这些转录本在转录水平上呈现差异调节。密度分析显示,与E. coli-LPS相比,P. gingivalis-LPS上调了42 - 100个基因,而下调了53 - 116个基因。测序鉴定结果表明,在P. gingivalis-LPS上调的克隆中存在 supervillin(SVIL)和血管内皮生长因子(VEGF)基因。因此,半定量分析证明,在P. gingivalis-LPS刺激的中性粒细胞中,SVIL和VEGF的mRNA水平显著高于其他细菌(大肠杆菌、伴放线放线杆菌、中间普氏菌)LPS或合成脂多糖A刺激的中性粒细胞。我们的研究结果表明,当受到P. gingivalis-LPS刺激时,SVIL的mRNA表达升高可能与中性粒细胞功能受损有关。此外,VEGF mRNA的过度表达可能与牙龈卟啉单胞菌相关性牙周炎的发病机制有关。本研究中使用的RAP-PCR技术使我们能够鉴定出许多迄今尚未报道的受P. gingivalis-LPS调控的基因。