92-kD gelatinase is released by the prolonged survival neutrophils in gingival crevicular fluid and cleaves the extracellar domain of recombinant 180-kD bullous pemphigoid autoantigen.

92-kD 明胶酶由龈沟液中延长存活的中性粒细胞释放，并裂解重组 180-kD 大疱性类天疱疮自身抗原的胞外结构域。

• 批准号: 07672087
• 负责人: SHIMOJIMA Takahiro
• 金额: $ 1.41万
• 依托单位: Meikai University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07672087/
• 关键词: periodontitis; junctional epithelium; hemidesmosome; neutrophils; 92-kD gelatinase; apoptosis; 92kゼラチナーゼ

This report presents the results of experiments designed to examine the possibility that the exposure of neutrophils to agents known to regulate inflammatory reactions can influence neutrophils survival and interfere with the physiologic process of apoptosis these cells. Moreovere, we examined whether neutrophils were a possible source of the 92-kD gelatinase and this enzyme cleaved hemidesmosomal elements, the extracelular domain of recombinant 180-kD bullous pemphigoid (BP) autoantigen, a transmembrane molecule of the epidermal hemidesmosome. After 72 hrs of culture, 100 ng/mL LPS treated neutrophils showed a survival of 96.7%(]SY+-[)10.5%, and also when neutrophils were treated with GCF samples, at 72 hrs the survival percentages were for AP-derived GCF 42.7%(]SY+-[)3.8%. Based on the migration patterns of GCF gelatinase on SDS-PAGE and the effects of APMA activation, the major GCF enzymes were identified as the latent 92-kD progelatinase that comigrated with progelatinase from lysed neutrophils, and the faster migrating activated forms. Patients with periodontitis showed higher levels of total (active plus latent) gelatinase activity than control patients (median gelatinase activity of 1843 units for AP,2241 units for EOP and 452 units for controls). Furthermore, to assess if 92-kD gelatinase contributes to the destruction of hemidesmosomal elements, we incubated purified, activated enzyme with a 42-kD recombinant fusion protein, including a large portion of the first and largest collagenous domain. Based on a decrease of -8-kD,92-kD gelatinase cleaved recombinant BP180 at the beginning of the collagenous ectodomain, and as for gelatin, degraded the collagenous sequences into fragments too small to be seen on the gel. In conclusion, our findings suggest that the release of 92-kD gelatinase by intact senescent neutrophils in GCF may be a critical event in pocket formation and its development seen in periodontal disease.

本报告介绍了旨在检查中性粒细胞暴露于已知调节炎症反应的药物可能影响中性粒细胞存活并干扰这些细胞凋亡的生理过程的可能性的实验结果。此外，我们检查了中性粒细胞是否是 92-kD 明胶酶的可能来源，并且该酶切割半桥粒元件，即重组 180-kD 大疱性类天疱疮 (BP) 自身抗原（表皮半桥粒的跨膜分子）的细胞外结构域。培养 72 小时后，100 ng/mL LPS 处理的中性粒细胞显示出 96.7%(]SY+-[)10.5% 的存活率，并且当中性粒细胞用 GCF 样品处理时，72 小时时 AP 衍生的 GCF 的存活率为 42.7%(]SY+-[)3.8%。根据 GCF 明胶酶在 SDS-PAGE 上的迁移模式和 APMA 激活的影响，主要的 GCF 酶被鉴定为与裂解的中性粒细胞中的原明胶酶共迁移的潜在 92-kD 原明胶酶，以及迁移更快的激活形式。牙周炎患者的总（活性加潜伏）明胶酶活性水平高于对照患者（AP 中位明胶酶活性为 1843 单位，EOP 为 2241 单位，对照为 452 单位）。此外，为了评估 92-kD 明胶酶是否有助于破坏半桥粒元件，我们将纯化的活化酶与 42-kD 重组融合蛋白（包括第一和最大胶原结构域的大部分）一起孵育。基于-8-kD的减少，92-kD明胶酶在胶原胞外域的起始处切割重组BP180，并且对于明胶，将胶原序列降解成太小而在凝胶上看不到的片段。总之，我们的研究结果表明，GCF 中完整的衰老中性粒细胞释放 92-kD 明胶酶可能是牙周病中牙袋形成及其发展的关键事件。