Studies on CGRP receptor subtypes and their specific intracellular signaling pathways in the periodontal tissue

牙周组织CGRP受体亚型及其特异性细胞内信号通路的研究

基本信息

批准号：
12671803
负责人：
KAWASE Tomoyuki
金额：
$ 2.11万
依托单位：
Niigata University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671803/
关键词：
CGRP Receptor Phosphorylation cAMP MAPK Calcium

项目摘要

Based on the published data about 3-D configuration of CGRP, we have attempted to produce some CGRP fragments and performed bioassay with those new peptides. However, we could not particularly find any bioactive peptides.1) Production of CGRP fragments and screening - In addition to commercially available human CGRP_<1-37>, CGRP_<8-37>, CGRP_<19-37>, CGRP_<22-37> and [Cys(Acm)^<2,7>]-CGRP, we have produced CGRP_<1-12>, reverse-CGRP_<1-8> and reverse-CGRP_<1-14>. On screening with cAMP production and other indeces, we have found that CGRP_<8-37>, CGRP_<22-37> and [Cys(Acm)^<2,7>]-CGRP have substantial agonistic or antagonistic action. 2) Effects of bioactive CGRP fragments on intracelllar signaling pathways - In osteoblastic MG63 cells, CGRP_<8-37> potently anta gonized CGRP_<1-37> action in cAMP production. CGRP_<22-37>, also had a weak antagonistic action. [Cys(Acm)^<2,7>]-CGRP showed both a medium antagonistic and a weak agonistic action. Similar results were obtained in either gingival fibroblastic Gin-1 or oral epithelial SCC25 cells. 3) Binding affinity of bioactive CGRP fragments - Fluoro-CGRP bound to its receptors in a concentration-dependent fashion, and its binding was specifically replaced with CGRP_<8-37>, but not clearly with [Cys(Acm)^<2,7>]-CGRP. 4) Effects of CGRP fragments on cell proliferation - Any CGRP fragments produced here failed to reproducibly influence the proliferation, but only CGRP_<1-37> weakly stimulated Gin-1 proliferation. 5) Effects of CGRP fragments on cell apoptosis - Any CGRP fragments did not apparently induce apoptosis in any cell types used here. 6) Expression of CGRP receptor subtypes in periodontal tissues - Expression of both CRLR and RAMP-I proteins were detected using those specific antibodies either in periodontal tissues or in any cell types used here.

基于有关CGRP的3-D配置的已发布数据，我们试图生成一些CGRP片段并使用这些新肽进行生物测定。但是，我们找不到任何生物活性肽。1）生产CGRP片段和筛选 - 除了市售的人CGRP_ <1-37>，CGRP_ <8-37>，CGRP_ <19-37>，CGRP_ <22 -37>和[cys（acm）^<2,7>] -CGRP，我们生产了CGRP_ <1-12>，反向cgrp_ <1-8>和反向cgrp_ <1-14>。在使用营地生产和其他indeces筛查时，我们发现CGRP_ <8-37>，CGRP_ <22-37>和[CYS（ACM）^<2,7>] - CGRP具有实质性的激动或拮抗作用。 2）生物活性CGRP片段对成骨细胞MG63细胞中的纤维化信号通路的影响，CGRP_ <8-37>在cAMP生产中有效的Anta Gonatized CGRP_ <1-37>作用。 CGRP_ <22-37>，也具有较弱的拮抗作用。 [CYS（ACM）^<2,7>] - CGRP既显示中等拮抗作用又显示出弱的激动作用。在牙龈成纤维细胞GIN-1或口服上皮SCC25细胞中获得了相似的结果。 3）生物活性CGRP片段的结合亲和力 - 以浓度依赖性方式与其受体结合的氟-CGRP，其结合被CGRP_ <8-37>专门替换，但并不清楚地用[Cys（acm）^<2， 7>] -CGRP。 4）CGRP片段对细胞增殖的影响 - 此处产生的任何CGRP片段都无法重现影响增殖，但仅CGRP_ <1-37>弱刺激的Gin-1增殖。 5）CGRP片段对细胞凋亡的影响 - 任何CGRP片段显然不会诱导此处使用的任何细胞类型的凋亡。 6）CGRP受体亚型在牙周组织中的表达 - 使用牙周组织中的那些特定抗体或此处使用的任何细胞类型中检测到CRLR和RAMP -I蛋白的表达。