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Physiological and pathological roles of adhesion molecules expressed on gingival fibroblasts in response to oral microbes

牙龈成纤维细胞表达的粘附分子对口腔微生物的生理和病理作用

基本信息

批准号：
10671762
负责人：
SHIBATA Ken-ichiro
金额：
$ 2.05万
依托单位：
HOKKAIDO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 1999
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10671762/
关键词：
Mycoplasma salivarium ICAM-1 gingival fibroblasts lipoprotein lipopeptide 口腔微生物接着因子

项目摘要

A lipoprotein with a molecular mass of 44 kDa (Lp44) in Mycopiasma lipoproteins possessed the ICAM-1 expression-inducing activity. The N-terminal amino acid sequence of Lp44 was suggested to be CGDPKH-PKSFTEWVA - and the amino group of the N- terminal cysteine is free. The activity of Lp44 was resistant to proteinase K and sensitive to lipoprotein lipase, suggesting that the lipid moiety, but not the proteinous moiety, of Lp44 is involved in the expression of the activity. The lipid moiety of Lp44 was purified by a reverse-phased HPLC. The activity was eluted at around 90% isopropanol. In order to obtain informations on the structure of the active entity, the fractions with the activity were collected and applied to the Infrared (IR) spectrometer. The IR spectra of the fractions revealed the presence of fatty acid alkyl chains and the typical ester bond. Therefore, the structure of the N-terminal lipo-peptide of Lp44 (Lpp44) is speculated to be S-(2,3 -bisacyloxypropyl)-CGDPKHPKSFTEWVA … More -, because it is well known that lipid is bound to SH group of the N-terminal cysteine in lipoproteins from prokaryotesAt present, it remains unknown what the fatty acid(s) is(are). In order to confirm whether Lpp44 possesses the ICAM-1 expression-inducing activity, a lipopeptide, S-(2,3 -bispalmitoyloxypropyl)-CGDPKHPKSF, analogous to Lpp44 was synthesized. The Lpp44 analogue was found to possess the ICAM-1 expression-inducing activity, and to induce IL-6 production by HGF and TNF-α production by THP-1 cells, a monocytic cell line as well. The ICAM-1 expression-inducing activity level of the Lpp44 analogue was **nd to be significantly higher than that of LPS purified from Escherichia coli and Salmonella enteritidis, but lower than that. Of human recombinant TNF-α.Thus, this study demonstrated that Lp44 was one of M. salivarium lipoproteins responsible for the ICAM-1 expression- and cytokine production-inducing activities and its N-terminal lipopeptide was the active entity.Lpp44 is different in the amino acid sequence from, but similar in the structure to MALP-2, S-(2,3 -bisacyloxypropyl)-CGNNDESNISFKEK, which is purified and identified from M. fermentans and is capable of activating monocytes/macrophages to induce production of nitric oxide and TNF-α. Synthetic analogues of N-terminal lipopeptide of E. coli murein lipoproteins are known to be potent macrophage and B cell activators. The structures of these lipopeptides were similar to those of MALP-2 and Lpp44. Therefore, lipoproteins from prokaryotes are speculated to be a potent activator for macrophages, B cells and fibroblasts. Less

Mycopiasma脂蛋白中分子量为44 kDa的脂蛋白（Lp 44）具有ICAM-1表达诱导活性。Lp 44的N-末端氨基酸序列被认为是CGDPKH-PKSFTEWVA -，并且N-末端半胱氨酸的氨基是游离的。Lp 44的活性对蛋白酶K具有抗性，而对脂蛋白脂酶敏感，这表明Lp 44的脂质部分而不是蛋白部分参与活性的表达。通过反相HPLC纯化Lp 44的脂质部分。活性在约90%异丙醇下洗脱。为了获得关于活性实体的结构的信息，收集具有活性的级分并应用于红外（IR）光谱仪。馏分的红外光谱显示存在脂肪酸烷基链和典型的酯键。因此，推测Lp 44（Lpp 44）的N-末端脂肽的结构为S-（2，3-双酰氧基丙基）-CGDPKHPKSFTEWVA ...更多信息 - ，因为众所周知，在原核生物的脂蛋白中，脂质与N-末端半胱氨酸的SH基团结合。为了确认Lpp 44是否具有ICAM-1表达诱导活性，合成了类似于Lpp 44的脂肽S-（2，3-双棕榈酰氧基丙基）-CGDPKHPKSF。发现Lpp 44类似物具有ICAM-1表达诱导活性，并诱导HGF产生IL-6和单核细胞系THP-1细胞产生TNF-α。Lpp 44类似物诱导ICAM-1表达的活性水平显著高于从大肠杆菌和沙门氏菌纯化的LPS，但低于从大肠杆菌和沙门氏菌纯化的LPS。因此，本研究证明Lp 44是M. Lpp 44与MALP-2（S-（2，3-bisacyloxypropyl）-CGNNDESNISFKEK）在氨基酸序列上不同，但在结构上相似，后者是从M.发酵素，并能够激活单核细胞/巨噬细胞以诱导一氧化氮和TNF-α的产生。E. N-末端脂肽的合成类似物。已知大肠杆菌胞壁蛋白脂蛋白是有效的巨噬细胞和B细胞活化剂。这些脂肽的结构与MALP-2和Lpp 44的结构相似。因此，推测来自原核生物的脂蛋白是巨噬细胞、B细胞和成纤维细胞的有效激活剂。少