Hirano F, Hirano H, Hino E, Takayama S, Saito K, Kusumoto Y, Shimabukuro Y, Murakami S, Okada H
Department of Periodontology and Endodontology, Osaka University Faculty of Dentistry, Japan.
J Periodontal Res. 1997 Nov;32(8):634-45. doi: 10.1111/j.1600-0765.1997.tb00573.x.
CD44 functions as a receptor for various extracellular matrices and plays crucial roles in homotypic and heterotypic cell-cell interactions. Recently, the molecular structure of CD44 has been extensively analyzed and multiple isoforms produced by alternative splicing of messenger RNA have been identified. In this study, we examined the expression of CD44 isoforms on different cell types isolated from periodontal tissue. In order to examine tissue differences in CD44 isoform expression, we established in vitro cell culture of human gingival fibroblasts (HGF), human periodontal ligament cells (HPDL) and human gingival epithelial cells (HGEC). These cells all expressed CD44 protein and messenger RNA. However, immunoprecipitation and Northern blot analysis revealed that HGEC expressed larger CD44 isoforms than HGF and HPDL. Reverse transcription-polymerase chain reaction with primers flanking the insertion site of alternatively spliced exons was used to study details of the heterogeneity. All cells examined expressed a major band in the absence of alternatively spliced exons and additional larger bands. In particular, HGEC contained more abundant high molecular mass species. In vitro stimulation by IL-1 beta, TNF alpha or phorbol 12-myristate 13-acetate induced an increase in total CD44 messenger RNA in HGF but not change in overall patterns of CD44 isoform expression. However, the isoform expression of HGEC was sensitive to cell density. The amount of larger isoform was decreased by culturing cells beyond confluence. These findings suggest that CD44 isoform expression is cell type-specifically regulated in periodontium and altered according to growth phase of HGEC.
CD44作为多种细胞外基质的受体，在同型和异型细胞间相互作用中发挥关键作用。近来，对CD44的分子结构进行了广泛分析，已鉴定出由信使核糖核酸可变剪接产生的多种异构体。在本研究中，我们检测了从牙周组织分离的不同细胞类型上CD44异构体的表达。为检测CD44异构体表达的组织差异，我们建立了人牙龈成纤维细胞（HGF）、人牙周膜细胞（HPDL）和人牙龈上皮细胞（HGEC）的体外细胞培养体系。这些细胞均表达CD44蛋白和信使核糖核酸。然而，免疫沉淀和Northern印迹分析显示，HGEC表达的CD44异构体比HGF和HPDL更大。使用位于可变剪接外显子插入位点两侧的引物进行逆转录-聚合酶链反应，以研究异质性的细节。所有检测的细胞在不存在可变剪接外显子时均表达一条主要条带以及额外的更大条带。特别是，HGEC含有更丰富的高分子量种类。白细胞介素-1β、肿瘤坏死因子α或佛波醇12-肉豆蔻酸酯13-乙酸酯的体外刺激可使HGF中总CD44信使核糖核酸增加，但CD44异构体表达的总体模式没有变化。然而，HGEC的异构体表达对细胞密度敏感。细胞汇合后继续培养会使更大异构体的量减少。这些发现表明，CD44异构体表达在牙周组织中受到细胞类型特异性调节，并根据HGEC的生长阶段而改变。
