Monical P L, Kefalides N A
Connective Tissue Research Institute, University City Science Center, Philadelphia, Pennsylvania 19104.
Exp Cell Res. 1994 Feb;210(2):154-9. doi: 10.1006/excr.1994.1023.
To investigate the role of cell-cell interactions between keratinocytes and fibroblasts at the dermal-epidermal junction in the regulation of extracellular matrix protein expression, we have developed a cell coculture model that allows both cell types to interact on a reciprocal basis and yet be isolated as pure populations for quantitative analysis. Using porous membrane inserts as a substrate for keratinocytes grown in coculture over fibroblast monolayers, we report that coculture stimulates cell growth and total protein synthesis in both cell types when compared to monocultured controls. Both keratinocytes and fibroblasts synthesize laminin B1 and B2 chains with an additional subunit comigrating with laminin M chain detected in keratinocytes but only faintly visible in fibroblasts. Laminin A chain synthesis could not be detected. Although laminin subunit composition did not change in either cell type, fractional laminin synthesis increases by 41.0 +/- 2.3% in cocultured keratinocytes and decreases by 73.8 +/- 8.1% in cocultured fibroblasts when compared to monocultured controls. In cocultured keratinocytes, steady-state mRNA levels for laminin B1, B2, and M chains increased by 69.4, 63.5, and 136.8%, respectively, when compared to monocultured controls. However, in cocultured fibroblasts, laminin B1 and B2 chain mRNA decreased by 74.2 and 72.7%, respectively. Laminin M chain mRNA could not be detected in fibroblasts. These results suggest that synthesis and expression of laminin is regulated through reciprocal cell-cell interactions in fibroblasts and keratinocytes grown in coculture.
为了研究真皮-表皮交界处角质形成细胞与成纤维细胞之间的细胞间相互作用在细胞外基质蛋白表达调控中的作用，我们建立了一种细胞共培养模型，该模型允许两种细胞类型相互作用，同时又能作为纯群体分离出来进行定量分析。使用多孔膜插入物作为角质形成细胞在成纤维细胞单层上共培养生长的底物，我们报告说，与单培养对照相比，共培养刺激了两种细胞类型的细胞生长和总蛋白合成。角质形成细胞和成纤维细胞都合成层粘连蛋白B1和B2链，在角质形成细胞中检测到一个与层粘连蛋白M链共迁移的额外亚基，而在成纤维细胞中仅隐约可见。未检测到层粘连蛋白A链的合成。尽管两种细胞类型中层粘连蛋白亚基组成均未改变，但与单培养对照相比，共培养的角质形成细胞中层粘连蛋白合成分数增加了41.0±2.3%，共培养的成纤维细胞中减少了73.8±8.1%。在共培养的角质形成细胞中，与单培养对照相比，层粘连蛋白B1、B2和M链的稳态mRNA水平分别增加了69.4%、63.5%和136.8%。然而，在共培养的成纤维细胞中，层粘连蛋白B1和B2链mRNA分别下降了74.2%和72.7%。在成纤维细胞中未检测到层粘连蛋白M链mRNA。这些结果表明，在共培养的成纤维细胞和角质形成细胞中，层粘连蛋白的合成和表达是通过相互的细胞间相互作用来调节的。
