Kubo M, Kan M, Isemura M, Yamane I, Tagami H
J Invest Dermatol. 1987 May;88(5):594-601. doi: 10.1111/1523-1747.ep12470207.
Extracellular matrices (ECMs) play an important role as components of basement membrane of normal human skin and in migrating epidermal cells in wound healing. We investigated the effects of various ECMs on human keratinocyte adhesion and growth as well as on its secretion and deposition of fibronectin (FN) in vitro using a serum-free, low-calcium culture system. Since cell adhesion is the first step of cell growth, we performed cell adhesion assay for 14 h. Human keratinocytes adhered best on FN and less well on types I/III collagen, type IV collagen, and heparan sulfate proteoglycan (HSPG) as compared with bovine serum albumin (BSA) (control) or laminin (La). Cell growth assayed for 7-8 days on the dishes coated with various extracellular matrices revealed significantly increased keratinocyte growth on FN and on types I/III collagen in comparison with that on type IV collagen, HSPG, BSA (control), or La. Morphology of keratinocytes and of their colonies on FN and types I/III collagen was strikingly different from that of the control; the colonies were not so compact as in the control, but rather loose and larger; each keratinocyte was spread out more on these substrata. These morphologic features seemed to correlate with the increased keratinocyte growth on these extracellular matrices. Both immunofluorescence study for FN with keratinocytes in 8-day culture on various extracellular matrices and enzyme-linked immunosorbent assay for FN measurement on substratum or in conditioned medium with keratinocytes in 5-day culture demonstrated that extracellular matrices modulated the secretion and deposition of FN by human keratinocytes in culture; the keratinocyte growth correlated with the amount of FN detected on substratum but not with that in medium. Based on the results of the present investigation, we think that the growth of human keratinocytes depends on the amount of FN on substratum.
细胞外基质（ECM）作为正常人皮肤基底膜的组成成分以及在伤口愈合过程中迁移的表皮细胞中发挥着重要作用。我们使用无血清、低钙培养系统在体外研究了各种细胞外基质对人角质形成细胞黏附、生长以及纤连蛋白（FN）分泌和沉积的影响。由于细胞黏附是细胞生长的第一步，我们进行了14小时的细胞黏附试验。与人血清白蛋白（BSA）（对照）或层粘连蛋白（La）相比，人角质形成细胞在FN上黏附最佳，而在I/III型胶原、IV型胶原和硫酸乙酰肝素蛋白聚糖（HSPG）上黏附较差。在涂有各种细胞外基质的培养皿上进行7 - 8天的细胞生长试验表明，与在IV型胶原、HSPG、BSA（对照）或La上相比，角质形成细胞在FN和I/III型胶原上的生长显著增加。角质形成细胞及其在FN和I/III型胶原上的集落形态与对照有显著差异；集落不像对照那样紧密，而是松散且更大；每个角质形成细胞在这些基质上展开得更多。这些形态学特征似乎与角质形成细胞在这些细胞外基质上的生长增加相关。在各种细胞外基质上进行8天培养的角质形成细胞对FN的免疫荧光研究以及在5天培养的角质形成细胞的基质或条件培养基中进行FN测量的酶联免疫吸附试验均表明，细胞外基质调节了培养的人角质形成细胞中FN的分泌和沉积；角质形成细胞的生长与在基质上检测到的FN量相关，而与培养基中的量无关。基于本研究结果，我们认为人角质形成细胞的生长取决于基质上FN的量。
