Clark R A, Nielsen L D, Howell S E, Folkvord J M
J Cell Biochem. 1985;28(2):127-41. doi: 10.1002/jcb.240280206.
Fibronectin and laminin production by human keratinocytes cultured in serum-free, low-calcium medium without a fibroblast feeder layer were examined by several techniques. By indirect immunofluorescence, fibronectin but not laminin appeared as short radial fibrils between the cells and the substratum, and in the pericellular matrix. Synthesis of fibronectin and laminin by 7-day keratinocyte cultures was determined by 18 hr 35S-methionine metabolic labeling followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and autoradiography. Fibronectin accounted for 2.9% of total synthesized protein, 26.5% of fluid phase protein secretion, and 4.3% of deposited ECM protein. In contrast, only 0.1% of the total synthesized protein was laminin, little (6.3%) of this product was secreted, and none of this product was deposited in the ECM. Our results indicate that human keratinocytes under culture conditions that prevent terminal differentiation in vitro can synthesize, secrete, and deposit fibronectin in the extracellular matrix. Although these cells synthesize laminin, they secrete very little and deposit no detectable laminin in the matrix under these culture conditions. From these data we believe that fibronectin may play an important role in the interaction of epidermal cells with connective tissue matrix during wound healing or morphogenesis in in vivo situations in which the epidermis is not terminally differentiated.
通过多种技术检测了在无血清、低钙培养基中培养且无成纤维细胞饲养层的人角质形成细胞中纤连蛋白和层粘连蛋白的产生情况。通过间接免疫荧光法，纤连蛋白而非层粘连蛋白表现为细胞与基质之间以及细胞周围基质中的短径向纤维。通过18小时的35S-甲硫氨酸代谢标记，随后进行十二烷基硫酸钠聚丙烯酰胺凝胶电泳和放射自显影，测定了7天角质形成细胞培养物中纤连蛋白和层粘连蛋白的合成情况。纤连蛋白占总合成蛋白的2.9%，液相蛋白分泌的26.5%，以及沉积的细胞外基质蛋白的4.3%。相比之下，层粘连蛋白仅占总合成蛋白的0.1%，该产物很少（6.3%）分泌，且没有该产物沉积在细胞外基质中。我们的结果表明，在体外防止终末分化的培养条件下，人角质形成细胞能够合成、分泌并在细胞外基质中沉积纤连蛋白。尽管这些细胞合成层粘连蛋白，但在这些培养条件下，它们分泌很少且在基质中没有可检测到的层粘连蛋白沉积。根据这些数据，我们认为纤连蛋白可能在伤口愈合或形态发生过程中表皮细胞与结缔组织基质相互作用中发挥重要作用，在体内情况下，表皮并非终末分化。
