Schor A M, Canfield A E, Sloan P, Schor S L
CRC Department of Medical Oncology, Christie Hospital and Holt Radium Institute, Manchester, United Kingdom.
In Vitro Cell Dev Biol. 1991 Aug;27A(8):651-9. doi: 10.1007/BF02631109.
We have previously reported that pericytes derived from retinal and brain microvessels aggregate into nodules soon after reaching confluence. Nodule formation involves a reorganization of the cells resulting in the presence of sparse cells, confluent monolayers, multilayers, sprouts, and nodules within the same culture dish. Extracellular calcification occurs only within the nodules, demonstrating that pericytes are capable of undergoing osteogenic differentiation in culture and that this differentiation is related to nodule formation. Using immunofluorescence we have now studied the distribution of laminin, type IV collagen, type X collagen, and tenascin in pericyte cultures during nodule formation. These matrix macromolecules were also identified by a combination of biochemical techniques, including Northern blot hybridization, immunoblotting and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A molecule that seems to be related to type X collagen was demonstrated by the presence of a pepsin-resistant, collagenase-sensitive polypeptide of molecular weight approximately 45 kDa. The production of laminin, type X-related collagen, and tenascin by pericytes has not been previously reported. Our results suggest that the synthesis or distribution or both of these molecules is dependent on the state of pericyte differentiation. The expression of laminin, type IV collagen, and type X-related collagen was maximal in multilayer areas, sprouts, and nodules. Tenascin appeared homogeneously distributed in monolayer and multilayer areas; when calcified nodules were present, the anti-tenascin serum preferentially decorated a discrete area circumscribing the nodules. Tenascin and type X collagen have been found transiently in vivo preceding calcification; their possible role in this process is not known. Our results also suggest an association between laminin, type IV collagen, and calcification. The in vitro experimental system described here may help to clarify the role of matrix macromolecules in the calcification process.
我们之前曾报道，源自视网膜和脑微血管的周细胞在达到汇合后不久就会聚集成结节。结节形成涉及细胞的重新组织，导致在同一培养皿中出现稀疏细胞、汇合单层、多层、芽和结节。细胞外钙化仅发生在结节内，表明周细胞在培养中能够进行成骨分化，且这种分化与结节形成有关。我们现在使用免疫荧光研究了结节形成过程中周细胞培养物中层粘连蛋白、IV型胶原、X型胶原和腱生蛋白的分布。这些基质大分子也通过包括Northern印迹杂交、免疫印迹和十二烷基硫酸钠-聚丙烯酰胺凝胶电泳在内的生化技术组合进行了鉴定。通过存在一种分子量约为45 kDa的耐胃蛋白酶、对胶原酶敏感的多肽，证明了一种似乎与X型胶原相关的分子。周细胞产生层粘连蛋白、X型相关胶原和腱生蛋白此前尚未见报道。我们的结果表明，这些分子的合成或分布或两者均取决于周细胞的分化状态。层粘连蛋白、IV型胶原和X型相关胶原的表达在多层区域、芽和结节中最高。腱生蛋白在单层和多层区域均匀分布；当存在钙化结节时，抗腱生蛋白血清优先标记围绕结节的离散区域。腱生蛋白和X型胶原在体内钙化之前已被短暂发现；它们在这个过程中的可能作用尚不清楚。我们的结果还表明层粘连蛋白、IV型胶原与钙化之间存在关联。这里描述的体外实验系统可能有助于阐明基质大分子在钙化过程中的作用。
