Characteristics of human chondrocytes, osteoblasts and fibroblasts seeded onto a type I/III collagen sponge under different culture conditions. A light, scanning and transmission electron microscopy study.
Hyaline cartilage has only a limited capacity of regeneration, thus, lesions of articular cartilage can lead to early osteoarthrosis. Current concepts in conservative orthopedic therapy do not always lead to satisfying results. As one new attempt to facilitate cartilage repair, autologous transplantation of articular chondrocytes is investigated in different assays. This study was designed to create a resistible and stable cell-matrix-biocomposite with viable and biosynthetically active human chondrocytes, osteoblasts or fibroblasts. This biocomposite might serve as an implant to treat deep osteochondral defects in the knee. We collected cartilage, spongiosa and skin probes from healthy patients undergoing hip-surgery and enzymatically liberated the chondrocytes, seeded them into culture flasks and cultured them until confluent. The spongiosa and the skin samples were also placed in culture flasks and cells cultured until confluent. After 4-6 weeks, cells were trypsinized and grown on a type I/III collagen matrix (Chondrogide, Geistlich Biomaterials, Wolhusen, Switzerland) for 7 days in standard Petri dishes and in a special perfusion chamber culture system. As controls, cells were seeded onto plastic surfaces. Then scaffolds were fixed and embedded for light microscopy and electron microscopy by routine methods. Light microscopically, chondrocytes grown on the surface of the scaffold form clusters or a dense layer of sometimes rather fibroblast-like and sometimes roundish, chondrocyte-like cells. Only a few cells grow deeper into the matrix. In transmission electron microscopy, the cells have a rather chondrocyte-like morphology which emphasizes the matrix-induced redifferentiation after dedifferentiation of chondrocytes in monolayer-culture in culture flasks. Chondrocytes on plastic surfaces have a spinocellular aspect with little signs of differentiation. Grown on Chondrogide, cells are more roundish and adhere firmly to the collagen fibrils of the scaffold. Osteoblasts grown on the collagen scaffold and examined by light microscopy form a thin cell-layer on the surface of the matrix with a reticular layer of dendritic cells underneath this sheet. Transmission electron micrographs show spinocellular and flat cells on the collagen fibrils. Scanning electron micrographs show large dendritic osteoblasts on plastic and a confluent layer of flattened, dendritic cells on the collagen scaffold. Fibroblasts form a thick multi-layer of typical spinocellular cells on the collagen matrix. Fibroblasts grown on plastic surfaces and examined by scanning electron microscopy also show a dense layer of fibroblast-like cells. For all three different types of cells no morphological differences could be seen when comparing cultivation in the perfusion culture system to cultivation in standard Petri dishes, although mechanical stress is believed to induce differentiation of chondrocytes. Especially the observed partially differentiated chondrocyte-matrix biocomposite might serve as an implant to treat deep cartilage defects, whereas osteoblasts and fibroblasts seem to be less suited.
Klinik für Orthopädie, Medizinischen Universität zu Lübeck, Ratzeburger, Germany.
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Fuss
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Ehlers
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Russlies
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Rohwedel
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Behrens
D000368:Aged; D000369:Aged, 80 and over; D019644:Arthroplasty, Replacement, Hip; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000648:ultrastructure; D046508:Culture Techniques / Q000379:methods; D005347:Fibroblasts / Q000166:cytology* / Q000648:ultrastructure; D006801:Humans; D008854:Microscopy, Electron; D008855:Microscopy, Electron, Scanning; D010006:Osteoblasts / Q000166:cytology* / Q000648:ultrastructure; D012867:Skin / Q000166:cytology*