Objective. To examine the ability of cartilage-like tissue, generated ectopically in a diffusion chamber using recombinant human bone morphogenetic protein 2 (rHuBMP-2), to repair cartilage defects in rats.Methods. Muscle-derived mesenchymal cells were prepared by dissecting thigh muscles of 19-day postcoital rat embryos. Cells were propagated in vitro in monolayer culture for 10 days and packed within diffusion chambers (10 6 /chamber) together with type I collagen (CI) and 0, 1, or 10 g rHuBMP-2, and implanted into abdominal subfascial pockets of adult rats. Tissue pellets were harvested from the diffusion chambers at 2 days to 6 weeks after implantation, and examined by histology, by reverse transcription-polymerase chain reaction (PCR) for aggrecan, CII, CIX, CX, and CXI, MyoD1, and core binding factor a1/runt-related gene 2, and by real-time PCR for CII. Tissue pellets generated in the chamber 5 weeks after implantation were transplanted into a full-thickness cartilage defect made in the patellar groove of the same strain of adult rat.Results. In the presence of 10 g rHuBMP-2, muscle-derived mesenchymal cells expressed CII messenger RNA at 4 days after transplantation, and a mature cartilage mass was formed 5 weeks after transplantation in the diffusion chamber. Cartilage was not formed in the presence of 1 g rHuBMP-2 or in the absence of rHuBMP-2. Defects receiving cartilage engineered with 10 g rHuBMP-2 were repaired and restored to normal morphologic condition within 6 months after transplantation.Conclusion. This method of tissue engineering for repair of articular defects may preclude the need to harvest cartilage tissue prior to mosaic arthroplasty or autologous chondrocyte implantation. Further studies in large animals will be necessary to validate this technique for application in clinical practice.
The dental follicle contains mesenchymal cells that differentiate into osteoblasts, cementoblasts, and fibroblasts. However, the characteristics of these mesenchymal cells are still unknown. α-Smooth muscle actin (α-SMA) is known to localize in stem cells and precursor cells of various tissues. In the present study, to characterize the undifferentiated cells in the dental follicle, immunohistochemical localization of α-SMA was examined during rat molar tooth development. Rat mandibles were collected at embryonic days (E) 15-20 and postnatal days (P) 7-28. Immunohistochemical stainings for α-SMA, periostin, Runt-related transcription factor-2 (Runx2), tissue nonspecific alkaline phosphatase (TNAP), and bone sialoprotein (BSP) were carried out using paraffin-embedded sections. α-SMA localization was hardly detected in the bud and cap stages. At the early bell stage, α-SMA-positive cells were visible in the dental follicle around the cervical loop. At the late bell to early root formation stage (P14), these cells were detected throughout the dental follicle, but they were confined to the apical root area at P28. Double immunostaining for α-SMA and periostin demonstrated that α-SMA-positive cells localized to the outer side of periostin-positive area. Runx2-positive cells were visible in the α-SMA-positive region. TNAP-positive cells in the dental follicle localized nearer to alveolar bone than Runx2-positive cells. BSP was detected in osteoblasts as well as in alveolar bone matrix. These results demonstrate that α-SMA-positive cells localize on the alveolar bone side of the dental follicle and may play a role in alveolar bone formation.
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