Cell-free Implants Research Articles

Stem cell based tissue engineering for meniscus repair

Defects of the meniscus greatly alter knee function and predispose the joint to degenerative changes. The purpose of this study was to test a recently developed cell-scaffold combination for the repair of a critical-size defect of the rabbit medial meniscus. A bilateral, complete resection of the pars intermedia of the medial meniscus was performed in 18 New Zealand White rabbits. A hyaluronan/gelatin composite scaffold was implanted into the defect of one knee of 6 rabbits and the contralateral defect was left untreated. Scaffolds loaded with autologous marrow-derived mesenchymal stem cells and cultured in a chondrogenic medium for 14 days were implanted in a second series of 12 rabbits. Empty scaffolds were implanted in the contralateral knees. Meniscii were harvested at 12 weeks. Untreated defects had a muted fibrous healing response. Defects treated with cell-free implants showed also predominantly fibrous tissue whereas fibrocartilage was present in some scaffolds. The cross-sectional width of the repair tissue after treatment with cell-free scaffolds was significantly greater than controls (p < 0.05). Pre-cultured implants integrated with the host tissue and 8 of 11 contained meniscus-like fibrocartilage, compared with 2 of 11 controls (p < 0.03). The mean cross-sectional width of the pre-cultured implant repair tissue was greater than controls (p < 0.004). This study demonstrates the repair of a critical size meniscal defect with a stem cell and scaffold based tissue engineering approach.

P211 Cell-free scaffold implantation in the horse

P211 Cell-free scaffold implantation in the horse

ALLOGENEIC MESENCHYMAL STEM CELLS REGENERATE BONE IN A CRITICAL-SIZED CANINE SEGMENTAL DEFECT

Mesenchymal stem cells from adult bone marrow are multipotent cells capable of forming bone, cartilage, and other connective tissues. In a previous study, we demonstrated that autologous mesenchymal stem cells could repair a critical-sized bone defect in the dog. The objective of this study was to determine whether the use of allogeneic mesenchymal stem cells could heal a critical-sized bone defect in the femoral diaphysis in dogs without the use of immunosuppressive therapy. A critical-sized segmental bone defect, 21 mm in length, was created in the mid-portion of the femoral diaphysis of twelve adult dogs that weighed between 22 and 25 kg. Each defect was treated with allogeneic mesenchymal stem cells loaded onto a hollow ceramic cylinder consisting of hydroxyapatite-tricalcium phosphate. A complete mismatch between donor stem cells and recipient dogs was identified by dog leukocyte antigen typing prior to implantation. The healing response was evaluated histologically and radiographically at four, eight, and sixteen weeks after implantation. The radiographic and histological results at sixteen weeks were compared with the historical data for the control defects, which included defects that had been treated with a cylinder loaded with autologous mesenchymal stem cells, defects treated with a cylinder without mesenchymal stem cells, and defects that had been left untreated (empty). The systemic immune response was evaluated by the analysis of recipient serum for production of antibodies against allogeneic cells. For defects treated with allogeneic mesenchymal stem cell implants, no adverse host response could be detected at any time-point. Histologically, no lymphocytic infiltration occurred and no antibodies against allogeneic cells were detected. Histologically, by eight weeks, a callus spanned the length of the defect, and lamellar bone filled the pores of the implant at the host bone-implant interface. Fluorescently labeled allogeneic cells were also detected. At sixteen weeks, new bone had formed throughout the implant. These results were consistent with those seen in implants loaded with autologous cells. Implants loaded with allogeneic or autologous stem cells had significantly greater amounts of bone within the available pore space than did cell-free implants at sixteen weeks (p < 0.05). The results of this study demonstrated that allogeneic mesenchymal stem cells loaded on hydroxyapatite-tricalcium phosphate implants enhanced the repair of a critical-sized segmental defect in the canine femur without the use of immunosuppressive therapy. No adverse immune response was detected in this model.

Autologous bone marrow stromal cells loaded onto porous hydroxyapatite ceramic accelerate bone repair in critical-size defects of sheep long bones.

The ability of marrow-derived osteoprogenitor cells to promote repair of critical-size tibial gaps upon autologous transplantation on a hydroxyapatite ceramic (HAC) carrier was tested in a sheep model. Conditions for in vitro expansion of sheep bone marrow stromal cells (BMSC) were established and the osteogenic potential of the expanded cells was validated. Ectopic implantation of sheep BMSC in immunocompromised mice led to extensive bone formation. When used to repair tibial gaps in sheep, cell-loaded implants (n = 2) conducted a far more extensive bone formation than did cell-free HAC cylinders (n = 2) over a 2-month period. In cell-loaded implants, bone formation was found to occur both within the internal macropore space and around the HAC cylinder while in control cell-free implants, bone formation was limited mostly to the outer surface and was not observed in most of the inner pores. As tested in an indentation assay, the stiffness of the complex HAC-bone material was found to be higher in cell-loaded implants compared to controls. Our pilot study on a limited number of large-sized animals suggests that the use of autologous BMSC in conjunction with HAC-based carriers results in faster bone repair compared to HAC alone. Potentially this combination could be used clinically in the treatment of extensive long bone defects.

Culture-Expanded, Bone Marrow-Derived Mesenchymal Stem Cells Can Regenerate a Critical-Sized Segmental Bone Defect

Bone marrow has been shown to contain a population of rare cells capable of differentiating into the cells that form bone, cartilage, and other connective tissues. These cells, referred to as mesenchymal stem cells (MSCs), can be purified and culture-expanded from bone marrow, and are capable of forming bone when implanted ectopically in an appropriate carrier. To test the ability of MSCs to heal a critical-sized segmental defect, syngeneic culture-expanded MSCs were loaded onto a porous cylinder of hydroxyapatite/tricalcium phosphate (HA/TCP) and implanted in an 8-mm segmental defect in the femora of adult rats. For comparison, other defects received either marrow-loaded or cell-free HA/TCP cylinders, or no implant. The healing response was evaluated at 4 and 8 weeks by radiography, histology, and quantitative histomorphometry. Defects that received no implant, cell-free HA/TCP cylinders, or cylinders loaded with fresh marrow failed to heal by 8 weeks. In contrast, defects that received MSC-loaded cylinders showed significantly more bone formation at 4 weeks, and were completely healed by 8 weeks with excellent union of the host-implant interfaces. Histomorphometry demonstrates that bone fill in MSC-loaded implants (43%) is greater (p < 0.05) than that obtained with marrow-loaded (19%) or cell-free implants (10%).

Transfilter Bone Induction by Chinese Hamster Ovary (CHO) Cells Transfected by DNA Encoding Bone Morphogenetic Protein-4

This study was undertaken to identify the factor responsible for classical transfilter bone induction by a murine osteosarcoma. Chinese hamster ovary (CHO) cells were transfected with the complementary DNA (cDNA) for bone morphogenetic protein-4 (BMP-4) that was purified from a murine (Dunn) osteosarcoma. Diffusion chambers were filled with the cells expressing the gene for BMP-4 and implanted subcutaneously into the flanks of ICR strain nude mice. Ectopic transfilter bone formation was seen consistently on the outer surfaces of the cellulose acetate membranes of chambers containing transfected cells at three weeks after implantation. Bone was not observed on chambers loaded with nontransfected CHO cells. The transfected CHO cells were inoculated into nude mice to form tumors, which were then homogenized, defatted, and bioassayed also in the ICR, nu/nu mice. The cell-free implants consistently elicited new bone and marrow within three weeks, whereas the control implants consisting of nontransfected tumor were not osteoinductive. These experimental results suggest that BMP-4 is one of the molecules responsible for the transfilter bone induction by vital Dunn osteosarcoma cells reported by Heiple and for the ectopic bone induction after implantation of devitalized, freeze-dried Dunn osteosarcoma tissue described originally by Amitani.