In skin regeneration therapy using a marrow mesenchymal cell/artificial dermis composite graft, skin regeneration is possible with bone marrow aspiration, a minimally invasive procedure. Compared with existing skin grafting techniques, the present technique is practical and much less invasive.

From 10 cases (9 patients, mean age, 65 years : range 17-75 years), bone marrow fluid was collected from the ilium in 9 cases, and from the proximal tibia in 1 case. The diagnosis was humeral fracture, femoral nonunion, metatarsal bone nonunion and osteoarthrosis of the hip joint in 1, 2, 1 and 5 patients, respectively. After 10-20 mL of bone marrow fluid was collected, it was immediately transferred to the culture room and incubated in a flask containing MEM with 15% fetal bovine serum and dexamethasone, etc.. In patients with fracture or femoral nonunion, porous beta-TCP blocks were impregnated with cultured mesenchymal cells and implanted at the site of fracture or nonunion. In 2 patient with osteoarthrosis, cultured bone marrow cells were impregnated into a porous ceramic block and that was grafted into the bone defect of the acetabular shelf or femur. In 3 patients with hip osteoarthrosis, cultured bone marrow cells were spread over the contact area between the artificial joint and the bone and arthroplasty was done without bone cement. Radiological studies revealed callus formation at 2 weeks after grafting in patients with bone fracture or nonunion. At 1 to 2 months after grafting, the grafted construct was adherent to the native bone in the patient in whom cultured artificial bone was grafted to the acetabular shelf or femur. In 3 patients with hip joint replacement, the stem of the artificial joint showed good integration with the bone of the medullary cavity. Treatment with bone marrow mesenchymal cells has a low invasiveness (only marrow aspiration), and therefore it is suitable for weak elderly patients as well as patients in good general condition, because its low invasiveness enables an early return to work. This treatment is expected to be applied more widely in the near future.

Bone tissue was produced when marrow cells were incubated in a medium containing dexamethasone. Bio-artificial bone with a high osteogenetic capacity can be produced by combining such cultured bone tissue with an artificial bone material. With respect to the osteogenetic capacity of adult human marrow cells, however, the potential for proliferation and differentiation varies from individual to individual, involving many factors. We succeeded in producing advanced bio-artificial bone with a greater osteogenetic capacity by this method. In this report, we discuss bone regeneration therapy using activated cultured bone constructs. The activated cultured bone construct was used for 25 bone regeneration treatments in 23 patients. Reasons for bone regeneration were prolonged bone fracture therapy in 11 cases, coxarthrosis in 12 cases, lumbar spondylosis in 1 case and tumor resection in 1 case. In all patients, 10 to 20 ml of bone marrow was collected from the ilium or the tibia and incubated in MEM containing autologous serum or fetal bovine serum and an antibiotic. After two weeks in primary culture, the marrow mesenchymal cells were seeded onto hydroxyapatite, beta-TCP, another ceramic material, or a prosthetic joint and cultured in the osteogenic medium as reported separately. The bio-artificial bone thus obtained was then implanted at the affected site. In patients with pseudoarthrosis, repeat joint replacement, or an osseous defect following tumorectomy, the implanted artificial bone survived and bone regeneration was detected radiographically. Shortand medium-term follow-up has shown that the bone implants were effective in all of the patients. The prosthetic joints which were implanted with cultured bone marrow cells showed good integration at the bone-joint interface.

Subjects were graft patients with pseudoarthrosis (average age, 60.3 years; range, 17-85 years). Pseudoarthrosis affected the thoracolumbar spine, the femur, the clavicle, the humerus and the metatarsal. From the ilium (tibia in one patient), 10-20 ml of bone marrow fluid was collected, and then, it was immediately transferred to the culture room and incubated in a flask containing MEM with 15% autologous or fetal bovine serum, etc.. After 2 weeks in primary culture, cells were released by trypsin treatment and were subsequently incubated with porous beta-TCP in order to prepare tissue-engineered artificial bone, according to the previously reported modified culturing technique. Tissue-engineered artificial bone was grafted around the non-union site of each affected long bone, while tissue-engineered artificial bone was grafted via the pedicle of each affected vertebral body. In all patients, favorable bone formation was seen at three months after surgery. In the patients with pseudoarthrosis of the spine, CT and MRI confirmed favorable vertebral body formation. In the patients with pseudoarthrosis of a long bone, the artificial bone was remodeled and favorable bone union was confirmed. In 2 patients in whom bone biopsy was performed during pin removal, bone regeneration was confirmed histologically. With present type of tissue-engineered artificial bone, an artificial material with a high bone regeneration capacity can be prepared by aspiration, which is minimally invasive, and thus when compared to iliac bone grafts, it is possible to radically reduce postoperative pain without damage of autologous bone.

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