Hypertrophy of vascularized bone isograft in rats treated with cyclosporine A.

The aim of this study was to investigate the effects of cyclosporine A (CsA) on vascularized tibio-fibula isograft between 12-week-old male Lewis rats. After transplantation, 45 rats were randomly allocated to one of the following 7 treatment groups: (1) 4-week vehicle (n = 5), (2) 4-week CsA (n = 5), (3) 8-week vehicle (n = 10), (4) 8-week CsA (n = 10), (5) 4-week CsA followed by 4-week vehicle (n = 5), (6) 16-week vehicle (n = 5), or (7) 4-week CsA followed by 12-week vehicle (n = 5). In soft X-ray and micro-computed tomography examination, hypertrophic change of the grafted bones was apparent in the 4- and 8-week CsA groups. Mineral apposition rate and bone formation rate of the grafted bones in the 4-week CsA group were markedly higher than those in the 4-week vehicle group. In the 4- and 8-week CsA groups, however, bone mineral density (BMD) of the grafted bones was lower and strength of the reconstructed bones was weaker than the 4- and 8-week vehicle groups. Urinary deoxypyridinoline (DPD) level was higher in the 4- and 8-week CsA groups than in the 4- and 8-week vehicle groups. The group of 4-week CsA followed by 4-week vehicle had a level of urinary DPD equal to the 8-week vehicle group, but their BMD of the grafted bones was lower and strength of the reconstructed bones was weaker than the 8-week vehicle group. By contrast, the group of 4-week CsA followed by 12-week vehicle had BMD of the grafted bones and strength of the reconstructed bones similar to the 16-week vehicle group. These findings demonstrate that short-term CsA treatment induces hypertrophic change of vascularized bone graft with high-turnover bone loss, and strength of the reconstructed bone is gradually restored after the cessation of CsA treatment.

Effects of cerivastatin on vascularized allogenic bone transplantation in rats treated with cyclosporine A.

The aim of the present study was to investigate the effects of oral cerivastatin (0.1 mg/kg/day) on vascularized allogenic transplanted bone that is treated with cyclosporine A (CsA) (10 mg/kg/day) and on vascularized isogenic transplanted bone that is not treated with CsA. Allogenic transplantation was performed on 12-week-old male DA rats with the major histocompatibility antigen (MHC) RT1a (as the donor) and age-matched male Lewis rats with MHC RT1l (as the recipient), and isogenic transplantation was performed on 12-week-old male Lewis rats. After transplantations, 20 rats (10 rats in each transplantation) were randomized into four groups to receive the following treatment for 16 weeks: (1) CsA plus cerivastatin vehicle or (2) CsA plus cerivastatin in the allogenic transplanted rats, and (3) CsA vehicle plus cerivastatin vehicle or (4) CsA vehicle plus cerivastatin in the isogenic transplanted rats. Bone biochemical markers, mineral density, and strength were measured at the end of the study period. Serum levels of osteocalcin (OC) and parathyroid hormone (PTH) and urinary deoxypyridinoline (DPD) level were higher in the allogenic transplanted rats than in the isogenic transplanted rats. In the allogenic transplanted rats, the cerivastatin treatment decreased urinary DPD levels, but not serum OC nor PTH levels. Furthermore, the cerivastatin treatment improved bone mineral density of the allogenic transplanted bones and bone strength of the allogenic reconstructed bones. In contrast, no effect of the cerivastatin treatment was observed in the isogenic transplanted rats. These results suggest that the cerivastatin treatment improves CsA-induced high-turnover osteopenia mainly through the inhibition of bone resorption.

Effects of vitamin D analog, 22-oxa-1,25-dihydroxyvitamin D(3), on bone reconstruction by vascularized bone allograft.

We previously reported that vascularized bone allograft using immunosuppressants, such as cyclosporine A (CsA), is one approach for reconstruction of large bone defects in both experimental animals (Microsurgery 15:663; 1994) and clinically in humans (Lancet 347:970, 1996). Because immunosuppressive agents such as CsA induce significant side effects, including bone loss, other therapeutic agents supporting successful vascularized bone allografts have been sought after. We investigated the effects of 22-oxa-1,25-dihydroxyvitamin D(3) (OCT) on vascularized bone allograft, and compared its effects with CsA. Twelve-week-old DA rats with the major histocompatibility antigen (MHC) RT-1(a) were used as donors and age-matched Lewis rats with MHC RT-1(l) used as recipients. Allografted bones in rats treated with vehicle were rejected completely. Soft X-ray examination demonstrated that administration of OCT (0.5 microg/kg per day) for 12 weeks after bone graft induced bone union as effective as treatment for 12 weeks with CsA (10 mg/kg per day). Transplanted bones in OCT-treated rats showed higher bone mineral density than that in CsA-treated rats. Histologically, transplanted bones in OCT-treated rats at 12 weeks were nonvital, but these bones united with recipient vital bones. After cessation of 12 week treatment with OCT, new bone formation occurred around the grafted nonvital bones during a 9 month period. Transplanted bones in CsA-treated rats were vital and formed union with recipient bones, whereas cortical bones became thin when compared with nonvital bones in OCT-treated rats. Urinary deoxypyridinoline levels in rats treated with CsA were significantly higher than levels in rats treated with OCT, suggesting accelerated bone resorption in CsA-treated rats. These results suggest that OCT exerts an anabolic action on bone reconstruction by allogeneic bone transplantation.

Involvement of bone morphogenic protein-2 (BMP-2) in the pathological ossification process of the spinal ligament.

OBJECTIVE: To investigate the function of bone morphogenic protein-2 (BMP-2) in the ossification of the spinal ligament (OSL). METHODS: Total RNA was prepared from the cultured spinal ligament cells from patients with OSL and analysed by reverse transcription-polymerase chain reaction using specific primers for BMP-2. BMP-2 mRNA expression in ligament tissues was examined by in situ hybridization. Spinal ligament cells from patients without OSL were treated with BMP-2 and examined for alkaline phosphatase activity. RESULTS: Expression of the BMP-2 gene was detected in cultured spinal ligament cells. In ligament tissues, BMP-2 mRNA was present in the chondrocyte-like cells in the fibrocartilage zone. Exogenous BMP-2 increased alkaline phosphatase activity in spinal ligament cells from patients without OSL. CONCLUSION: The BMP-2 gene is expressed in the spinal ligaments of OSL patients, and exogenous BMP-2 stimulates osteogenic differentiation of spinal ligament cells. The expression of BMP-2 in the spinal ligaments could be a clue in elucidating how heterotrophic osteogenesis develops in ligament tissue.

Bioremediation of trichloroethylene and cis-1,2-dichloroethylene-contaminated groundwater by methane-utilizing bacteria.

Experimental studies on the bioremediation of groundwater contaminated with low concentration trichloroethylene (TCE) and cis1,2-dichloroethylene (DCE) were performed with two sets of bioreactors. Reactors No. 1 and No. 2 were operated without and with methane supplement, respectively. No inoculum was used. The concentrations of TCE and DCE in the effluent and the off gas from reactor No. 2 were much lower than those from reactor No. 1. When air and an H2O2 solution were supplied to reactor No. 2, concentrations of TCE and DCE in the effluent and the off gas were lower than the lowest detectable limit. The population of methane-utilizing bacteria in reactor No. 2 was 1,000 times higher than that in groundwater or in the effluent from reactor No. 1. These methane-utilizing bacteria were apparently attributable to the treatment of TCE.

Some characteristics of bacteria found in a bioreactor to treat trichloroethylene-contaminated groundwater.

A mixture of bacteria, having a methane-utilizing ability, was separated from a bioreactor supplied with air and methane gas. The bioreactor was operated to treat trichloroethylene (TCE)-contaminated groundwater. The mixture was composed of an obligate methane-utilizing bacterium and a heterotroph, identified as Methylomonas methanica and Pseudomonas sp., respectively. The mixed culture of these two strains removed TCE. In addition, it appeared that a cooperative metabolic interaction of these strains enabled Meth.methanica to maintain the TCE degradation ability.