Skeletal growth is tightly coupled to energy balance via complex and incompletely understood mechanisms. Leptin-deficient ob/ob mice are obese and develop multiple pathologies associated with the metabolic syndrome. Additionally, ob/ob mice have skeletal abnormalities. The objective of this study was to evaluate the effects of leptin deficiency and long-duration selective central leptin repletion via recombinant adeno-associated virus-leptin (rAAV-lep) gene therapy on bone in growing ob/ob mice. The ob/ob mice were injected in the hypothalamus with either rAAV-lep or rAAV-GFP (control vector). Treated ob/ob and untreated wildtype (WT) mice were then maintained on a normal diet for 15 weeks. In a second experiment, similarly treated mice along with a group of pair-fed mice were maintained for 30 weeks. Leptin was not detected in blood of either rAAV-lep or rAAV-GFPtreated mice although rAAV-lep treated mice displayed leptin transgene expression in the hypothalamus. As expected, rAAV-lep normalized body weight and food intake. Compared to WT mice, rAAV-GFP-treated ob/ob mice had decreased femoral length (by 1.6 mm or 10 %, P<0.001), decreased total femur bone volume (by 3.3 mm 3 or 19%, P<0.001), but increased cancellous bone volume in the distal femur (by 0.04 mm 3 or 60%, P<0.09) and lumbar vertebrae (by 0.26 mm 3 or 118%, P<0.001). Treatment with rAAV-lep rescued the ob/ob skeletal phenotype by increasing femoral length and total bone volume, and decreasing femoral and vertebral cancellous bone volume, so that at 15 weeks post-rAAV-lep injection the ob/ob mice no longer differed from WT mice. No further skeletal changes in either the femur or lumbar vertebra were observed at 30 weeks post-rAAVlep administration. The results suggest that hypothalamic leptin functions as an essential permissive factor for normal bone growth.

HLU suppressed bone formation and resulted in bone loss in the tibial metaphysis of 6-monthold male rats. A human therapeutic dose of intermittent PTH (1 µg/kg/day) prevented the skeletal changes associated with HLU.Introduction: Skeletal unloading of skeletally mature rats results in trabecular thinning in the proximal tibial metaphysis, which is in part caused by a decrease in bone formation. We examined the efficacy of PTH in preventing the detrimental skeletal effects that occur with hindlimb unloading (HLU). Materials and Methods: Six-month-old male Fisher 344 rats were HLU and treated with vehicle or recombinant human PTH(1-34) at 1, 5, 20, or 80 g/kg/day for 2 weeks. The bone response was measured by CT analysis of bone structure, histomorphometric analysis of static and dynamic bone parameters, and Northern blot analysis of mRNA levels for bone matrix proteins. The PTH-treated HLU animals were compared with vehicle-treated HLU and pair-fed normal weight-bearing controls. Results: Unloading resulted in a decrease in cancellous bone volume that was caused in part by a dramatic 83% decrease in bone formation. All dose rates (1-80 g/kg/day) of human PTH(1-34) significantly increased bone formation rates compared with vehicle-treated HLU controls. There was a dose response, and the highest dose rate of the hormone increased bone formation compared with normal weight-bearing rats by 708% (p < 0.0001). The increases in bone formation were accompanied by increases in mRNA levels for type 1 collagen, osteocalcin, and osteonectin. Also, treatment with PTH resulted in increases in mineral apposition rate and double-labeled perimeter, but the latter was disproportionally increased at high dose rates. A therapeutic dose of PTH (1 g/kg/day) prevented disuse-induced trabecular thinning, whereas high-dose PTH (80 g/kg/day) increased trabecular thickness compared with normal weight-bearing rats. Conclusions: These findings reveal that administration of a therapeutic dose of PTH to HLU rats prevents the decrease in bone formation and trabecular thinning, whereas high dose rates of the hormone increase bone formation and trabecular thickness to values that exceed normal values.