For this approach to be valid, it is necessary to be sure that the adaptive response of the loaded bones is confined Trichostatin A to those bones and does not influence their contra-lateral controls. This assumption has been challenged by the work of Sample et al. [30] who recently reported that in rapidly growing male rats a single period of dynamic high-magnitude axial loading of the ulna on one side was associated with significant levels of new cortical bone formation at the periosteal surface of the contra-lateral non-loaded ulna and in the cortical regions of adjacent bones in the loaded limbs. These responses were prevented by neuronal blockade.

The authors [30] inferred from this that mechanically adaptive bone (re)modelling is controlled by processes with substantial systemic and central nervous components. If this inference were true, the focus of research into the mechanisms of mechanically adaptive bone (re)modelling would need to shift away from local responses and toward systemic and central regulation. Although their inference did not accord with our experience [31], we could find no published studies specifically directed to establishing that the (re)modelling of bones contra-lateral to those which had been loaded buy Nivolumab was not different from that in bones in comparable animals where no bones had received artificial loading. Since use of

the contra-lateral non-loaded limb as a control has become accepted practice, we undertook the present study to assess whether this was indeed the case. C57BL/6 mice are extensively used as the background of genetically modified animals in the field of bone research, and therefore we used the C57BL/6 mouse unilateral tibia/fibula axial loading model [12], [27] and [29]. This model has the advantage over the ulna loading model [2], [8] and [3] of enabling the study of trabecular as well as cortical bone compartments. Virgin, female C57BL/6 mice at 8 weeks of age were purchased from Charles River Laboratories, Inc.

(Margate, UK) and group-housed in sterilized polypropylene cages with PtdIns(3,4)P2 free access to water and a maintenance diet containing 0.73% calcium, 0.52% phosphorus and 3.5 IU/g vitamin D (RM1; Special Diet Services Ltd., Witham, UK) in a 12:12-h light/dark cycle, with room temperature at 21 ± 2 °C. Body weight was measured once a week until sacrifice at 21 weeks of age. All procedures complied with the UK Animals (Scientific Procedures) Act 1986 and were reviewed and approved by the ethics committee of the Royal Veterinary College (London, UK). At 19 weeks of age, 21 mice were randomly allocated to three equal numbered groups. In the NOLOAD group, neither left nor right tibiae/fibulae received any artificial load. In the STATIC group, the right tibia/fibula received a small (2.