Problems attributable to skeletal muscle during limb lengthening include muscle contractures, joint luxation, axial deviation, and joint stiffness. Foremost is the classic equinus contracture that frequently occurs with significant lengthening of the tibia, and requires extensive daily rehabilitation. Presumably, the equinus contracture develops because the longitudinal growth of the plantar flexor muscles lags behind the distraction of the tibia. The primary objective of this proposal is to address the underlying basis for the development of the equinus contracture by focusing upon the following use: What mechanism(s) regulates muscle fiber length in the plantar flexor muscles during tibial distraction? To address this issue, three fundamental hypotheses will be tested: i) the satellite cell hypothesis; ii) the transcriptional control hypothesis; and iii) the IGF-1 hypothesis. The """"""""satellite cell"""""""" hypothesis states that longitudinal growth of muscle fibers can only occur during distraction by activating satellite cells. This hypothesis will be tested in Phase I by distracting the tibia, and making measurements of muscle length, muscle fiber length, sarcomere length, sarcomere number, myonuclei, myonuclei derived from satellite cells, and cell volume/myonuclei ratio.
The specific aim of Phase II is to test both the """"""""satellite cell"""""""" and """"""""transcriptional control"""""""" hypotheses. Irradiation of the hind limb will be used to destroy the mitotic capacity of satellite cells. Hence, if satellite cell activation is a prerequisite for the longitudinal growth of muscle fibers during distraction, then irradiation will prevent this process from occurring. If the longitudinal growth of muscle fibers occurs via the upregulation of transcription (i.e., the transcription control hypothesis), then gel mobility shift assays will be used to identify potential regulatory elements of the actin promoter that respond to distraction, and DNA binding proteins that act on these elements.
The specific aim of Phase III is to test the IGF-1 hypothesis which states that the progressive chronic stretch imposed upon skeletal muscle by distraction will produce a local increase in IGF-1 that mediates the longitudinal growth of muscle fibers either by satellite cell activation or upregulation of transcription. This hypothesis will be tested by: i) examining the effects of distraction on the local concentrations of IGF-1; and ii) local infusion of a monoclonal antibody (MAb) that binds to the IGF-1 receptor and inhibits the action of IGF-1. In summary, the long term goal of this proposal is to provide a mechanistic understanding of factors mediating the longitudinal growth of muscle fibers during distraction so that interventions can be developed to minimize the occurrence of so-called contractures.
