Cloning of local growth factors involved in the determination of muscle mass.

Unlike the cells of many tissues, muscle and neuronal cells do not replicate throughout life. Therefore there has to be an eVective mechanism of inducing local repair and preventing cell death. We have identified and cloned two growth factors that are expressed by muscle when it is subjected to activity which are derived from the insulin like growth factor-I (IGF-I) gene by alternative splicing. 2 One isoform, muscle L.IGF-I, is very similar to the liver type of IGF-I. The other is a new growth factor that could only be detected in exercised or stretched muscle. This has been called mechano growth factor (MGF) to distinguish it from liver IGF-I which has a systemic mode of action. The structure of the cDNA of this isoform indicates that it has diVerent exons from the liver types and is not glycosylated. Therefore it is smaller and has a shorter half life than liver IGF-I unless it is bound to its tissue binding protein. Unlike hormones, the site and mode of action of growth factors are determined to a large extent by their specific binding proteins. Both muscle isoforms are upregulated by stretch and overload and the evidence indicates that, during exercise, muscle L.IGF-I contributes significantly to circulating levels of IGF-I. 3 MGF, however, appears to be designed for local action and does not enter the blood stream in any quantity. It has a 52 base insert in the E domain which alters the reading frame of the 3’ end, which results in it binding to a diVerent binding protein which exists in the interstitial tissue spaces of muscle and neuronal tissue. This would be expected to localise its action as it would be unstable in the unbound form; this is important as its production would not unduly perturb blood sugar levels. Although MGF production in response to stretch during exercise is apparently to induce local repair, its overexpression in situations that involve pronounced overload—for example, weight lifting—results in hypertrophy. Therefore it has an adaptative as well as a protective function. The mechanism whereby cells respond to mechanical signals must involve a mechanochemical transduction mechanism to link the physical signal with the activation or repression of certain genes. Recently, there have been a number of studies on other cell types that implicate the cytoskeleton in mechanochemical transduction. These have been reviewed by Ingebar. Experiments have shown that in dystrophic muscle, including that in mdx mice (a model for dystrophin deficient dystrophies) and dydy mice (model for autosomal dystrophies), MGF is not expressed as it is in normal mice muscle when stretched. 6 In both types of dystrophy, the cytoskeletal dystrophin complex is defective and so apparently is the mechanotransduction system. At the C-terminus, dystrophin is attached to an elaborate array of diVerent proteins, which are in turn attached to the extracellular matrix through laminin (merosin) one of which is missing in the autosomal dystrophies. Also associated with this complex are neuronal nitric oxide synthase and a tyrosine kinase, therefore it seems inconceivable that this elaborate structure is present merely to stiVen the membrane. The defects in the rather specialised cytoskeleton in muscle apparently results in inadequate production of local IGF-I, and the failure to maintain muscle mass is a possible cause for all the dystrophies. Using an isolated heart preparation, we were able to detect the production of the MGF peptide using a specific antibody just two hours after the heart had been subjected to a pressure overload. This growth factor appears to be involved in protecting heart as well as skeletal muscle by inducing local repair and preventing apoptosis. There is also evidence that it is also involved in maintaining nervous tissue, as IGF-I is known to be transported within neurones. Therefore the possibility exists that motor neurone maintenance is facilitated by IGF-I produced by the active muscles that they innervate. Using a specific antibody that we have generated, we found the MGF splice variant in the central nervous system, which is another tissue in which there is no cell replacement and in which continuous local repair is very important. In addition, the systemic type of IGF-I produced by active muscles (L.IGF-I) and its beneficial eVect on other types of tissue, as well as the protective and adaptative eVects of the autocrine variant (MGF), provides a basis for understanding the beneficial eVects of exercise on general health. Recently, we placed MGF cDNA in an engineered gene and injected it into muscles of the laboratory mouse. Somewhat to our surprise, we found a 20% increase in muscle mass in two weeks. At the same time, a group in Philadelphia introduced the liver type IGF-I into muscles using a similar approach and they also reported a 20% increase but only after four months. Interestingly, this group reported greater increases within four months in the muscles of older mice as compared with the normal aging controls, indicating that the age related loss of muscle mass (sarcopenia) is associated with decreased IGF-I levels. As systemic levels of growth hormone and liver IGF-I decrease with age, this can be supplemented by the potent MGF version produced by active muscles. This emphasises the need to remain active during later years. Clearly MGF is very potent and much more eVective in increasing muscle mass than the liver type of IGF-I. This may be good news for the elderly suVering from advanced sarcopenia and for children suVering from muscular dystrophy and other musculoskeletal problems, but it will be open to abuse. If one intramuscular injection results in a 20% increase in that muscle within a short time, it may be predicted that it will be misused—for example, for resculpturing the body for athletic performance or other non-medical purposes. As MGF and muscle L.IGF-I are produced naturally by the body during activity, it would be diYcult to detect whether an individual performance had been enhanced by gene therapy or achieved by hard training. At present, there is apparently considerable misuse of recombinant liver type IGF-I, and it is naive to think that this would not be extended to include the more potent form. It is to be hoped that the method we have in mind for testing for this form of abuse (which cannot be disclosed at this juncture) will be eVective in detecting exogenous MGF.