Chapter 7. Conclusions

This study confirms the previous findings of decreased collagen biosynthesis, and brings new knowledge of type IV collagen turnover during immobilization. While the synthesis of collagens is decreased during immobilization, the degradation of type IV collagen is simultaneously increased, leading to decreased type IV collagen concentration. The substantially increased expression of MMP-2 and its proenzyme form is mediated by increased gene expression of the enzyme itself, possibly together with the increased activation of MMP-2 by diminished TIMP-2 levels. These results suggest strictly regulated net degradation of the major basement membrane component, type IV collagen, during the first week of immobilization. It would be interesting to see the early responses of type IV collagen, MMP-2 and TIMP-2 to remobilization. Future studies are also needed to explore the role of TIMP-2 in the activation and inhibition of MMP-2 in muscle tissue during normal turnover and in diseases.

The effects of glucocorticoids on skeletal muscle collagens have not been studied previously. The gene expression of type IV collagen decreases during 10-day dexamethasone treatment. Diminished gene expression of MMP-2 causes the quantity of proMMP-2 to be decreased, although the inhibitory activity of TIMP-2 towards proMMP-2 and MMP-2 is also decreased. The content of type IV collagen is not changed due to the decrease in both gene expression and degradation of this protein. This deceleration of type IV collagen turnover is probably caused by the dexamethasone-induced inhibition of overall protein synthesis, and the inhibition of transcription factors inducing the synthesis of MMPs. Thus, while they are clinically used as anti-inflammatory agents, glucocortoids do not seem to cause excessive loss of the basement membrane protein, type IV collagen.

The present study confirms the previous results of increased gene expression of collagens in polymyositis and polyneuropathy. The results further suggest type IV collagen accumulation not only in polymyositis but also in polyneuropathy. This accumulation of type IV collagen is physiologically disadvantageous, since thickening of the basement membranes of capillaries and muscle fibers complicates the transportation of nutrients and cellular excreta between blood and muscle cells. The increase in type IV collagen concentration seems to be caused by increased gene expression of type IV collagen with constant degradation by MMP-2. Underlying diseases, e.g. diabetes, in polyneuropathy patients may have a role in the observed increase in type IV collagen concentration. Further research could be conducted to find out the possible roles of systemic diseases on skeletal muscle connective tissue. Unexpectedly, increased quantity of proMMP-9 with increased staining intensity of MMP-9 in some cells was observed in polyneuropathy muscles. The possible role of MMP-9 in the pathogenesis of polyneuropathy could be an interesting area for future research.

The changes in collagen synthesis and degradation in rats can be partly prevented by immobilizing muscles in lengthened positions. Stretch applied in leg muscles during immobilization has preventive effects especially on the plantarflexors, which are the muscles most sensitive to immobilization. The exercise used in this study did not prevent the anti-anabolic effects of 10-day dexamethasone treatment in rats, even though exercise itself had enhancing effects on type IV collagen gene expression and TIMP-2 quantity. To find out whether exercise has any compensatory effects on the glucocorticoid-induced changes, more strenuous or prolonged exercise could be used in future studies. It would also be interesting to find out the effects of exercise prior to glucocorticoid treatment.