Alterations of fibronectin and laminin and expression of matrix metalloproteinases following skeletal muscle ischemia-reperfusion injury

Abstract

Aims: To investigate alterations in flrbronectin (FN) and laminin (LN) levels, and the expression of associated caseinolytic MMPs in tissues following skeletal muscle (SM) ischemia and reperfusion (VR). Materials and Methods: The rats were subjected to 4 h hindlimb ischemia, followed by reperfusion for 0, 4, 24 or 72 h. Two further groups was administered doxycycline before ischemia was induced. The rats were then sacrificed and samples were processed. ELISA, Immunohistochemical techniques,Zymography and Western blotting were employed. Results: Plasma FN (pFN) decreased following hindlimb ischemia and subsequently increased with reperfusion. FN accumulated in the basement membrane (BM) of SM, whereas degradation occurred in the lungs and kidneys after SM I/R. FN decreased in the liver after SM ischemia and then increased with reperfusion. However, LN was degraded in all BM. Following doxycycline treatment, levels of FN increased in lungs, kidneys, and liver. Degradation of LN was inhibited in all tissues by doxycycline. In addition, a 20 l<Da caseinolytic species was identified in lung tissues, which was inhibited by EDTA and 1,10- phenanthroline, but not inhibited by PMSF. This species was down-regulated by anesthesia only and then up-regulated immediately following bilateral hindlimb yR. An increased production of this MMP was also found after treatment of the rats with doxycycline. Conclusions: FN has a specific affinity to injured tissue, as was minored by rapid depletion of pFN, which was secreted by the liver. Moreover, the degradation of FN in lungs and kidneys and LN in all BM was suggested to be due to the effects of MMPs. In addition, doxycycline reduced the degradation of FN in lungs and kidneys and LN in all tissues studied by inhibiting the activity of MMPs. However, degradation of FN was not evident in SM, because there was an over deposition of FN in SM following VR. Furthermore, a caseinolytic MMP, rather than a serine protease, was involved in lung injury following SM I/R Doxycycline increased the production of this MMP. Although it has not been identified, this MMP was suggested to be a lower molecular weight MMP, possibly MMP-7 or MMP-12. Further study is needed for confirmation of its identity.