Study of costameric proteins in soieus muscle longitudinal sections of female rats immobilized and rehabilitated by intermittent passive manual stretching

Abstract

The interface between the sarcolemma and the extracellular matrix in skeletal muscle tissue is constituted by specialized proteins, responsible for the transmission of transversal and longitudinal forces to the myofiber, or mechanotransduction. Adaptations of skeletal muscle to physiological forces, and pathological ones like segmental immobilization and rehabilitation exercises, may contribute to cellular perception of mechanical signals and thus induce changes in flexibility and muscular force. The objective of this study is to evaluate functional alterations in gait and morphological aspects of the soleus muscle in female rats having the right hind leg immobilized and later submitted to two training protocols: (1) intermittent passive manual stretching during 3 days and (2) during 10 days. Experiments are planned with 48 female Wistar rats, weighing around 250g divided into 6 groups: Immobilized (IG); Immobilized and stretched for 3 days (ISG (3)); Immobilized and stretched for 10 days (ISG(10)); Immobilized and free for 3 days (IFG(3)); Immobilized and free for 10 days (IFG(10)); Immobilization control (ICG). Female rat hind limb will be immobilized for 10 days in a plantar flexion to keep the soleus muscle in a shortened position. Care will be taken to prevent ulceration and edema of limb endings. The animals will be further submitted to intermittent passive manual stretching. Animal gait will be functionally analyzed before immobilization and during the period of stretching or free movement. After the stated periods of time and sacrifice the soleus muscle will be removed, frozen and further processed for reactions (H.E., Gomori trichrome stain) and immunohistochemistry tests for dystrophin, larninin, fibronectin, satellite cells and macrophages (Antibodies, rabbit anti-rat dystrophin, rabbit anti-rat larninin, rabbit anti-rat fibronectin, goat anti-rat NCAM, mouse anti-rat PAX7, mouse anti-rat CD68). Qualitive and quantitative results will be obtained with a Light Microscope and a system of image analysis. Inter- and intra-group data will be treated by statistical techniques like: statistical calculations of the central position (mean, median, mode) and of dispersion (standard and quartile deviations), Student t test for hypothesis, the Wilcoxon-Mann -Whitney test, F (ANOVA and the Kruskal-Wallis test besides graphical methods of description and comparison. (AU)