Isolation, cultivation and immunostaining of single myofibers: An improved approach to study the behavior of satellite cells.

Satellite cells are myogenic cells responsible for muscle growth shortly after birth and muscle repair/regeneration during adulthood. Therapies based on satellite cells hold promise for treating muscular dysfunctions. Studying satellite cells is technically challenging owing to their low abundance, small size and anatomical dispersed location between the basal lamina and the sarcolemma of myofibers. In this article, we present three improved protocol strategies for studying the properties of satellite cells of the mouse during the different stages of muscle regeneration: (1) immunostaining of freshly isolated single myofibers to facilitate the study of quiescent satellite cells, (2) cultivation of single myofibers on Matrigel®-coated dish to study the myogenesis programs initiated by satellite cell activation, and (3) cultivation of single myofibers in floating conditions to analyze activated satellite cells or the doubling time of satellite cells in myofibers. In brief, when compared to previously published protocols, this article presented an improved protocol that requires shorter experimental time and less laborious approach for higher yield of intact single myofibers for downstream analyses.

Defects in nerve conduction velocity and different muscle fibre-type specificity contribute to muscle weakness in Ts1Cje Down syndrome mouse model.

BACKGROUND: Down syndrome (DS) is a genetic disorder caused by presence of extra copy of human chromosome 21. It is characterised by several clinical phenotypes. Motor dysfunction due to hypotonia is commonly seen in individuals with DS and its etiology is yet unknown. Ts1Cje, which has a partial trisomy (Mmu16) homologous to Hsa21, is well reported to exhibit various typical neuropathological features seen in individuals with DS. This study investigated the role of skeletal muscles and peripheral nerve defects in contributing to muscle weakness in Ts1Cje mice. RESULTS: Assessment of the motor performance showed that, the forelimb grip strength was significantly (P<0.0001) greater in the WT mice compared to Ts1Cje mice regardless of gender. The average survival time of the WT mice during the hanging wire test was significantly (P<0.0001) greater compared to the Ts1Cje mice. Also, the WT mice performed significantly (P<0.05) better than the Ts1Cje mice in the latency to maintain a coordinated motor movement against the rotating rod. Adult Ts1Cje mice exhibited significantly (P<0.001) lower nerve conduction velocity compared with their aged matched WT mice. Further analysis showed a significantly (P<0.001) higher population of type I fibres in WT compared to Ts1Cje mice. Also, there was significantly (P<0.01) higher population of COX deficient fibres in Ts1Cje mice. Expression of Myf5 was significantly (P<0.05) reduced in triceps of Ts1Cje mice while MyoD expression was significantly (P<0.05) increased in quadriceps of Ts1Cje mice. CONCLUSION: Ts1Cje mice exhibited weaker muscle strength. The lower population of the type I fibres and higher population of COX deficient fibres in Ts1Cje mice may contribute to the muscle weakness seen in this mouse model for DS.