Résumé :
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Adult stem cells reside in all tissues, where they maintain homeostatic conditions and respond to injuries. These cells are regulated and supported by the surrounding microenvironment, called stem cell "niche", composed by cellular and molecular factors, that interact with and regulate stem cell fate. Key niche components are represented by growth factors, cell-cell interactions and cell-matrix adhesion. The interplay of these factors is very important to understand the in vivo stem cell behaviour, both in physiological and pathological conditions. Also in muscle tissue niches are present, especially in myofiber basal lamina, where a network of extracellular matrix components (such as collagen, laminin and fibronectin) and secreted growth factors (i.e. HGF, IGF-1and EGF) stimulate muscle stem cell survival, activation and/or proliferation. In order to better understand the influence of muscle tissue secreted factors on stem cells, we developed a new technical approach to perform a muscle tissue-stem cells co-culture; in particular we focused on a specific peripheral blood derived population, expressing stemness surface marker, known to possess a myogenic potential. The development of a culture system that create a physical separation between a fresh muscle tissue section and stem cells allows soluble factors exchange and prevent tissue-cell contamination; the absence of cell mix was evaluated through cytogenetic analysis of cell karyotype; the reproducibility of the technique have been demonstrated through muscle slice weight monitoring. Preliminary proliferation experiments have been performed, with results showing an increase in CD133+ stem cells rate of proliferation in presence of muscle tissue; cell immunophenotype has been monitoring to confirm the maintenance of stemness. Future experiments are focused on identifying the factors released and correlating them to specific pathological or physiological effects on stem cells. In conclusion, we have developed a reproducible and standardized technique for ex vivo muscle tissue slice and designed a culture system that guarantees, spatial division of culture environment, no cellular contaminations between culture compartments, in vitro tissue survival, without necrosis morphology for 7 days, stem cell viability and proliferation, communication between tissue and cells through chemical signals (factors release).
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