Myogenic Differentiation of Mesenchymal Stem Cells is Induced by Striated Muscle Influences in vitro

Anna      Bajek; Tomasz      Drewa; Romana      Joachimiak; Zaneta      Spoz; Maciej      Gagat; Magdalena      Bodnar; Robert      Debski; Alina      Grzanka; Andrzej      Marszalek

Abstract

Introduction and Hypothesis: Regeneration of external urinary sphincter can improve stress urinary incontinence following surgical treatment of prostate cancer. One of the most promising approaches is to force transformation of injected undifferentiated stem cells into muscle phenotype by surrounding tissues, which release paracrine factors. The aim of the study was to determine whether different conditioned media and co-culture system may induce transdifferentiation of bone marrow mesenchymal stem cells (MSCs) in muscle cells.

Methods: MSCs were harvested from bone marrow, and cultured for 14d prior differentiation to myogenic lineage protocol. Conditioned media were prepared from skeletal muscle cell line (CRL 1458), and primary muscle precursor cells culture. Co-culture system was prepared from bone marrow mesenchymal stem cells and muscle precursor cells. Mesenchymal stem cells and primary culture of muscle precursor cells phenotype were confirmed by flow cytometry and immunofluorescent labeling. The transdifferentiation process was detected by immunofluorescence studies for calponin, α-actin, desmin, sarcomeric actin and myogenin.

Results: At 2 weeks after culturing MSCs in media conditioned with growth factors showed the expression of specific skeletal muscle markers. The level of expression of muscle differentiation markers was higher in comparison to the control (MSCs without conditioned media supplementation). The expression of sarcomeric actin and myogenin was comparable to muscle precursor cells. Skeletal muscle markers as well as polyploid cells formation were observed in MSCs when co-cultured with muscle precursor cells.

Conclusion: Transdifferentiation of bone marrow MSCs into skeletal muscle cells can be evoked by growth factors released by neighboring cells building rhabdosphincter.

Keywords: Bone marrow mesenchymal stem cells, external urinary sphincter regeneration, stress incontinence, immunofluorescence, calponin, cytometry, desmin, α-actin, chondrogenic, Adipogenic