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The use of mesenchymal stromal cells (MSCs) differentiated toward a smooth muscle cell (SMC) phenotype may provide an alternative for investigators interested in regenerating urinary tract organs such as the bladder where autologous smooth muscle cells cannot be used or are unavailable. In this study we measured the effects of good manufacturing practice (GMP)-compliant expansion followed by myogenic differentiation of human MSCs on the expression of a range of contractile (from early to late) myogenic markers in relation to the electrophysiological parameters to assess the functional role of the differentiated MSCs and found that differentiation of MSCs associated with electrophysiological competence comparable to bladder SMCs. Within 1–2 weeks of myogenic differentiation, differentiating MSCs significantly expressed alpha smooth muscle actin (αSMA;  ACTA2 ), transgelin ( TAGLN ), calponin ( CNN1 ), and smooth muscle myosin heavy chain (SM-MHC;  MYH11 ) according to qRT-PCR and/or immunofluorescence and Western blot. Voltage-gated Na +  current levels also increased within the same time period following myogenic differentiation. In contrast to undifferentiated MSCs, differentiated MSCs and bladder SMCs exhibited elevated cytosolic Ca 2+  transients in response to K + -induced depolarization and contracted in response to K +  indicating functional maturation of differentiated MSCs. Depolarization was suppressed by Cd 2+ , an inhibitor of voltage-gated Ca 2+ -channels. The expression of Na + -channels was pharmacologically identified as the Na v 1.4 subtype, while the K +  and Ca 2+  ion channels were identified by gene expression of  KCNMA1 ,  CACNA1C  and  CACNA1H  which encode for the large conductance Ca 2+ -activated K +  channel BK Ca  channels, Ca v 1.2 L-type Ca 2+  channels and Ca v 3.2 T-type Ca 2+  channels, respectively. This protocol may be used to differentiate adult MSCs into smooth muscle-like cells with an intermediate-to-late SMC contractile phenotype exhibiting voltage-gated ion channel activity comparable to bladder SMCs which may be important for urological regenerative medicine applications.

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Smooth Muscle-Like Cells Generated from Human Mesenchymal Stromal Cells Display Marker Gene Expression and Electrophysiological Competence Comparable to Bladder Smooth Muscle Cells