Journal: Stem Cell Research & Therapy

Article Title: Generation of myogenic progenitor cell-derived smooth muscle cells for sphincter regeneration

doi: 10.1186/s13287-020-01749-w

Figure Lengend Snippet: Characterization of MPC to MPC-SMC differentiation. a Expression of intracellular muscle-specific proteins performed by immunostainings using antibodies against aSMA, smoothelin, and desmin, each combined by nuclear staining with Hoechst dye. Representative images of cells, derived from three individual human biopsies, are shown. Scale bar (white) = 100 μm for all images. b Quantification of percentage MPC and MPC-SMC positive for myogenic markers aSMA, smoothelin, and desmin. c Fold change in cell number calculated from an initially seeded number of 500,000 cells in T175 flasks after 6 days in either growth (MPC) or smooth muscle differentiation medium (MPC-SMC). Flow cytometric quantification of d CD146 and e CD49a in populations of MPC cultivated in growth (MPC) or smooth muscle differentiation medium (MPC-SMC) for 6 days. All data ( b – e ) demonstrated as mean ± SD and derived from at least three individual human muscle biopsies. Statistical comparison performed between MPC and MPC-SMC by paired t test ( c – e ) or two-way ANOVA with multiple testing applying Holm-Sidak correction ( b ) considering a p < 0.05 as statistically significant (n.s., not significant)

Article Snippet: Primary human bladder smooth muscle cells (HBdSMC-c) were obtained from PromoCell® (Cat. No.: C-12571) and cultivated in growth medium (Ham’s F10, 10% FCS, bFGF, gentamicin) for expansion to reach synthetic smooth muscle cells (s-hBd-SMC).

Techniques: Expressing, Staining, Derivative Assay

Journal: Stem Cell Research & Therapy

Article Title: Generation of myogenic progenitor cell-derived smooth muscle cells for sphincter regeneration

doi: 10.1186/s13287-020-01749-w

Figure Lengend Snippet: Change in the gene expression of MPC and MSC during SMC differentiation. Changes in the gene expression were assessed by microarray analysis of RNA isolated from MSC, MPC, MSC-SMC, and MPC-SMC. Cells derived from two individual human muscle biopsies were cultivated in growth (MSC and MPC) or smooth muscle differentiation medium (MSC-SMC and MPC-SMC) for 6 days, respectively. a Hierarchical clustering (Euclidean distance) of Log2 fold changes of smooth muscle-associated genes in MSC vs MSC-SMC (MSC) and MPC vs MPC-SMC (MPC). The cluster of genes similarly b upregulated or c downregulated in both MSC and MPC upon SMC differentiation obtained by k -means clustering. *Genes which in both cell types were either log2 FC ≥ 1 or log2 FC ≤ − 1 up- or downregulated. d Percent of tested genes calculated to be upregulated or downregulated during MSC to MSC-SMC or MPC to MPC-SMC differentiation. Statistical comparison was performed by chi-squared test considering a p value below 0.05 as significant

Article Snippet: Primary human bladder smooth muscle cells (HBdSMC-c) were obtained from PromoCell® (Cat. No.: C-12571) and cultivated in growth medium (Ham’s F10, 10% FCS, bFGF, gentamicin) for expansion to reach synthetic smooth muscle cells (s-hBd-SMC).

Techniques: Expressing, Microarray, Isolation, Derivative Assay

Journal: Stem Cell Research & Therapy

Article Title: Generation of myogenic progenitor cell-derived smooth muscle cells for sphincter regeneration

doi: 10.1186/s13287-020-01749-w

Figure Lengend Snippet: Stability of smooth muscle differentiation program of MPC-SMC. a Smooth muscle lineage marker (aSMA) expression stained by fluorescent immunostaining was compared between MPC, MPC-SMC, and MPC-SMC de-differentiated by cultivation for 3 days in growth medium following cultivation in smooth muscle differentiation medium (De-diff MPC-SMC). Representative images of MPC, MPC-SMC, and De-diff MPC-SMC derived from at least three different skeletal muscle biopsies are shown. Scale bar = 100 μm. b Formation of myotubes (red arrows) observed by fluorescence microscopy following Hoechst33342 staining for visualization of the nuclei as well as c quantification of fusion index (FI) and the number of nuclei was compared between MPC, MPC-SMC, and De-diff MPC-SMC each cultivated in skeletal muscle differentiation medium for 6 days. Scale bar = 200 μm. d Quantification of percent aSMA-positive cells in MPC-SMC and De-diff MPC-SMC was performed. e Comparison of AChE and CK enzyme activity between MPC, MPC-SMC, and MPC in skeletal muscle differentiation medium (MPC in SKDiff) and MPC-SMC in skeletal muscle differentiation medium (MPC-SMC in SKDiff). All data presented as mean ± SD of cells derived from at least three individual human muscle biopsies. Statistical analysis performed by paired t test considering p < 0.05 as significant

Article Snippet: Primary human bladder smooth muscle cells (HBdSMC-c) were obtained from PromoCell® (Cat. No.: C-12571) and cultivated in growth medium (Ham’s F10, 10% FCS, bFGF, gentamicin) for expansion to reach synthetic smooth muscle cells (s-hBd-SMC).

Techniques: Marker, Expressing, Staining, Immunostaining, Derivative Assay, Fluorescence, Microscopy, Activity Assay

Journal: Stem Cell Research & Therapy

Article Title: Generation of myogenic progenitor cell-derived smooth muscle cells for sphincter regeneration

doi: 10.1186/s13287-020-01749-w

Figure Lengend Snippet: Engraftment of mMPC-derived SMC into smooth muscle tissue in vivo. a Quantification of percent desmin- and aSMA-positive cells performed by immunofluorescence staining of two preparations of mMPC-SMC. b Fluorescence signal detection of co-injected fluorescent beads and TdTomato transgene expressing cells localized in the intact pyloric sphincter muscle by IVIS imaging ex situ. c Tissue engraftment and aSMA protein expression of TdTomato-positive mMPC-SMC in pyloric sphincter histological sections of intramuscular-injected SMC 12 weeks after implantation. Representative images of n = 8 injected mice

Article Snippet: Primary human bladder smooth muscle cells (HBdSMC-c) were obtained from PromoCell® (Cat. No.: C-12571) and cultivated in growth medium (Ham’s F10, 10% FCS, bFGF, gentamicin) for expansion to reach synthetic smooth muscle cells (s-hBd-SMC).

Techniques: Derivative Assay, In Vivo, Immunofluorescence, Staining, Fluorescence, Injection, Expressing, Imaging, Ex Situ