Figure 3.

Characterization of USC fractions from DMD and control individuals

(A) Comparison of cellular morphology among fractions. No differences in the morphology are observed between the total fraction (Ftot) and the sorted fractions F1, F2, and F3: both elongated and rounded cells are present before and after Celector separation. The morphology of the third control sample was like CTRL4.

(B) Evaluation of CD surface markers expressed in the control (upper graph) and DMD (lower graph) USCs by flow cytometry. Bars represent the average of expression between controls (3 samples) and the two DMD samples (unpaired t test: ∗p < 0.05). CD73 and CD105 were highly expressed in all control USC fractions, while significantly decreased in some fractions of the DMD USCs. CD73 is significantly lower both in F1 and F2 of DMD USCs as compared to F1 and F2 of control (p < 0.05 and p < 0.01, respectively) and in F2 compared to F1 in DMD USCs (p < 0.05). CD105 is significantly lower in Ftot, F2, and F3 (p < 0.05) of DMD fractions as compared to controls. CD146 has low expression in USCs of the control and is almost not expressed in DMD USCs.

(C and D) Gene microfluidic card (FluiDMD) profiles of native USCs from DMD IH carrying the deletion of exons 50–52 (C) and DMD AF with the deletion of exons 46–47 (D). FluiDMD shows amplification of the DMD isoforms for the three fractions (F1 blue bar, F2 red bar, and F3 green bar) obtained following the separation by Celector. M and B isoforms are represented both in F2 and F3, while P and Dp140 isoforms are present only in DMD individual AF and in F3. Dp71 is expressed in all the fractions, including F1. Dp260 and Dp116 isoforms are inconsistently expressed across fractions. DCt, delta Ct corresponding to the difference between Ct from DMD isoforms and Ct of the housekeeping gene (β-actin).