FIG. 5.

Inhibition of FTRI-induced photoreceptor differentiation of hMSC by TGFβ1. (A) Culture of MIO-M1 cells with FTRI caused an increase in WNT2B mRNA expression, but addition of TGFβ1 to the differentiation medium inhibited this increase; n = 4. ANOVA test, **P < 0.01; ***P < 0.001. FTRI alone did not modify WNT5B mRNA expression, but addition of TGFβ1 to the differentiation cocktail increased WNT5B mRNA expression (similar to that shown above with TGFβ1 alone). Histograms represent the mean ± SEM from UV spectrophotometer readings of gel bands. Representative bands are shown above the histograms; n = 3. ANOVA test, **P < 0.01; ***P < 0.001. (B) Addition of TGFβ1 to hMSC undergoing photoreceptor differentiation with FTRI inhibited the mRNA expression of NR2E3, recoverin, and rhodopsin as compared with FTRI alone; n = 5–8. ANOVA test, *P < 0.05; ***P < 0.001. (C) Immunostaining for NR2E3 and recoverin confirmed that while FTRI alone caused a marked increase in the expression of this photoreceptor protein, addition of TGFβ1 to hMSC cultured with FTRI caused inhibition of photoreceptor differentiation (Alexa 488, fluorescent cells). Cell nuclei counterstained with DAPI (non-fluorescent cell structures). Scale bars 50 μm. Histograms on the right represent the proportion of cells immunostaining for each of the markers following 7-day culture under the different conditions; n = 3. ANOVA test, **P < 0.01.