Fig. 3

Osteogenic, adipogenic, myofibroblastic and neurosphere formation capacity of L-GSCs. A The osteogenic capacity was confirmed by Alizarin Red S staining after 28 days of culture of S-GFs, S-GSCs, L-GFs and L-GSCs in either FCS or PL osteogenic medium. B The adipogenic potential of L-GSCs and S-GSCs in both PL and FCS adipogenic media was confirmed by Oil Red O staining of the intracellular lipid vacuoles. C The spectrometry of Alizarin Red S staining showed that L-GFs and L-GSCs had 2–2.5-fold and fourfold higher values than S-GFs and S-GSCs in FCS and PL-osteogenic media, respectively. The spectrometry of Oil Red O staining showed no significant difference of the adipogenic capacity of L-GSCs and S-GSCs in FCS adipogenic medium, but a higher capacity of L-GSCs in PL-adipogenic medium. D Xylenol orange showed mineral deposits at day 15 of osteogenic differentiation in both S-GSCs and L-GSCs. E SEM imaging showed that both S-GSCs and L-GSCs were attracted to the allogenic Cancellous Particulate Allograft (ZIMMER-BIOMET) and formed mineral nodules around and between the particles of this matrix. F Immunostaining with Nestin and β3-tubulin antibodies confirmed the phenotype of neurospheres formed from S-GSCs and L-GSCs after 7 days of culture in untreated neurogenic medium. G Immunofluorescence labelling with α-SMA/Actin/Dapi antibodies in both groups of GSCs confirmed their capacity to undergo myofibroblastic differentiation after 5 days of culture under suitable conditions. Scale bars: A 1000 µM, B 100 µM, D 400 µM, E 400 µM, F 400 µM, G 25 µM