Fig. 4
Osteogenic effects of Osteo- and Naïve-EVs in cultured hMSC. a Representative images of ALP after 14 days. b Representative images of ARS after 21 days. Matrix mineralization was greater after Osteo-EVs treatment than after treatment with the Naïve-EVs and control medium. Quantification of the ARS experiment from three independent donors indicated that Osteo-EVs enhanced matrix mineralization of hMSC after 21 days. c Osteogenic gene expression array heatmap among the three donors of each group: control-medium, Naïve- and Osteo-EVs, after 14 days. d Prediction ellipses showed a new observation of donors from the same group falling within the prediction ellipse (confidence interval 95%). e Gene expression level (fold change) between the control medium and EV groups showed upregulation of several genes related to osteogenesis in the EV groups. Genes with red colours are identified in the Osteo-EVs and those with green colours are identified in the Naïve-EVs; green boxes refer to the downregulated genes in the Naïve-EVs f Gene expression level (relative fold change) between the Naïve- and Osteo-EVs showed upregulation of several genes related to osteogenesis in the Osteo-EVs group. g Validation of gene array findings with RT-qPCR: Osteo-EVs triggered higher mRNA expression levels of osteogenesis-related markers SPP1, BSP and BGLAP. *p < 0.05, **p < 0.01, ****p < 0.0001