Fig 1.
Immunocytochemical detection of SOX2 expression four days after transduction.
BJ fibroblasts were stained with anti-SOX2 antibody (A), counterstained with 4’,6-diamidino-2-phenylindole, DAPI (B) and results of staining were merged (C). Images were captured using Eclipse Ci-S epifluorescence microscope with 40x objective; scale bar = 50 μm.
Fig 2.
Characterization of induced pluripotent stem cells (iPSCs) derived from BJ fibroblasts by means of lentiviral transduction.
iPSCs were spontaneously differentiated via embryoid body formation into cells from three germ layers. Differentiated cells expressed markers of ectoderm—MAP2 and presented neuronal morphology (A); mesoderm—alpha-smooth muscle actin (αSMA) and features of cytoskeleton (B); and endoderm—SOX17 (C). Images were captured using Eclipse Ci-S epifluorescence microscope with 20x objective; scale bar = 100 μm.
Fig 3.
The results of real-time PCR analysis.
Expression of each gene in each analyzed sample was firstly normalized to cDNA from all analyzed samples (pooled cDNA) and compared to normalized expression in BJ fibroblasts (SOX2, MSI1, nestin) and iPSCs (NKX2.2). The p-value was indicated in the selected samples (* p < 0.05; ** p < 0.01; *** p < 0.001; SEM), all statistical data are summarized in S4 Table.
Fig 4.
The expression of three neural stem cell markers SOX2, nestin and SOX1 in NSC (A, E), ebiNSc (B, F), SiNSc-like cells (C, G) and SMiNSc-like cells (D, H).
Images were captured using Eclipse Ci-S epifluorescence microscope with 20x (A, B, E, F) and 40x objective (C, D, G, H); scale bar = 50 μm.
Fig 5.
The effectiveness of induced neural stem cell generation presented as the percentage of NSC, ebiNSc, SiNSc-like cells and SMiNSc-like cells expressing simultaneously SOX2 and nestin (SEM).
Fig 6.
The expression of MAP2 and GFAP in NSC (A, E), ebiNSc (B, F), SiNSc-like cells (C, G) and SMiNSc-like cells (D, H) after 7 and 14 days of differentiation in the neural induction medium.
Images were captured using Eclipse Ci-S epifluorescence microscope with 20x (A, B, E, F) and 40x objective (C, D, G, H); scale bar = 50 μm.
Fig 7.
The expression of MAP2 and TH in NSC (A, E), ebiNSc (B, F), SiNSc-like cells (C, G) and SMiNSc-like cells (D, H) after 7 and 14 days of differentiation in the neural induction medium.
Images were captured using Eclipse Ci-S epifluorescence microscope with 20x (A, B, E, F) and 40x objective (C, D, G, H); scale bar = 50 μm.
Fig 8.
The expression of Synapsin I and Tau in NSC (A, E), ebiNSc (B, F), SiNSc-like cells (C, G) and SMiNSc-like cells (D, H) after 7 and 14 days of differentiation in the neural induction medium.
Images were captured using Eclipse Ci-S epifluorescence microscope with 20x (A, B, E, F) and 40x objective (C, D, G, H); scale bar = 50 μm.
Fig 9.
The expression of VGLUT1 in NSC (A, C) and ebiNSc (B, D) lines after 7 and 14 days of differentiation in the neural induction medium.
Images were captured using Eclipse Ci-S epifluorescence microscope with 20x objective; scale bar = 100 μm.
Fig 10.
The potency of neuronal cells generation.
Comparing the total number of MAP2-positive cells (A) and MAP2 expressing cells with distinctive neuronal morphology (B) within the population of NSC and ebiNSc after seven days of differentiation (SEM).
Fig 11.
The results of BrdU incorporation assay and Senescence-Associated (SA)-β-galactosidase staining obtained for NSC (A, E), ebiNSc (B, F), SiNSc-like cells (C, G) and SMiNSc-like cells (D, H) after 48 h of incubation.
Images were captured using Eclipse Ci-S epifluorescence microscope with 20x objective; scale bar = 100 μm.
Fig 12.
Incorporation of BrdU in analyzed cell lines after 48 h of incubation (SEM).