The flexible neuronal conversion of malignant C6 glioma through the action of small molecules.

<p>(A and B) Representative images of C6 glioma before neural induction. (C) Representative fluorescence image of Tuj1-positive cells 7 days after neural induction of glioma with CHIR99021 (2 μM) and forskolin (10 μM). (D) Representative image of Tuj1/DAPI staining at 7 days after treatment of mouse embryonic fibroblasts with a combination of CHIR99021 (2 μM) and forskolin (10 μM). (E) Representative image of Tuj1-positive cells 7 days after treatment of glioma with a variety of concentrations of CHIR99021 and forskolin. Scale bars represent 500 μm (A), 50 μm (B), 100 μm (C), 100 μm (D), and 100 μm (E).</p

Anti-cancer effects of combined CHIR99021 and forskolin in a severe IMSCT model.

<p>(A) Neurologic function tests. (B) Representative image of dissected spinal cord at 16 days post-transplantation with RFP-expressing C6 glioma. (C) Representative optical imaging of dissected spinal cord at 16 days post-transplantation with RFP-expressing C6 glioma. (D) Representative image of sagittal sectioned-spinal cord at 16 days post-transplantation with RFP-expressing C6 glioma. (E) Quantitative result of ROI at 16 days post-transplantation with RFP-expressing C6 glioma. (F) Quantitative result of tumor area at 16 days post-transplantation with RFP-expressing C6 glioma. C, CHIR99021 (12.5 mg/kg); F, forskolin (10 mg/kg). * indicates <i>p</i> < 0.05. Asterisk means a significant difference to control. Data are presented as the mean ± S.E.M. Control (n = 11), CF (n = 7). Scale bars represent 1 cm (B) and 2 mm (D).</p

Gene expression profile of a malignant glioma and SMiN.

<p>(A) Heat map of significant gene in C6 glioma and SMiNs. (B) Up- or down-regulated gene ontology term in SMiNs. (C) Comparison of neuronal differentiation-related genes in C6 glioma and SMiNs. (D) Comparison of cell cycle-related genes in C6 glioma and SMiNs. (E) Comparison of ECM-related genes in C6 glioma and SMiNs. (F) Comparison of vessel development-related genes in C6 glioma and SMiNs. (G) Enriched gene list in C6 glioma and SMiNs.</p

Direct reprogramming of C6 glioma to neurons using two small molecules.

<p>(A) Representative fluorescence images of small molecule-induced neurons (SMiNs) 35 days after neural induction with CHIR99021 (20 μM) and forskolin (100 μM). (B) Quantitative result of Tuj1, MAP2, GFAP, PDGFR, and NG2 staining in SMiNs 35 days after neural induction. (C) Representative image of Tuj1- or MAP2-positive SMiNs 35 days after neural induction with CHIR99021 (20 μM) and forskolin (100 μM). (D) Quantitative result of morphological complexity of Tuj1-positive SMiNs 35 days after neural induction. (E) Representative band image of SSEA1 (a cancer stem cell marker) and C-Myc (an oncogene marker) 35 days after neural induction with CHIR99021 (20 μM) and forskolin (100 μM). (F) Representative fluorescence images of SMiNs 7 days after neural induction with lithium carbonate (3 mM) and forskolin (100 μM). (G) Representative fluorescence images of SMiNs 7 days after neural induction with CHIR99021 (2 μM) and dbcAMP (0.5 mM). (H) Representative images of patch-clamp in SMiNs. (I) Representative traces of sodium current of SMiN 84 days after neural induction before (left) and after (right) 0.5μM TTX bath application. (J) Representative traces of action potential spike of SMiN 84 days after neural induction before (left) and after (right) 0.5μM TTX bath application. (K) Representative traces of sodium current (left) and action potential spike (right) of undifferentiated C6 glioma in proliferation media. (L) Representative traces of sodium current (left) and action potential spike (right) of undifferentiated C6 glioma in neural differentiation media without small molecules. Data are presented as the mean ± S.E.M. Scale bars represent 50 μm (A), 20 μm (C), 100 μm (F), and 100 μm (G).</p

Growth suppression effect of a combination of CHIR99021 and forskolin in C6 glioma.

<p>(A) The proliferation rate of normal C6 glioma in neural induction media without small molecules. Cells were transferred to a new large dish at day 7. (B and C) Quantitative results of cell density at day 7 after neural induction. (D) Quantitative results of cell density at day 7 after neural induction. C, CHIR99021 (20 μM); F, forskolin (100 μM). (E) Quantitative results of cell density at days 7 and 14 days after neural induction with CHIR99021 (20 μM) and forskolin (100 μM). (F and G) Representative image and percentages of Ki67-positive cells in C6 glioma and SMiN cultures 35 days after neural induction with CHIR99021 (20 μM) and forskolin (100 μM). (H) Quantitative results of cell density at 7 days post-treatment. C, CHIR99021 (20 μM); F, forskolin (100 μM); T, temozolomide (50 μM). * indicates <i>p < 0</i>.<i>05</i>. Data are presented as the mean ± S.E.M. Scale bars represent 100 μm (F and G).</p

Change of gene expression relating to cell cycle, glioma development, cytokine-cytokine receptor interaction, ECM-receptor interaction, cell adhesion molecule, PI3K-AKT, MAPK and RAS signaling pathway after treatment of small molecules.

<p>Change of gene expression relating to cell cycle, glioma development, cytokine-cytokine receptor interaction, ECM-receptor interaction, cell adhesion molecule, PI3K-AKT, MAPK and RAS signaling pathway after treatment of small molecules.</p

Growth suppression effect of a combination of CHIR99021 and forskolin in patient-derived GBM.

<p>(A) Quantitative results of cell density 7 days after neural induction. (B) Quantitative results of cell density 7 days post-treatment. C, CHIR99021 (10 μM); F, forskolin (50 μM); TMZ, temozolomide (50 μM). (C) Representative fluorescence image of Tuj1-positive cells 7 days after neural induction with CHIR99021 and forskolin. * indicates <i>p < 0</i>.<i>05</i>. Data are presented as the mean ± S.E.M. Scale bars represent 200 and 100 μm.</p