Fig 2: Suppression of epithelial stem/progenitor and CSC properties of OM-1 by an intracellular p75 signal inhibitor TAT-Pep5. (A) OM-1 formed tumor spheres. (B) The average number of tumor spheres obtained from 200 cells with or without the p75 signal inhibitor TAT-Pep5. Double asterisks indicate a statistical difference (p < 0.01). (C) Immunofluorescences (green: CK13, red: CK14, blue: DAPI) of single cell-derived colonies formed under treatment with TAT-Pep5. Outer cells lacked CK14 failed to express CK13. . (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig 3: OM-1 cells maintained stratified epithelial progenitor and CSC properties. (A) The mRNA expression profiles of p75, CK13, CK14, andG3PDH in various squamous cell carcinoma cell lines are shown. A431 cells were used as the positive control for CK13 and CK14 expression [23]. (B) Immunocytochemistry using anti-p75 antibody in growing OM-1 cell cultures (green: p75, blue: DAPI). Some cells expressed p75 with the characteristic clusters. (C) Immunofluorescence of CK14 and CK13 in growing OM-1 cultures (green: CK13, red: CK14). CK14 and CK13 expression were exclusive in a given cell. (D, E) Immunofluorescence of a colony of OM-1 cells derived from a single cell. The cells with the characteristic p75 cluster were located at the center. CK14-negative cells can be found at the outer layer of the colony in D (green: p75, red: CK14, blue: DAPI). Outer cells displayed CK13 expression in E (green: CK13, red: CK14, blue: DAPI). (F) Distinct slow-growing small colonies with p75 clustering. (G) Histological analysis with HE staining of air-exposed 3D cultures of OM-1 cells on a dermis-mimic collagen gel. OM-1 cells built a multi-layered stratified epithelium. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig 4: Ketamine regulates the BDNF pathway in neurons and astrocytes. Human neurons were treated with ketamine and the expression of BDNF mRNA was determined using RT-PCR (A) and BDNF secretion by ELISA (B). Pro-BDNF secretion was analyzed using ELISA (C) and the expression of p75NTR and TrkB was analyzed using Western blot analysis (D). The effect of ketamine on BDNF (F) and pro-BDNF (G) secretion in astrocytes was analyzed using ELISA. The expression of the lncRNA BDNF-AS was analyzed using RT-PCR in both neurons (E) and astrocytes (H). The results are the means ± SD of three independent experiments analyzed in quadruplets or are a representative of three independent experiments (D). *P < 0.05, **P < 0.01, and ***P < 0.001.

Fig 5: Schematic images and immunohistochemical analysis of in vivo cell transplantation in experimental rats with nerve defects. (A) The sciatic nerve was exposed on the thigh of the experimental animal; (B) The sciatic nerve was resected and a 5-mm nerve gap developed; (C) After tubulization with biodegradable collagen bovine dura mater (Lyoplant®) in the nerve gap, a total of 1 × 106 cells (DF-chN) were transplanted into the collagen tube with a 0.1-mL fibrin glue scaffold (Greenplast®). The tube was then sealed with sutures; (D) Photograph of the experimental nerve at eight weeks after DF-chN transplantation. In the cell transplantation site, the regeneration of nerve defect was detected with continuity of the nerve gap; (E–I) Histological and immunohistochemical analysis of low-affinity nerve growth factor receptor (p75NGFR) in regenerated motor nerve fibers at eight weeks post cell transplantation. (E) In non-cell-transplanted control specimens, regenerated nerve fibers were not detected in abundance and p75NGFR expression was weak. (F) In the DF-chN transplanted group, significantly increased regenerating nerve fibers were detected in the nerve gap (arrows). The asterisk (*) indicates the proximal end of the nerve gap. (G,H) High magnification of generated nerve fibers at the DF-chN graft site. High expression of p75NGFR was detected in newly generated nerve fibers, especially in the perineurium and endoneurium of regenerating nerve fibers. (I) Toluidine blue staining of regenerating nerve fibers showed blue-colored positive staining of the myelin sheath, which was detected in abundance in the outer layer of newly generated axonal fibers (Scale bar = 100 µm).