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Corneal grafts are the imperative clinical treatment for canine corneal blindness. To serve the growing demand, this study aimed to generate tissue-engineered canine cornea in part of the corneal epithelium and underlying stroma based on canine limbal epithelial stem cells (cLESCs) seeded silk fibroin/gelatin (SF/G) film and canine corneal stromal stem cells (cCSSCs) seeded SF/G scaffold, respectively. Both cell types were successfully isolated by collagenase I. SF/G corneal films and stromal scaffolds served as the prospective substrates for cLESCs and cCSSCs by promoting cell adhesion, cell viability, and cell proliferation. The results revealed the upregulation of tumor protein P63 (P63) and ATP-binding cassette super-family G member 2 (Abcg2) of cLESCs as well as Keratocan (Kera), Lumican (Lum), aldehyde dehydrogenase 3 family member A1 (Aldh3a1) and Aquaporin 1 (Aqp1) of differentiated keratocytes. Moreover, immunohistochemistry illustrated the positive staining of tumor protein P63 (P63), aldehyde dehydrogenase 3 family member A1 (Aldh3a1), lumican (Lum) and collagen I (Col-I), which are considerable for native cornea. This study manifested a feasible platform to construct tissue-engineered canine cornea for functional grafts and positively contributed to the body of knowledge related to canine corneal stem cells., Competing Interests: The authors have declared that no competing interests exist.

Objective: An investigation on neuronal differentiation capacity of human dental pulp stem cells (DPSCs) was still lacking. In this study, two different neuronal induction protocols were investigated and compared., Methods: The neuronal differentiation was induced using chemical or growth factor induction protocol. The differentiation was confirmed by the neurogenic mRNA and protein expression using polymerase chain reaction and immunocytochemistry, respectively., Results: Chemical-induced neuronal differentiation protocol promoted morphological change and β3-TUBULIN protein expression. Though, SOX2, SOX9, and β3-TUBULIN mRNA levels were not different compared with the control, indicating a defective differentiation. For growth factor induction protocol, the cells were exhibited neurite-like cellular process and positively stained with β3-TUBULIN. In addition, the increase in intracellular calcium was noted upon NMDA stimulation, implying the neuronal function. A dramatic increased mRNA expression of neurogenic markers [SOX2, SOX9, β3-TUBULIN, and gamma-aminobutyric acid (GABA receptors)] was noted as compared to the control. In addition, a remarkable increased expression of Notch signaling target gene, HEY1, was observed in growth factor-induced DPSCs derived neuronal-like cells compared with the control., Conclusion: These data indicate that growth factor induction method is a preferable protocol for neuronal differentiation by DPSCs., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)