| The central nervous system has a limited capacity for self-repair since mature neuronslack the ability to regenerate. Stem cell-based therapies have introduced new possibilities forrepair and restoration of neuronal function after spinal cord injury. It has been demonstratedthat transplantation of BMSCs is a promising therapy for spinal cord injury and some studieshave reached the stage of clinical investigations. However, BMSCs must be isolated by bonemarrow aspiration, which is traumatic and painful. Moreover, the percentage of stem cells inbone marrow is very low and it decreases with aging. These problems have limitedapplication of BMSCs to clinical treatments. For these reasons, there is a need to find otherautologous source of adult stem cells.1Isolation, cultivation and characterization of rat vibrissa dermal papilla cellsIn this study, we isolated and cultured DPCs. As a control, BMSCs were isolated fromthe same donor rat. Then we analysed and compared their characteristics. RT-PCR analysisand Immunofluorescent staining showed that DPCs strongly expressed papilla markers ofALP and Sox2, as well as NCSCs markers P75and Nestin. Immunofluorescent staining andflow cytometer analysis showed that DPCs were positive for the MSC markers. Based on thesimilar expression patterns of MSC markers with BMSCs, we speculated that DPCs may havesimilar multipotent differentiation potential with BMSCs. Under specific inducing conditions,DPCs differentiated into adipocytes, osteoblasts, smooth muscle cells and neurons. Inconclusion, our data demonstrate that DPCs from craniofacial hair follicle dermal papillahave adult stem cell properties.2Assessment of neurotrophic factors secretion and bioactivityTo investigate the potential for DPCs to secrete NTFs, we selected several NTFs,including NGF, BDNF and GDNF. We found that these factors demonstrate similar gene expression patterns in DPCs and BMSCs at the mRNA level. Moreover, we detected theexpression of NGF in DPCs and BMSCs by Western Blot analysis. In comparison withBMSCs, the higher expression level of NGF was detected in DPCs. In addition, DPC-CMstimulated neuron-like differentiation of PC12cells. Finally, using ELISA assay, we detectedthe secretion of NGF, BDNF and GDNF in DPC-CM. Unexpectedly, we found that the levelsof NGF in DPC-CM were undetectable. In contrast, the amount of BDNF and GDNF secretedper ten thousand cells in DPC-CM for24hours were significantly higher than in BMSC-CM.Thus, we considered that in DPC-CM, at least BDNF and GDNF, but not NGF, were crucialNTFs that played important roles in stimulating neuronal differentiation of PC12cells.We suspected that within the monolayer-expanded DPCs, the neurosphere-forming cellswere major producers of BDNF and GDNF. To confirm our hypothesis, DPCs were culturedin suspension at low density in serum-free sphere-forming medium. After7days,neurosphere-like spheres were generated, which were identified with double immunostainingby co-expression of the NCSCs markers P75and Nestin. RT-PCR analysis revealed that,compared with total DPCs and non-sphere-forming DPCs, sphere-forming DPCs showedhigher levels of Nestin and P75expression. At the mRNA level, we found that thesphere-forming DPCs expressed NTFs at levels similar to those of other cells. In contrast, byELISA analysis, similar amounts of BDNF and a more abundant amount of GDNF weredetected in the supernatants from monolayer cultured sphere-forming DPCs compared withtotal DPCs. These results demonstrated that sphere-forming DPCs might be a majorsubpopulation of cells for secreting GDNF.3Transplantation of rat vibrissa dermal papilla cells into the lesion site of rats subjected to spinalcord injuryFinally, we transplanted DPCs and BMSCs from the same GFP transgenic rat into thelesion site of rats underwent T8complete transection, and further compared the therapeuticeffects of both cell types. On the21stday after transplantation, we observed that thefluorescence intensity of the spinal cord transplanted with DPCs was stronger than the others.In addition, the axon sprouting was more significantly stimulated by DPCs.In summary, our data demonstrate that DPCs from craniofacial hair follicle dermalpapilla have adult stem cell properties and a greater capacity to secrete BDNF and GDNF than do BMSCs. We suggest these cells are a promising source for autologous cell therapy intreatments for spinal cord injury. |
PDF Full Text Request