Effects Of Melatonin On The Secretion Of Exosomes Derived From Hair Follicle Stem Cells In Inner Mongolian Cashmere Goat

The purpose of this experiment was to investigate the effects of melatonin stimulation on the morphology,particle size,protein expression and secretion of exosomes derived from hair follicle stem cells in Inner Mongolian cashmere goat,and to study the effects of exosomes to hair follicle development related receptor cells.1.The optimal concentration and duration of melatonin stimulation to the secretion of exosomes from hair follicle stem cells were measured.The melatonin concentrations of 0,100,300,500,and 700 pg/mL were set to act on hair follicle stem cells,and the proliferation was observed to extract exosomes of the cell supernatant.The results showed that melatonin at a concentration of 500 pg/mL was the optimal proliferation concentration of hair follicle stem cells,and the exosomes extracted from the cell supernatant were optimal after 3 days treated by 500 pg/mL melatonin.2.The morphology,size,protein expression and secretion quality were observed to identificate the extracted exosomes treated by 500 pg/mL melatonin.The groups including 0 pg/mL,500 pg/mL melatonin,500 pg/mL melatonin+500 pg/mL Luzindole(MTI+MT2 receptor block),500 pg/mL melatonin-5500 pg/mL 4-P-PDOT(MT2 receptor blockade),were set to stimulates hair follicle stem cells and extracts exosomes from each group.The morphology and size of exosomes were observed by transmission electron microscopy.The BCA protein was quantified and the protein expression was detected by Western blot.The results showed that the exosome morphology and size of the stimulated group and the unstimulated group were not significantly different.The exosome surface markers,HSP70,CD9 and CD81,were detected in each group.The secretion of exosomes from hair follicle stem cells was increased by melatonin stimulation.3.The effects of exosomes derived from hair follicle stem cells in each group on the hair follicle development related receptor cells(hair follicle stem cells and dermal papilla cells)were investigated.The exosomes of each group were applied to hair follicle stem cells and dermal papilla cells according to concentration gradient.The proliferation of recipient cells was detected by MTT assay,and the migration of recipient cells was detected by Transwell chamber.The RNAs of Od,ld,3d,5d,7d,14d and 21d receptor cells were extracted,and cDNA was used as a template to detect the hair follicle development related genes ?1-integrin,SOX9,FGF5,P-caintein,KAP7 and KAP8.The results showed that exosomes promoted the proliferation,migration and development of genes involved in hair follicle stem cells and dermal papilla cells,which could promote the differentiation of hair follicle stem cells into hair follicle cells.The exosome-promoting effect after melatonin stimulation was more pronounced and increased with increasing concentration gradient.