Study Of The Relationship Between The Hypertrophic Scarring And Melatonin

ObjectiveTo study the relationship between the hypertrophic scarring and melatonin, melatonin receptor. To investigate the effect of melatonin on proliferation and apoptosis of fibroblasts from human hypertrophic scar, as well as to explore the new approach of preventing hypertrophic scar formation.Methods1. The melatonin of the serum, the scar blisters liquid, as well as the tissues lysate supernatant which obtained from hypertrophic scar was quantified by ELISA method.2. The expression of melatonin receptorGPR50 was located with immunohistochemistry and the content of melatonin receptor(MT1,MT2) were assessed with RT-PCR method.The positive production was sequenced with auto sequencing instrument.3. Fibroblasts from hypertrophic scar were cultured and incubated with melatonin at the concentration of 10-5 mmol/L,10-3 mmol/L,1mmol/L for 24h, cell morphology was observed by microscope, and XTT-PMS assay were used to measure the proliferation of fibroblasts. Flow cytometry was used to assess the cell cycle distribution and apoptosis. The content of cell cyclinE,P53 and FasmRNA were determined with RT-PCR method.Results1. The serum melatonin concentrations of hypertrophic scar patients was significantly lower than control group during daytime (P<0.05), there were no significant difference of serum melatonin concentration in patients between the day and night (P>0.05); the melatonin concentration in scar blisters liquid was significantly lower than the serum (P<0.05); and compared with the normal skin tissue, the melatonin in hypertrophic scar was significantly decreased (P<0.05)2. Positive signals of melatonin receptor could be found in the cell membrane and cytoplasm.In normal skin,melatonin receptorGPR50 was located in the epithelial basal cells,sweat gland cells and hair follicle.In human hypertrophic scar,the expression of melatonin receptorGPR50 was mainly existed in the epidermal cells, fibroblasts, survived hair follicle and sweat glands. RT-PCR analysis identified that the expression of melatonin receptor(MT1,MT2)mRNA in hypertrophic scar was significantly stronger than normal skin(P<0.05).In normal skin and hypertrophic scar group, the expression of MT1mRNA was 0.99081±0.26485(copy number/μ1 cDNA) and 1.16584±0.21829(copy number/u1cDNA) respectively; the expression of MT2mRNA was 0.77083±0.15927(copy number/u 1 cDNA) and 0.99550±0.14624(copy number/μu 1 cDNA), respectively. Sequencing results indicated that the positive product coincided with cDNA of human melatonin receptor in GeneBank.3. Compared with the control,melatonin at the concentration of 10-5 mmol/L 10"3 mmol/L,1mmol/L could depress the proliferation of fibroblasts (P<0.05),and the nuclear pyknosis and nuclear fragmentation increased which is concentration-dependent.With the increasing of melatonin concentration, G1 phase cells and the apoptotic rate increased, but S and G2 phase cells decreased (P< 0.05),and the expression of cell cyclin E mRNA were suppressed,but the expression of p53 & FasmRNA were elevated (P<0.05)Conclusions:1. The abnormalities of melatonin and melatonin receptors may be the factors of hypertrophic scarring.2. Melatonin can inhibit the proliferation and induce the apoptosis of hypertrophic scar fibroblasts in vitro, suggesting that melatonin may be effective on preventing hypertrophic scar.