Experimental Study On The Treatment Of Melasma Rat Modal By Adipose-derived Stem Cell-conditioned Medium By Antioxidant Mechanism

Objective: To explore the effect and antioxidant mechanism of adipose-derived stem cell-conditioned medium on melasma and to provide experimental basis for further study on adipose-derived stem cell-conditioned medium treatment of melasma.Melasma rat models were established,rat adipose-derived stem cells to obtain the conditioned medium and act on melasma rats,observe their apparent indexes and pathological changes,detecte the contents of skin tyrosinase(TYR),superoxide dismutase(SOD)and malondialdehyde(MDA).Methods : 1.Rat adipose tissue was taken to obtaine adipose-derived stem cells through collagenase digestion for culture and identification.2.Culture the third to fifth generation cells to cover the bottom area of about 80-90%,replacing the serum-free DMEM culture,cultivate after 24 ~ 72 h,clear fluid collection.3.Melasma rat models were prepared by systemic progesterone attack + ultraviolet radiation method.4.Identification of melasma rat models were established by observing the apparent indexes,pathological changes,and detecting the skin TYR,MDA and SOD related biochemical indicators.5.Melasma rat models were randomly divided into a blank control group,a treatment group and a positive control group,and the serum-free DMEM medium,adipose-derived stem cell-conditioned medium and tranexamic acid were injected into the skin lesions respectively.6.By observing the apparent indexes and pathological changes of skin lesions in melasma rats,the contents of TYR,SOD and MDA in the skin of rats in each group by ELISA and the results were statistically analyzed according to the groups.Results:1.Under inverted phase difference microscope,the adipose-derived stem cells cultured in the primary started to stick to the wall after about 6h.were round,polygonal,fusiform,fibrous,and whirlpool,after passage were relatively uniform,showing long fusiform and whirlpool growth.These cells were passed on to the third generation for further experiments.2.Immunophenotypic results of the third generation adipose-derived cells showed that CD29 and CD90 were positively expressed and CD45 and CD106 were negatively expressed.3.The identification of melasma rat models showed that the skin of the rats presented brown patches of uniform size,and pathology suggested epidermal hyperplasia with different thickness.Focal inflammatory cell infiltration,local collagen degeneration,hair follicle and sebaceous gland hyperplasia were observed in the dermis.Biochemical tests showed that SOD decreased,MDA and TYR increased,and oxidation and antioxidant imbalance.4.After 4 weeks of intervention,the skin of melasma rats in the blank control group still showed uniform brown patches,while the skin patches in the treatment group and the positive control group were lighter,smaller,scattered and uneven than before treatment.5.Pathological findings: in the normal group,the epidermis was relatively thin,with 2-3 layers,the cuticle and basal layer visible,and the dermal collagen fiber bundles stained light red,with a small number of inflammatory cells and blood vessels visible.After treatment,there was no significant difference between the pathological manifestations of rats in the blank control group and melasma rat model.The epidermal hyperplasia and thickness of rats in the treatment group and the positive control group were less than those in the melasma rat model,the hyperplasia of hair follicles and sebaceous glands were less than those in the melasma rat model and the inflammatory cells in the dermis were reduced.6.After treatment,compared with the melasma rat model,SOD,MDA and TYR in the skin of the blank control group showed no statistically significant difference,while there was a statistical difference in SOD increase,MDA and TYR decrease in the skin of the treatment group and the positive control group showed a statistically significant difference(P<0.05).Conclusion: 1.The adipose-derived stem cell-conditioned medium has a therapeutic effect on melasma rat model.2.The therapeutic effect of fat stem cell-conditioned on melasma rat model may be that it can inhibit oxidative stress injury and further inhibit TYR activity by reducing MDA generation and promoting SOD expression.It has been preliminarily confirmed that fat stem cell-conditioned has the effect of fighting oxidative stress injury and thus treating melasma.