| Object: By constructing chloasma rat model,isolating and culturing adipose-derived stem cells(ADSCs),to collect the conditioned medium from adipose-derived stem cells(ADSC-CM)which is used to treat the lesions of the rat model of chloasma,and observe its curative effect.Moreover,to explore its effect on related inflammatory factors by detecting the contents and comparing their changes of the Prostaglandin 2(PGE-2),the Stem Cell Factor(SCF)and the Nitric Oxide(NO)of the skin homogenate of these model rats,so as to provide more experimental basis for new strategies of treatment of chloasma.Methods: Adipose-derived stem cells were isolated from rat inguinal adipose tissue and the purity was analyzed by immunocytochemistry method.After the ADSCs passage to the third generation,Covering the bottom of the culture bottle was about 80-90%.Serum-free DMEM medium was replaced and the ADSC-CM was collected.Forty-two clean grade healthy female SD rats aged 8 weeks old were selected,12 of them were fed normally,and 30 of them were used to make models.The chloasma rat model was established by progesterone intramuscular injection combined with ultraviolet irradiation.The success of the chloasma model was judged by naked eye,pathological examination and determination of superoxide dismutase(SOD)and malondialdehyde(MDA)in skin homogenate.At the end of the modeling time,6 rats were randomly selected from 30 rats participating in the modeling and 6 rats were randomly selected from 12 rats normally fed for model identification.The remaining 6 normally fed rats were taken as normal group,and the remaining 24 modeling rats were randomly divided into four groups: model group,blank control group,positive control group and treatment group.The model group did not intervene after successful modeling;the blank control group used serum-free DMEM culture medium to treat;the positive control group used tranexamic acid injection;the treatment group used ADSC-CM to treat,and the treatment adopted the method of multi-point injection in the dorsal lesion area of rats.After treatment finished,the lesion area of chloasma rats was observed by naked eye,biopsy specimen was analysed pathologically and the contents of PGE-2,SCF and NO in skin homogenate were examined to evaluate the curative effect and the effect on inflammatory reaction comprehensively.Results: 1.The ADSCs were found to be spindle-shaped under the inverted phase-contrast microscope,and the cells were vortex-like when the density of the cells was higher.2.The expression of immunophenotype of the ADSCs was detected by flow cytometry,the CD29 and CD90 were positive,the CD45 and CD106 were negative.3.The identification of rat model of chloasma: The naked skin on the back of rats showed erythema,pigmentation and dryness,and the gloss of hair decreased.Skin pathology showed epidermal hyperplasia with varying thickness.Inflammatory cells infiltrated in the superficial dermis,capillaries increased and dilated obviously,and hair follicle sebaceous gland hyperplasia.Biochemical tests showed that SOD decreased and MDA increased.4.After the treatment,the naked skin of each group was observed,and no obvious skin lesions were found in the positive control group and the treatment group,but the color of the skin of the normal group was deep,the hair glossiness was low,and the difference between the two groups was small;the color of the skin of the model group was still more red,and scattered Pigmentation;the blank control group was found to be scattered on erythema and the skin of the model group was thinner,but still thick in the remaining groups.There were still large number of capillary expansion,infiltration of the inflammatory cells,and hyperplasia of the sebaceous glands of the hair follicle in the dermis.The skin of the blank control group was thin but thicker than that of the positive control group,and in the treatment group,the pathological manifestations of the positive control group and the treatment group are similar,the epidermis is obviously thinner,the infiltration of the inflammatory cells is reduced,the capillary expansion is lessened.5.The analysis of PGE-2,SCF and NO in skin homogenate shows that the model group by their own recovery after the risk factors been removed,the inflammatory factors were higher than normal group,the differences were statistically significant(P < 0.05),the inflammatory factors of the positive control group and the treatment group were both reduced significantly after the intervention respectively compared with the blank control group,the differences were statistically significant(P < 0.05).Conclusions: 1.The conditioned medium from adipose-derived stem cells is used for the model of the chloasma rat has curative significance;2.The conditioned medium from adipose-derived stem cells can down-regulate inflammatory factors PGE-2,SCF and NO of the skin lesions in the model of chloasma. |
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