| Objective The aim at this study is to identify the influence of age factors of the biological characteristics of human adipose-derived stem cells(ADSCs).Compare the differences in morphology,aging characteristics,differentiation potential,surface marker expression,and proliferation ability of human ADSCs with different ages;and the effect of human ADSCs of different ages on proliferation and migration of fibroblasts.Methods Adipose tissues from 27 healthy patients,aged 0-60 years,were collected from Burns and Plastic Surgery of the 940 th Hospital of Joint Logistics Support Force from January 2020 to December 2020.According to the age of the donor,they were divided into 6groups.Group A: 0-9 years old(4 cases),Group B: 10-19 years old(4 cases),Group C: 20-29 years old(5 cases),Group D: 30-39 years old(5 cases),E group: 40-49 years old(5 cases),F group: 50-59 years old(4 cases).To obtain and culture the ADSCs of each group,subculture to obtain the third generation ADSCs,phase contrast microscope was employed to detect their morphological characteristics,the proliferation ability of each group of ADSCs was detected by CCK-8 method,the differentiation potential of each group of ADSCs was detected by in vitro adipogenesis and osteogenic induction.Flow cytometry was used to detect the surface marker expression levels of each group of ADSCs,and q PCR was used to detect the expression levels of HGF,PPAR-?,type ? collagen and Db X2 in each group.Collected the skin tissues of a healthy patient from the Burns and Plastic Surgery of the 940 th Hospital of Joint Logistics Support Force in 2020,obtained and cultured fibroblasts,subcultured to obtain the second generation of fibroblasts,and prepared each group of ADSCs conditioned medium(ADSCs-CM),the proliferation ability of fibroblasts cultured in each group of ADSCs-CM was detected by CCK-8 method,and the migration ability of fibroblasts cultured in each group of ADSCs-CM was detected by cell scratch method.Using SPSS 22.0 software for data analysis,P<0.05 is considered statistically significant.Results The cells of all age groups were in a typical spindle shape.The cells of the younger age group showed small nuclei and the cell morphology were uniform and similar;the cells of the older age group had larger nuclei and uneven morphology.With aging,the proliferation ability of ADSCs gradually decreases.In vitro adipogenic differentiation induction results: with the increase in age,the ability of ADSCs to differentiate into adipocytes gradually decreases.In vitro osteogenic differentiation induction results: with the increase in age,the ability of ADSCs to differentiate into Osteogenesis gradually decreases.The expression rates of positive surface markers CD29,CD73,CD90,and CD105 of ADSCs in each group of ADSCs were all above 93%,and the expression rates of negative surface markers CD34 and CD45 were all below 2%.With aging,the expression levels of PPAR?,HGF,and type ? collagen decreased,and the expression levels of Db X2 showed an upward trend with age.With aging,the number of ?-galactosidase staining positive cells increased,while the difference between group A,B and C was not statistically significant,and the difference between group A and D,E,and F was statistically significant(P< 0.05).The ADSCs conditioned medium of each group can promote the proliferation of fibroblasts,but the difference between the groups is not statistically significant.With aging,the ability of ADSCs conditioned medium to promote the migration of fibroblasts gradually decreases.Conclusion The proliferation ability and multidirectional differentiation ability of human ADSCs and the ability to secrete a variety of cytokines that promote the proliferation and migration of fibroblasts show a downward trend,but the adipogenic differentiation ability of ADSCs declines significantly after the age of 40.ADSCs in all age groups could promote the proliferation and migration of fibroblasts. |
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