Improvement Of Skin Ageing By Overexpression Of Gstm2 In Engineered Exosomes

Objective:The dermal tissue and its main component cells（dermal fibroblasts）in the skin gradually decline in function with age.The role of extracellular vesicles（EVs）has attracted considerable attention in age-related diseases.By using engineering techniques to modify vesicle surface proteins and contents,engineered exosomes targetedly transport specific inclusions to specific tissues.The ability of engineered exosomes to perform bioregulatory functions more efficiently than naturally secreted exosomes has attracted significant interest from researchers.Glutathione S-transferase mu 2（Gstm2）is a cytoplasmic enzyme that catalyzes the coupling of GSH and reduces the damage caused by ROS.Therefore,we constructed engineered EVs overexpressing Gstm2(EVs^Gstm2).The aim of this study was to construct engineered EVs^Gstm2 to research its efficiency in regulating skin aging and to provide new ideas for clinical skin anti-aging.Methods:（1）Engineered extracellular vesicles(EVs^Gstm2)were constructed by transient transfection,extracted by ultracentrifugation and assayed for transfection efficiency.（2）A fibroblast senescence model was constructed by passaged senescence and verified byβ-Galactosidase（β-Gal）staining,western blot,q RT-PCR,Ed U staining and scratch assay to see if the ageing model was successfully constructed.（3）EVs^Gstm2 was used to treat with senescent FBs,and the anti-aging effect of EVs^Gstm2was evaluated in terms of cell proliferation,migration and changes in senescence-related indicators respectively.（4）48-weeks-old ICR mice were selected as the animal model of senescence,and EVs^Gstm2 was injected into the model mice by needle-free injection to assess the anti-aging effect of EVs^Gstm2 in vivo in terms of collagen levels and changes in senescence-related indexes respectively.Results:（1）EVs^Gstm2 was successfully constructed,and the amount of Gstm2 in EVs^Gstm2was higher in the overexpression group than in the empty vector group.（2）Primary FBs was passed to the 10th generation（Old group）,and compared with the 3rd generation（Young group）,the ability of proliferation and migration in senescent cells in the old group was significantly weakened,and the expression of senescence-related indexes was significantly increased,and the cellular senescence model was successfully constructed.（3）EVs^Gstm2 enhances proliferation and migration of senescent FBs and reduces senescence-associated phenotypes thus exerting anti-aging effects at the cellular level.（4）EVs^Gstm2 was injected into mouse skin by needle-free injection and exerted anti-aging effects in vivo by increasing dermal thickness,improving aging-related indicators,and increasing collagen expression levels.Conclusion:EVs^Gstm2 improves skin aging by reducing oxidative stress-related damage induced by aging,decreasing the expression of aging-related indicators,and enhancing the proliferation and migration ability of fibroblasts.