stemmacs trilineage differentiation kit  (Miltenyi Biotec)

Journal: Journal of Sport and Health Science

Article Title: Long-term aerobic exercise enhances circulating exosomal miR-214-3p to promote endothelial progenitor cell-mediated repair of endothelial damage induced by obesity

doi: 10.1016/j.jshs.2025.101094

Figure Lengend Snippet: Eight weeks of aerobic exercise improved the proliferation and migration capabilities of circulating EPC in both humans and rats with obesity through circulating exosomes. (A) Representative transmission electron microscopy image of exosomes derived from human peripheral blood. Scale bar = 200 nm. (B) Exosome characterization and identification. Exosomes derived from human peripheral blood express TSG101 and CD63. (C) Nanoparticle tracking analysis confirms the presence of exosomes with a peak diameter of 100 nm, characteristic of exosomal size. Quantitative analysis of exosomes derived from human peripheral blood revealed no statistically significant difference in the number of exosomes isolated from equal volumes of circulating blood between the control group and the exercise group ( n = 30 for each group). (D) Cell proliferation assay results showed that exosomes derived from the exercise group significantly enhanced the proliferative capacity of human EPC compared to those from the control group, as measured by the CCK-8 method ( n = 20 for each group). *** p < 0.001, Exercise vs . Control. (E) Scratch assay results showed that exosomes derived from the exercise group significantly promoted the migratory ability of human EPC compared to those from the control group ( n = 5 for each group). * p < 0.05, Exercise vs . Control. (F) Representative images of wound healing in the scratch assay, showcasing the migratory response of human EPC. (G) Characterization of circulating exosomes from rat peripheral blood. (H) Quantitative analysis of exosomes derived from rat peripheral blood revealed no statistically significant difference in the number of exosomes isolated from equal volumes of circulating blood among all groups ( n = 3 for each group). (I) Cell proliferation assays revealed that exosomes derived from the HC group exhibited a diminished capacity to promote EPC proliferation compared to those from the NC group in rats. In contrast, exosomes induced by 8 weeks of aerobic exercise significantly enhanced EPC proliferation ( n : 5–6 for each group). * p < 0.05, HC vs . NC; ## p < 0.01, HE vs . HC. (J) Scratch assays indicated that exosomes derived from the HC group exhibited a diminished capacity to enhance EPC migration rates compared to those from the NC group in rats. In contrast, exosomes induced by 8 weeks of aerobic exercise significantly enhanced EPC migration rates ( n = 4 for each group). ** p < 0.01, HC vs . NC; ## p < 0.01, HE vs . HC. (K) Representative images of wound healing in the scratch assay, showcasing the migratory response of rat EPC. CCK-8 = cell counting kit-8; CD63 = cluster of differentiation 63; EPC = endothelial progenitor cells; HC = the high-fat diet with sedentary group; HE = the high-fat diet with exercise group; NC = the normal diet with sedentary group; TSG101 = tumor susceptibility gene 101.

Article Snippet: The primary antibodies used included PI3K (SC-365290, 1:1000; Santa Cruz Biotechnology, Dallas, TX, USA), Akt1 (SC-5298, 1:1000; Santa Cruz), p-Akt (Ser473) (66444-1-lg, 1:1000; Proteintech Group, Rosemont, IL, USA), phosphatase and tensin homolog (PTEN) (60300-1-Ig, 1:1000; Proteintech), tumor susceptibility gene 101 (TSG101) (DF8427, 1:1000; Affinity Biosciences, Cincinnati, OH, USA), cluster of differentiation 63 (CD63) (AF5117, 1:1000; Affinity), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (GB15002-100, 1:4000; Servicebio).

Techniques: Migration, Transmission Assay, Electron Microscopy, Derivative Assay, Isolation, Control, Proliferation Assay, CCK-8 Assay, Wound Healing Assay, Cell Counting