Abstract:

The bioactivity of the exosomes from Bletilla striata (B-exo) on human immortalized keratinocytes (Hacat) in vitro was investigated to provide data for the development of skincare products. The B-exo was extracted by polyethylene glycol precipitation and identified by transmission electron microscopy. The protein concentration was measured by the BCA Protein Assay Kit. Then, the survival rate of Hacat cells, the apoptosis rate and the apoptosis-related pathway proteins were detected by the Cell Counting Kit-8 (CCK8) method, flow cytometry and western blotting, respectively. The B-exo, a double-layer membrane with a teacup-like shape, had an average particle size of 69.63 nm. Each gram of the fresh Bletilla striata contained 5.24E+8 Particles of the B-exo and 19.53 μg protein. The B-exo significantly increased the survival rate of Hacat cells in all treatment groups, compared to the control (P<0.05). The ratios of early, late and total apoptosis induced by H₂O₂ oxidation were all significantly decreased in Hacat cells after being treated with 10 μg/mL B-exo (P<0.01). In addition, the expressions of Caspase-9 and Caspase-3 protein were significantly increased in the control group, while a decrease was detected in the B-exo treatment. However, the level of BCL-2 protein expression had no difference among the groups (P<0.05). The B-exo isolated by PEG precipitation can promote Hacat cell proliferation and inhibit H₂O₂-induced apoptosis that is modulated by the changed Caspase-9 and Caspase-3 protein expression in vitro.

Key words: Bletilla striata, exosome, human immortalized keratinocyte (Hacat), apoptosis, pathway proteins