[Isolation of rabbit aqueous humor-derived exosomes and their immunosuppression function]
Zhi-xiong Liao1, Zheng-qing Lou, Zhong Xiao
1Hangzhou Traditional Chinese Medicine Hospital, Hangzhou 310007, China. liaozx611@163.com
Objective:
To isolate exosomes from rabbit aqueous humor and to investigate their immunosuppression function.
Methods:
Aqueous humor was collected from 40 New Zealand rabbits and exosomes were isolated by fractional separation and ultracentrifugation methods; the morphology was studied with electron microscopy. The immunosuppressive-related proteins of exosomes were detected with Western blotting; their inhibitory effect on ConA-induced proliferation of T lymphocyte was estimation with CCK-8 cells proliferation assay.
Results:
Eight milliliters of aqueous humor were collected from 40 New Zealand rabbits and 200 μg exosomes was yielded. Under electron microscope, the exosomes had typical structure of lipid bi-layer with a diameter of 50-100 nm. The results of Western blotting showed that these exosomes expressed Hsp70, CD9 and Alix but not Grp94, presenting a typical exosomes protein profile. Moreover, exosomes expressed high level of TGF-β and significantly inhibited the proliferation of T lymphocytes.
Conclusion:
Immunosuppressive exosomes can be isolated from rabbit aqueous humor, which may be involved in immunotolerance of the eye.
