[A proliminary study on the regulatory function of human plasma exosomes-like vesicles].
OBJECTIVE
To identify the exosomes-like vesicles from the plasma and study their biologic characteristics and regulatory effect.


METHODS
The exosomes-like vesicles were purified from healthy donors plasma with a series of high-speed centrifugations and ultrafiltration. Morphology was identified by transmission electron microscopy and biologic characteristics by Western blot and flow cytometry. CD4(+)T cells and CD4(+)CD25(+)CD127low Treg cells were purified from peripheral blood mononuclear cells (PBMCs) by Magnetic cell sorting. After exosomes-like vesicles cultured with CD4(+)T cells or CD4(+)CD25(+)CD127low Treg cells, cell proliferation and apoptosis were assayed. Phosphorylated β-catenin level in Wnt signaling by phosflow.


RESULTS
Exosomes-like vesicles from plasma were similar to previously described exosomes in shapes and size and expressed exosome marker proteins CD63 and CD81 as well as the MHC-II molecule, costimulatory molecules CD86 etc. After co-cultured with CD4(+) T cells, exosomes-like vesicles inhibited the proliferation of the T cells in a dose-dependent manner. After Treg cells cultured with exosomes-like vesicles for 14 days, the survival rate of the Treg cells was 57.07%, while that of the control Treg was 30.91%. Frizzled receptors 2, 3, 4and LRP6 gene mRNA expressed (the relative gray value was 48.50, 34.84, 23.85, 49.73) in the Treg cells by RT-PCR, and Wnt molecular expressed in exosomes-like vesicles. After Treg cells co-cultured with exosomes-like vesicles, the MFI of phosphorylated β-catenin decreased (from 20.06 ± 2.99 to 12.41 ± 2.08), and the expression of Bcl-2 mRNA was upregulated significantly (the relative gray value from 0.45 to 84.97).


CONCLUSIONS
Exosomes-like vesicles existed in human plasma and express immune regulatory molecules. They can suppress the proliferation of activated CD4(+) T cells induce their apoptosis and pro-long the survival of natural Treg cells via Wnt signaling pathway.
Department of Clinical laboratory medicine, Huashan Hospital of Fudan University, Shanghai 200032, China.
Yue-E
YE
Ma
Ya-Na
YN
Ren
Jie
J
Yang
Dan
D
Li
Chen-Zhi
CZ
Guo
Juan
J
Guo
Bing
B
Ni
Hua-Hua
HH
Fan
D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D002478:Cells, Cultured; D055354:Exosomes; D005434:Flow Cytometry; D006801:Humans; D007155:Immunologic Factors; D053645:Interleukin-2 Receptor alpha Subunit; D007963:Leukocytes, Mononuclear / Q000276:immunology; D050378:T-Lymphocytes, Regulatory / Q000276:immunology
D007155:Immunologic Factors; D053645:Interleukin-2 Receptor alpha Subunit