Extraction and identification of exosomes in follicular fluid from patients with polycystic ovary syndrome and isolation and detection of miRNAs in exosomes
10.3969/j.issn.1674-8115.2017.08.007
- VernacularTitle:多囊卵巢综合征患者卵泡液外泌体的提取鉴定及其miRNAs的提取和检测
- Author:
Qian YANG
;
Lanxin LIU
;
Hefeng HUANG
- Keywords:
polycystic ovary syndrome;
follicular fluid;
exosome;
biomarker;
identification;
miRNAs
- From:
Journal of Shanghai Jiaotong University(Medical Science)
2017;37(8):1085-1089
- CountryChina
- Language:Chinese
-
Abstract:
Objective · To extract and identify exosomes in follicular fluid from patients with polycystic ovary syndrome in order to determine the existence of exosomes in follicular fluid, to isolate and extract miRNAs in exosomes, and to detect relative expression of miRNAs. Methods · Exosomes in follicular fluid were collected with membrane affinity chromatography and their size and morphology were observed with the transmission electron microscope. Exosome protein markers CD63 and CD81 were detected with flow cytometry. miRNAs in purified exosomes were extracted and expressions of miR-125b, miR-19b, and miR-222 were measured with TaqMan real-time PCR. Results · Exosomes in follicular fluid were circular or elliptic under the transmission electron microscope with diameters of around 30-100 nm. They had complete membrane structure and contained low density matter. Flow cytometry showed that CD63 and CD81 were positively expressed in exosomes. Real-time PCR detected expressions of miR-125b, miR-19b, and miR-222. Conclusion · Exosomes can be collected in follicular fluid from patients with polycystic ovary syndrome. Transmission electron microscopy and flow cytometry can be used to identify exosomes in follicular fluid. miR-125b, miR-19b, and miR-222 can be detected in exosomes.
