The Study Of Exosome Detection In The Clinical Diagnosis Of Epithelial Ovarian Malignancy

BackgroundOvarian cancer has the highest mortality rate among gynecological tumors in China,which has the characteristics of low 5-year survival rate,low early diagnosis rate and easy recurrence,which seriously threatens the health of Chinese women.The diagnosis and treatment of ovarian cancer has always been a difficult clinical problem,because the early symptoms of ovarian cancer are not obvious,the rate of admission to the doctor is low.Because of the lack of effective early diagnosis means,2/3 of the patients at the first diagnosis has been advanced,losing the opportunity of surgery.Currently,the independent and combined application of serum markers such as human epididymal protein(HE4)and carbohydrate antigen(CA125,CA199,CA153)plays an important role in the auxiliary diagnosis,treatment monitoring and prognosis evaluation of ovarian cancer.However,due to the unknown etiology of ovarian cancer,markers for more reliable and effective early diagnosis and prognosis evaluation are currently lacking.Therefore,in the era of precision medicine,it is crucial to explore new markers for early diagnosis of ovarian cancer to improve patient survival.Exosomes are small vesicles with an extracellular diameter of 30-150 nm released from intracellular polyvesicles.As an important carrier of intercellular information and genetic material,exosomes contain DNA,RNA,proteins,lipids and other active substances,which are widely distributed in blood,urine and other body fluids.It has been found that most cells can produce exosomes,especially tumor-derived exosomes,which play an important role in tumorigenesis and development and are potential tumor markers.Epithelial cell adhesion molecules(EpCAM)belong to the family of adhesion molecules,is shifted from tumor related calcium signal 1 gene encoding a single transmembrane protein involved in regulating cell adhesion,mediated signal transduction,cell migration,proliferation and differentiation,and other functions.EpCAM,as a candidate protein for tumor diagnosis and treatment,is attracting more and more attention.V type 2 Collagen chain(COL5A2)is a kind of low abundance of Collagen fiber alpha chains.COL5A2 is the main component of interstitial extracellular matrix and plays an active role in many biological processes such as cell morphology,proliferation,migration,differentiation,apoptosis and carcinogenesis.Therefore,tumor-derived plasma exosomal EpCAM and COL5A2 proteins are expected to be new biomarkers for non-invasive screening of tumors and prognostic assessment.PurposeTo compare the expression levels of total plasma exosomal protein,plasma exosomal COL5A2 and EpCAM protein in patients with primary epithelial ovarian cancer,benign ovarian disease group and healthy control group to explore the application value of exosomal COL5A2 and EpCAM protein in early diagnosis and prognosis evaluation.MethodsInto the object of study of 57 cases,of which 24 cases of ovarian cancer before surgery,9 cases after the operation,14 cases of ovarian benign disease,8 cases of healthy controls and 2 cases of borderline ovarian cancer disease.This research adopts the aggregate precipitate and ultracentrifugation combined method to extract the plasma exosomes,and the morphology,particle size and surface markers were characterized by transmission electron microscopy(TEM),nano particle and ZETA potentiometer and Western blot.The expression of COL5A2 and EpCAM proteins in exosomes was detected by Western blot.Results1.Under transmission electron microscope,the exosomes were observed to be round saucer-shaped,with visible membrane structure and diameter between 30-150 nm,the exosome particle size was detected by nano particle size and ZETA potentiometer,and the peak value was about 100 nm.Western blot was used to detect the expressions of TSG101 and LAMP-1 in plasma exosomes extracted in this study.2.There were significant differences in plasma exosomal total protein concentrations between the ovarian cancer group,the benign disease group and the normal group(P<0.05).Plasma exosome total protein concentration was significantly higher in the preoperative group than in the postoperative group(P<0.05).There was a negative correlation between plasma exosome protein concentration and its staging in patients with ovarian cancer(r=-0.519,P<0.01**).3.Plasma exosomal EpCAM protein expression was significantly higher in the ovarian cancer group than in the benign disease group and the normal group(P<0.05).4.The expression of plasma exosome COL5A2 protein in the ovarian cancer group was significantly higher than that in the benign disease group and the normal group(P<0.05).The expression of plasma exosome COL5A2 protein in the preoperative group was significantly higher than that in the postoperative group(P<0.05).Ovarian cancer on early stage(?,?)and late stage(?,?)had significant difference(P < 0.001).Conclusion1.Plasma exosome protein concentration and EpCAM and COL5A2 protein expression were significantly increased in patients with epithelial ovarian malignancy.2.Plasma exosome protein concentration and COL5A2 protein expression level in patients with epithelial ovarian malignancy were correlated with the stage of epithelial ovarian malignancy.3.Plasma exosome protein concentration and COL5A2 protein expression level were significantly decreased in patients with epithelial ovarian malignancy after surgery.