Detection Of GA733 MRNA By RT-PCR In Exfoliated Cells Of Pleural And Peritoneal Effusions And Its Clinical Values

Objectives Pleural and peritoneal fluids are common clinical symptoms. Recently, the incidence of malignant pleura and peritoneal fluids is increasing with the increase of malignant tumor. The differentiation between malignant and benign effusions is critical for the diagnosis, therapy and prognosis evaluation of the patients. Currently the diagnosis of malignant effusions mainly depends on the routine cytological examination, but unfortunately, its sensitivity is low, and the cytological distinction of reactive mesothelial cells from epithelial carcinoma cells is often difficult. Thereby, looking for ideal diagnosis methods is always concerned about. GA733 genes are a family, which encodes tumor-associated antigens and are widely expressed on the surface of epithelial cells and epithelial tumors. More than 80% of human tumors are carcinomas of epithelial origin, and more than 90% of malignant pleural or peritoneal effusions are caused by epithelial carcinoma metastasis to pleura and peritoneum. Because no epithelial cells in benign pleural and peritoneal fluids, detection of new epithelial makers is valuable for establishing new methods to diagnose malignant effusions. In the present study, in order to establish a voluble diagnostic method, RT-PCR was used to detect GA733 mRNA of exfoliated cells of pleural and peritoneal effusions in pleural and peritoneal fluids, and its values of differentiation between malignant and benign effusions was evaluated. Methods Sixty specimens of fresh pleural and peritoneal fluids were collected from 60 patients. All specimens were routinely centrifuged to prepare smears which were stained by Papanicolaou method for cytological diagnosis. The clinical data of these patients were collected for clinical diagnosis. According to clinical diagnosis, these specimens were divided into malignant tumor group, benign disease group and cause-unknown group. According to cytological diagnosis, these specimens were divided into positive group, negative group and suspicious group. GA733-1 and GA733-2 mRNA in the exfoliative cells of effusions were detected by RT-PCR: extraction of total RNA, reverse transcription of cDNA and PCR. PCR products were electrophoresed on 8% polypropylene gel and stained with silver nitrate. The positive rates of GA733-1 and GA733-2 mRNA were calculated, and their sensitivities, specificities, positive predictive values, negative predictive values and diagnosis accordance rates were evaluated. Results Of 60 specimens, 36 specimens were obtained from patients with epithelial cancers, 13 from benign disease and 11 from cause-unknown disease. According to cytological examination, 26 specimens were diagnosed as malignant effusions, 28 as benign effusions, 6 specimens as suspicious. GA733-2 mRNA was detected in 27 (75.0%) of 36 samples from cancer cases and in 7 (63.3%) of 11 cause-unknown cases. All specimens from benign cases are negative for GA733-2 mRNA. The differences of GA733-2 mRNA expression among three groups of specimens were significant (P<0.005). GA733-2 mRNA was detected in 22 (84.6%) of 26 cytological positive specimens, in 6 (21.4%) of 28 cytological negative specimens, and in all cytological suspicious specimens. The 5 specimens with reactive mesothelial cells were all negative for GA733-2 mRNA. The differences of GA733-2 mRNA positive rates among three groups of specimens were significant (P<0.005). GA733-1 mRNA was detected in 5 (13.9%) of 36 samples from cancer patients, in one of 11 cause-unknown cases and none in benign cases. The differences of GA733-1 mRNA positive rates among three groups were not significant (P>0.05). GA733-1 mRNA was detected in 4 (15.4%) of 26 cytological positive specimens, in 1 of 28 cytological negative specimens and 1 of 6 cytological suspicious specimens. The differences of GA733-1 mRNA positiverates among three groups were not significant (P>0.05). Sensitivity of RT-PCR detection for GA733-2 mRNA in diagnosis of malignant effusions is 75.0%, specificity is 100%, positive predictive value is 100%, negative predictive value is 59.1%, diagnosis accordance rate is 81.6%. The good specificity and positive predictive value are its prominent features, and sensitivity, negative predictive value and diagnosis accordance rate were better than cytology. The positive rate of the expression of GA733-1 mRNA is low, and its clinical value for diagnosing malignant effusions needs further studies. Conclusions 1. GA733-2 mRNA is highly expressed in malignant in pleural and peritoneal effusions causes by epithelial carcinomas, and rarely in benign effusions. 2. The detection of GA733-2 mRNA by RT-PCR is sensitive and specific for diagnosis of malignant pleural and peritoneal effusions, and very useful for the differential diagnosis between malignant and benign effusions. 3. The detection of GA733-2 mRNA by RT-PCR has potential value on determining the nature of cytological suspicious effusions and discriminating epithelial tumor cells from reactive mesothelial cells, and more studies should be performed for its complementary value to cytological examination. 4. The expression of GA733-1 mRNA was infrequent in exfoliative cells of pleural and peritoneal effusions, and its clinical values on diagnosing malignant effusions of epithelial origin and differentiating malignant effusions with benign ones need more studies.