| Background Pleural effusion is one of the most common clinical manifestations of pleural diseases,according to the different factors and the prognosis of patients with clinical effect,pleural effusion is often divided into two categories,namely benign pleural effusion(BPE)and malignant pleural effusion(MPE).The etiology and prognosis of two types of pleural effusion there is a big difference,so it is very important for accurate identification of the two.But for a long time,the differential diagnosis of benign and malignant pleural effusion is always a big problem for clinicians.Although the tumor cell found in the pleural effusion is the gold standard for the diagnosis of pleural effusion,the positive rates are reported to vary greatly between 40%-87%.In addition,for the protection of the pleural effusion hydrothorax cytology negative,some of theindicators,such as lactate dehydrogenase(LDH),adenosine dehydrogenase(ADA),carcinoembryonic antigen(CEA),have certain differential value to some extent,but the specificity and sensitivity of these markers is not very ideal.So to explore a kind of high sensitive and specific identification of benign and malignant pleural effusion auxiliary detection method,provide the basis for clinical diagnosis is very necessary.In recent years,the detection of circulating tumor cells(CTCs)has gradually entered the era of accurate medicine.At present,many researchers have made great contributions to explore the diagnostic value of CTC,prognosis,value,efficacy monitoring and prediction of tumor recurrence.As "rare cells" in the blood,the detection of CTC is technically demanding.The Cellsearch system has long been used as the “gold standard for CTC testing".However,in the process of in-depth research and clinical use,the limitations of the Cellsearch system are gradually emerging,mainly because of the inability to detect epithelial mesenchymal transition(EMT),resulting in false negative results.With the continuous innovation of technology,immunomagnetic sorting has quietly entered the scientists' field of vision.This cell enrichment method is different from the Cellsearch system and does not depend on the expression of epithelial markers and can be enriched into EMT tumor cells.Immunomagnetic beads sorting is divided into positive and negative sorting.Because the positive sorting method is affected by the heterogeneity of tumor cells,it may cause the activation of intracellular signaling pathways in tumor cells.While the negative sorting method via removing samples of red and white blood cells and other components,the enrichment to CTC in the natural state,and does not depend on the expression of surface molecules in tumor cells,thus the negative selection method is applicable to any type of tumor CTC enrichment,applied more widely.By means of methodological improvement,the recovery rate can reach 100%at present.After enrichment,the cell components can be removed by immunofluorescence staining,and further identification by fluorescence in situ hybridization can be used for subsequent functional analysis.The methods of CTC identification are mainly based on the expression of tumor cell surface antigen,cell specific m RNA expression and chromosomal abnormalities.Fluorescence in situ hybridization(FISH)has been confirmed for prostate cancer,colorectal cancer,breast cancer,lung cancer,bladder cancer cell.FISH expression and identification of epithelial tumor does not depend on the tumor cell surface molecules marker,which can detect epithelial cell specific markers of weakening or disappearance of CTC,at the same time can be directly observed the CTC of malignant tumor cells under the microscope.Objective Our study unite immunofluorescence staining,magnetic beads negative enrichment method and FISH technology--SOP?im FISH technique[2],to explore the value of chest hydrocele desquamate cell(pleural effusion “CTC”)for the diagnosis of malignant pleural effusion.Patients and Methods(a)Participants: pleural effusions in the Guangzhou first people's hospital from March2015 to January 2017,and met the inclusion and exclusion criteria.(b)Methods: using Macs sorting enrichment of pleural effusion and peripheral blood CTC,CD45 antigen identified by the enrichment of CTCs.cells are also collected pleural effusion cytology in traditional were analyzed by FISH hybridization signal and immunofluorescence staining.(c)experimental design Sample group: divided into two groups,namely malignant pleural effusion group(experimental group)and benign pleural effusion group(control group).1.1 Cases of malignant pleural effusion1.1.1 Acception criteria(1)Any age ? 18 years,gender(except during pregnancy,lactation women);(2)by histopathology or cytological diagnosis of malignant tumor;(3)physical examination and(or)imaging examination confirmed that the pleural effusion;(4)can tolerate pleural puncture pumping liquid;(5)volunteered for the researchers.1.1.2 Exclusion criteria(1)Pregnancy women and lactation women;(2)the histopathology and cytology has not confirmed as malignant tumor;(3)Before and after operation,radiotherapy and chemotherapy within 3 months;(4)with severe diseases such as cardiovascular,liver and kidney should not accept thoracentesis;(5)Poor adherence or refused to participate in the study.1.2 Cases of benign pleural effusion1.2.1 Acception criteria(1)Any age ? 18 years,gender(except during pregnancy,lactation women);(2)show the exudate of malignant pleural effusion,including tuberculous pleurisy,pneumonia and other diseases caused by pleural effusion;(3)no history of malignant tumor;(4)volunteered for this study,and can actively cooperate with patients.1.1.2 Exclusion criteria(1)pregnancy women and lactation women;(2)because of heart failure,hypoalbuminemia,nephrotic syndrome caused by other diseases such as pleural effusion show the leakage liquid;(3)pleural effusion to wrap or due to other reasons such as less pleural puncture drainage performer.(4)ever suffered from cancer.2.Specimen collection The experimental group was selected from Guangzhou No.1 People's Hospital in March 2015 to January 2017,and 46 cases of malignant pleural effusion were confirmed by histopathology or cytology.The primary causes include 18 cases of lung adenocarcinoma,4 cases of breast cancer,3 cases of squamous cell carcinoma,3cases of small cell lung cancer,2 cases of non Hodgkin's lymphoma,1 cases of malignant pleural mesothelioma,1 cases of nasopharyngeal carcinoma,7 cases of malignant tumors of the digestive tract(1 cases,2 cases of esophageal carcinoma,1cases of gastric cancer,pancreatic cancer 1 cases of sigmoid colon,1 cases of liver cancer,1 cases of rectal cancer),7 cases of pelvic malignant tumors(1 cases,1 cases of endometrial cancer,cervical cancer,1 cases of renal carcinoma,1 cases,1 cases of bladder carcinoma,1 cases of prostate cancer,pelvic cystadenocarcinoma in 1 cases of ovarian cancer).The control group selected the same period of hospitalized patients,pleural effusion showed pulmonary effusion benign diseases,27 cases of hospitalized patients,including tuberculous pleural effusion in 12 cases,pneumonia,pleural effusion in 15 cases.Blood and pleural effusion were collected in the two groups.3.Statistical methods The experimental data were analyzed by SPSS13.0 software packet,two-sided test,inspection standard for 0.05.Measurement data using the mean and standard deviation(X ± S);Classification variables: four tables using Pearson 2or 2 consecutive test correction formula.Diagnostic test: diagnosis accuracy test,by drawing the receiver operating characteristic curve(ROC),calculate the sensitivity,specificity,positive predictive value,negative predictive value.Sensitivity,specificity,positive predictive value,and negative predictive value are calculated.Sensitivity = true positive number /(true positive number,false negative number)*100%;Specificity = true negative number /(true negative number,false positive number)*100%;Positive predictive value = true positive number /(true positive number,false positive number)*100%;Negative predictive value = true negative number /(true negative number,false negative number)*100%.Results1.Comparison of general data This study included 73 cases of patients,aged 29~91years old,average 66.9 ± 14.6 years.Among them,46 cases in the experimental group,28 cases were male,18 were female,aged 41~91 years old,average 68.17 ±13.22 years.The control group were 27 cases,21 cases were male,6 female,age29~90,average 64.19.At the age of 16.71.The experimental group and the control group in gender,age,clinical manifestation,no difference in pleural fluid properties and imaging examinations(P >0.05),the data of the two groups were comparable.2.The experimental group included 46 patients with pleural effusion specimens,which CTC was positive in 34 cases,12 cases were negative;traditional pleural effusion cytology positive in 21 cases,25 cases were negative;peripheral blood CTC positive20 cases,26 cases were negative.The control group included 27 cases of pleural effusion specimens,including "CTC" and 3 false positive cases,24 cases were negative;traditional pleural effusion cytology 1 false positive cases,26 cases were negative;peripheral blood CTC 2 false positive cases,25 cases were negative.3.The experimental group and the control group pleural water "CTC" comparison: the experimental group chest water "CTC" positive rate(73.9%,34/46),compared with the control group(11.1%,3/27),the positive rate was significantly higher,2=26.846,P=0.000<0.5,the difference was statistically significant.4.Compared with CTC of peripheral blood,the positive rate of pleural effusion "CTC" in experimental group was higher than that of peripheral blood CTC(43.5%,20/46),and the difference was statistically significant(2=8.788,P=0.003).5.Compared with the experimental group "CTC" pleural effusion and pleural fluid cytology traditional positive rate: the positive rate of pleural effusion in "CTC" is 73.9%,the positive rate of pleural fluid cytology(21/46),the traditional 45.7%compared to 2=7.64,P=0.006,the difference was statistically significant.6.The comparison among pleural effusion CTC,peripheral blood CTC,and pleural effusion cytology: the positive rate of pleural effusion of "CTC" and the highest sensitivity and specificity of pleural effusion cytology traditional minimum;the highest specificity,sensitivity and positive rate of pleural effusion after "CTC";the positive rate of CTC in peripheral blood the sensitivity was inferior to pleural effusion,"CTC",but the specificity is higher than that of hydrothorax in "CTC" 7.The source of pleural malignant tumor and other parts of the source of malignant pleural effusion,pleural effusion compared the positive rate of "CTC" : the positive rate of pleural effusion from malignant tumor group "CTC" was 58.7%(27/46),from other parts of the malignant tumor group of hydrothorax CTC positive rate was 15.2%(7/46),the comparison between the two 2=14.988,P=0.000,the difference was statistically significant.Conclusion The SOP-im FISH technology can detect pleural effusion exfoliated cells,and compared with the traditional pleural effusion cytology examination and detection of CTC in peripheral blood and pleural effusion CTC detection positive rate was the highest.In general,the "CTC" pleural effusion diagnosis of malignant pleural effusion of the highest efficiency,the positive rate of diagnosis was 73.9%,sensitivity 73.9%,the specificity was 88.9%.and compared with the detection of CTC in peripheral blood,has higher diagnostic value of malignant tumor of chest water "CTC" test. |
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