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Detection of Micrometastases in Peripheral Blood of Patients with Gastrointestinal Cancer

Author: FangWeiJia
Tutor: LiuJian
School: Zhejiang University
Course: Oncology
Keywords: Negative immunomagnetic separation Nested reverse transcription-polymerase chain reaction Gastrointestinal cancer Peripheral blood Cytokeratin Immunocytochemistry Micrometastataes
CLC: R735
Type: Master's thesis
Year: 2005
Downloads: 46
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Abstract


Background: Gastrointestinal cancer is common malignant tumor. Metastasis especially the metastasis via blood stream is the most characteristic feature in malignant tumor. The patient, whose metastasis lesion can be found by clinical examination lesion, always is in late stage. Most of patients with gastrointestinal cancer were died from metastasis. As most metastasis is via blood stream, that detection of the micrometastases in peripheral blood of patients with gastrointestinal cancer is significance to surveillance and diagnosis of the metastasis early.Objectives: To evaluate the sensitivity and specificity and clinical significance of the methods of detection of micrometastases in peripheral blood of patients with gastrointestinal cancer, which examining the expression of CK19 mRNA , CK20 mRNA and CK19 mRNA or CK20 mRNA using nested reverse transcription-polymerase chain reaction （RT-PCR） with or without negative immunomagnetic separation （IMS）, and examining the expression of BerEP4 using immnocytochemistry（ICC） technique with or without negative IMS. Look for sensitive and liable methods to detect micrometastases in peripheral blood of patients with gastrointestinal cancer.Material and Methods: Thirty two patients with gastrointestinal cancer without detectable metastases by clinical examination before operation were enrolled for the detection of CK19 mRNA、 CK20 mRNA, and 13 patient with benign gastrointestinal disease and 15 health volunteer were as control. Another 12 patients with gastrointestinal cancer without detectable metastases by clinical examination were enrolled for the detection of expression of BerEP4. Samples of blood were obtained from the patients before 1 to 24 before operation. The expression of CK19 mRNA and CK20 mRNA and CK19 mRNA or/and CK20 mRNA were detected by nested RT-PCR without negative IMS and with negative IMS. And the expression of BerEP4 was detected by without negative IMS and with negative IMS. The positive rate and the correlation with clinical pathological stage after operation were analyzed and compared in with and without negative IMS patients respectively. And the positive rate and the correlation with clinical pathological stage after operation in expression of CK19 mRNA or/and CK20 mRNA while compared with in expression of CK19 mRNA and CK20 mRNA both in nested RT-PCR with negative IMS and nested RT-PCR without negative respectively.Results: 1. Both of the sensibility of detection of the SW60 cells by examination of CK19 mRNA and CK20 mRNA by nested RT-PCR were 10-6. The positive rate of expression of CK19 mRNA and CK20 mRNA and CK19 mRNA or/and CK20 mRNA by nest RT-PCR was 28%, 34% and 50% in the peripheral blood of patients respectively and were negative in the control. The positive rate in the ⅢⅣ clinical pathological stage patients （with lymph or blood metastasis lesion） was more high than that in ⅠⅡ clinical pathological stage patients （without lymph or blood metastasis lesion）（ 37%, 47%, and 63% to 15% , 15% and 31% ）. And the ratio of ⅢⅣ clinical pathological stage in the positive expression of CK19 mRNA, CK20 mRNA and CK19 mRNA or/and CK20 mRNA patients were more higher than in thenegative patients（78% , 82% and 75% to 52% , 48% and 44%）, respectively. 2.The depleted rate of negative IMS was 71-82%, The positive rate of expression of CK19 mRNA , CK20 mRNA and CK19 mRNA or/and CK20 mRNA by nested RT-PCR after negative IMS was 47%, 56%和81 %, were more higher than by nested RT-PCR without negative IMS respectively. The positive rate in the ⅢⅣ clinical pathological stage patients was more high than that in ⅠⅡ clinical pathological stage patients （ 63% , 68% and 95% to 23% , 38% and 62% ）. And ratio of Ⅲ—Ⅳ clinical pathological stage patients after operation in the positive expression of CK19 mRNA and CK20 mRNA and CK19 mRNA or/and COO mRNA patients were more higher than in the negative patients（ 80% , 72% and 69% to 41% , 43% and 17% ）. 3.The positive rate of combination of IMS and ICC was 33%, and that of ICC was negative, the difference betwe

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