| IntroductionWith the high incidence of gastric cancer,the mortality of gastric cancer is on the first place in our country.The main treatment method of gastric cancer is gastric radical resection,and the postoperative 5-year survival rate after surgery is about 40%.Lethal peritoneal metastasis happened in 30%to 50%gastric cancer patients.And once the metastatic focus comes into being,it is hard to achieve radical cure.After dropping into abdominal cavity,gastric cancer cells produce kinds of molecules because of their biological activities.So it is possible to detect the existence of cancer cells in abdominal cavity by investigating those molecules.To increase the detection rate of exfoliate gastric cancer cells in abdominal cavity,many molecular biological methods were used to investigate kinds of molecules relating with peritoneal metastasis in peritoneal washing.But the results were not coincident and it was indefinite which molecule was the best to predict the existence of exfoliate cancer cells in abdominal cavity.Among kinds of molecular biological methods,the better sensitivity of Real-time RT-PCR makes it possible to detect peritoneal micrometastasis.In this study,we selected 8 biological molecules,the mRNA expression of these molecules in 115 peritoneal washing samples were investigated by Real-time RT-PCR, to find more sensitive molecules to increase the detection rate of exfoliate cancer cells in abdominal cavity.Materials and Methods1.Peritoneal washing samples115 peritoneal washing samples were collected from the patients admitted to the Department of Surgical Oncology in the First Affiliated Hospital of China Medical University.2.Grouping of the samplesAccording to the results of Peritoneal Lavage Cytology(PLC) examination,the samples were assigned to the following groups:PLC(-) group;PLC(+) group;PLC(±) group and control group which peritoneal washing samples were collected from non-cancer patients.3.Total RNA extraction from peritoneal washing samplesTotal RNA of each sample was extracted using Trizol reagent.1.5μl of the extracted RNA was used to assess quantity and purity,and the remaining were conserved at-80℃.4.The mRNA was amplified by Real-time RT-PCR(1)The total RNA was reverse-transcribed by using a SuperscriptPreamplification System for First Strand cDNA Synthesis Kit.(2)Real-time RT-PCR was carried out using primers designed.(3) Rotor-Gene 3000 system was used to analysis the product of PCR.5.Statistics analysisData were analyzed by X~2-test,Mann-Whitney test and Kruskal-Wallis test of SPSS 13.0 software.Results1.PLC results and groupingPLC(-) group:66 cases,57.3%(66/115);PLC(+) group:31 cases,27.0% (31/115);PLC(±) group:18 cases,15.7%(18/115).2.Real-time RT-PCR positive results for the markers in peritoneal washings samplesCEA:39.1%(45/115);DDC:34.8%(40/115);HPA:33.0%(38/115);MMP-7: 31.3%(36/115);Ki-67:30.4%(35/115);TGF-β1:28.7%(33/115);PCNA:25.2% (29/115);L-3-PP:24.3%(28/115). 3.The results of ROC Curve analysisThe diagnosis power of each marker was evaluated by the area under the ROC curve.Markers:CEA＞DDC＞HPA＞MMP-7＞Ki-67＞TGF-β1＞PCNA＞L-3-PP.4.The relationship between CEA,DDC,HPA expression and the clinicopathological features of peritoneal metastasisIn the 31 PLC(+) samples,CEA positive rate was 93.5%(29/31),DDC positive rate was 90.3%(28/31),HPA positive rate was 87.1%(27/31),MMP-7 positive rate was 77.4%(24/31),Ki-67 positive rate was 71.0%(22/31),TGF-pi positive rate was 61.3%(19/31),PCNA positive rate was 54.8%(17/31) and L-3-PP positive rate was 45.2%(14/31).Among the 8 molecules,the positive rate of CEA,DDC,HPA expression which were 93.5%,90.3%,87.1%respectively,and higher than the other 5 molecules and PLC results(P＜0.05),which indicated their better sensitivity.In the 14 samples with positive peritoneal metastasis,CEA positive rate was 100% (14/14),DDC positive rate was 100%(14/14),HPA positive rate was 92.9%(13/14), MMP-7 positive rate was 78.6%(11/14),Ki-67 positive rate was 71.4%(10/14), TGF-β1 positive rate was 64.3%(9/14),PCNA positive rate was 64.3%(9/14) and L-3-PP positive rate was 57.1%(8/14).Among the 8 molecules,the positive rate of CEA,DDC,HPA expression which were 100%,100%,92.9%respectively,and significantly higher than the other 5 molecules and PLC results(P＜0.05),which indicated their better sensitivity.The relative quantity of CEA,DDC and HPA expression was related significantly with invasive depth,serosal types,growth modes and gross types(P＜0.05).Conclusions(1)As three markers related with peritoneal metastasis of gastric cancer,CEA, DDC and HPA are sensitive and specific to predict subclinical peritoneal metastasis;(2)Real-time RT-PCR is sensitive to detect exfoliate cancer cells in peritoneal cavity of gastric cancer patients and may widely used in clinical detection;(3)CEA,DDC and HPA are correlated to the clinicopathologic factors of peritoneal metastasis of patients with gastric cancer.
|
PDF Full Text Request