| Objective In order to evaluate the clinical significance of circulating cell free DNA size distribution state and DNA integrity(the ratio of long segment gene to short segment gene)in the diagnosis and treatment of primary gastric cancer,and investigate the application value of serum tumor marker and its corresponding cell free DNA in the diagnosis and treatment with gastric cancer.To further explore the application value of plasma cell free DNA and blood routine detection index in distant metastasis of gastric cancer.Methods First Part Construction the standard curve of ALU(Alu repetitive sequence,ALU)gene standard plasmid,set up a method for the detection of SYBR Green real time fluorescence quantitative PCR.Design 115 bp,247 bp specific primers of the amplification product length to the ALU reference gene.Collected the peripheral blood of 81 cases of gastric cancer,19 cases of gastric precancerous lesion,18 cases of gastric benign tumors,43 cases of healthy control group,36 cases gastric cancer patients followed up for postoperative 3 day and 26 cases followed up for postoperative 2 months,detected the content of two free fragments of plasma and serum circulating cell free DNA,at the same time,combined with clinical pathological data of the patients,to evaluate the value of clinical application of cell free DNA and integrity in the diagnosis and treatment of gastric cancer,comparison of plasma and serum selection in the detection of cell free DNA.Second Part Design the specific primers of CEA target gene.Collected the peripheral blood of 81 patients with gastric cancer,19 cases of gastric precancerous lesion,43 cases of healthy control group,36 cases gastric cancer patients followed up for postoperative 3 day and 26 cases followed up for postoperative 2 months.Detected the content of their serum CEA and its corresponding cell free DNA(CEA cf-DNA)in their plasma.Then,combined with clinical and pathological data of the patients,analyzed its potential application value in the diagnosis and treatment of gastric cancer.Third Part Objective to investigate the clinical value of plasma free DNA and routine blood test in 81 cases of primary gastric cancer and 43 cases of healthy control group,and compared the value and correlation of these indexes in the metastasis and distant metastasis of gastric cancer.Results First Part(1)The standard curve of ALU gene standard plasmid were successfully constructed,set up a method for the detection of SYBR Green real-time fluorescence quantitative PCR.(2)The content and integrity of cell free DNA had statistically significant difference in four groups,in pre-and post-operation and follow-up specimens,in staging,differentiated degree and with or without metastasis(P < 0.05).The content of cf-DNA in plasma and serum was relevant and the former had less discrete trend,plasma was more suitable for quantitative detection of cell free DNA than serum.The area under the curve(AUC)of ALU247 was 0.985.Its sensitivity and specificity for the diagnosis of gastric cancer were higher than those of CEA,CA724,CA199 and other traditional tumor markers.Second Part(1)The content of plasma CEA cf-DNA was higher than that of gastric precancerous lesions group and healthy control group,the difference between the three groups was statistically significant(H = 16.600,P < 0.001),postoperative gastric cancer CEA cf-DNA gene was lower than that before surgery,the difference was statistically significant(U = 2.016,P = 0.044).(2)The content of CEA cf-DNA in plasma of patients with gastric cancer had significant correlation with patients’ gender,age,smoking,hypertension,clinical stage,differentiation and tumor size,had no significant correlation with metastasis(divided into metastasis,only proximal lymph node metastasis and distal organ metastasis three group).the content of CEA in serum of patients with gastric cancer had no significant correlation with the presence of hypertension,differentiation,metastasis and tumor size,had no significant correlation with gender,age,smoking status and clinical stage.(3)For the ROC curve,the efficacy of plasma CEA cf-DNA in the diagnosis of gastric cancer was significantly higher than that of serum CEA,The area of ROC curve of CEA cf-DNA and serum CEA in the diagnosis for gastric cancer were 0.848.Third Part(1)cf-DNA,neutrophil count(NE#),the percentage of lymphocytes(LY%),mean platelet volume(MPV),D-Dimer(D-D),fibrinogen degradation products(FDP),Globulin ratio(A/G)and Pre albumin(PAB)in gastric cancer metastasis had significant difference(P < 0.05).(2)There was a certain correlation between the content of cell free DNA and D-D,FDP and A/G three indexes.It is suggested that the megakaryocyte was stimulated to produce a large number of platelets,and the destruction of DNA may release a large number of DNA into the peripheral blood,which is one of the sources of free plasma.Conclusion1.There is a highly application value of cell free DNA in the early diagnosis and treatment of gastric cancer,which is higher than the traditional tumor markers.The content and integrity of cell free DNA in blood had some reference value in the evaluation of the prognosis of gastric cancer patients.2.Although there was a correlation between plasma cell free DNA and serum free DNA in the same patients,the discrete trend in the serum of cell free DNA was greater than that in plasma,so the plasma was better than that of serum and was suitable for the detection of cell free DNA.3.CEA cf-DNA had some value in the early diagnosis of gastric cancer,and it had some significance in the evaluation of gastric cancer metastasis.4.Plasma cell free DNA content and some index of routine detection had a certain value in clinical judgment and prediction of gastric cancer metastasis,the plasma cell free DNA and pre albumin levels were independent risk factors for distant metastasis of gastric cancer.5.Detection of peripheral blood cell free DNA has broad prospects in clinical application for its advantages of convenient and simple operation,small and easy to get and carry out large-scale. |
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