Screening Of Colorectal Cancer By Fecal Shedding Cells And Peripheral Blood Markers

BackgroundColorectal cancer is one of the common malignant tumors in the world,and its incidence rate and mortality rate are the third of all the malignant tumors.In recent years,the incidence of colorectal cancer increased year by year,which has become a threat to public health and life safety of major diseases.Studies show that early effective screening and treatment can reduce the incidence and mortality of colorectal cancer.Currently,fecal occult blood test?FOBT?and Colonoscopy are the most widely used metholds for detection of colorectal cancer.Fecal occult blood test is commonly used,but its sensitivity and specificity are relatively low,and the false negative rate and false positive rate were higher.Although the colonoscopy is a gold standard for clinical screening and diagnosis,but its cost is also high,and can not be promoted as a first-line screening method.Stool detection which is simple,high specificity,noninvasive,and without special detection before preparation,can be used as an important tool for early screening and diagnosis of colorectal cancer.However,there are a lot of impurities such as bacteria,food residue,bile and intestinal mucus in the feces,which leads to the low purity of extraction the cells.The commonly used cell extraction methods including cell centrifugal elutriation method,cell density gradient centrifugation and cell immune magnetic bead method,but the three extraction methods for smear background unclear,time-consuming and laborious,high cost,complicated operation and other shortcomings greatly limits the screening bed application in clinical.Therefore,we used nano Fe304 folic acid beads alternative immunomagnetic beads to collect exfoliated cells,and to further explore the application feasibility and clinical screening efficiency in clinical diagnosis of colorectal cancer in the first part of this study.In addition,recent studies have found that the methylation of SEPT9 gene can be used as a molecular marker for early detection of colorectal cancer,but early studies used the most early detection methods of the SEPT9,and the experimental population are.the western countries.By improving the process and the experimental process,the new generation of SEPT9 gene methylation detection method improve the level of analysis of trace free DNA,which can improve the detection performance of colorectal cancer,but the relevant research is still very little.Therefore,in the second part of this study,we evaluated the efficiency of the second generation of the SEPT9 gene in diagnosis of colorectal cancer in the Chinese population node,and compared with the method of immune fecal occult blood test?FIT?.PurposesThe first aim is to explore the viability and clinical screening efficiency of enrichment by the Fe304 folic acid magnetic beads in stool of patients with colorectal cancer.The second part is to explore the role of SEPT9 gene methylation in the early diagnosis of colorectal cancer in Chinese population,and to compare with FIT.Method1.To explore the clinical significance and viability of nano Fe304 folic acid magnesphere beads in extraction of fecal shedding cell.First,to assess the feasibility of nano Fe304 folic acid magnesphere beads in extraction of fecal shedding cell by enriched faecal shedding cells test of colorectal cancer patients;the second,to assess the viability of nano Fe304 folic acid magnesphere beads in extraction of fecal shedding cell by enriched HCT116 cells test of colorectal cancer patients.2.To investigate the sensitivity and specificity of SEPT9 in detecting colorectal cancer.First,a retrospective case control study using colonoscopy and pathological diagnosis as gold standard is used.The study subjects were healthy and normal subjects for the diagnosis of colorectal cancer by colonoscopy and pathological diagnosis.Second,the detection performance of FIT and SEPT9 is compared by using the method of paired study.ResultsThe first aim is to explore the viability and clinical screening efficiency of enrichment by the Fe304 folic acid magnetic beads in stool of patients with colorectal cancer.1.The efficiency of magnetic beads detection system in enrichment of vitro cultured colorectal cancer cell1.1 Fe304 folic acid magnetic beads surrounded by high glucose compound does not appear bead and liquid layer after suspension placeing for a long time.suspension is stability and magnetic beads will not precipitate in the bottom of the tube.The cells of the internal structure can be more clearly observed,and cells of the internal structure can not be affected.1.2 The Optimal reaction conditions for cultured colorectal cancer cell in vitro.The Optimal reaction conditions for cultured colorectal cancer cell in vitro is that 100ul of HCT116 cells suspension was added to the 40ulFe₃O₄ folic acid magnetic beads surrounded by high glucose compound,and water bath incubation for 60min in 3 7 ?,then 15min adherent reaction will be done in the mixed suspensions with nano folic acid beadsand cells.2.The exfoliative experiment of Fe304 folic acid magnetic beads in enrichment of fecal shedding cells in patients with colorectal cancer.2.1 The sensitivity of fecal cytology in the detection of colorectal cancer.The sensitivity of the cells in the examination of the large intestine was 66%?33/50?.The positive rate of the abnormal cells was significantly correlated with the distribution of colorectal cancer?P<0.05?.2.2 The specifity of fecal cytology and fecal occult blood test in the detection of colorectal cancer.The specifity of fecal cytology in examination of colorectal cancer can be as high as 100%;The specifity of fecal cytology for detecting colorectal cancer was significantly higher than that of fecal occult blood test?P<0.05?.2.3 The results of bowel feces and natural stool cytology in patients with colorectal cancer.The sensitivity of the combined bowel feces and natural stool cytology for the detection of ocolorectal cancer is much higher than that of single bowel feces or natural stool cytology;the detection sensitivity of combined cytology can be as high as 92%?46/50?.2.4 The effectiveness of the nano folic acid magnesphere beads and three screen integration method to collect exfoliated cytology.The three layers of screen meshes of various method to collect the exfoliated cells often are uneven distribution,large amounts of waste residue,seriously affected the cell morphology,but nano folic acid magnesphere beads method enrichment of shedding cells are uniformly dispersed and only a very small amount of waste residue,cell morphology was not affected.The sensitivity and specificity of SEPT9 for detecting colorectal cancer by peripheral blood markers.1.The result of case control study between colorectal cancer patients by colonoscopy and pathological diagnosis and healthy controls were retrospectively reviewed.1.1.In tumor lesions,The positive rate of SEPT9 increased with the progression of the disease.1.2.the sensitivity of SEPT9 to colorectal cancer was 74.8%?95%67.0%-81.6%Cl:?,and the specificity was 87.4%.The positive rate of SEPT9 increased with the progression of colorectal cancer staging.1.3 There were no significant differences in the positive rates of SEPT9 in different age,gender and tumor location.2.The detection performance of FIT and SEPT9 was compared by paired study.2.1 SEPT9 compared with the FIT test for colorectal cancer has better detection performance.2.2 SEPT9 and FIT combined detection of colorectal cancer can significantly improve the detection rate.Conclusion1.Nano folic acid magnetic beads method can significantly improve the detection sensitivity of exfoliative cells in stool,which provide a reliable and effective method for colorectal cancer early detection,and Nano folic acid magnetic beads method detection method not only can increase detection efficiency,but also not affect the cellular structure and background picture.2.SEPT9 has good detection performance,which can be used for colorectal cancer screening.The second generation SEPT9 test method has better performance than the FIT in colorectal cancer detection,and it is better than the first generation SEPT9 for detection performance.Combined use of SEPT9 and FIT to detect colorectal cancer may improve the detection rate and the screening effect.