Analysis Associated With Prediction Model Of Cervical Intraepithelial Neoplasia Progression

Chapter1 Research status associated with progress of cervical intraepithelial neoplasiaCervical intraepithelial neoplasia(CIN)is a precursor lesion for cervical cancer。The occurrence and progress of CIN is a multi-factor,multi-level process.Numerous studies confirm that high-risk HPV persistent infection is an important tumorigenesis cause of CIN and cervical cancer,closely associated with the development of cervical cancer.With advances in molecular biology and the development of sequencing technology,the association of uterine cervical microbiota with HPV in the development of cervical intraepithelial neoplasia and cervical cancer is gradually being recognized.It has become a hot research of cervical lesions to find and explore the prognostic biomarkers of CIN.Early intervention of CIN can block the progression of CIN to cervical cancer.Appropriate treatment and testing instruments according to the characteristics of the different stages of CIN has become the key concern of CIN diagnosis,treatment and follow-up.Therefore clinically effective treatment and follow-up methods blocking the progress of CIN become necessary.However,the best approach is still controversial.This paper reviews the etiology and progression of CIN and its influencing factors,markers and clinical status of other aspects,in order to properly regulate diversion and cervical intraepithelial management.Chapter2 Meta-analysis of cold-knife conization versus loop electrosurgical excision procedure for preventing cervical intraepithelial neoplasia progressObjective: This meta-analysis aims to compare the superiority of loop electrosurgical excision procedure(LEEP)or large loop excision of the transformation zone(LLETZ)and cold-knife conization(CKC)in the prevention of cervical intraepithelial neoplasia(CIN)progress.Methods: Systematic searches were performed in the MEDLINE,EMBASE,Cochrane databases,and the China National Knowledge Infrastructure Databases to identify all potential articles involving patients with CIN treated with LEEP/LLETZ or CKC published up to February 2016.Risk ratios(RR)or weighted mean difference(MD)with a 95% confidence interval(95%CI)were calculated.Results: Seven randomized controlled trials,one prospective cohort study and twelve retrospective cohort studies were included in this meta-analysis.There were no significant differences following LEEP/LLETZ compared with CKC in recurrence rate(RR 1.75,95%CI0.99-3.11,P=0.06),positive margin rate(RR 1.45;95%CI 0.85-2.49,P=0.17),residual disease rate(RR 1.17,95%CI 0.75-1.85,P=0.49),secondary hemorrhage(RR 1.19,95%CI 0.75–1.87;P=0.46),or cervical stenosis.Moreover,subgroup analyses based on randomized trials also revealed that no statistically significance was observed in the above outcomes.Small studies showed CKC has increased the risk of preterm birth over LLETZ/LEEP.However,women treated with CKC had a significantly deeper cervical cone than those treated with LLETZ/LEEP(MD-5.71,95%CI-7.45 to-3.96;P<0.001).Conclusions: LEEP/LLETZ is as effective as CKC regarding recurrence rate,positive margin rate,residual disease rate,secondary hemorrhage,and cervical stenosis for the surgical treatment of CIN.Further large-scale studies are needed to confirm our findings.Charpter3 Clinic Research for preventing cervical intraepithelial neoplasia progressObjective: To evaluate the short-term and long-term efficacy after therapy for cervical intraepithelial neoplasia,and explore the influencing factors of recurrence.Methods: A multi-center retrospective study of 1226 cases about high grade intraepithelial neoplasia between 2009 and 2014 was involved from3 third-grade class-A communal hospitals in Guangxi Zhuang Province.The clinical efficacy of surgical cone,pathologic positive margins,and postoperative recurrence were assessed with univariate and multivariate analysis.Results: There was significant difference in estimated blood loss and intra-operative time between CKC group and LEEP group(P<0.05).There was no significant difference in surgical margins in the two groups(P > 0.05).Univariate analysis showed that age,menopause,the range of lesions,colposcopy whether satisfaction may be related to risk factors as affecting surgical margin status(P <0.05).Logistic regression analysis revealed that menopause,the range of lesions,colposcopy whether satisfaction were risk factors of postoperative positive margin.Cox regression analysis showed that margin status,persistent infection of high-risk HPV and menopause were risk factors for recurrence after surgical cone.Conclusions: Both CKC and LEEP are safe and effective methods for prohibition of CIN progress.Positive margin,high-risk HPV persistent infection and menopause are risk factors for CIN recurrence.Repeat conization may be choosen for positive margins.Charpter4 Bioinformatics analysis related to the progress of cervical intraepithelial neoplasiaObjective: The aim of this study was to excavate the potential genes associated with the progress of cervical intraepithelial neoplasia(CIN)through microarray expression profiling data analysis and bioinformatics approaches.Methods: Gene expression profile were downloaded from Gene Expression Omnibus database and the dysregulated genes were screened with the GEO2 R.Subsequently,the correlation between the differentially expressed genes and the regulation of progress in CIN was investigated through comprehensive bioinformatic approaches,including pathway enrichment,biological process annotation,text mining and protein-protein interaction analysis.Results: 1,Two mRNA expression microarray datasets that were related to progress in CIN were obtained from the GEO database.Through analysis of the microarray data,we obtained 144 differentially expressed upregulated genes and 219 differentially expressed down-regulated genes.2,Pathway enrichment analysis of the differentially expressed upregulated genes identified 3 signaling pathways associated with progress in CIN,including Wnt signaling pathway,Endocytosis signaling pathway and Vibrio cholerae infection signaling pathway.Fourteen differentially expressed genes were also identified including CCND2,NFATC1,PRKCB,PPP2R5 D,WNT2B,SH3KBP1,HSPA6,PIKFYVE,RABEP1,TGFBR2,KDELR1,MUC2,KCNQ1 and TJP2.3,Biological annotation through COREMINE and text mining showed that,except PPP2R5 D and PIKFYVE,CCND2,TGFBR2 and MUC2 were directly related to progress in CIN and another nine genes,while not directly related to CIN progression,were associated with tumor progression and recurrence.These genes Also significantly correlated with each other,such as TGFBR2 and WNT2 B.4.Twenty-four reported genes related to CIN progression were mined by PUBMED.Gene Mania tool analysis indicated that protein interaction network formed between all the differentially expressed genes and the reported genes in direct or indirect way.Wherein CCND2 and TGFBR2 formed direct interaction with many reported genes.Conclusions: 1.We found 14 differentially expressed genes that associated with progress in CIN through analyzing microarray data.2,CCND2 and TGFBR2 may be the important candidate genes related to progress in CIN.Wnt signaling pathway may be an important pathway in the development of CIN,and research based on this pathway may provide new solutions to block the progress of CIN.Chapter5 Expression analysis of P16,Ki-67 and the gene based on bioinformatics associated with cervical intraepithelial neoplasia progressObjective: Nine differentially expressed genes based on bioinformatics screening and gene P16,Ki-67 were verified for the function related to CIN progress.Methods: Tissue microarray was made included CIN1(n=14),CIN2(n=23),CIN3(n=31),cervical cance(33)and normal tissues(n=29).P16,Ki-67 and the nine gene based on bioinformatics associated with cervical intraepithelial neoplasia progress were detected with immunohistochemistry.The clinical and pathological analysis was conducted.Results: Expression of NFATC1,WNT2 B,SH3KBP1,PIKFYVE,RABEP1 and TGFBR2 were not detected in CIN and cervical carcinoma.CCND2,P16 and Ki-67 expression showed a gradual increasing trend in normal,CIN and cervical cancer lesions.CCND2,P16 and Ki-67 expression in hr HPV positive CIN tissues was significantly higher than that of high-risk HPV-negative CIN tissue.Factors affecting CCND2 expression was CIN grade and high-risk HPV infection(P <0.05).Logistic diagnostic screening model found that Ki-67,P16 and high-risk HPV was the three valuble factors of CIN diagnosis.Conclusions: CCND2 may be able to predict CIN progression.CCND2,P16 and Ki-67 may play a role in synergy of HPV during the progression from CIN to cervical cancer.CCND2 is expected to be the new markers to predict CIN progression except P16 and Ki-67.Chapter6 The composition and diversity of cervical microbiome in patients with cervical intraepithelial neoplasiaObjective: To describe the 16 S r DNA gene profiling of cervical microbiome patients with CIN and cervical cancers,and in health controls.Methods: From May 2015 to July 2015,cervical tissues confirmed by histopathology were collected from the patients in Tumor Hospital of Guangxi Medical University and Guangxi Liuzhou People’s Hospital without receiving any treatment before sampling.Bacterial 16 S r DNA V4 gene was sequenced on the Illumina Miseq high-throughput sequencing platform.Sequences were analyzed with the QIIME software package;in addition to custom Perl scripts to analyze alpha and beta diversity.Rarefaction curves were generated for complexity analysis of sample.QIIME calculated both weighted and unweighted unifrac,which are phylogenetically aware measures of beta diversity.We used unweighted unifrac to do principal coordinate analysis(PCo A)to differentiate species number and variety.Results: Sixty subjects without receiving any treatment were enrolled included normals(n=14),CIN1(n=9),CIN2(n=11),CIN3(n=18)and cervical cancers(n=10).No differences in age,parity,contraceptive methods were found between the groups.High-risk HPV infection rates increased gradually with cervical lesions(P<0.05).Alpha diversity analysis based on rarefaction curves suggested a good sequenced coverage,and a good balance between the depth of sequencing and the species richness among normal,CIN and CC group.In general,seventy-three bacterial genus from twenty-three bacterial class were identified on the cervical tissues.Lactobacillus were present in a vast majority in all groups,but there were still some normal samples without dominance of lactobacilli.The dominant genus in CIN1/2 group was Stenotrophom,Chitinophaga and Acinetobacter.The dominant genus in CIN3 and cervical cancer was Halomonas,Shewanella,Acinetobacter.Conclusions: The present study used 16 S rDNA sequencing to detect the cervical microflora indicated that different grades of CIN presented diversity of microflora.Combined with high-risk HPV infection and the relationship between the degree of the disease,it may suggest that cervical microbial diversity was associated with high-risk HPV infection and disease severity.