Study On The Aquaporin Expression Profile And The Relationship With Clinical Prognosis In Cervical Carcinoma Of Uygur Women

Objective: It is very significant for patients of cervical carcinoma that earlydiagnosis, normalized treatment and efficient survey. To find the soecufuc aquaporinssubtypes which were differential expressed in occurrence, invasion and metastasis ofcervical squamous carcinoma.To analysis the significance and expression of theseaquaporin subtypes in gene transcription level and protein expression level.To investigatethe mechanism of the specific aquaporin subtypes in cervical carcinogenesis, invasionand metastasis of uygur patients.Select the marriage-pregnant factors, clinicalpathological factors and specific aquaporin expression factors which may be related withthe prognosis and analyzing the follow-up survival data of uygur patients with cervicalcarcinoma. To find the independent factors which affected the prognosis of cervicalcarcinoma and the relationship of specific AQPs expression with prognosis.To provideevidence and new approach for the molecular mechanism, servings standardizedaquaporin molecular index for early diagnosis, prognosis monitoring and curative effect.Methods:1) Total RNA was isolated from2fresh sample of early stage cervicalcarcinoma,2sample of late stage cervical carcinoma,2sample of cervicitis tissues byTRIzol. The mRNA expression of these aquaporin subtypes were detected by RealtimeRT-PCR on gene transcription level which were existed in female reproductive system(AQP1, AQP2, AQP3, AQP4, AQP5, AQP8).2) Select10sample of cervitis18sample ofcervical carcinoma.To screen the protein expression of aquaporin subtypes which wereexpressed in female reproductive system by immunofluorescence technique on proteinlevel.3) The mRNA expression of relavant aquaporin subtypes were detected by real timeRT-PCR on gene transcription level in10sample of cevitis tissue,10sample of earlystage cervical carcinoma and10sample of late-stage cervical carcinoma.4) The proteinexpression of these special aquaporin subtypes were detected by Western blot in12sample of cevitis tissue,12sample of early-stage cervical carcinoma and12sample of late-stage cervical carcinoma in protein level.5) Immunohistochemistry was used todetecte specific aquaporin subtypes protein expression in36sample of cevitis,31sampleof CINⅠ-Ⅱ,36sample of CINⅢ and98sample of cervical carcinoma.And then variousclinical and pathological parameters in cervical carcinoma were investigated andanalysed with specific aquaporin subtypes protein expression.6) Choose196cases ofcervical squamous carcinoma who were treated in hospital and select the clinical andprognostic data.Survival curves were prepared according to the method of Kaplan andMeier, and statistical significance between curves was tested using the log-rank test. TheCox proportional hazard model was also performed to evaluate the independentprognostic value to investigate the correlation with specific aquaporin subtypes. Results:1) AQP1, AQP2, AQP3, AQP5, AQP8mRNA were expressed in cervitis and cervicalcarcinoma in gene transcription level. AQP4mRNA was not detected.The relativeabundance of AQP1were1.005±0.0217，1.94±0.18and3.45±1.22in cervitis, earlystage cervical carcinoma and late stage cervical carcinoma.The relative abundance ofAQP2were1.001±0.0049，1.13±0.0604and1.65±0.134in cervitis, early stagecervical carcinoma and late stage cervical carcinoma. The relative abundance of AQP3were1.04±0.17，1.82±0.0667and4.10±0.12in cervitis, early stage cervical carcinomaand late stage cervical carcinoma. The relative abundance of AQP5were1.016±0.0662，0.899±0.169and1.82±0.815in cervitis, early stage cervical carcinoma and late stagecervical carcinoma. The relative abundance of AQP8were1.000±0.000312，2.33±1.32and5.29±5.04in cervitis, early stage cervical carcinoma and late stage cervicalcarcinoma.The difference of AQP1, AQP2, AQP3, AQP8had no significant in cervitis,early stage cervical carcinoma and late stage cervical carcinoma (P＞0.05). The relativeabundance of AQP5in cervitis was higher than that of cervitis and late stage cervicalcarcinoma. The difference of AQP5was not significant between three groups.2) AQP1,AQP3and AQP8protein expressed in cervitis and cervical carcinoma tissues.AQP2,AQP4and AQP5did not express.AQP1was localized on vascular endothelial. TheAQP1/MVD (microvessel density, MVD) were49.94±4.07and76.51±3.31in cervitisand cervical carcinoma. AQP3was localized on tumor cell membrane. The positive rateof AQP3was20%and66.67%in cervitis and cervical carcinoma.AQP8was localized ontumor cell membrane. The positive rate of AQP8was30%and72.22%in cervitis andcervical carcinoma. The expression of these three protein were all significant different (P＜0.05).3) The relative abundance of AQP1mRNA were1.0067±0.01581,1.770±0.005518and3.954±0.2617; The relative abundance of AQP3mRNA were1.008±0.01886,1.999±0.06361and3.721±0.1660; the relative abundance ofAQP8mRNA were1.0106±0.02255,1.874±0.02854,3.5447±0.1878. The relativeabundanceof AQP1, AQP3and AQP8mRNA were increased and had significantdifference (P＜0.05);4)The protein expression concentration of AQP1were0.9195±0.1351,1.460±0.2963and2.624±0.6603; The protein expression concentration ofAQP3were0.0056±0.002930,0.6362±0.1452,0.9926±0.2173; The protein expressionconcentration of AQP8were1.113±0.2302,1.725±0.2362,2.607±0.6830. Theexpression of AQP1, AQP3and AQP8protein were increased and had significantdifference(P＜0.05).5).AQP1and AQP3protein expression showed a increased tendencygradually from cervitis, precancerous process and cervical cancer.AQP1/MVD were37.51±14.62,40.1±15.32,56.05±27.51,69.84±34.93in in cevitis, CINⅠ-Ⅱ, CINⅢ,cervical carcinoma (P＜0.05). The positive rate of AQP3was16.67%,18.6%,27.78%and44.9%in cervitis, CINⅠ-Ⅱ, CINⅢ and cervical carcinoma (P＜0.05). The positiverate of AQP8was38.38%,41.93%,91.67%and56.12%in cevitis, CINⅠ-Ⅱ, CINⅢ andcervical carcinoma (P＜0.05). The AQP1protein expression was correlated with clinicalstage, diameter of tumor, invasive depth, lymph node metastasis and pathologicalclassification.AQP3protein expression was correlated with clinical stage, diameter oftumor, invasive depth and lymph node metastasis. AQP8protein expression wascorrelated with clinical stage, diameter of tumor, invasive depth and lymph nodemetastasis.6) The proportional hazard factors were clinical stage, treatment,diameter oftumor, depth of invasion, lymph nodal metastasis, HPV infection, SCC-Ag, AQP1, AQP3and AQP8expression. Cox model with stepwise multivariable analysis showed lymphnodal metastasis, clinical stage and diameter of tumor were important factor affecting theprognosis of cervical carcinoma (P＜0.05). The survival rates of AQP1, AQP3and AQP8negative-expression group and positive-expression had significant differences (P＜0.01).AQP1,AQP3and AQP8were one of the factors which affected prognosisi of cervicalcarcinoma,but were not the independent factors.Aquaporins might affect the prognosis ofuygur patients of cervical carcinoma by promoting lymph nodal metastasis and tumorgrowth.Conclusion:1) The study found that AQP1, AQP3, AQP8play important role incervical carcinogenesis of uygur patients. These three special aquaporin subtypes wereselected from the aquaporin subtypes that existed in reproductive system.2) Theexpression of AQP1,AQP3and AQP8upregulated and translated corresponding proteinupregulated,too.And all play important role in carcinogenesis, tumor growth andmetastasis.AQP1were expressed in the endothelial cells of tumor vascular, which increase the expression the angiogenesis and promote tumor growth, mesenchymalinvasion and lymph node metastasis. AQP3and AQP8were expressed in cellularmembrane and/or cytoplasm in tumor cells.which enhanced the movement and invasionability and promote the invasion and metastasis of tumor.These three subtypes showed asynergism.AQP1, AQP3and AQP8may be oncongene and play significant role incarcinogenesis and metastasis of uygur patients and may be a molecular index fordiagnosis, disease survey of uygur patients of cervical carcinoma.3) The survival rates ofAQP1, AQP3and AQP8negative-expression group and positive-expression hadsignificant differences (P＜0.05). Aquaporin may affect the prognosisi of uygur patientsof cervical carcinoma by promoting lymph nodal metastasis and tumor growth.Althoughthey were not independent factor, but provided molecular biology evidence for judgmentand treatment.