ABSTRACT
Objective To explore the infection status of high risk human papilloma virus (hrHPV) and its effects and clinical significance on expression of cysteinyl aspartate specific protease 1 (Caspase-1) and interleukin 1 β(IL-1β)in tissues of human cervical carcinoma. Methods A total of 102 patients with cervical carcinoma (cervical carcinoma group), 60 patients with cervical intraepithelial neoplasia (CIN group) and 30 patients with normal cervix (control group) were used as the research objects. PCR reverse dot hybridization method combined with DNA chip technique were used to detect hrHPV. The expressions of Caspase-1 and IL-1β were detected by immunohistochemical technique. Data were analyzed between hrHPV positive group and hrHPV negative group, between single type of hrHPV infection group and multiple type of hrHPV infection group. The relationship between caspase-1 and IL-1βexpression and clinicopathological parameters in cervical carcinoma patients were observed. Results HrHPV infection was detected in 75 cases(73.5%)in cervical carcinoma group and 11 types of hrHPV were detected. In these 11 cases, single type and multiple type of hrHPV infection were 61 cases(81.3%)and 14 cases(18.7%) separately. HrHPV infection rate was much higher in cervical carcinoma group than those in CIN group and control group(36.7%and 6.7%). Caspase-1 and IL-1βpositive rates were significantly higher in cervical carcinoma group(61.8%and 51.0%)than those in control group(26.7%and 23.3%). The positive rate of Caspase-1 was significantly higher in cervical carcinoma group than that in CIN group(40.0%, all P0.05). The difference of positive expressions of Caspase-1 and IL-1βwere significantly related with cell differentiation, tumor size, lymphatic metastasis and clinical stage(P<0.05 or P<0.01). Conclusion There are single and multiple types of hrHPV infection in cervical carcinoma and the infection rate is high. HrHPVs may promote the progression of cervical carcinoma by restraining the expressions of Caspase-1 and IL-1β.
ABSTRACT
Objective To investigate the effects of p53 on expression and activity of protein kinase R (PKR) as well as biological characters of HeLa cells from cervical carcinoma patients. Methods Recombinant plasmid vector pEGFP-C1/p53 was constructed to over-express p53 then it was transfected into HeLa cells. Transcription levels of p53 and PKR mRNA were detected by reverse transcriptase polymerase chain reaction (RT-PCR) among pEGFP-C1/p53 transfection group, pEGFP-C1 transfection group and blank control group(only transfection reagent was added);Protein expression lev?els of p53, PKR, phosphated PKR(p-PKR)and phosphatedαsubunit of eukaryotic initiation factor 2(p-eIF2α)which is the downstream substrate of PKR were detected by Western Blot among three groups;Proliferation of HeLa cell were deter?mined by methyl thiazolyl tetrazolium(MTT)assay;Invasion of HeLa cell were determined by Transwell cell assay. Re?sults Recombinant plasmid vector pEGFP-C1/p53 was successfully constructed to overexpress p53;Transcription level of p53 and PKR mRNA in pEGFP-C1/p53 transfection group were higher than those in pEGFP-C1 transfection group and in blank control group (P<0.05),and there were no significant difference between their levels in pEGFP-C1 transfection group and in blank control group;Protein expression levels of p53, PKR, p-PKR andp-eIF2α in pEGFP-C1/p53 transfection group were higher than those in pEGFP-C1 transfection group and in blank control group (P<0.05),and there were no sig?nificant difference between those expression levels in pEGFP-C1 transfection group and in blank control group;MTT and Transwell cell results showed that proliferation and invasion of HeLa cells in pEGFP-C1/p53 transfection group were weaker than those in pEGFP-C1 transfection group and in blank control group (P<0.05),and there were no significant difference between proliferation and invasion of HeLa cells in pEGFP-C1 transfection group and in blank control group. Conclu?sion p53 can up-regulate the expression and activity of PKR, promote activation of PKR/eIF2αsignal transduction pas?sage and restrain cell proliferation and invasion of HeLa cells.
ABSTRACT
Objective To identify the significance of expression and phosphorylation of protein kinase R(PKR) and nuclear factor NF-κB p65 in cervical lesions, and the effect of high-risk human papilloma virus(hsHPV) on expression and phosphorylation of R(PKR) and NF-κB p65. Methods A total of 67 patients with cervical cancer, 149 patients with cervi-cal intraepithelial neoplasia (CINⅠ-Ⅲ) and 15 normal control were included in this study. The expression levels of PKR, phosphorylated PKR (p-PKR), NF-κB p65 and phosphorylated NF-κB p65 (p-NF-κB p65) were detected by immunohisto-chemical SP method in three groups. Results The positive expression rates of PKR and p-PKR in cytoplasm were signifi-cantly lower in hsHPV positive group than those in hsHPV negative group (27.2% and 11.0% vs 41.1% and 21.1%,χ2 =4.858 and 4.371,P＜0.05). The positive expression rates of NF-κB p65 and p-NF-κB p65 in cytoplasm and nucleus were significantly higher in hsHPV positive group than those in hsHPV negative group (46.3%, 25.7%, 22.8% and 12.5% vs 32.6%, 14.7%, 11.6%and 4.2%,χ2=4.345,4.048,4.729 and 4.650 respectively,P＜0.05). The positive expression rates of PKR in kytoplasm and karyon were significantly lower in NF-κB p65 (+) group than those in NF-κB p65 (-) group (25.5%vs 38.0%and 20.4%vs 36.3%,χ2=3.898 and 4.396 respectively, P＜0.05). The positive expression rate of PKR in kyto-plasm was significantly lower in p-NF-κB p65 (+) group than those in p-NF-κB p65 (-) group (19.0%vs 36.0%,χ2=4.462, P＜0.05). Conclusion hsHPV may inhibit the expression and phosphorylation of PKR but promote the expression and phosphorylation of NF-κB p65. The expression and phosphorylation of NF-κB p65 may inhibit the expression of PKR. Regu-lating effects of three may be associated with the generation and progression of cervical cancer.
ABSTRACT
Objective To explore the relationship of signal transduction among human papillomavirus 18 E6 oncoprotein (HPV18E6), signal transducers and activators of transcription 1 (STAT1), protein kinase R( PKR )/α subunit of eukaryotic initiation factor 2 ( eIF2α ), nuclear factor-kappa Bp65 ( NF-κBp65 ), mitogen-activated protein kinase( MAPK)/c-Jun N-terminal kinase(JNK) ,and possible molecular mechanism. Methods Construct two lentiviral vectors which contain shRNA interfering sequence aiming at the targets of HPV18E6 oncogene and NC sequence( HPV18E6-RNAi-LV, NC-GFP-LV), based on the transduction with HPV18E6-RNAi-LV and NC-GFP-LV into HeLa cell to interfere the expression of HPV18E6 oncogene and NC sequence,the expressions of mRNA and protein( including phosphating patem)of HPV18E6, STATI, PKR, eIF2α, NF-κBp65, MAPK, JNK are measured with RT-PCR and Western blot, the difference of proliferation and sensitivity to carboplatin of HeLa cell are determined with Transwell cell methods and MTT among every groups. Results The expression of HPV18E6 oncogene can affect the expression level of mRNA and protein of NF-κBp65 and PKR genes, also affect phosphating levels of phosphating protein p-STAT1, p-PKR and p-eIF2α;the restraining rates of proliferation and sensitivity to carboplatin of HeLa cell are higher in HPV18E6-RNAi-LV group than the other groups( P＜0. 05 or P＜0.01 ). Conclusion HPV18E6 oncoprotein not only reduces the expression of PKR but dephosphorylates p-STAT1, pPKR and p-eIF2α to restrain activation of PKR/eIF2α signal transduction passage, maintain the proliferation and invading ability of HeLa cell and restrain apoptosis. The signal transduction among HPV18E6, MAPK/JNK are not clear.
ABSTRACT
Objective:To identify the relationship between expression of protein kinase R(PKR), phosphating PKR, EIF2α(p-PKR, p-EIF2α) in PKR→EIF2α signal transduction passage and the grades of cervical lesions, the role in generation and progression of cervical tumor and their effects to prognosis of cervical cancer patients. Methods:The expressions of PKR, p-PKR and p-EIF2α in human cervical cancer tissue of 63 cases, cervical intraepithelial neoplasia(CINⅠ-Ⅲ) of 114 cases and normal cervical epithelium of 15 cases were detected by immunohistochemical technique. Results:With the increase in grades of cervical lesions, the positive-expression rate of PKR increased and significantly correlated with the grades of cervical lesions(P < 0.05). With the increase in grades of cervical lesions, the positive-expression rates of p-PKR and p-EIF2α increased firstly, and then decreased. In cervical cancer group, the positive-expression rate of PKR was much higher than that of p-PKR(P < 0.01). The development and progression was quicker in later clinical stages of cervical cancer than that of earlier clinical stages of cervical cancer (P < 0.01). The development and progression of cervical cancer was quicker in patients with negative-expression of p-PKR and p-EIF2α than that in patients with positive-expression of p-PKR and p-EIF2α(P < 0.05). Conclusion:The positive-expression rate of PKR was correlated with the grades of cervical lesions. There are some factors which can impede PKR and EIF2α to be phosphorylated or make p-PKR and p-EIF2α dephosphorylate in high level cervical lesions, which promotes the development and progression of cervical lesions, worsens the prognosis of cervical cancer.