Regulation Of ERK On The Expression Of Nuclear Transcriptin Factor In Cervical Cancer Cells

Objective:Epidemiological studies have shown that the onset of cervical cancer is associated with HPV infection,but HPV infection is not the only factor for cervical cancer happening.Recent studies found that mitogen-activated protein kinases(MAPKs)signaling pathway plays an important role in the process of malignant transformation and the invasion and metastasis of tumor.As a subtribe of the MAPK family,extracellular signal-regulated kinase(ERK) could transduct extracellular signals into the nucleus through a series of cascade transduction system,participate in and promote the phosphorylation of a variety of transcription factors,and then launch transcription and expression of multiple Oncogenes and proliferation-associated genes.Previous studies have shown that in cervical cancer,the expression rate of ERK tends to increase with the deterioration of the cervical lesions,but it is not clear how the ERK in cervical cancer cells participate in the phosphorylation of nuclear transcription factors and regulate the expression mechanism,and then affect the biological function of cervical cancer cells.Using the in vitro experimental method, this study applys ERK inhibitor U0126 on cervical cancer cells Siha and C33 A,evaluates the effect of ERK on hn RNP E1,c-Fos and c-Jun and the influence on the biological functions of cervical cancer cells,and explores the mechanism of ERK in cervical cancer.Methods:Select HPV16-positive and HPV-negative C33 A Siha cervical cancer subculture,adding inhibitor U0126 in two kinds of cells, building ERK inhibition system,and forming the control group and intervention group.Employing living cells count and CCK-8 method to detect cell growth situation and cell activity;using Flow Cytometry to detect cell proliferation and apoptosis;adopting Real-time PCR method to detect m RNAexpression level of ERK1/2,hn RNP E1,c-Fos and c-Jun;using Western blot method to detect protein expression of ERK1/2,hn RNP E1,c-Fos and c-Jun.Using SPSS17.0 to conduct t test ahd F test.Results:1.The comparison of general biological characteristics of Siha and C33A:① Except for 24 hours,there are significant differences between the two cells in terms of the remaining number at each time point(P<0.05);②For Siha cells,there is higher proliferation index(PI)(P<0.001)when compared with C33 A cells.③The apoptosis rate is higher in C33 A cells than those in the Siha cells(P=0.024).2.(1)Comparison of expression of ERK1/2 genes m RNA and protein: For Siha cells,there are less relative expression of ERK1 m RNA when compared with C33 A cells(t=-17.489,P<0.001),there was no statistically significant difference between two kinds of cells in terms of ERK2 m RNA relative expression(t=1.003,P=0.372);ERK1/2and p-ERK1/2 protein expression levels of Siha cells were higher than C33 A cells,and the differences were statistically significant(ERK1/2:t=16.309,P<0.001;p-ERK1/2:t=4.776,P=0.028).(2)Comparison of m RNA and protein expression of hn RNP E1 genes:hn RNP E1 m RNA relative expression of Siha cells were higher than C33 A cells(t=3.376,P<0.001),Siha cells hn RNP E1 protein relative expression was higher than C33 A cells(t=4.821,P<0.001).(3)Comparison of m RNA and protein expression of c-Fos genes :For Siha cells,the c-Fos m RNA relative expression were significantly higher than those of C33 A cells(t=3.590,P=0.013).(4)Comparison of m RNA and protein expression of C-Jun genes:c-Jun m RNA relative expression amount of Siha cells were higher than C33 A cells(t=4.713,P<0.001);phosphorylation c-Jun protein expression levels of Siha cells were higher than C33 A cells,and the difference was statistically significant(t=10.209,P<0.001).3.Effect of ERK on Siha and C33 A cervical cancer biological characteristics:(1) the evaluation of ERK inhibitory effect:C33A cells ERK2 m RNA inhibition was 56.89 ±3.08%, higher than Siha cells54.08 ± 4.05%, but the difference was not statistically significant(t = 1.004, P = 0.107), C33 A cell ERK1 / 2 protein inhibition rate of 63.16 ±7.24%, higher than 51.32 ± 6.51% Siha cells(t =-4.281, P = 0.007), two kinds of cells have reached a good inhibitory effect.(2) the ERK impact on Siha and C33 A cervicalcancer cell cycle:compared with control group, the two kinds of cell proliferation index in intervention group were lower,(Siha:t=6.863,P<0.001;C33A:t=0.001,P<0.001),Siha cells intervention group proliferation index is higher than C33 A cells in the intervention group, and the difference was statistically significant(t=7.066, P<0.001).(3) ERK effect on Siha and C33 A cervical cancer cell apoptosis: compared with control group, the two kinds of cell apoptosis rate in intervention group were increased, and the differences were statistically significant(Siha:t=-5.201,P=0.006;C33A:t=-4.335,P=0.005).4.ERK’s impact on the hn RNP E1, c-Fos, c-Jun gene expression of cervical cancer cells Siha and C33A:(1) compared with control group, hn RNP E1 m RNA relative expression of the Siha cells intervention group decreased, and the difference was statistically significant(t=2.407,P<0.001);Siha cells in the intervention group hn RNP E1 protein expression levels were lower than the control group(t=2.539,P=0.004).(2) For control group, the c-Fos m RNA relative expression were significantly lower than those in the intervention group(t=4.129, P <0.001),compared with control group, p-c-Fos protein expression levels of the two kinds of cells in the intervention group were lower, and the differences were statistically significant(Siha: t=2.753, P=0.038; C33A: t=2.547, P=0.041).p-c-Fos protein expression levels of Siha cells in the intervention group were higher than C33 A cells in the intervention group(t=2.910, p<0.001).(3)For Siha cells control group, there are less c-Jun m RNA relative expression when compared with intervention group(t=4.037, P<0.001);c-Jun protein expression level of Siha cells in the intervention group decreased, and the difference was statistically significant(t=2.059,P=0.017).c-Jun protein expression levels of Siha cells in the intervention group was below those of C33 A cells in the intervention group,and the difference was statistically significant(t=-2.917, P<0.001);for control group,Siha cells intervention group p-c-Jun protein expression levels were significantly lower than those of intervention group(t=4.209,p<0.001),Siha cells intervention group p-c-Jun protein expression levels were higher than C33 A cells in the intervention group(t=3.106, p=3.106),Siha cell inhibition rate of p-c-Jun protein in C33 A cells(t = 11.734, P <0.001).Conclusions:1.ERK activation promote cervical cancer proliferation, reduce the role of apoptosis,ERK activation and the presence of HPV infection may promote synergy in cervicalcancer cell growth process.2.ERK activation may increase the expression of hn RNP E1 cervical cancer cells,thus contributing to the growth of cervical cancer, HPV infection may be involved in the regulation of hn RNP E1 role of ERK.3.ERK activation can activate c-Fos, c-Jun biological activity of effector proteins to promote its phosphorylation, thus contributing to the proliferation of cervical cancer,HPV infection may be involved in the ERK activation of c-Fos, c-Jun in.