Relationship Between SENP1and Human Gliomas Progression And Its Molecular Action Mechanism

Objective: To detect the difference of SENP1mRNA expression levelbetween the normal control and different grade of astrocytic glioma samples，analyze the relationship between SENP1and human gliomas progression, theknockdown of SENP1expression in U87MG cell by RNA interference, tostudy the influence on proliferation and apoptosis of glioma cell, as well as itsprobablely molecular mechanism.Methods: Real time-quantitative PCR(RT-qPCR) was used to analyze thedifference of SENP1mRNA expression level between the normal control(3cases) and astrocytic glioma sample(28cases), and the difference of SENP1atprotein level between the normal tissue and various grades of the tumor wasdetected by protein blotting. The siRNA specific for SENP1was synthesizedand transfected into U87cells. U87MG cells were inoculated in the6-wellplate respectively according to2.5×105cells/well in2ml，after transfectionwith si-NC(control group), si-1and si-2for12hrs,Lipofectamine2000(Invitrogen,USA) was used to transfect the cells when thecell fusion degree reached to over90%, and the cells were incubatedcontinually for48hrs for the detection at both mRNA and protein level. Theknockdown of SENP1expression in U87MG cells, the number of apoptoticcells was estimated by detecting the cell surface phosphotidyl serine(PS) usingdouble staining with Annexin V-FITC/Propidium Iodide(PI). At last, theknockdown of SENP1expression in U87MG cells,to decrease the proteinexpression level of IκBα, p65, bcl-2and p38, as well as IκBα and p38phosphorylation level by protein blotting.Results: It was found that the mRNA expression level of SENP1inhuman brain astrocytic glioma was significantly raised in31cases of braintissue samples collected by qPCR detection in comparison with normal brain tissue (P<0.05) and increased with the increase of malignancy degree of thetumor, especially much more significantly in the higher grades(gradeIII,VI,WHO) of astrocytic glioma(P<0.01). The difference of SENP1atprotein level between the normal tissue and various grades of the tumor wasdetected by protein blotting, the result was similar with the changes at mRNAlevel,.After transfection, the expression level of SENP1mRNA wassignificantly decreased(P<0.01),and the protein content of SENP1was alsoobviously reduced. By analysis of the cell apoptosis, it was found that the cellapoptosis of si-1and si-2transfected U87MG group was notably increasedand significantly different from that of the si-NC control group(P<0.01), Theknockdown of SENP1expression in U87MG cells, and analysis of the proteinexpression level of IκBα, p65and bcl-2demonstrated that IκBαphosphorylation level was decreased significantly in comparison with thecontrol group and the expression of its downstream target factor p65andanti-apoptotic protein bcl-2was thus decreased p38phosphorylation levelafter kocking-down SENP1expression in U87MG cells was not as clear as theabove-mentioned pathways.Conclusion:1the expression difference of SENP1in the brain glioblastoma sampleswas closely related to the genesis and progress of the tumor, and can be usedas the important indication for clinical surveillance and prognosis.2SENP1can influence the expression of its downstream gene bcl-2through regulating NF-κB pathway-mediated anti-apoptosis route bydeSUMOylation, further modulating the tumor cell apoptosis or survival, so asto further reveal a new possible molecular mechanism of SENP1acceleratingthe genesis and progress of brain glioma.