Study On Expression Of Cyclin Kinase Subunit1(Cks1) In Human Glioma And Its Significance

ObjectiveHuman gliomas are by far the most commonly intracranial malignant tumors in thecentral nervous system.Up to now, the traditional treatment of this neoplasm was operationand chemotherapy and radiation therapy. Gliomas can not be cured because of tumor cellproliferation and aggressive charactic. Gene therapy acts as a new way to treat glioma andstudies were focused on particular genes and proteins,in order to find tumor targets toinhibit grown or eliminate its body. This is not muture and need further study.Findingtumor target of gliomas and investigating the theories of tumor development andprogression have become a hot point.The aim of this study is to exam the protein level ofcyclin kinase subunit1(Cks1) in human gliomas,together with collecting the clinical datas.then analysis the relationship between Cks1expression、tumor grade and prognosis. Nextan over-expression Cks1lentiviral vector named LV5（EF1-GFP/Puro）was build up byPCR. In A172glioma cell line,cell cycle phase were analyzed in order to explore thepreliminary function of Cks1in tumor initiation and progression.May be we could find anew biomarker for glioma treatment.MethodImmunohistochemical staining and Western blot were used to examine the proteinexpression level of Cks1、Skp2、p27in human glioma samples and normal brainslices.Together with follow-up data,the clinical pathological features also wereanalysed.Then an over-expression Cks1lentiviral vector named LV5（EF1-GFP/Puro）was build up. The Cks1protein level were analysised by Western blotting aftertransinfected glioma cell line A172. Flow cytometry analysised the cell cycle phase in theglioma cell line A172in order to explore the preliminary function of Cks1in gliomasdevelopment and progression.ResultsThe expression of Cks1、 Skp2and p27were located at nucleus. The positive ratesof Cks1expression in normal brain tissue、 low grade gliomas and high grade gliomaswere58.54±15.66%、31.25±27.50％and19.62±18.75％. Skp2protein was negative inall10cases of normal brain tissues，and the positive rates of Skp2expression in low and high grade gliomas were6.97±5.50%and17.51±17.68％,it increased in amalignancy-dependent manner. The positive rates of p27expression in normal braintissue、 low grade gliomas and high grade gliomas were85.50±7.17%、51.11±19.87％and23.78±17.53％, p27expression was decreased as the tumor gradedevelopment,respectively.There were significant differences in the expression levels ofthis three proteins among normal brain tissues，low and high grade gliomas(p<0.05)．ByWestern blotting experiment we can see Cks1protein expression in normal brain tissue、low grade gliomas and high grade gliomas were6.43±1.03、3.61±1.45and1.21±0.87,also there were significant differences in this three sections.Significantly positivecorrelation was found only between expression of Cks1and p27(r=0.38, p <0.05). Therewere no significant correlations between expression of Cks1and Skp2（r=0.08，p>0.05）,together with the expression of p27and Skp2（r=-0.16，p>0.05）.Cox univariateregression analysis shew low tumor grade、total resection rate、low Skp2expression andhigh p27expression were associate with better overall survival rates(p <0.05),but Cks1expression had no significant difference with glioma survival (p>0.05). Further study usedCox multivariate analysis in high grade glioma layer revealed age bellow55years old、total tumor removal and high expression of Cks1can have better prognosis.Anover-expression Cks1lentiviral vector named LV5（EF1-GFP/Puro）was build upsucceedly by PCR. Western blot did show Cks1expression increase highly in glioma cellline A172after transinfection（5.77±0.92vs2.46±0.12,p<0.05）. Flow cytometry analysisthat tumor cells mostly concentrate on G2/M phase. Significantly difference was foundbetween cells transinfected with LV5-Cks1and cells transinfected with LV5-NC gliomacell line A172(38.13±0.72vs15.23±0.33，p<0.05).ConclusionThe expression of Cks1was located at nucleus. Cks1expression decreased withglioma grade progression. Significantly positive correlation was found between expressionof Cks1and p27in gliomas. High expression of Cks1can predict better prognosis in highgrade glioma.Cks1may inhibit glioma progression through G2/M cell cycle checkpoint.It is necessary that we should make further studies to explore the exact function ofCks1in future.