Effect Of Bortezomib And Arabinoside On Proliferation And Apoptosis Of Jurkat Cell

Objective：Non-Hodgkin lymphoma (NHL) is one of the commonmalignant tumors,which includes B-cell lymphoma and T-cell lymphoma.Compared with B cell lymphoma, the invasion of T cell lymphoma is morepowerful, the prognosis is even worse, the survival period is shorter.T celllymphoma is a serious threat to the health and life of human being. Todaythere is no standard first-line therapy for the treatment of T-cell lymphoma.The majority of patients have relapsed in a few months or years afterremission of the application of conventional chemotherapy, short remission.Increasing the dose of conventional chemotherapy drugs can make part of thepatients to ease the extension,but the side effects of the drugs are more serious.Conventional chemotherapy for late effect of T-cell lymphoma is poor, sofinding the ideal chemotherapy to improve the efficacy and prognosis of thedisease is very urgent.Bortezomib(BTZ) makes the treatment of hematologic malignancies fullof hope, bortezomib as a proteasome inhibitor is the first clinical application.The unique anti-tumor mechanism makes it has been a great deal of attentionin the treatment of hematologic malignancies, It is reported in the literaturethat bortezomib for NHL has a certain effect, but bortezomib combining withother traditional chemotherapy drugs is less reported. BTZ mainly decreasesthe level of NFκB or regulates cyclin and the signal apoptosis of tumor cell,and the anti-tumor effects of T-cell lymphoma is not very clear. To investmentthe effect of bortezomib(Bor) alone and in combination with cytarabine(Ara-c)on proliferation and apoptosis of lymphoma cell line Jurkat.we observecytarabine (Ara-C) or BTZ single and combined treatment of T-cell lymphomaJurkat cells in different ways, Analysis the effect of proliferation and apoptosisinduced by their combination, and its possible mechanism preliminary. Then can provide a theoretical guide to help us choose the better treatment regimenof T-cell lymphoma.Methods:1First of all, we used MTT assay to examine The effects of BTZ orAra-c with different concentrations on the growth of Jurkat cells wereexamined by MTT assay.2We used MTT assay to examine The effects of bortezomib combinedwith Ara-c on the growth of Jurkat cells were examined by MTT assay.3Then,we used flow cytometric analysis detection to examine Theeffects of BTZ or Ara-c with different concentrations and their combinationson the growth of Jurkat cells were examined by flow cytometric analysisdetection.4The RT-PCR to assay the mRNA expression of the NF-KB.5Synergetic effects of BTZ and Ara-c was analyzed by median-effectprinciple.Results:1Effect of BTZ alone or Ara-c on the proliferation of Jurkat cellsDates from MTT assays show that1-100nmol/L BTZ or0.05-5ug/ml treatedcells, inhibition rates enhanced in a time-and dose-dependently manner.(Fig.1,Fig.2, Table1, Table2).2Bortezomib on Jurkat cells for48h cell proliferation inhibition of50%of the drug concentration (IC50) is51.001nmol/L (Table8). Cytarabine onJurkat cells for48h cell proliferation inhibition of50%of the drugconcentration (IC50) is1.183ug/ml (Table8).3We selected a concentration of30nmol/L (less than48h IC50) ofbortezomib and a concentration of0.2ug/ml (less than48h IC50) ofcytarabine, Jurkat cells were treated with30nmol/L BTZ or0.2ug/ml Ara-c incombination for48h.The experiment is divided into six groups (1) Blankcontrol group: not a member of any drugs;(2) BTZ group: Jurkat cells weretreated with30nmol/L BTZ alone for48h;(3) Ara-c group: Jurkat cells weretreated with0.2ug/ml Ara-c alone for48h;(4) Combination therapy group a: Jurkat cells were treated with30nmol/L BTZ or0.2ug/ml Ara-c incombination for48h;(4) Combination therapy group b: Jurkat cells weretreated with0.2ug/mlAra-c pretreated for6h and then give30nmol/L BTZfor a total of48h;(6) Combination c: Jurkat cells were treated with0.2ug/mlAra-c pretreated for6h and then give30nmol/L BTZ for a total of48h; inhibition rates were (2±1)%,(44.95±4.9)%(42.09±2)%,(64.98±4)%,(87.69±2)%(55.44±5)%, The inhibition rates of Jurkat cells incombination groups were higher than those in the two single treatmentgroups(P<0.05),espically in the combined treatment group in which Jurkatcells were treated first with Ara-c for6h then with BTZ combined.(table-3Fig.3).4Flow cytometry results showed that single-agent bortezomib orcytarabine can be time-dose-dependent manner to induce apoptosis in Jurkatcells (Fig.6, Fig.7). Flow cytometry results showed that: blank control group,BTZ group of Ara-C group, the combination therapy group a combinationgroup b, c group apoptosis rate of the combination therapy (4.2±3)%,(35.5±5.3)%(36.6±6.6)%,(67.5±7.2)%,(84.2±4.1)%(55.3±5.15)%, andcombined treatment group compared with the monotherapy group differencewas statistically significant (P <0.05)(table-4Fig.4). The apoptosis rates ofJurkat cells in combination groups were higher than those in the twosingletreatmentgroups(P<0.05),espically in the combined treatment group inwhich Jurkat cells were treated first with Ara-c for6h then with BTZcombined..5By RT-PCR results show that NF-kB gene expression changes in Jurkatcells: RT-PCR results showed that: blank control group, the BTZ group Ara-cgroup, the combination of a group of the combination therapy group b, thejoint drug c of NF-kB gene expression levels were2.02±0.01,0.76±0.02,1.57±0.01,0.59±0.03,0.42±0.02,0.67±0.02. Three joint mode ofadministration of NF-kB expression than those in the monotherapy group,especially in the combination therapy group b was the lowest statisticallysignificant (P <0.05)(table7, Fig.8.) Test. Conclusion:1Bortezomib or cytarabine has a dose-and time-dependentantiproliferation and proapoptotic effect on Jurkat lymphoma cells invitro.The50%inhibitory concentration of bortezomib or cytarabinerespectively on Jurkat cells for48h,is51.001nmol/L and1.183ug/ml.2BTZ can effecttively inhibit Jurkat cell proliferation,and induced itsapoptosis.This effect was enhanced significantly when in combination withAra-c. The inhibition and apoptosis rates of Jurkat cells in combination groupswere higher than those in the two singletreatmentgroups(P<0.05),espically inthe combined treatment group in which Jurkat cells were treated first withAra-c for6h then with BTZ combined.Pretreatment of Jurkat cells with Ara-clead to the increased activity of NF-kB.Bortezomib is a treatment for T-celllymphoma which is very promising targeted drugs, bortezomib in combinationwith cytarabine therapy for T-cell lymphoma clinical studies provides anexperimental basis.