Study Of The Effect And Mechanism Of Arsenic Trioxide Combined With Gemcitabine On EL-4Cell

Objective:T cell lymphoma (T-cell lymphoma, TCL) is a group ofheterogeneous diseases with unique histopathologic types and clinicalcharacteristics,whose incidence is not exactly the same at home andabroad. The incidence of TCL takes25%~35%in non-hodgkin’slymphoma (Non-Hodgkin ’s lymphoma, NHL) in our country, and theoverall prognosis of TCL is poor than B lymphoma cells.Although thetreatment level has been improved in recent years,5-year survival rate formost patients with TCL is still less than30%. To enhance TCL efficacyand prolong patient survival are still hot topics to study for scholars athome and abroad. Arsenic trioxide (Arsenic Trixide, ATO) made theexact curative effect in acuting promyelocytic leukemia (acutepromyelocytic leukemia, APL). In recent years, it has been studied andfound that antitumor effect also exsit in some solid tumors, meanwhileGemcitabine (Gemcitabine, GEM) has been applied to clinical as a NHLsecond-line chemotherapy drugs, therefore, the ATO and GEM arechosed as the study drug in this experiment, it is observed that the studydrug’s impact on proliferation and apoptosis of murine T-cell lymphomacell line EL-4cell, and whether the two-drug combination has a synergistic effect, to provide the theoretical basis for treatment of T-celllymphoma with ATO combined with GEM.Methods:①It was detected that EL-4cell’s Inhibition of proliferation, drugConcentration draw and time effect curve drawing impacted by ATOalone trioxide and GEM alone via MTT method, Impact on EL-4cell wasobserved by chosing drug Concentration and time of curve drawing ofATO combined with GEM.②EL-4cell apoptosis rate was determinedby flow cytometry (flow cytometry, FCM) when ATO and GEM are usedrespectively and together.③cell proliferation inhibition rate wascalculated: inhibition rate=(1－A value/controls A value)×100%.ToCalculate whether the two drugs have a synergistic effect. according tothe literature[46], it is assumed that the growth inhibition rate of the twodrugs were A and B, and A＞B. Based on the maximization model, thetwo drugs combined effects of the expected growth inhibition rate of Cshould be equal to A. If C＜A, the two drugs antagonism; if C＞1－(1－A)×(1－B), the two drugs synergies; C additive effect in between.Statistically using SPSS17.0software for data analysis, the results asx±s; each group were compared using the t-test, P＜0.05was consideredstatistically significant, P＞0.05for the difference was not statisticallysignificant. Results:(1) EL-4cell were cultured and treated with different concentrationsof ATO for24h、48h、72h.The results showed ATO could effectivelyinhibit EL-4cell in a time-dependent manner, at the same concentrationof ATO for24h、48h、72h cell inhibition rate comparison (P＜0.05), hastime-dependent manner. Moreover, The inhibitory effect of ATO(0.625μmol/L~10μmol/L for24, and72hours) could effectively inhibitEL-4cell in a dose-dependent manner.(2) GEM EL-4cell was treated with concentration of GEM0.01μg/ml、0.1μg/m1、1μg/ml、10μg/ml、100μg/ml,respectively.Theresults showed that the GEM concentration above1ug/ml for48h cellinhibition higher more apparent than24h, between24h~72h,the inhibitionrate were enhanced along with the increase of treatment time,exceptconcentration of GEM0.01ug/ml, the other concentration GEM24h,48hand72h cell inhibition rate between were statistically significant (P＜0.05). GEM concentration0.01ug/ml and0.1ug/ml for24h under theconditions of weak inhibition of cell,48h and72h inhibition significantlyenhanced; higher than1ug/ml GEM for24h with increasing drugconcentration, inhibition gradually enhanced，48h and72h with the GEMconcentration increases, the cell inhibition rate without a correspondingincrease.(3) To investigate the effect ATO combination with GEM,EL-4cells was treated with ATO at the concentration of for0.15625μmol/L、 0.3125μmol/L、0.625μmol/L combination with GEM the concentrationfor0.1μg/ml and1μg/ml,respectively，for24h、48h and72h.The resultsshowed:①There are significantly inhibit proliferation that theconcentration of three groups ATO Combination with GEM theconcentration for0.1μg/ml or GEM the concentration1μg/mg for24h,The differenc was statistically significant（P＜0.05）,Compared withsingle ATO group, Although not statistically significant, proliferation ofEL-4inhibited significant.Compared with single ATO group,combinationof concentration of ATO group and GEM0.1μg/ml has similar effect.②combination of concentration of ATOgroup and GEM group for48h,inhibited significant the statistically significantly difference the (P＜0.05);ATO incombination GEM1μg/ml group the proliferation inhibition rateswere significantly higher than GEM1μg/ml group, the ATO concentrationof0.15625μmol/L and0.625μmol/L combined group difference wassignificant (P＜0.05).③combination of concentration of ATO groupand GEM group for72h,the proliferation inhibition rates weresignificantly higher combination of three group concentration of ATOand GEM concentration of0.1μg/ml and1μg/ml than ATO alonerespectively,inhibited significant the statistically significantly differencethe (P＜0.05); the proliferation inhibition rates were significantly higherthan GEM1μg/ml group, the differences were significantly (P＜0.05).④combination ATO with GEM, the inhibition rate were enhanced alongwith the increase of treatment time,the proliferation of EL-4inhibitedsignificantly, compare with24h,48h and72h, the differences were significantly (P＜0.05).⑤EL-4cells was treated with ATO at theconcentration of for0.15625μmol/L、0.3125μmol/L、0.625μmol/Lcombination with GEM the concentration for0.1μg/ml and1μg/ml，respectively， under the conditions of the calculation found that ATO theconcentration24h growth inhibition rate is synergistic,48h is additiveeffect,72h Neither synergistic nor additive.⑥Through the FCM detectiondrugs the role of48h rate apoptosis of EL-4cells,ATO and GEM alonecan induce apoptosis of EL-4cells apoptosis0.3125μmol/L、1.25μmol/Lcombined with the GEM0.1μg/ml and1μg/ml,respectively, rate apoptosishigher than the ATO and GEM alone.Conclusion:(1) ATO alone or GEM alone can restrain proliferation of the EL-4cells in vitro, In a certain condition of concentration and time, Inhibitionof proliferation depends on time and concentration in evidence.(2) More inhibitory effect on EL-4cells was showed when ATO wascombined with GEM48h and72h than ATO alone or GEM alone, In acertain condition of concentration and time, it is proved to be synergy oradditive effect.(3) ATO alone and GEM alone or combined can induce in EL-4cellsapoptosis，and at lower concentration, ATO combined with GEM showedmore inhibitory effect on EL-4...