Experimental Study Of The Inhibitory Effect Of Curcumin On Nasopharyngeal Carcinoma

Introduction: Study the effect of curcumin and its combination with5-FU onnasopharyngeal carcinoma CNE-2Z cell growth inhibition and apoptosis, study itsmechanism, and analyze whether the two drugs have combined synergies, provide anexperimental basis for searching new ways of combination chemotherapy onnasopharyngeal carcinoma.Methods:1. Using different concentrations of curcumin in NPC CNE-2Z cells atdifferent times, test cell growth conditions by MTT method, test apoptosis ofnasopharyngeal carcinoma cells by TUNEL method and flow cytometry, study theeffect of curcumin on nasopharyngeal carcinoma CNE-2Z cell growth inhibition andapoptosis. Western blot test on Bcl-2, Bax, caspase-3, Fas, and NF-κB proteinexpression levels, Real time PCR test on caspase family including caspase-3,caspase-7, caspase-8and caspase-9mRNA expression level. Explore curcumin’smechanism on inhibiting the growth of nasopharyngeal carcinoma cells.2. Curcumin and5-FU’s individual and their combined effects onnasopharyngeal carcinoma CNE-2Z cells, test cell growth conditions by MTT method,test apoptosis of nasopharyngeal carcinoma cells by flow cytometry, study the effectof curcumin and5-FU combined effects on nasopharyngeal carcinoma CNE-2Z cellgrowth inhibition and apoptosis. Western blot test on Bcl-2, Bax, caspase-3, andNF-κB protein levels. Explore the combination mechanism of the two on inhibitingthe growth of nasopharyngeal carcinoma cells.Results：1. MTT test results showed that curcumin can inhibit the growth ofnasopharyngeal carcinoma CNE-2Z cells significantly, the effect was dependent oftime and dose. Using TUNEL method, we can observe under fluorescence microscope that the green fluorescence was released by apoptotic cells. With increasingconcentrations of curcumin, an increase in the number of apoptotic cells was observed.Test the apoptosis of CNE-2Z cells by flow cytometry, the results showed:0,50,100,and200μM of curcumin for48hours, the apoptosis rates were3.00±0.50%,2.70±0.20%,22.00±2.00%and52.80±4.00%. For100μM and200μM group, theapoptosis rate was significantly higher (P<0.01), while the apoptosis rate of50μMclose to the control group (P>0.05). Curcumin (50μM,100μM,200μM) for24hours,Western blot analysis showed that: The expression levels of Bcl-2protein in CNE-2Zcells and NF-κB protein in the nucleus and cytoplasm declined with the increasing ofthe drug concentration, Cleaved caspase-3protein and Bax protein levels increasedwith increasing of drug concentration, and Fas protein expression did not changesignificantly; Real time PCR analysis showed that: With the curcumin concentrationincreased, caspase-3, caspase-7, and caspase-9in the mRNA expression levelsgradually increased, while the mRNA expression of caspase-8level did notsignificantly change.2. Curcumin,5-FU and the two drug combination treatment on nasopharyngealcarcinoma CNE-2Z cells for72hours, MTT results showed that curcumin and5-FUcould significantly inhibit the growth of NPC cells, after the combination of two drugs,inhibition was significantly enhanced. Through calculating, the value of q of the twodrugs in combination is greater than1.15. It indicates that curcumin in combinationwith5-FU to inhibit the growth of NPC cells have synergies. The apoptosis rate of thetest results by flow cytometry showed: the control group, curcumin group,5-FUgroup and the drug combination group, the apoptosis rates were4.52±1.37%,28.50±4.16%,34.30±2.84%and72.80±3.34%individually. After application of twodrugs in combination, the tumor cell apoptosis was significantly higher than the singledrug group (p<0.01). It reveals that drug combination enhanced apoptosis effect.Western blot test shown that: curcumin group,5-FU group and the drug combinationgroup of expression of levels of Bcl-2protein in CNE-2Z cells and NF-κB protein the nucleus and cytoplasm decreased, levels of protein expression of Bax protein andCleaved caspase-3increased and protein levels of drug combination group has moresignificant changes.Conclusion：1. Curcumin can inhibit the growth of nasopharyngeal carcinomaCNE-2Z cells in vitro in dependence of dose and time. Curcumin realizes theinhibition on the growth of nasopharyngeal carcinoma CNE-2Z cell by inducingapoptosis of cell. By inhibiting the expression of anti-apoptosis gene Bcl-2, curcuminenhanced expression of pro-apoptotic gene Bax; and through the mitochondrialpathway and inhibition of NF-κB activity to induce apoptosis of CNE-2Z cells; thusto inhibit the growth of CNE-2Z cells.2. Curcumin and5-FU in synergistic inhibition of nasopharyngeal carcinomaCNE-2Z cells in vitro. By inducing apoptosis of cell, curcumin and5-FU combinedachieve inhibition of nasopharyngeal carcinoma CNE-2Z cell growth. The effect ofthe two drugs combination was better than that of single drug. Curcumin and5-FUcombined inhibited the expression of Bcl-2gene, increased expression of Bax gene.The inhibition role on NF-κB activity was significantly better than that of single drug.Drug combination inhibited the growth of nasopharyngeal carcinoma CNE-2Z cellsmay be realized by inhibiting the expression of Bcl-2gene, increasing the expressionof Bax gene, and inhibiting the induction of apoptosis on NF-κB activity. Curcuminmay enhance the effect on the sensitivity of nasopharyngeal carcinoma cells toradiotherapy and chemotherapy. It may be an effective way to add curcumin intocombination chemotherapy and concurrent radiotherapy&chemotherapy treatment ofnasopharyngeal carcinoma.