Study On The Resistance Mechanism Of Carcinoma Of Pacreas Induced By TRAIL

Objective Our experiments use different dose TRAIL and/or cisplatin treated the pancreatic cell line PANC-1 and PACA-2, and then detected the sensitivity after treated withTRAIL and/or cisplatin, analysed the resistance mechanisms of carcinoma of pacreas induced by TRAIL.Methods Our experiments use PACA-2 and PANC-1 cell lines as the research object. We devided TRAIL group into different dose (0.1 ng/ml,0.3 ng/ml,1ng/ml,3 ng/ml,10 ng/ml, 30 ng/ml,100 ng/ml,300 ng/ml), and then treated the cells with different concentrated TRAIL and/or cisplatin, detected the percentate of cell death; analyzed the expression of DR5, FADD, caspase-8, RIP, TRAF2, A20 protein; and study the expression caspase-8 cleavage; we got the PANC-1 A20-/- cell line by siRNA, and detected the sensitivity of A20-/- cell after treated with TRAIL.Results①Our results showed that PACA-2 was sensitive to TRAIL, we devided TRAIL to different group (0.1 ng/ml,0.3 ng/ml, lng/ml,3 ng/ml,10 ng/ml,30 ng/ml,100 ng/ml, 300 ng/ml), the apoptosis rate was 5.90%,17.39%,26.76%,48.97%,68.00%,70.06%, 72.81%,71.85% and 71.71% respectively, so we conclude that PACA-2 was sensitive to TRAIL. And in spite of 0.1 ng/ml, the others were sighhnificance when compared with control group(P<0.05).②We treated the PANC-1 cell line with different concentrated TRAIL, the apoptosis rate was 2.36%,4.15%,2.27%,5.10%,11.44%,21.26%,25.47%, 26.08%,26.56%, then compared with control group, from 10 ng/ml group was sighnificant(P<0.05), so the PANC-1 was resistant cell line to TRAIL. After that we choosed cisplatin and cotreated with different dose TRAIL to PANC-1 cell line, the apoptosis rate was 4.78%,7.51%,7.39%,8.79%,22.47%,27.22%,30.65%,47.65%. It suggested that the co-treatment group of cisplatin and TRAIL sighificantly kill tumor cells when compared with TRAIL alone group(P<0.05). Thus, PACA-2 had sighnificant apoptosis with TRAIL alone, PANC-1 was not, but in co-treatment group, after cisplatin treated to PANC-1, they can improve the sensitivity of TRAIL, and promoted the PANC-1 activity of killing tumor cells.③We detected the DR5, FADD, TRAF2, RIP,caspase-8 and A20 protein of PACA-2 and PANC-1 cell lines by Western blot, we found that A20 expression was so different. The PACA-2 expressed A20 weakly, but PANC-1 was overexpressed A20. We showed there was two steps hydrolysis after TRAIL alone or cotreatment with cislatin, the p10 and p18 fragments expressed on the memberance, suggested that the PACA-2 cell lines treated with TRAIL, or PANC-1 cell lines co-treated with TRAIL and cisplatin combination, and through the activity with caspase-8, triggered the apoptosis response.④We got the PANC-1 A20-/- cell lines by siRNA, and then detected the sensitity to TRAIL, the percentage of cell death was 8.53%,13.67%,19.16%, 24.33%,25.24%,31.51%,38.26%,42.52%,50.07%. The results showed that the percentage of apoptosis was significantly impoved after knock down A20 gene(P<0.05). So that, A20 overexpression has a role in PANC-1 resistance to TRAIL, after elimited A20, the sensitivity was much higher than before. Conclusions Besed on the above experimental results, we had a conlclusion, firstly, the PACA-2 and PANC-1 have a different response, the PACA-2 was a sensitive cell line to TRAIL, and PANC-1 was a resistant cell line. Secondly, TRAIL combined with cisplatin induce PANC-1 apoptosis by TRAIL pathway, they have sighificant synergies. Thirdly, the mechanism of resistance to TRAIL of PANC-1 cell lines would overexpress A20, otherwise DR5, FADD, TRAF2, RIP and caspase-8.