冬凌草甲素逆转耐吉西他滨胰腺癌PANC-1／Gem细胞耐药性的研究

目的分析冬凌草甲素对耐吉西他滨胰腺癌PANC-1／Gem细胞的影响，探讨其逆转耐药的机制。方法诱导建立耐吉西他滨胰腺癌PANC-1／Gem细胞；CCK-8法检测吉西他滨对人胰腺癌PANC-1／Gem及PANC-1细胞增殖的影响；采用Chou—Talalay计算药物联合指数（CI）；Westernblot检测干预前后耐药相关蛋白的表达水平建立皮下异种移植瘤模型，观测不同药物组别对肿瘤生长情况的影响；免疫组化法检测不同组别移植瘤中相关耐药蛋白的变化差异。结果PANC-1／Gem细胞较PANC-1细胞对吉西他滨的敏感性降低（P〈0．01）；冬凌草甲素能明显抑制胰腺癌细胞增殖；冬凌草甲素与吉西他滨具有协同抗肿瘤作用；与空白组和吉西他滨组比较，冬凌草甲素及联合组中MRP1和GST—π表达量降低；冬凌草甲素对移植瘤具有抑制作用且能够降低移植瘤中MRP1蛋白的表达。结论冬凌草甲素在体内外能够抑制人耐吉西他滨胰腺癌PANC-1／Gem细胞的增殖，通过降低MRP1及GST-π耐药蛋白的表达逆转PANC-1／Gem对吉西他滨的耐药，增强其对吉西他滨的敏感性。

Reversal of the phenotype by K-ras^(val12) silencing mediated by adenovirus-delivered siRNA in human pancreatic cancer cell line Panc-1

AIM: To investigate the in vitro antitumor effect of adenovirus-mediated small interfering RNAs (siRNAs) on pancreatic cancer and the associated mechanism. METHODS: A 63-nucleotide (nt) oligonucleotide encoding K-rasval12 and specific siRNA were introduced into pSilencer 3.1-H1, then the H1-RNA promoter and siRNA coding insert were subcloned into pAdTrack to get plasmid pAdTrackH1-Avasval12. After homologous recombination in bacteria and transfections of such plasmids into a mammalian packaging cell line 293, siRNA expressing adenovirus Adh1-K-rasval12 was obtained. Stable suppression of K-rasval12 was detected by Northern blot and Western blot. Apoptosis in Panc-1 cells was detected by flow cytometry. RESULTS: We obtained adenovirus AdHl-K-rasval12 carrying the pSilencer 3.1-H1 cassette, which could mediate gene silencing. Through siRNA targeted K-rasval12, the oncogenic phenotype of cancer cells was reversed. Flow cytometry showed that apoptotic index of Panc-1 cells was significantly higher in the AdH1-K-rasval12-treatment group (18.70% at 72 h post-infection, 49.55% at 96 h post-infection) compared to the control groups (3.47%, 3.98% at 72 and 96 h post-infection of AdH1-empty, respectively; 4.21%, 3.78% at 72 and 96 h post-infection of AdHl-p53, respectively) (P<0.05). CONCLUSION: These results demonstrate that adenoviral vectors can be used to mediate RNA interference (RNAi) to induce persistent loss of functional phenotypes. In gene therapy, the selective down-regulation of only the mutant version of a gene allows for highly specific effects on tumor cells, while leaving the normal cells untouched. In addition, the apoptosis of pancreatic cancer cell line Panc-1 can be induced after AdH1-K-rasval12 infection. This kind of adenovirus based on RNAi might be a promising vector for cancer therapy.

THE AUTOCRINE REGULATORY EFFECT OF VASOACTVE INTESTINAL PEPTIDE ON THE GROWTH OF HUMAN PANCREATIC CARCINOMA CELLS

In the present study, the effects of VIP on the growth of two human pancreatic carcinoma cell lines PU-PAN-l and PANC-I were determined using tritiated thymidine incorporation. VIP receptors. intracellular cAMP and polyamines were investigated. The results indicated that VIP at a concentration of 10-8 mol/L to 10-7 mol/L can significantly stimulate the growth of PU-PAN-I cells but not PANC-1 cells. This effect is dose-dependent and abolished by VIP receptor antagonist, [4-C1-Phe6 . Leu17] VIP, suggesting VIP receptors in PU-PAN-I cells may mediate this effect. VIP can markedly elevate the levels of intracellular cAMP and polyamines in PU-PAN-I cells.indicating that the growth-promoting effect stimulated by VIP may be via a rapid increase in the biosyntheses of cAMP and polyamines. In addition, the VIP-antibody inhibited the growth of PU-PAN-I cells in serum-free culture mediurn. The results above suggested that VIP has an autocrine regulatory effect on this pancreatic carcinoma cell line (PU-PAN-1).

去甲斑蝥素对人胰腺癌PANC-1细胞增殖及蛋白激酶B蛋白的影响

目的观察去甲斑蝥素（NCTD）对人胰腺癌PANC-1细胞增殖及蛋白激酶B（Akt）蛋白的干预效应。方法实验分NCTD组和对照组，分别应用噻唑蓝比色法（MTY）和免疫组织化学法测定NCTD对人胰腺癌PANC-1细胞杀伤效应和对Akt蛋白表达的影响。结果NCTD在浓度为4μg／L作用24h时，即对PANC-1细胞增殖有抑制作用，抑制率达到35％以上，且随作用时间的延长、药物浓度升高，呈剂量-时间效应关系。与对照组比较，实验组Akt蛋白表达的阳性棕色反应颗粒明显减少（P〈0．05）；实验组16μg/L浓度下胞质的棕色颗粒表达比1μg／L减少更为明显（P〈0．05）。结论NCTD能抑制人胰腺癌PANC-1细胞的生长和Akt蛋白的表达。