mtt-formazan and Pancreatic-Neoplasms

Zinc finger RNA binding protein (ZFR) is involved in the regulation of growth and cancer development. However, little is known about ZFR function in pancreatic cancer.. Herein, to investigate whether ZFR is involved in tumor growth, Oncomine microarray data was firstly used to evaluate ZFR gene expression in human pancreatic tumors. Then short hairpin RNA (shRNA) targeting ZFR was designed and delivered into PANC-1 pancreatic cancer cells to knock down ZFR expression. Cell viability, cell proliferation and cell cycle analysis after ZFR knockdown were determined by MTT, colony forming and FACS, respectively. In addition, cell migration and invasion were assessed using the Transwell system.. The expression of ZFR was significantly higher in pancreatic tumors than normal pancreas tissues by Oncomine database analysis. Knockdown of ZFR by shRNA-expressing lentivirus significantly decreased the viability and invasion ability of pancreatic cancer cells. Moreover, FACS analysis showed that knockdown of ZFR in PANC-1 cells caused a significant cell cycle arrest at G0/G1 phase. Furthermore, knockdown of ZFR decreased the levels of CDK2, CDK4, CyclinA and CyclinD1 and enhanced the expression of p27, which has evidenced by qRT-PCR and Western blot analysis.. Knockdown of ZFR might provide a novel alternative to targeted therapy of pancreatic cancer and deserves further investigation.

Topics: Animals; Blotting, Western; Cattle; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cells, Cultured; Flow Cytometry; Formazans; Gene Knockdown Techniques; Humans; Lentivirus; Molecular Targeted Therapy; Neoplasm Invasiveness; Pancreatic Neoplasms; Real-Time Polymerase Chain Reaction; RNA-Binding Proteins; RNA, Small Interfering; Tetrazolium Salts

To examine STI-571-induced radiosensitivity in human pancreatic cancer cells in vitro.. Three human pancreatic cancer cell lines (Bxpc-3, Capan-1, and MiaPaCa-2) exhibiting different expression levels of c-Kit and platelet-derived growth factor receptor beta (PDGFRbeta) and showing different K-ras mutation types were used. For evaluation of the antitumor activity of STI-571 in combination with radiation, clonogenic survival assays, Western blot analysis, and the annexin V/propidium iodide assay with microscopic evaluation by 4',6-diamidino-2-phenylindole were conducted.. Dramatic phosphorylated (p)-c-Kit and p-PDGFRbeta attenuation, a modest dose- and time-dependent growth inhibition, and significant radiosensitization were observed after STI-571 treatment in view of apoptosis, although the levels of growth inhibition and increased radiosensitization were different according to cell lines. The grades of radiosensitivity corresponded to the attenuation levels of p-c-Kit and p-PDGFRbeta by STI-571, particularly to those of p-c-Kit, and the radiosensitivity was partially affected by K-ras mutation in pancreatic cancer cells. Among downstream pathways associated with c-Kit or PDGFRbeta, p-PLCgamma was more closely related to radiosensitivity compared with p-Akt1 or p-extracellular signal-regulated kinase 1.. STI-571 enhances radiation response in pancreatic cancer cells. This effect is affected by the attenuation levels of p-c-Kit or p-PDGFRbeta, and K-ras mutation status. Among them, p-c-Kit plays more important roles in the radiosensitivity in pancreatic cancer compared with p-PDGFRbeta or K-ras mutation status.

Topics: Analysis of Variance; Apoptosis; Benzamides; Cell Line, Tumor; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Extracellular Signal-Regulated MAP Kinases; Formazans; Genes, ras; Humans; Imatinib Mesylate; Mutation; Pancreatic Neoplasms; Phospholipase C gamma; Phosphorylation; Piperazines; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-kit; Pyrimidines; Radiation Tolerance; Radiation-Sensitizing Agents; Receptor, Platelet-Derived Growth Factor beta; Tetrazolium Salts; Thiazoles; Time Factors

Although the physiological actions of many herbs are gradually being elucidated at the molecular level, it remains unclear how individual components of herbs contribute to their biological activities. In the present study, the antiproliferative activity of Coptidis rhizoma, a medicinal herb, and the major component berberine was investigated in 8 human pancreatic cancer cell lines. Gene expression patterns associated with sensitivities to each agent were analyzed with oligonucleotide arrays that comprised approximately 11,000 genes. We used a tetrazolium dye (MTT) assay to determine ID(50) values after the 8 cell lines were exposed to the 2 agents for 72 hr. The ID(50) value for berberine was correlated positively with that for C. rhizoma (r=0.725, p=0.0401). C. rhizoma killed tumor cells more effectively than purified berberine when normalized to the level of berberine present in the herb. From the oligonucleotide array data, we selected 20 and 13 genes with strong correlations (r(2)>0.81) to ID(50) values for berberine and C. rhizoma, respectively. Among these 33 genes, the levels of expression of 12 were correlated with the ID(50) values of both agents, suggesting that these genes are associated with tumor-killing activity of berberine in C. rhizoma. Expression of the remaining 21 genes was correlated with the ID(50) value of either purified berberine or C. rhizoma. Thus, we identified common and distinct genes responsible for anti-proliferative activities of purified berberine and C. rhizoma. This strategy may improve our understanding of the actions of herbs with antitumor activities.

Topics: Antineoplastic Agents; Berberine; Biomarkers, Tumor; Cell Division; Coptis chinensis; DNA Primers; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Drugs, Chinese Herbal; Flow Cytometry; Formazans; Gene Expression Profiling; Humans; Neoplasm Proteins; Oligonucleotide Array Sequence Analysis; Pancreatic Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; RNA Probes; Tetrazolium Salts; Tumor Cells, Cultured

Retinoids are natural differentiation-inducing compounds that are promising as anticancer agents. Cancer cell lines are valuable in the investigation of the potential of retinoids for the treatment of specific cancers. However, using different treatment conditions but the same cell lines, investigators have produced markedly contradictory results for the effectiveness of retinoids. The present study examined different factors in the treatment conditions that may have masked or interfered with the effects of retinoids and, thereby, resulted in this conflict. Our studies revealed that the effects of retinoids on cancer cell proliferation were influenced by serum, the choice of vehicle (DMSO vs ethanol) and its concentration, phenol red, the degree of cellular confluence, and the method of assessing proliferation (cell number or [3H]thymidine uptake vs the MTT assay). Optimized conditions were the use of serum-free, ethanol-free, and phenol red-free media, investigating cells in the log phase of growth, using

Topics: Adenocarcinoma; Antineoplastic Agents; Carbonic Anhydrases; Cell Count; Cell Differentiation; Cell Division; Culture Media, Serum-Free; Dimethyl Sulfoxide; DNA; Dose-Response Relationship, Drug; Ethanol; Fixatives; Formazans; Freezing; Humans; Light; Pancreatic Neoplasms; Phenolsulfonphthalein; Retinoids; Solvents; Tetrazolium Salts; Tumor Cells, Cultured