RESUMO
PURPOSE: Gemcitabine resistance is the main problem in pancreatic adenocarcinoma patients. Hence, we aimed to identify the correlation between expression of RRM1 and CDA as the resistance genes and their predicted targeting miR-608 in the resistant pancreatic cancer cell lines to gemcitabine. METHODS: Dual luciferase assay was performed to determine whether both RRM1 and CDA are targeted by miR-608 in 293T and pancreatic cancer cell lines. AsPC-1 and MIA PaCa-2 cell lines became gradually resistant to gemcitabine by exposing to the increasing doses of gemcitabine. After RNA and miRNAs extraction and cDNA conversion, the expressions of RRM1, CDA and miR-608 in all cell lines were studied by quantitative PCR. Pre-miR-608 transfection to the cell lines was done by calcium phosphate method. MTT assay was performed for analyzing the chemo sensitivity of different cell lines to gemcitabine. RESULTS: Luciferase assays showed that miR-608 targeted RRM1 and CDA genes in 293T, AsPC-1 and MIA PaCa-2 cell lines. Compared to parental cell line, resistant MIA PaCa-2 and AsPC-1 cells demonstrated increased expression of RRM1 and CDA. On the other hand the expression of miR-608 in resistant MIA PaCa-2 and AsPC-1 cells was lower than parental cells. Furthermore, transfection of MIA PaCa-2 and AsPC-1 cells by miR-608 lead to decreased expression of RRM1 and CDA and lowered viability of the cells in comparison with scrambled microRNA transfected cells. CONCLUSION: During resistance induction in pancreatic cancer cells, miR-608 which is targeting RRM1 and CDA is downregulated which leads to upregulation of these genes.
Assuntos
Adenocarcinoma/tratamento farmacológico , Antimetabólitos Antineoplásicos/farmacologia , Desoxicitidina/análogos & derivados , MicroRNAs/genética , Neoplasias Pancreáticas/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Citidina Desaminase/genética , Desoxicitidina/farmacologia , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Luciferases/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Ribonucleosídeo Difosfato Redutase , Transfecção , Proteínas Supressoras de Tumor/genética , Regulação para Cima , GencitabinaRESUMO
BACKGROUND: Multiple sclerosis (MS) is a multifactorial disease. Positive genetic background could predispose individuals to this chronic disabling disease. In order to investigate the role of some proinflammatory cytokines (interleukin (IL)-2, IL-12, and interferon-gamma (IFN-gamma)) as a risk factor for MS, this study was performed. METHODS: Two hundred and eleven patients with relapsing-remitting form of MS were enrolled in this study and compared with 359 healthy individuals. Using polymerase chain reaction based on sequence-specific primer method, the cytokine genes were amplified, and alleles and genotypes were detected on gel electrophoresis. RESULTS: Significant increases for IFN-gamma AT (+874) genotype (54.5% vs. 37.8%, p = 0.0002) and IL-12 AA (-1188) genotype (60.8% vs. 49.7%, p = 0.014) were found in MS patients in comparison with healthy controls. A significant decrease in IFN-gamma TT (+874) genotype (17.7% vs. 27.5%, p = 0.01) and IL-12 CA (-1188) genotype (30.9% vs. 45%, p = 0.001) in MS patients was also detected. No significant differences of IL-2 G/T (-330) and IL-2 G/T (+166) in alleles and genotypes were observed between MS patients and normal subjects. CONCLUSIONS: It could be suggested that the genetic variation in IL-12 A/C (-1188) and IFN-gamma A/T (+874) cytokine genes could be risk factors for MS patients.
Assuntos
Citocinas/genética , Esclerose Múltipla/genética , Polimorfismo Genético , Adulto , Estudos de Casos e Controles , Feminino , Genótipo , Humanos , Mediadores da Inflamação , Interferon gama/genética , Interleucina-12/genética , Interleucina-2/genética , Masculino , Polimorfismo de Nucleotídeo Único , Fatores de Risco , Adulto JovemRESUMO
Different research groups have extensively studied the associations of cytokine gene polymorphisms in different diseases. The role of cytokines gene polymorphisms in multiple sclerosis (MS), as a chronic Immune-mediated neurodegenerative disease, has been previously reported in the various populations. For determining pro-inflammatory cytokine gene polymorphisms, 100 relapsing remitting multiple sclerosis (RRMS) Iranian patients and 140 normal individuals as control enrolled in this study. DNA of each sample was extracted by a modified salting out method. Cytokine single gene nucleotide polymorphisms including IL-1alpha -889, IL-1beta (-511 and +3962), IL-1R pst1 1970, IL-1RA mspal 11100, and TNF-alpha (-308 and -238) were determined by using the PCR-SSP method. The results of our data indicate the decrease in frequency of IL-1alpha TC-889 genotype (p=0.002), IL-1beta TC +3962 genotype (p=0.004), IL-1R T pst1 1970 allele (p= 0.0001), IL-1 RA TC Mspa1 11100 genotype (p=0.009), TNF-alpha A-308 allele (p=0.0002) and AG genotype (p=0.00001) in the patients group versus normal subjects. On the other hand the frequency of IL-1alpha TT -889 genotype (p=0.028), IL-1R C pst1 1970 allele (p=0.0001) and CC genotype (p=0.00006), TNFalpha G -308 allele (p=0.0002) and GG genotype (p=0.000001) decreased significantly in the patients versus normal subjects.These results suggest that polymorphic variations of these pro-inflammatory cytokines may play an important role in susceptibility of Iranian multiple sclerosis patients.
