RESUMO
BACKGROUND: HeberFERON is a co-formulation of α2b and γ interferons, based on their synergism, which has shown its clinical superiority over individual interferons in basal cell carcinomas. In glioblastoma (GBM), HeberFERON has displayed promising preclinical and clinical results. This led us to design a microarray experiment aimed at identifying the molecular mechanisms involved in the distinctive effect of HeberFERON compared to the individual interferons in U-87MG model. METHODS: Transcriptional expression profiling including a control (untreated) and three groups receiving α2b-interferon, γ-interferon and HeberFERON was performed using an Illumina HT-12 microarray platform. Unsupervised methods for gene and sample grouping, identification of differentially expressed genes, functional enrichment and network analysis computational biology methods were applied to identify distinctive transcription patterns of HeberFERON. Validation of most representative genes was performed by qPCR. For the cell cycle analysis of cells treated with HeberFERON for 24 h, 48 and 72 h we used flow cytometry. RESULTS: The three treatments show different behavior based on the gene expression profiles. The enrichment analysis identified several mitotic cell cycle related events, in particular from prometaphase to anaphase, which are exclusively targeted by HeberFERON. The FOXM1 transcription factor network that is involved in several cell cycle phases and is highly expressed in GBMs, is significantly down regulated. Flow cytometry experiments corroborated the action of HeberFERON on the cell cycle in a dose and time dependent manner with a clear cellular arrest as of 24 h post-treatment. Despite the fact that p53 was not down-regulated, several genes involved in its regulatory activity were functionally enriched. Network analysis also revealed a strong relationship of p53 with genes targeted by HeberFERON. We propose a mechanistic model to explain this distinctive action, based on the simultaneous activation of PKR and ATF3, p53 phosphorylation changes, as well as its reduced MDM2 mediated ubiquitination and export from the nucleus to the cytoplasm. PLK1, AURKB, BIRC5 and CCNB1 genes, all regulated by FOXM1, also play central roles in this model. These and other interactions could explain a G2/M arrest and the effect of HeberFERON on the proliferation of U-87MG. CONCLUSIONS: We proposed molecular mechanisms underlying the distinctive behavior of HeberFERON compared to the treatments with the individual interferons in U-87MG model, where cell cycle related events were highly relevant.
Assuntos
Glioblastoma , Neoplasias Cutâneas , Humanos , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Apoptose , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Interferon-alfa/farmacologia , Anáfase , Interferon gama/farmacologiaRESUMO
Hepatitis C virus (HCV) is a significant global public health problem with >185 million infections worldwide. A series of genome-wide association studies (GWAS) has identified IL-28B polymorphisms as a predictor of sustained virologic response (SVR), as well as spontaneous clearance in chronic HCV genotype 1 patients. The objective of this work was to evaluate the prevalence of IL-28B rs12979860 and rs8099917 polymorphisms in Cuban chronic HCV patients. The study cohort included 73 chronic HCV patients treated with concomitant administration of CIGB-230 and nonpegylated IFN-α plus ribavirin (non-pegIFN-α/R) antiviral therapy. The genotype distribution of IL-28B rs12979860CC, -CT, and -TT was 29, 41, and 30%, respectively, and the distribution for rs8099917TT, -TG, and -GG was 63, 31, and 5%, respectively. The allele frequencies for rs12979860C and -T alleles were 51 and 49%, respectively, and for rs8099917G and -T alleles, the values were 21 and 79%, respectively. SVR rates were 55, 42, and 35% for rs12979860CC, -CT, and -TT, respectively, and 52, 30, and 25% for rs8099917TT, -GT, and -GG, respectively. The combined assessment of both single nucleotide polymorphisms (SNPs) resulted in 3 major genotypes (rs12979860CC/rs8099917TT, rs12979860CT/rs8099917TT, and rs12979860CT/rs8099917GG) with a frequency of 30.1, 21.9, and 20.5%, respectively. In patients with heterozygous variant rs12979860CT, the additional genotyping of rs8099917 contributed to increase the SVR rate. It is concluded that in Cuban HCV-infected patients, the responder homogeneous variant rs8099917TT is the most frequent genotype. The simultaneous genotyping of 2 IL-28B SNPs could improve the prediction of SVR contributing to better therapeutic decisions and treatment management.
Assuntos
Hepatite C Crônica/genética , Interleucinas/genética , Polimorfismo de Nucleotídeo Único , Adolescente , Adulto , Antivirais/uso terapêutico , Estudos de Casos e Controles , Estudos de Coortes , Cuba , Feminino , Frequência do Gene , Hepatite C Crônica/tratamento farmacológico , Humanos , Interferon-alfa/uso terapêutico , Interferons , Masculino , Pessoa de Meia-Idade , Ribavirina/uso terapêutico , Resultado do Tratamento , Vacinas de DNA/uso terapêutico , Vacinas contra Hepatite Viral/uso terapêuticoRESUMO
BACKGROUND: Recent studies suggest that celiac disease (CD) is common in many developing countries. Because the disease may be under diagnosed in Cuba, we studied the presence of the disease in a group of apparently healthy adult. AIMS/HYPOTHESIS: It was to assess for the first time, the presence of silent CD in a cohort of healthy Cuban adults individuals and to evaluate the tools for diagnosis of CD in this group. METHODS: A total of 200 healthy Cuban adult from Havana City were evaluated. Tissue transglutaminase antibodies (tTGA) were determined by one-step immunochromatographic assay and by commercial ELISA kit. CD specific human leukocyte antigen (HLA) typing was performed by polymerase chain reaction amplification, using sequence-specific primers. In the subject positive for tTGA, the CD was confirmed by intestinal biopsy. RESULTS: From the 200 studied individuals, only one subject was identified as positive by both assays, being submitted to duodenal biopsy. Morphological changes consistent with CD were found and also supported by HLA-DQ2 (HLA-DQA1*0501-DQB1*02). In the follow-up after one year, histological recovery was assessed by a second intestinal biopsy and the serological marker became negative. CONCLUSIONS: This study confirms the existence of silent CD among healthy adult in Cuba and highlights the importance of mass screening for this disease among them. The one-step immunochromatographic assay is a good tool for this purpose.
Assuntos
Doença Celíaca/diagnóstico , Doença Celíaca/epidemiologia , Saúde , Adolescente , Adulto , Idoso , Anticorpos/sangue , Anticorpos/imunologia , Doença Celíaca/dietoterapia , Doença Celíaca/imunologia , Estudos de Coortes , Cuba/epidemiologia , Dieta Livre de Glúten , Duodeno/patologia , Feminino , Proteínas de Ligação ao GTP/imunologia , Antígenos HLA-DQ/imunologia , Cadeias alfa de HLA-DQ , Cadeias beta de HLA-DQ , Humanos , Imunoglobulina A/sangue , Imunoglobulina A/imunologia , Masculino , Pessoa de Meia-Idade , Proteína 2 Glutamina gama-Glutamiltransferase , Transglutaminases/imunologia , Adulto JovemRESUMO
The extreme polymorphism found at some of the human leukocyte antigen (HLA) system loci makes it an invaluable tool for population genetic analyses. In the present study the genetic polymorphism of the Cuban population was estimated at HLA-A, -B, and -Cw loci by DNA typing. HLA class I allele and haplotype diversity were determined in 390 unrelated Cuban individuals (188 whites and 202 mulattos) from all over the country. In whites 19, 27, and 14 allele families for the HLA-A, -B, and -Cw loci, respectively, were identified. In mulattos, for the same loci, 20, 18, and 14 allele families were identified. Allele and haplotypes frequencies, comparisons with other worldwide populations based on genetic distances, neighbor-joining dendrograms, and correspondence analyses were estimated. Most of the identified allele groups and haplotypes are also common to sub-Saharan African and Europeans populations. However, Amerindian and Asian alleles were also detected at lower frequencies. The results clearly reveal the high diversity and interethnic admixture of the studied population. Our results provide useful information for the further studies of the Cuban population evolution and disease association in terms of HLA class I genes.
Assuntos
Antígenos de Histocompatibilidade Classe I/genética , Polimorfismo Genético , Alelos , Cuba , Frequência do Gene , Haplótipos , Humanos , FilogeniaRESUMO
BACKGROUND: The angiotensin converting enzyme (ACE) is a key protein of the renin angiotensin system, whose main function is the conversion of angiotensin I to II. ACE is involved in the physiological control of blood pressure and it is a candidate gene for essential hypertension in humans. We tested the relevance of the ACE insertion/deletion (I/D) polymorphism in our population. METHODS: We recruited 243 hypertensive and 407 normotensive subjects in the city of Havana, matched according to age, sex and ethnic group. The ACE (I/D) polymorphism was determined by the polymerase chain reaction (PCR) technique. The fit of genotype frequencies to Hardy-Weinberg proportions was evaluated in all groups analyzed. The possible association between the ACE I/D polymorphism and hypertension status was tested by chi2 and odds ratio tests. RESULTS: All groups but black female cases were in Hardy-Weinberg equilibrium. The frequencies of the D allele in hypertensive/normotensive subjects were 0.61/0.59 in white males, 0.58/0.58 in white females, 0.47/0.59 in black males and 0.58/0.54 in black females. The distribution of ACE genotypes differed significantly between cases and controls only in black women according to the additive model (chi2p=0.04) but the adjusted OR did not show significant association (OR 1.14 95% CI 0.62 to 2.10). CONCLUSION: The ACE I/D polymorphism was not associated with hypertension in our multiethnic sample. While the chi2 test for additive model in black women suggested a marginal significance, the adjusted OR did not show any significant association.
Assuntos
Hipertensão/genética , Peptidil Dipeptidase A/genética , Polimorfismo Genético , Adulto , População Negra , Cuba , Feminino , Genótipo , Humanos , Hipertensão/etnologia , Masculino , Pessoa de Meia-Idade , População BrancaRESUMO
Celiac disease (CD) susceptibility has been strongly associated with HLA-DQ2 and HLA-DQ8. The main objective of this study was to assess the distribution of HLA DQA1*0501 and DQB1*02 alleles (DQ2) for the first time in a group of Cuban celiac patients. We evaluated 22 patients, 54 first-degree relatives, and 60 controls for detection of antitissue transglutaminase (tTG)-specific antibodies in serum. We found that 100% of the probands and 19% of the first-degree relatives were positive for the antibodies in serum. We did not detect any specific response for the healthy control individuals. We observed a significant over-representation of DQ2 heterodimer, both in patients and relatives. In the group of patients, 86.3% were positive for DQA1*0501, 90.2% were positive for DQB1*02, and 86.3% were positive for both alleles. The frequencies in relatives and controls were as follows: 70%, 90%, and 70%; and 56.6%, 45%, and 20%, respectively. In conclusion, we found that the proportion of our celiac patients carrying DQ2 was similar to the proportion of CD patients reported in populations with different genetic backgrounds. These results underline the primary importance of HLA-DQ alleles in susceptibility to celiac disease.
Assuntos
Doença Celíaca/genética , Predisposição Genética para Doença , Antígenos HLA-DQ/genética , Adolescente , Adulto , Criança , Pré-Escolar , Cuba , Feminino , Cadeias alfa de HLA-DQ , Cadeias beta de HLA-DQ , Humanos , MasculinoRESUMO
Los leucocitos humanos fueron inducidos con Le-IFN y virus Sendai. Las células fueron recolectadas después de 5 ó 6 horas de incubación. El ARN de estas células fue extraído por el procedimiento de tiocianato de guanidinio y cloruro de guanidinio. El ARN poli A fue obtenido por cromatografía sobre oligo(dt)-celulosa y esta preparación fue fraccionada sobre Sephacryl (300, 500 y 1 000). La actividad de ARNm del IFN fue determinada en extractos de oocitos de Xenopus laevis, que fueron inyectados con las muestras de ARN. La actividad de ARNm del IFN encontrada con el Sephacryl 500 llegó a ser superior a 5 000 U/*g de ARN, por lo que este procedimiento de fraccionamiento resultó rápido, seguro y poco costoso en la purificación del ARNm del IFN
Assuntos
RNA/isolamento & purificação , Cromatografia em Gel , Interferon Tipo IRESUMO
Los leucocitos humanos fueron inducidos con Le-IFN y virus Sendai. Las células fueron recolectadas después de 5 ó 6 horas de incubación. El ARN de estas células fue extraído por el procedimiento de tiocianato de guanidinio y cloruro de guanidinio. El ARN poli A fue obtenido por cromatografía sobre oligo(dt)-celulosa y esta preparación fue fraccionada sobre Sephacryl (300, 500 y 1 000). La actividad de ARNm del IFN fue determinada en extractos de oocitos de Xenopus laevis, que fueron inyectados con las muestras de ARN. La actividad de ARNm del IFN encontrada con el Sephacryl 500 llegó a ser superior a 5 000 U/*g de ARN, por lo que este procedimiento de fraccionamiento resultó rápido, seguro y poco costoso en la purificación del ARNm del IFN