RESUMO
PURPOSE: Non-alcoholic fatty liver disease (NAFLD) is a complex disease, resulting from a variety of genetic and environmental factors. The aim of this case-control study was to evaluate the effect of selected genetic polymorphisms, nutrition aspects and their interaction on the risk of NAFLD. METHODS: The sample consisted of 134 patients with NAFLD and 217 controls. Disease was diagnosed by liver ultrasound and volunteers were clinically and nutritionally assessed. Food groups were extracted from a 172 food-item FFQ questionnaire. Three genetic polymorphisms were assessed: PNPLA3 rs738409, TM6SF2 rs58542926 and GCKR rs780094. RESULTS: We replicated the effect of previously reported risk factors for NAFLD, such as elevated liver enzymes, obesity and metabolic syndrome. Food groups rich in simple sugars, fat and especially saturated fat were positively associated with NAFLD risk, whereas food groups rich in polyunsaturated fatty acids were reversely associated with the possibility of developing the disease (p < 0.05). Only the PNPLA3 genetic variant was statistically significantly associated with the disease (padditive = 0.015). However, it was found that a one-portion increase in fish intake increased the risk of NAFLD in carriers of the risk allele of TM6SF2 rs58542926 polymorphism compared to non-carriers, after adjusting for age, gender, energy intake, pack-years, PAL, TM6SF2 genotype and fish consumption (ORdominant = 1.503, 95% CI 1.094-2.064). CONCLUSIONS: Fish intake exerts an additive effect on NAFLD risk for carriers of the TM6SF2 polymorphism. This novel finding provides further rationale on the need for personalized nutritional advice, based on the genetic background of NAFLD patients.
Assuntos
Proteínas de Membrana/genética , Hepatopatia Gordurosa não Alcoólica/genética , Polimorfismo de Nucleotídeo Único/genética , Alimentos Marinhos , Adulto , Estudos de Casos e Controles , Feminino , Humanos , Fígado/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Avaliação Nutricional , Fatores de Risco , Inquéritos e Questionários , UltrassonografiaRESUMO
BACKGROUND: Sleeve gastrectomy (SG) and Roux-en-Y gastric bypass (RYGB) are the most frequent bariatric surgery procedures worldwide. In this prospective study, we examined the association of a genetic risk score (GRS) with loss of excess weight after bariatric surgery. METHODS: A total of forty-seven morbidly obese Greek patients who underwent SG (81%) or RYGB were recruited, followed up for 2 years and genotyped. Weight loss after surgery was reported as the percentage of excess weight that was lost (%EWL) at 12 and 24 months after surgery. A GRS was constructed based on previously BMI- and WHR-related single nucleotide polymorphisms (SNPs) that were found significantly correlated with weight loss after bariatric surgery in our population. The level of post-surgery %EWL after 12 and 24 months was estimated through two multiple linear regression models that considered the effects of relevant genetic risk variants. RESULTS: The first proposed model suggested that the predictor variables of GRS, age, and BMI had a significant effect on %EWL12m. GRS was significantly associated with %EWL12m, indicating a 4.618% decrease of %EWL12m per score unit. The second model indicated a positive correlation between %EWL24m and %EWL12m, suggesting that while post-surgery weight loss increased during the first 12 months, an increase was expected in the next 12 months as well. GRS was also significantly associated with %EWL24m, indicating approximately 3% decrease of %EWL24m per score unit. CONCLUSION: GRS can be used in the future together with other preoperative parameters in order to predict the outcome of bariatric surgery.
Assuntos
Cirurgia Bariátrica , Derivação Gástrica , Obesidade Mórbida , Gastrectomia , Humanos , Obesidade Mórbida/cirurgia , Estudos Prospectivos , Estudos Retrospectivos , Fatores de Risco , Resultado do Tratamento , Redução de PesoRESUMO
BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD), which is caused by mutations in polycystins 1 (PC1) and 2 (PC2), is one of the most commonly inherited renal diseases, affecting ~1 : 1000 Caucasians. MATERIALS AND METHODS: We screened Greek ADPKD patients with the denaturing gradient gel electrophoresis (DGGE) assay and direct sequencing. RESULTS: We identified a patient homozygous for a nucleotide change c.1445T > G, resulting in a novel homozygous substitution of the non-polar hydrophobic phenylalanine to the polar hydrophilic cysteine in exon 6 at codon 482 (p.F482C) of the PKD2 gene and a de-novo PKD1 splice-site variant IVS21-2delAG. We did not find this PKD2 variant in a screen of 280 chromosomes of healthy subjects, supporting its pathogenicity. The proband's parents did not have the PKD1 mutation. Real-time PCR of the PKD2 transcript from a skin biopsy revealed 20-fold higher expression in the patient than in a healthy subject and was higher in the patient's peripheral blood mononuclear cells (PBMCs) than in those of her heterozygote daughter and a healthy subject. The greater gene expression was also supported by Western blotting. Inner medullar collecting duct (IMCD) cells transfected with the mutant PKD2 mouse gene presented a perinuclear and diffuse cytoplasmic localization compared with the wild type ER localization. Patch-clamping of PBMCs from the p.F482C homozygous and heterozygous subjects revealed lower polycystin-2 channel function than in controls. CONCLUSIONS: We report for the first time a patient with ADPKD who is heterozygous for a de novo PKD1 variant and homozygous for a novel PKD2 mutation.
Assuntos
Rim Policístico Autossômico Dominante/genética , Proteínas/genética , Animais , Células Cultivadas , Análise Mutacional de DNA , Eletroforese/métodos , Feminino , Homozigoto , Humanos , Masculino , Camundongos , Mutação , Reação em Cadeia da Polimerase , Canais de Cátion TRPPRESUMO
Several naturally occurring constituents have received considerable attention because of their potential antioxidant activity. Consuming a diet rich in natural antioxidants has been associated with prevention from and/or treatment of atherosclerosis. Bioactive components of food, which are of special interest, include the Vitamins E and C, polyphenols, carotenoids-mainly lycopene and beta-carotene, and coenzyme Q10, featured by antioxidant properties. Antioxidant therapy is supposed to be effective in the early stages of atherosclerosis by preventing LDL oxidation and the oxidative lesion of endothelium. This review focuses on the effect of dietary antioxidants pertained to LDL oxidation and to the vascular endothelial dysfunction. Now that the human genome has been completely sequenced, genetic factors involved in oxidation may open new horizons to identify persons at risk for cardiovascular disease, allowing effective dietary intervention strategies to recover normal homeostasis and to prevent diet-related implications. On this basis, current studies on the action of selected antioxidant nutraceuticals on the activity of transcription factors, such as final targets in the signal transduction cascade and gene regulation, may emerge into new treatment concepts.
Assuntos
Antioxidantes/farmacologia , Aterosclerose/prevenção & controle , Dieta , Regulação da Expressão Gênica , Animais , Antioxidantes/metabolismo , Aterosclerose/tratamento farmacológico , Modelos Animais de Doenças , Endotélio Vascular/metabolismo , Humanos , Lipoproteínas LDL/metabolismo , Modelos Biológicos , Estresse Oxidativo , Oxigênio/metabolismo , Transcrição Gênica , Vitaminas/metabolismoRESUMO
Familial hypercholesterolaemia is a genetic disorder characterised by high low-density lipoprotein (LDL) cholesterol concentrations, which frequently gives rise to premature coronary artery disease (CAD). The clinical expression of familial hypercholesterolaemia is highly variable even in patients carrying the same LDL receptor gene mutation. This variability may be due to environmental and other genetic factors. Apolipoprotein E (Apo-E) has been extensively studied for its effects on the phenotype of familial hypercholesterolaemia. In this study we examined the influence of Apo-E genotype on lipid parameters and the incidence of CAD in 93 Greek patients with familial hypercholesterolaemia. Apo-E E2, E3 and E4 allele frequencies were 0.06, 0.86 and 0.09 respectively. The levels of total cholesterol, LDL cholesterol, HDL cholesterol, triglycerides, apolipoproteins A and B and lipoprotein alpha did not differ significantly among carriers and non-carriers of the E4 allele. The prevalence of CAD and hypertension did not differ either. Our results suggest that the E4 allele is not associated with lipid levels or with the prevalence of CAD among familial hypercholesterolaemia patients of the Greek population.
Assuntos
Apolipoproteínas E/genética , Doença da Artéria Coronariana/genética , Hiperlipoproteinemia Tipo II/genética , Lipídeos/sangue , Polimorfismo Genético , Adulto , Feminino , Humanos , Hiperlipoproteinemia Tipo II/sangue , MasculinoRESUMO
We investigated the effect of intracellular glutathione (GSH) levels on Natural Killer-mediated apoptosis in cisplatin-resistant K562 cells. K562/B6 and K562/C9 are cisplatin-resistant K562 cells less susceptible to lysis by natural killer cells. Cisplatin-resistant K562 cells did not present the apoptotic pattern of DNA fragmentation as it was observed for their maternal counterparts. K562/B6 and K562/C9 cell lines produce 1.6- and 1.9-times more GSH than K562 cells. Treatment of both cell lines with D,L-buthionine-(S,R)-sulfoximine (BSO, a gamma-glutamyl cysteine synthetase inhibitor) decreased GSH levels and augmented cell death induced by NK cells via a necrotic rather than an apoptotic process. Proliferating cell nuclear antigen (PCNA) expression was elevated in cisplatin-resistant K562 subclones, and the reduction of GSH levels after treatment with BSO decreased the expression of PCNA. These results suggest that the GSH level affects the NK cell-mediated cell death of cisplatin-resistant K562 cells by inducing necrosis rather than apoptosis.
Assuntos
Apoptose , Glutationa/metabolismo , Células Matadoras Naturais/fisiologia , Necrose , Antígeno Nuclear de Célula em Proliferação/metabolismo , Antineoplásicos/farmacologia , Butionina Sulfoximina/farmacologia , Separação Celular , Cisplatino/farmacologia , Técnicas de Cocultura , Testes Imunológicos de Citotoxicidade , Fragmentação do DNA , Resistencia a Medicamentos Antineoplásicos , Inibidores Enzimáticos/farmacologia , Citometria de Fluxo , Corantes Fluorescentes/metabolismo , Humanos , Células K562RESUMO
The use of the chromium-release assay to determine cytotoxicity of effector against target cells has various limitations mostly due to the inherent properties of the radioactive substance. We have developed an improved flow cytometric method that is able to measure cytotoxicity, based on two fluorescent dyes. Calcein-AM, a non-fluorescent substance which is intracellularly converted to the green fluorescent calcein by esterase activity in viable cells, is initially used to stain target cells. After incubating targets with effectors for 2 h, ethidium homodimer-1, a red DNA stain non-permeable to viable cells, is added. Dead target cells are distinguished by their double (green-red) staining. Data analysis is performed by gating the regions of living target, dead target and living effector cells, based on appropriate controls. Non-specific events are subtracted from the dead target region and the ratio of specific dead target events to total target events is expressed as percent cytotoxicity. The method is used to quantify natural killer (NK) and lymphokine-activated killer (LAK) activities against the human K562 and Daudi cell lines and the murine YAC-1 and L1210 cell lines respectively, as well as cell-mediated lympholysis (CML) exerted by tumor-infiltrating lymphocytes (TIL) against autologous and allogeneic human breast cancer tumor cells. The method is fast, reliable and correlates well with the standard 51Cr-release assay.
Assuntos
Testes Imunológicos de Citotoxicidade , Citometria de Fluxo/métodos , Animais , Relação Dose-Resposta Imunológica , Fluoresceínas , Corantes Fluorescentes , Humanos , Imunidade Celular , Técnicas In Vitro , Células Matadoras Ativadas por Linfocina/imunologia , Células Matadoras Naturais/imunologia , Camundongos , Camundongos Endogâmicos BALB CRESUMO
The expansion of myeloma cells is regulated by cytokines, among which IL-6 is a major growth factor. It has been recently suggested that serum transforming growth factor beta 1 (TGF beta 1), a cytokine found in large amounts in alpha-granules of platelets, might play a role in multiple myeloma (MM). It was the purpose of this study to determine serum TGF beta 1 levels in MM patients and to seek a correlation with disease parameters. Measurements were done by ELISA. We studied 35 MM patients (19 stage II, 16 stage III, 20 IgG, 8 IgA and 6 BJ, 1 IgD) in different phases of the disease, 27 healthy individuals and 17 thrombocytopenic patients with other haematological diseases (three MDS, three congenital thrombocytopenia, 11 ITP). Overall samples from MM patients were included: 10 at diagnosis, 18 in remission and 32 in relapse. In normal controls TGF beta 1 serum levels ranged from 1 to 33 ng/ml (median 16.5 ng/ml). In both thrombocytopenic controls with other diseases and thrombocytopenic MM patients (seven samples), TGF beta 1 serum levels were very low (median 3.2 and 4.5 ng/ml respectively). In MM patients with PLT > 100 x 10(9)/L (53 samples), TGF beta 1 serum levels were in the normal range in patients without immunoparesis (1 to 27 ng/ml, median 16.6 ng/ml), whereas they were higher in patients with immunoparesis (polyclonal immunoglobulins (Igs) below lower normal reference values) ranging from 10.2 to 45 ng/ml (median 26.8 ng/ml) (P < 0.01). Serum TGF beta 1 levels fluctuated in the same patient at different times but not according to relapse or remission. Correlation was found only between serum TGF beta 1 levels and immunoparesis and not between serum TGF beta 1 levels and disease stage or Ig subtype nor with prognostic factors for MM (serum CRP, beta 2M or IL-6). This finding suggests that the remaining normal plasma cells are sensitive to the inhibitory action of TGF beta 1 on Ig production. In conclusion TGF beta 1 serum levels are very low in thrombocytopenic patients confirming that platelets are the major source of this cytokine. Furthermore, a strong correlation was found between TGF beta 1 serum levels and immunoparesis in MM patients.
Assuntos
Mieloma Múltiplo/imunologia , Fator de Crescimento Transformador beta/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulinas/sangue , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/sangue , Contagem de Plaquetas , PrognósticoRESUMO
Epirubicin is an anthracyclinic antibiotic that has been increasingly used in the treatment of a variety of malignancies. A hybridoma producing monoclonal antibody (MAb) against the drug was obtained by cell fusion. The MAb is of the IgM isotype and has an affinity constant of 1.4 x 10(-7) M. Inhibition analysis showed that the antibody recognizes an epitope related to the C 4'-hydroxyl group in the amino sugar moiety, distinguishing epirubicin from the closely related doxorubicin. Since the precise mechanism of anthracycline action as well as its immunomodulating effects are still under scrutiny, powerful tools for their study are clearly needed. Moreover, this MAb can be useful in monitoring the levels of epirubicin in treated patients, as well as for the construction of bispecific antibodies in tumor-targeting immunotherapy.
Assuntos
Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/química , Epirubicina/imunologia , Animais , Especificidade de Anticorpos , Epirubicina/química , Feminino , Hibridomas/metabolismo , Imunoconjugados/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Soroalbumina Bovina/imunologiaRESUMO
BACKGROUND/OBJECTIVES: Height is a classic polygenic trait, with a number of genes underlying its variation. We evaluated the prospect of gene-to-diet interactions in a children's cohort, for the insulin-like growth factor II (IGF) rs680 polymorphism and height variation. SUBJECTS/METHODS: We screened 795 periadolescent children (424 girls) aged 10-11 years old from the Gene and Diet Attica Investigation (GENDAI) pediatric cohort for the IGF rs680 polymorphism (rs680). RESULTS: Children homozygous for the common allele (GG) were taller (148.9+/-7.9 cm) compared with those with the A allele (148.1+/-7.9 cm), after adjusting for age, sex and dairy intake (beta+/-s.e.: 2.1+/-0.95, P=0.026). A trend for rs680 x dairy intake interaction was also revealed (P=0.09). Stratification by IGF rs680 genotype revealed positive significant (P=0.014) association between dairy product intake and height in A-allele children adjusted for the same confounders. A daily increase of four dairy servings was associated with a 0.4 cm increase in height. On grouping dairy intake into low (1.9+/-0.7 servings per day) and high dairy product consumption (4.4+/-1.5 servings per day), children with the A allele who were high dairy product consumers were taller compared with the low dairy product consumers (148.8+/-7.9 vs 147.4+/-7.7 cm, respectively, P=0.05). CONCLUSIONS: A higher consumption of dairy products is associated with increased height depending on the rs680 IGF2 genotype.
Assuntos
Estatura/genética , Laticínios , Dieta , Fator de Crescimento Insulin-Like II/genética , Polimorfismo Genético , Fatores Etários , Alelos , Estatura/fisiologia , Criança , Estudos de Coortes , Feminino , Humanos , Masculino , Nutrigenômica , Fatores SexuaisRESUMO
Wilson disease (WD) is an autosomal recessive disorder of copper metabolism. The disorder is caused by mutations in the ATP7B gene, encoding a copper transporting P-type ATPase. The worldwide incidence is in the order of 30 cases per million, with a gene frequency of 0.56% and a carrier frequency of 1 in 90. The increased number of Wilson disease patients in the island of Crete led us to study the spectrum of mutations in a small village close to the city of Heraklion, from where many patients have been referred during the last 25 years. In order to estimate the frequency of the disease, we firstly investigated the number of births and the number of WD patients in the village since 1978. Six out of 90 births were diagnosed as WD patients, presenting the highest prevalence of WD reported so far. Analysis of the whole gene in three Wilson disease patients, and relatives of a boy who died from WD, led to the detection of 4 different point mutations. Two of them were missense (p.I1148T and p.G1176R) and cosegregated in cis in the same patient; the other allele of this patient carried a nonsense mutation (p.Q289X). This is the first report in the literature of three mutations co-segregating in the same WD patient. The fourth mutation identified was a novel frameshift mutation (c.398delT) with documented cosegregation. When screening 200 inhabitants originating from the same area, 18 were found to be carriers of one of these mutations. These findings indicate the need for health education intervention, genetic counselling and newborn screening for the Wilson disease.
Assuntos
Adenosina Trifosfatases/genética , Proteínas de Transporte de Cátions/genética , Degeneração Hepatolenticular/epidemiologia , Degeneração Hepatolenticular/genética , Idade de Início , Criança , Pré-Escolar , ATPases Transportadoras de Cobre , Análise Mutacional de DNA , Feminino , Mutação da Fase de Leitura , Grécia/epidemiologia , Humanos , Masculino , Mutação de Sentido Incorreto , Linhagem , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , PrevalênciaRESUMO
We studied a family in which two inherited defects of the non-alpha-globin cluster segregate: Greek hereditary persistence of fetal hemoglobin (HPFH) and beta-thalassemia. The compound heterozygote is a healthy man with 43% HbF, Ggamma/Agamma ratio (27:73) differing from that of 10 simple heterozygotes for the Greek HPFH (92:8), normal levels of total Hb (13.3 g/dl), and reduced HbA2 levels comparing with the levels of beta-thal heterozygotes for the same mutation. Molecular analysis of the beta-globin genotype revealed the presence of the IVSII-745 (C-->G) beta+ RNA splice mutation in trans with the -117 G-->A Greek HPFH. The beta+ mutation was linked to haplotype VII and the Greek HPFH was associated with haplotype Ia. The father of the compound heterozygote carries the Greek HPFH in trans with the -158 C-->T on the Ggamma promoter, which is linked with haplotype IV. He presented 13.5% HbF with a Ggamma/Agamma ratio 75:25. His daughter was a compound heterozygote for the IVSII-745 mutation in trans with the -158 C-->T, while her HbF levels were 3.7% with a Ggamma/Agamma ratio 31:69.
Assuntos
Hemoglobina Fetal/genética , Hemoglobina Fetal/metabolismo , Heterozigoto , Talassemia beta/genética , Adulto , Sequência de Bases/genética , Feminino , Globinas/genética , Haplótipos/genética , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Capuzes de RNA , Splicing de RNARESUMO
The splicing defect at IVS-I-110 is by far (43.15%) the most common beta-thalassaemia mutation in Greece. The - 117 (G-->A) Agamma hereditary persistence of fetal hemoglobin (Greek HPFH) is also the most frequent nondeletional HPFH in Greece. We report a case in which these two defects co-segregates. She is a healthy female where the total Hb is 12.3 g/dl with 51% HbF and normal HbA2. Her Ggamma/Agamma ratio is 35:65 differing from that of 10 simple heterozygotes for the Greek HPFH who have ratio of 8:92. Molecular analysis of the beta-globin genotype revealed the presence of the IVS-I-110 beta+ mutation in trans to the -117 G-->A Greek HPFH. Both mutations are linked to Ia. Her father has Greek HPFH in trans to the -158 C-->T on the Ggamma promoter, which is linked with haplotype IIIalpha. He has 13% HbF with a Ggamma/Agamma ratio 32:68. Her sister is a compound heterozygote for the IVS-I-110 mutation in trans to the - 158 C-->T, with HbF levels of 3% and a Ggamma/Agamma ratio 72:28.
Assuntos
Hemoglobina Fetal/biossíntese , Talassemia beta/genética , Adulto , Sequência de Bases , beta-Globulinas/genética , Feminino , Hemoglobina Fetal/genética , Haplótipos/genética , Hemoglobina A2/genética , Heterozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Mutação Puntual/genética , Regiões Promotoras Genéticas/genética , Splicing de RNA , Talassemia beta/sangueRESUMO
The aim of the study was to investigate the association of leptin with hematological parameters in beta-thalassemia patients in Greece. We measured plasma levels of soluble transferrin receptor (sTfR) and leptin by enzyme-linked immunosorbent assay (ELISA) in 40 beta-thalassemia patients (21 transfusion dependent and 19 not transfused or sporadically transfused), in 20 beta-thalassemia carriers, and in 30 healthy individuals (HI). The percentage of reticulocytes (RET) was measured by the NE 9500 Sysmex automated method. Body mass index (BMI) was calculated by dividing body weight (kg) by square height (m). Endocrine measurements including sex hormones were also determined. sTfR concentrations were significantly higher in both transfusion-dependent (females 10.5+/-2.9, males 9.1+/-3.1) and non-transfusion-dependent patients (females 15.8+/-5.4, males 19.8+/-13.7) as compared to carriers (females 3.1+/-2.5, males 3.8+/-1.8) and to HI (females 1.5+/-1.2, males 2.5+/-2.1). Leptin levels were lower both in female and in male transfusion-dependent patients (0.5+/-0.3 and 1.2+/-1, respectively) and in non-transfused males (1.9+/-2) compared to carriers (females 7.9+/-2.7, males 13.1+/-9.1) and HI (females 14.6+/-6, males 7.5+/-3). There was a negative correlation between leptin and sTfR levels in transfused patients (R=-0.61, p<0.05). A stronger negative correlation (R=-0.7, p=0.006) was found in hypogonadic men and women with beta-thalassemia. These findings enhance previous results indicating that leptin may play some role in hematopoiesis and could associate the pathophysiology of thalassemic patients with the triggering effect of leptin in reproductive ability.
Assuntos
Leptina/sangue , Receptores da Transferrina/sangue , Talassemia beta/metabolismo , Adulto , Estudos de Casos e Controles , Feminino , Grécia/epidemiologia , Heterozigoto , Homozigoto , Humanos , Hipogonadismo/sangue , Hipogonadismo/metabolismo , Leptina/fisiologia , Masculino , Pessoa de Meia-Idade , Receptores para Leptina , Solubilidade , Estatísticas não Paramétricas , Talassemia beta/sangueRESUMO
Serum levels of the interleukins (IL)-2, -1 beta, and -6, tumor-necrosis factor alpha (TNF-alpha), and prostaglandin E2 (PGE2) were measured serially before and after operation in patients undergoing various surgical procedures. Peripheral blood mononuclear cells (PBMC) from the same patients were analyzed before and after surgery for proliferative responses in the autologous mixed lymphocyte reaction (AMLR) and for cytolytic activity against the natural killer (NK)-sensitive K562 cell line. In patients who had major surgery, a decrease in IL-2 levels and increases in IL-1 beta, TNF-alpha, IL-6, and PGE2 levels were observed up to 9 days after the operation compared to those of the preoperative values. Decreased AMLR responses and NK activity were also observed in PBMC collected 5 days after surgery. All these changes were more intense in patients who had undergone major surgical procedures of increasing severity (i.e., cancer patients). Similar, although weaker, changes in cytokine serum levels, AMLR responses, and NK activity were also observed in patients undergoing minor surgery. Our data suggest that changes in cytokine serum levels may cause cellular immune dysfunctions, particularly in patients undergoing major surgery, and provide the basis for immune intervention in order to avoid infections occurring after major surgery.
Assuntos
Citocinas/sangue , Complicações Pós-Operatórias/imunologia , Citotoxicidade Imunológica , Dinoprostona/sangue , Humanos , Imunidade Celular , Interleucina-1/sangue , Interleucina-2/sangue , Interleucina-6/sangue , Células Matadoras Naturais/imunologia , Teste de Cultura Mista de Linfócitos , Complicações Pós-Operatórias/sangue , Fator de Necrose Tumoral alfa/análiseRESUMO
We studied the presence of the XmnI site and the beta-globin haplotype in 24 individuals, carriers of the IVSII-1 G-->A beta(0)-globin mutation, of whom fourteen had no detectable levels of HbF, while ten coming from 5 families, presented HbF levels ranging from 1.7 to 9% of the total Hb. Of these beta-thalassaemia heterozygotes with fetal hemoglobin, 6 were females and 4 were males with median HbF levels of 4.85% and 4% respectively, and an excess of (G)gamma chains (range (G)gamma/(A)gamma: 55/45-70/30). Of the group of carriers of beta-thalassaemia with HbF < 0.1, in all cases except one, IVSII-1 mutation was found associated with XmnI polymorphic site. Haplotype analysis in these individuals revealed that in 10 cases IVSII-1 was linked with haplotype IIIb, in 1 case with haplotype IIIa, and in 3 cases with haplotype IX. On the other hand, in the group of carriers with measurable levels of HbF, IVSII-1 was always associated with haplotype IIIa and the XmnI site was either in-homozygous or the heterozygous state in-cis or in-trans with the mutated beta-globin gene. In conclusion the results of the study of these families seem that XmnI site in-cis with the IVSII-1 does not induce HbF production when this beta(0)-thalassaemia mutation is associated with IIIb or IX haplotype. On the other hand the (G)gamma -158 C-->T mutation is associated with small amounts of HbF in IVSII-1 heterozygotes, when the beta-globin mutation is linked to haplotype IIIa.
Assuntos
Hemoglobina Fetal/biossíntese , Hemoglobina Fetal/genética , Globinas/genética , Talassemia beta/genética , Talassemia beta/metabolismo , Feminino , Haplótipos , Heterozigoto , Humanos , Masculino , Linhagem , Mutação PuntualRESUMO
Using peritoneal fluid or pleural effusion obtained from 20 patients with lung, ovarian or metastatic breast cancer, we separated tumour cells from malignant effusion-associated mononuclear cells (MEMNCs) using discontinuous Ficoll-Hypaque density gradients. CD3+ T lymphocytes represented the main population of MEMNCs. The mean +/- s.d. CD4/CD8 ratio of MEMNC suspensions was 1.18 +/- 0.40. MEMNCs proliferated and expanded in vitro with human interleukin 2 (IL-2) either as CD3+ CD8+ cells or as CD3+ CD4+ cells or as mixed populations of CD8+ and CD4+ cells. Preferential cytolytic activity against autologous tumour cells was demonstrated in IL-2-activated MEMNC cultures with excess CD3+ CD8+ cells. In contrast, effectors derived from IL-2-activated cultures with excess CD3+ CD4+ cells lysed both autologous and allogeneic tumour target cells. The addition on day 0 of interleukin 1 beta (IL-1 beta) to MEMNCs cultured in the presence of IL-2 was effective in promoting the growth of CD3+ CD8+ cells and augmenting the cytotoxicity against autologous tumour. Simultaneously, the production of gamma-interferon (IFN-gamma) was increased in these cultures. This is the first report suggesting that IL-1 beta synergises with IL-2 to induce autologous tumour-specific CD8+ cytotoxic T lymphocytes (CTLs) within the MEMNC population. Selective enrichment in T-cell subsets by IL-1 beta may be useful in cellular adoptive immunotherapy using cells isolated from malignant effusions.
Assuntos
Linfócitos T CD8-Positivos/metabolismo , Interleucina-1/farmacologia , Interleucina-2/farmacologia , Linfócitos T Citotóxicos/efeitos dos fármacos , Adulto , Idoso , Líquido Ascítico/citologia , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Sinergismo Farmacológico , Feminino , Humanos , Interferon gama/metabolismo , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Pessoa de Meia-Idade , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Derrame Pleural Maligno/citologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismoRESUMO
As several possible prognostic and therapeutic applications of interleukin-6 are currently under trial, the available methods for its quantification in biological fluids should be evaluated. In this report, the 7TD1 hybridoma bioassay is compared to an enzyme immunoassay for the determination of interleukin-6 in serum and plasma of normal subjects and patients with cancer, sepsis, and systemic lupus erythematosus, as well as in malignant pleural effusions and culture supernatants. The results show a good correlation between the two methods in all cases. Mean values of the examined groups were statistically different between the assays only in the case of septic patients. This may be attributed either to the influence of other molecules on the assays or to the nonlinearity of the dose-response curves. Since immunoassays are easier to perform, it seems that they are more suitable for routine use, the bioassay being preferable in cases where increased sensitivity is required.
Assuntos
Bioensaio , Técnicas Imunoenzimáticas , Interleucina-6/análise , Derrame Pleural/química , Linfócitos B/citologia , Divisão Celular , Células Cultivadas , Meios de Cultura/química , Relação Dose-Resposta a Droga , Humanos , Hibridomas , Interleucina-6/sangue , Interleucina-6/farmacologia , Leucócitos Mononucleares/química , Lúpus Eritematoso Sistêmico/sangue , Neoplasias/sangue , Neoplasias/química , Análise de Regressão , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Sepse/sangueRESUMO
Cisplatin is an effective chemotherapeutic agent that elicits its antineoplastic activity by binding to DNA and disrupting template functions. IL-6 is a cytokine which has been shown to play a central role in host immunological defense mechanisms. Although K562 leukemic cells have been shown to secrete IL-6, little is known of whether there exists a correlation between the expression of IL-6 and the resistance of these cells to anticancer chemotherapeutic agents. To determine the contribution of IL-6 to the regulation of cisplatin-induced apoptosis in K562 cells, we examined whether treatment of K562 cells and cisplatin-resistant K562 subclones with anti-IL-6 mAb enhances their sensitivity to cisplatin. The results show that cis-diamminedichloroplatinum (CDDP) resistance was overcome by treatment with nontoxic doses of CDDP in combination with anti-IL-6 mAb. When we tested if the synergistic effect of anti-IL-6 and cisplatin could restore the ability of K562 mutant cells to undergo apoptosis, we found the typical DNA laddering in these cells, even in the presence of a nontoxic dose of the drug. Treatment of cells with anti-IL-6 reduced the levels of glutathione. The current studies show that anti-IL-6 mAb sensitized CDDP-resistant K562 cells to CDDP by induction of apoptotic death and the reduction of glutathione levels might be implicated in the enhanced cytotoxicity observed.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Cisplatino/farmacologia , Interleucina-6/fisiologia , Células K562/fisiologia , Anticorpos Monoclonais/farmacologia , Células Clonais , Resistencia a Medicamentos Antineoplásicos/imunologia , Humanos , Interleucina-6/imunologia , Interleucina-6/metabolismo , Células K562/citologia , Células K562/metabolismoRESUMO
BACKGROUND: Familial hypercholesterolaemia (FH) is caused by mutations in the low-density lipoprotein receptor gene and the gene encoding apolipoprotein B-100, affecting one in 500 individuals. METHODS: One hundred and eighty-three Greek FH patients were screened for mutations on the LDLR and ApoB genes. RESULTS: We identified mutations in 67 probands and 11 relatives. Sixteen mutations located in eight different exons and the promoter of the LDLR were discovered. Among them 10 were missense mutations (C6W, S265R, A370T, Q363P, D365E, V408M, A410T, A517T, G528D, G571E), two were nonsense mutations (Q363X and C660X), three were splice defects (2140 + 5G-->A and 2140 + 9C-->T, 1706 - 10G-->A), and one was a nucleotide substitution (- 45delT) on the promoter. None of the subjects carried any apoB mutation. The detection rate of mutations in this study was 43%. From the above mutations, A410T, A519T and the splice site defects 2140 + 9C-->T were detected for the first time in the Greek population. Among them V408M, G528D, C6W and S265R account for 73% of heterozygous FH probands. V408M mutation is more common in Central West, while C6W is more common in Central East. Separating the patients into two groups (receptor defective and receptor negative) we found that the receptor negative group had higher levels of total cholesterol, low-density lipoprotein cholesterol and higher prevalence of tendon xanthomas compared with the receptor-defective group. DISCUSSION: The homogenous molecular basis of familial hypercholesterolaemia in Greece facilitates the application of a DNA diagnostic strategy based on the origin of the patient. The early mutation analysis would add valuable information on the severity of the disease.