RESUMO
BACKGROUND: The aim of this study was to investigate the distribution of full-length and deleted variants of KIR2DS4 in a population of southern Brazil and compare the results with other populations, as well as comparing two techniques, PCR-SSP and PCR-SSO, for typing of variants. METHODS: 258 individuals from southern Brazil were analysed by PCR-SSO ("polymerase chain reaction-sequence specific oligonucleotides", One Lambda, Inc., Canoga Park, CA), of which 161 were also analysed by PCR-SSP. RESULTS: The study population showed similarities with other Caucasian populations; 46.5% of individuals had only KIR2DS4 variants, 21.3% had the full-length form and 25.1% had both forms. CONCLUSION: The frequencies found in both groups (genotyped by PCR-SSP and PCR-SSO) were 100% concordant.
Assuntos
Receptores KIR/genética , Deleção de Sequência , Brasil , Genótipo , Humanos , Taxa de Mutação , Reação em Cadeia da Polimerase/métodos , Receptores KIR/químicaRESUMO
The genetic variability of the host contributes to the risk of human papillomavirus (HPV)-related cervical disease. Immune response genes to HPV must be investigated to define patients with the highest risk of developing malignant disease. The aim of this study was to investigate the association of polymorphic immune response genes, namely KIR, HLA class I and II, and single-nucleotide polymorphisms (SNPs) of cytokines with HPV-related cervical disease. We selected 79 non-related, admixed Brazilian women from the state of Paraná, southern region of Brazil, who were infected with high carcinogenic risk HPV and present cervical intraepithelial neoplasia grade 3 (CIN3), and 150 HPV-negative women from the same region matched for ethnicity. KIR genes were genotyped using an in-house PCR-SSP. HLA alleles were typed using a reverse sequence-specific oligonucleotide technique. SNPs of TNF -308G>A, IL6 -174G>C, IFNG +874T>A, TGFB1 +869T>C +915G>C, and IL10 -592C>A -819C>T -1082G>A were evaluated using PCR-SSP. The KIR genes were not associated with HPV, although some pairs of i(inhibitory)KIR-ligands occurred more frequently in patients, supporting a role for NK in detrimental chronic inflammatory and carcinogenesis. Some HLA haplotypes were associated with HPV. The associations of INFG and IL10 SNPs potentially reflect impaired or invalid responses in advanced lesions.
Assuntos
Antígenos HLA-D/genética , Polimorfismo de Nucleotídeo Único/genética , Canais de Potássio Corretores do Fluxo de Internalização/genética , Displasia do Colo do Útero/genética , Displasia do Colo do Útero/virologia , Alelos , Brasil , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença , Antígenos HLA/genética , Haplótipos , Humanos , Papillomaviridae/genética , Papillomaviridae/patogenicidade , Displasia do Colo do Útero/imunologiaRESUMO
INTRODUCTION: The present study was designed to investigate a possible role of HLA (histocompatibility leucocyte antigen) class-I alleles (HLA-A, -B, and -C) in leprosy patients from Southern Brazil. METHODS: Two hundred and twenty-five patients with leprosy and 450 individuals for the control group were involved in this research. HLA genotyping was performed through PCR-SSO protocols (One Lambda, USA); the frequency of these alleles was calculated in each group by direct counting, and the frequencies were then compared. RESULTS: There was an association between HLA-A*11 (6.9% vs 4.1%, p=0.0345, OR=1.72, 95% CI=1.05-2.81), HLA-B*38 (2.7% vs. 1.1%, p=0.0402, OR=2.44, 95% CI=1.05-5.69), HLA-C*12 (9.4% vs. 5.4%, p=0.01, OR=1.82, 95% CI=1.17-2.82), and HLA-C*16 (3.1% vs. 6.5%, p=0.0124, OR=0.47, 95% CI=0.26-0.85) and leprosy per se. In addition, HLA-B*35, HLA-C*04, and HLA-C*07 frequencies were different between lepromatous (LL) and tuberculoid (TT) patients. However, after adjusting for the number of alleles compared, Pc values became nonsignificant. CONCLUSIONS: Although our results do not support the previous findings that HLA class-I alleles play a role in leprosy pathogenesis, we suggest new studies because of the importance of the association between the HLA and KIR in the innate immune response to leprosy.
Assuntos
Antígenos de Histocompatibilidade Classe I/genética , Hanseníase/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Brasil , Estudos de Casos e Controles , Feminino , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Humanos , Hanseníase/imunologia , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
INTRODUCTION: The present study was designed to investigate a possible role of HLA (histocompatibility leucocyte antigen) class-I alleles (HLA-A, -B, and -C) in leprosy patients from Southern Brazil. METHODS: Two hundred and twenty-five patients with leprosy and 450 individuals for the control group were involved in this research. HLA genotyping was performed through PCR-SSO protocols (One Lambda, USA); the frequency of these alleles was calculated in each group by direct counting, and the frequencies were then compared. RESULTS: There was an association between HLA-A*11 (6.9 percent vs 4.1 percent, p=0.0345, OR=1.72, 95 percent CI=1.05-2.81), HLA-B*38 (2.7 percent vs. 1.1 percent, p=0.0402, OR=2.44, 95 percent CI=1.05-5.69), HLA-C*12 (9.4 percent vs. 5.4 percent, p=0.01, OR=1.82, 95 percent CI=1.17-2.82), and HLA-C*16 (3.1 percent vs. 6.5 percent, p=0.0124, OR=0.47, 95 percent CI=0.26-0.85) and leprosy per se. In addition, HLA-B*35, HLA-C*04, and HLA-C*07 frequencies were different between lepromatous (LL) and tuberculoid (TT) patients. However, after adjusting for the number of alleles compared, Pc values became nonsignificant. CONCLUSIONS: Although our results do not support the previous findings that HLA class-I alleles play a role in leprosy pathogenesis, we suggest new studies because of the importance of the association between the HLA and KIR in the innate immune response to leprosy.
INTRODUÇÃO: O presente estudo foi desenhado para investigar um possível papel para os alelos HLA (histocompatibility leucocyte antigen) de classe I (HLA-A, -B, and -C) em pacientes com hanseníase do sul do Brasil. MÉTODOS: Duzentos e vinte e cinco pacientes com hanseníase e 450 indivíduos para o grupo-controle foram envolvidos nesse estudo. O genótipo HLA foi determinado por protocolos PCR-SSO (One Lambda, USA) e, a frequência desses alelos foi calculada em cada grupo por contagem direta e, após, comparadas. RESULTADOS: Houve associação entre HLA-A*11 (6,9 por cento vs 4,1 por cento; p = 0,0345; OR = 1,72; CI = 1,05 - 2,81), HLA-B*38 (2,7 por cento vs 1,1; p = 0,0402; OR = 2,44; CI 95 por cento = 1,05-5,69), HLA-C*12 (9,4 por cento vs 5,4 por cento; p = 0,01; OR = 1,82; CI 95 por cento = 1,17-2,82) e HLA-C*16 (3,1 vs 6,5 por cento; p = 0,0124; OR = 0,47; CI 95 por cento = 0,26-0,85) e hanseníase per se. Além disso, as frequências de HLA-B*35, HLA-C*04 e HLA-C*07 foram diferentes entre os pacientes com as formas lepromatosa (LL) e tuberculoide (TT). Contudo, após o ajuste para o número de alelos comparados, os valores de p se tornaram não significativos. CONCLUSÕES: Embora nossos resultados não sustentem as conclusões anteriores de que os alelos HLA de classe I desempenham um papel na associação com a patogênese da hanseníase, sugerimos novos estudos devido à importância da associação entre HLA e KIR na resposta imune inata à hanseníase.
Assuntos
Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Antígenos de Histocompatibilidade Classe I/genética , Hanseníase/genética , Alelos , Brasil , Estudos de Casos e Controles , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Hanseníase/imunologiaRESUMO
Receptores killer cell immunoglobulin-like (KIRs) são moléculas localizadas na superfície de células natural killer (NK) e em subpopulações de linfócitos T codificadas por genes do cromossomo 19q13.4. A interação entre receptores KIR e moléculas antígeno leucocitário humano (HLA) de classe I determina se células NK exercerão ou não sua função citotóxica e/ou secretora de citocinas ou se esta será inibida. Este trabalho teve por finalidade otimizar a metodologia para a genotipagem KIR, baseando-se nas condições descritas por Martin (2004). A técnica utilizada foi a reação em cadeia da polimerase com primers de sequência específica (PCR-SSP) com iniciadores sintetizados pela Invitrogen® e visualização do produto amplificado em gel de agarose a 2 por cento com brometo de etídio. Adaptações foram realizadas e a concentração de alguns reagentes foi alterada, como a do controle interno de 100 nM para 150 nM, iniciadores específicos senso e antissenso de KIR12.5/12.3, KIR13.5/13.3, KIR14.5/14.3, KIR22.5/22.3 e KIR36.5/36.3 de 500 nM para 750 nM e da solução de MgCl2 de 1,5 mM para 2 mM. As concentrações dos demais reagentes e temperaturas de amplificação foram mantidas. Nessas condições, o uso da Taq DNA polimerase recombinante (Invitrogen®) foi satisfatório. Os resultados das genotipagens de 70 indivíduos foram confirmados por rSSO-Luminex® (One Lambda, Canoga Park, CA, EUA). A tipagem de genes KIR por essa técnica apresentou sensibilidade, especificidade, reprodutibilidade e baixo custo.
The killer cell immunoglobulin-like receptors (KIRs) are molecules expressed on natural killer (NK) cells surface and in T-cell subsets encoded by genes located in chromosome 19q13.4. The interaction between KIR receptors and HLA class I molecules determines if the NK cells will fulfill their cytotoxic function and/or cytokine secretion or if this function will be inhibited. The objective of this work was to optimize KIR genotyping method described by Martin (2004). It was used PCR-SSP (polymerase chain reaction-sequence-specific primers) with primers synthesized by Invitrogen® and visualization of the amplified products on 2 percent agarose gel electrophoresis, containing ethidium bromide. Some adaptations were made and the reagents had their concentrations increased: the internal control from 100 nM to 150 nM, forward and reverse specific primers KIR12.5/12.3, KIR13.5/13.3, KIR14.5/14.3, KIR22.5/22.3 and KIR36.5/36.3 from 500 nM to 750 nM, and MgCl2 solution from 1.5 mM to 2 mM. Other reagent concentrations and amplification temperatures were maintained. Satisfactory results were obtained with Taq DNA Polymerase Recombinant (Invitrogen®). The results of seventy samples were confirmed by rSSO-Luminex® (One Lambda, Canoga Park, CA, USA). This KIR typing method proved to be accurate, specific, reproducible and cost effective.
RESUMO
OBJECTIVE: To determine whether cytokine polymorphisms are associated with leprosy and/or their subtypes in a Brazilian population. METHODS: Genotyping using polymerase chain reaction with sequence-specific primers (PCR-SSP) was performed for: TNF(-308/-238), IL2(-330/+166), IL6(-174), IFNG(+874), TGFB1(+869/+915), and IL10(-592/-819/-1082) in 240 healthy controls and 167 patients with leprosy. RESULTS: For TNF(-308), a higher frequency of GG genotype (85.5% vs. 74.1% in healthy controls, p = 0.009), along with a decreased frequency of GA/AA genotypes was observed among leprosy patients as compared to the control group (14.5% vs. 25.9%, p = 0.009). The GG genotype was particularly higher in patients with tuberculoid (TT) and borderline (BB) leprosy (90.5% and 89.8%, respectively). Analysis of IL10 genotypes revealed a lower frequency of GCC/GCC haplotype in lepromatous leprosy (LL) patients (6.2%) in comparison to controls (15.4%). CONCLUSION: It is suggested that the G-->A substitution at position -308 in the TNF promoter region plays an important role in leprosy patients.
Assuntos
Interleucina-10/genética , Hanseníase/genética , Polimorfismo Genético , Polimorfismo de Nucleotídeo Único , Fator de Necrose Tumoral alfa/genética , Brasil/epidemiologia , Etnicidade , Feminino , Frequência do Gene , Genótipo , Humanos , Hanseníase/epidemiologia , Hanseníase/imunologia , Masculino , Reação em Cadeia da Polimerase , Valores de ReferênciaRESUMO
Killer cell immunoglobulin-like receptors (KIR) are encoded by polymorphic genes and have as binding human leukocyte antigen (HLA) class I molecules. The aim of this study was to investigate the distribution of KIR genes and inhibitory KIR/HLA pairs in a population from Southern Brazil, in the state of Paraná, and to compare the results with results from other populations. The genotyping of 16 KIR genes and HLA class I alleles of 289 unrelated individuals was accomplished by reverse sequence-specific oligonucleotide Luminex (One Lambda, Inc., Canoga Park, CA). This Brazilian population demonstrated several similarities to Caucasian populations with regard to the frequency of KIR genes. Thirty-eight genotypes were defined in which the most frequent was the homozygous haplotype A (33.2%). Therefore, it was possible to define two new genotypes. Most of the individuals demonstrated at least one inhibitory KIR/HLA pair. Two pairs were the most frequent (40.4%), followed by three pairs (38.2%), one pair (14.6%), and four pairs (6.4%). The KIR2DL2/3 + HLA-C1 pair was the most frequent (79.9%) and the least frequent pair was KIR3DL2 + HLA-A3/11 (25.0%). This study demonstrated the diversity of KIR genes in a population of Paraná, as well as the characteristic pattern of Caucasians with racial admixture, which enabled the definition of two new genotypes and the identification of one individual without the inhibitory KIR/HLA pair.
Assuntos
Antígenos HLA/genética , Células Matadoras Naturais/metabolismo , Polimorfismo Genético , Receptores KIR/genética , Adulto , Brasil , Feminino , Frequência do Gene , Antígenos HLA/imunologia , Antígenos HLA/metabolismo , Humanos , Células Matadoras Naturais/imunologia , Masculino , Pessoa de Meia-Idade , Ligação Proteica , Receptores KIR/imunologia , Receptores KIR/metabolismoRESUMO
As células NK (natural killer) são uma subpopulação de linfócitos que desempenham função essencial na resposta imune inata. As moléculas KIR (killer immunoglobulin-like receptor) são receptores expressos na superfície dessas células com função inibitória ou ativatória e contribuem para a regulação da função dascélulas NK. Os genes KIR fazem parte do Complexo de Receptores Leucocitários, localizado no cromossomo 19q13 e apresentam alto polimorfismo. Os ligantes de KIR são moléculas HLA de classe I, e a regulação dascélulas NK está relacionada à variação da expressão dessas moléculas na superfície das células-alvo, principalmente células infectadas, tumorais e alogênicas. O objetivo desse trabalho foi proceder a uma revisão bibliográfica sobre os receptores KIR. O levantamento foi realizado nos sites Pubmed/medline e ScienceDirect,e foram utilizadas como palavras-chave receptor, NK e KIR. A estrutura molecular desses receptores, a nomenclatura e classificação de KIR, a variabilidade gênica, alélica e haplotípica e os ligantes foram apresentados. Ênfase foi dada à regulação da expressão dos genes KIR e sua relação com a função das célulasNK.
NKC (natural killer cells) are a population of lymphocytes that play an essential role in innate immunity. KIR molecules are receptors expressed on the surface of these cells with an inhibitory or activating function that contributes to the regulation of NK cells. The KIR genes are located on chromosome 19q13 at the Leukocyte Receptor Complex, and exhibit high polymorphism. The KIR ligands are HLA class I molecules. NK cell functionsare related to the variation of the expression of these molecules on the surface of the target cells especially infected, allogeneic and tumor cells. The aim of this work was to make a review about KIR receptors. The Pubmed/medline and ScienceDirect online databases were accessed, using receptor, NK and KIR as keywords. KIR molecular structure, nomenclature and classification, gene diversity, allelic and haplotypic variability and itsligands were described. Regulation of KIR genes expression and NK cell function were also presented.
Las células NK (natural killer) son una subpoblación de linfocitos que desempeñan una función esencial en la respuesta inmune innata. Las moléculas KIR (killer immunoglobulin-like receptor) son receptores expresos en la superficie de esas células con función inhibidora o activadora y contribuyen para la regulación de la función delas células NK. Los genes KIR hacen parte del Complejo de Receptores Leucocitarios, localizado en el cromosoma 19q13 y presentan alto polimorfismo. Los ligantes de KIR son moléculas HLA de clase I, y laregulación de las células NK está relacionada a la variación de la expresión de esas moléculas en la superficiede las células blanco, principalmente células infectadas, tumorosos y alogénicas. El objetivo de ese trabajo fue proceder una revisión bibliográfica sobre los receptores KIR. La pesquisa fue realizada en los sites Pubmed/medline y ScienceDirect, y fueron utilizadas como palabras clave receptor, NK y KIR. La estructura molecular de esos receptores, la nomenclatura y clasificación de KIR, la variabilidad génica, alélica y haplotípicay los ligantes fueron presentados. Énfasis fue dada a la regulación de la expresión de los genes KIR y surelación con la función de las células NK.