RESUMO
BACKGROUND: Filaggrin-derived natural moisturizing factors (NMF) play an important role in skin barrier function and in atopic dermatitis (AD). Its deficiency is associated with dry skin and increased surface pH. Studies on childhood environmental exposures and associations with NMF levels are scarce. OBJECTIVES: To investigate previous exposures and genetic factors and their associations with NMF levels in young children. METHODS: In a case-control study nested in a prospective birth cohort (Odense Child Cohort), 169 healthy controls (HC) and 99 children with AD were included consecutively at the age of 7 years based on previous responses from questionnaires administered at 18 months, 3 years and 5 years, pertaining to past medical history, including allergy-specific questions. NMF levels were measured via a stratum corneum tape-stripping technique, genotyping for filaggrin (FLG) gene variants was performed and data on external exposures, including usage of moisturizer and topical steroids, antibiotics and early pet exposures, were obtained from questionnaires. RESULTS: Natural moisturizing factors levels were significantly lower in AD participants compared to HC (P < 0.001). This significance persisted after stratifying for AD subgroups of present AD, current AD during the last year and previous AD (P < 0.001, P = 0.039, P = 0.009 respectively). There was a significant association between NMF and FLG genotype (P = 0.016, P = 0.002 for HC, AD respectively). NMF levels were negatively correlated with early age moisturizer use (<18 months, P = 0.001) in HC but not significant in AD. CONCLUSIONS: We found decreased levels of NMF with early moisturizer use and a genetic influence of the FLG variant on these levels. NMF was decreased in the AD subgroup with previous AD compared with HC, which could suggest the persistence of a Th2 cytokine milieu suppressing these levels.
Assuntos
Eczema , Proteínas Filagrinas , Coorte de Nascimento , Estudos de Casos e Controles , Criança , Pré-Escolar , Eczema/genética , Humanos , Lactente , Proteínas de Filamentos Intermediários/genética , Proteínas de Filamentos Intermediários/metabolismo , Estilo de Vida , Mutação , Estudos Prospectivos , Pele/metabolismoRESUMO
OBJECTIVES: To investigate whether antibody response patterns against Klebsiella pneumoniae capsular serotypes can discriminate patients with axial spondyloarthritis (axSpA) from patients with non-specific low back pain (LBP). METHOD: Immunoglobulin (Ig)G and IgA antibodies against K. pneumoniae capsular serotypes K2, K26, K36, and K50 were measured, and antibody seropositivity compared between groups and analysed for patient correlation in five different groups: (a) 96 patients fulfilling the Assessment of SpondyloArthritis International Society (ASAS) classification criteria for axSpA; (b) 38 patients with either a positive magnetic resonance imaging (MRI) scan as defined by ASAS or a positive human leucocyte antigen (HLA)-B27 status plus one clinical SpA feature, characterized as 'non-axSpA'; (c) 82 non-specific LBP patients; (d) 40 healthy blood donors and (e) 43 patients with diagnosed ankylosing spondylitis (AS) served as the negative and positive control groups. RESULTS: There was no difference in IgG and IgA seropositivity against all serotypes between the axSpA, non-axSpA, and LBP groups. No significant correlations were found between anti-Klebsiella antibodies and age, gender, HLA-B27, or high-sensitivity C-reactive protein (hsCRP). IgG seropositivity against K50 was more frequent in AS (25.6%) than in axSpA (13.5%, p < 0.05). axSpA patients with radiographic sacroiliitis and AS controls concordantly had the highest frequency of seropositivity for ≥ 2 serotypes (21%). CONCLUSIONS: The antibody patterns against K. pneumoniae serotypes K2, K26, K36, and K50 did not discriminate between early axSpA and non-specific LBP.
Assuntos
Anticorpos Antibacterianos/imunologia , Klebsiella pneumoniae/imunologia , Dor Lombar/imunologia , Sacroileíte/imunologia , Espondiloartropatias/imunologia , Adolescente , Adulto , Cápsulas Bacterianas/imunologia , Proteína C-Reativa/imunologia , Estudos de Casos e Controles , Dinamarca , Feminino , Antígeno HLA-B27/genética , Humanos , Imunoglobulina A/imunologia , Imunoglobulina G/imunologia , Imageamento por Ressonância Magnética , Masculino , Sacroileíte/diagnóstico por imagem , Sacroileíte/genética , Sorogrupo , Espondiloartropatias/diagnóstico por imagem , Espondiloartropatias/genética , Espondilite Anquilosante/diagnóstico por imagem , Espondilite Anquilosante/genética , Espondilite Anquilosante/imunologia , Adulto JovemRESUMO
BACKGROUND: While much is known about childhood atopic dermatitis, little is known about persistence of atopic dermatitis into adult life. We report, to our knowledge for the first time, the clinical course of atopic dermatitis in an unselected cohort of adolescents followed into adulthood. METHODS: The course of atopic dermatitis from adolescence to adulthood was studied prospectively in a cohort of unselected 8th-grade schoolchildren established in 1995 and followed up in 2010 with questionnaire and clinical examination. RESULTS: The lifetime prevalence of atopic dermatitis was high (34.1%), and a considerable number of adults still suffered from atopic dermatitis evaluated both by questionnaire (17.1%) and clinical examination (10.0%). Persistent atopic dermatitis was found in 50% of those diagnosed in school age, and persistent atopic dermatitis was significantly associated with early onset, childhood allergic rhinitis and hand eczema. A close association was also found with allergic contact dermatitis and increased specific IgE to Malassezia furfur, but not with filaggrin gene defect. CONCLUSION: Persistence of atopic dermatitis in adulthood is common and affects quality of life. Persistent atopic dermatitis is particularly prevalent in those with early onset, allergic rhinitis and hand eczema in childhood. It is important to recognizing atopic dermatitis as a common and disabling disease not only in children but also in adults.
Assuntos
Dermatite Atópica/epidemiologia , Adolescente , Adulto , Fatores Etários , Asma/epidemiologia , Estudos de Casos e Controles , Criança , Pré-Escolar , Estudos de Coortes , Comorbidade , Estudos Transversais , Dermatite Alérgica de Contato/epidemiologia , Dermatite Atópica/diagnóstico , Dermatite Atópica/etiologia , Eczema/epidemiologia , Feminino , Proteínas Filagrinas , Seguimentos , Humanos , Incidência , Lactente , Recém-Nascido , Proteínas de Filamentos Intermediários/genética , Masculino , Mutação , Razão de Chances , Prevalência , Qualidade de Vida , Rinite Alérgica/epidemiologia , Fatores de Risco , Testes Cutâneos , Inquéritos e Questionários , Adulto JovemRESUMO
CONTEXT: In pregnancy, vitamin D insufficiency and deficiency, defined as serum 25-hydroxyvitamin D (25(OH)D) <50 nM, and <25 nM, respectively, may have adverse effects for both mother and child. Prevalence estimates, and identification of subgroups at special risk, may be useful for the planning of preventive strategies. OBJECTIVE: To study the prevalence and risk factors of hypovitaminosis D in early pregnancy. DESIGN AND METHODS: In a cross-sectional study of 1348 women in early pregnancy from the Odense Child Cohort, Denmark, 25(OH)D was determined and correlated to demographic and lifestyle variables (age, nationality, skin tone, parity, prepregnancy body mass index (BMI), smoking and sun exposure), using multiple linear and logistic regression analyses for all year, or stratified for summer and winter. The risk of vitamin D insufficiency was expressed as odds ratios (OR) with 95% confidence intervals in brackets. RESULTS: The prevalence of vitamin D insufficiency and deficiency was estimated to 27·8% and 3·5% respectively. In adjusted analyses, vitamin D insufficiency was directly associated with winter season, OR = 1·89 (1·35-2·63); increasing prepregnancy BMI, OR = 1·06 (1·03-1·10); and smoking, OR = 2·7 (1·34-5·41); but was less frequent in nulliparous, OR = 0·47 (0·33-0·68) and tanned Caucasians, OR = 0·63 (0·41-0·97). Season-specific associations having parental origin from outside Europe in summer, OR = 4·13 (1·41-12·13); in winter smoking, OR = 3·15 (1·19-8·36); and prepregnancy BMI, OR = 1·12 (1·06-1·18). CONCLUSIONS: Vitamin D insufficiency was widespread in early pregnancy. Associations to smoking, prepregnancy BMI and origin outside Europe varied with season. Multiparity and not being tanned in Caucasians represent new risk factors of vitamin D insufficiency.
Assuntos
Paridade , Bronzeado , Deficiência de Vitamina D/sangue , Vitamina D/análogos & derivados , Adolescente , Adulto , Índice de Massa Corporal , Estudos de Coortes , Estudos Transversais , Dinamarca , Feminino , Humanos , Razão de Chances , Gravidez , Complicações na Gravidez , Prevalência , Fatores de Risco , Estações do Ano , Luz Solar , Vitamina D/biossíntese , Deficiência de Vitamina D/epidemiologia , População Branca , Adulto JovemRESUMO
Chronic granulomatous disease (CGD) is a rare inherited disorder of the innate immune system caused by a defect in NADPH oxidase, leaving the granulocytes unable to kill invading microorganisms. CGD is caused by mutation in one of the five components gp91phox, p22phox, p47phox, p67phox and p40phox, encoded by the X-linked CYBB gene and the autosomal CYBA, NCF1, NCF2 and NCF4 genes respectively. We have collected samples from all Danish patients with known CGD followed in the clinic or newly diagnosed during a 5-year period, a cohort of 27 patients, and characterized them genetically. The cohort includes 10 male patients with X-linked CGD and one female with extremely lyonized expression of a defective CYBB allele. Six patients had mutation in CYBA. Seven of 10 patients with a defect in NCF1 were homozygous for the common GT deletion, one was compound heterozygous for the GT deletion and a splice-site mutation, and two patients were homozygous for a nonsense mutation in exon 7. Three novel mutations were detected, a deletion of exon 6 in CYBA, a duplication of exon 8-13 in CYBB and a splice site mutation in intron 7 of NCF1.
Assuntos
Doença Granulomatosa Crônica/genética , NADPH Oxidases/genética , Adolescente , Adulto , Criança , Pré-Escolar , Dinamarca , Feminino , Humanos , Lactente , Masculino , Glicoproteínas de Membrana/genética , Mutação , NADPH Oxidase 2RESUMO
Considering the female predominance in most of the autoimmune disorders that associate with the PTPN22 Trp620 variant and the complexity by which this variant influences immunologic tolerance, the objective of this study was to ascertain if the allele-specific expression of the disease-associated Arg620Trp polymorphism is affected by cis-acting or sex-specific trans-acting factor/s (e.g. sex-hormones). The use of the allele-specific transcript quantification of the Arg620Trp encoding 1858T polymorphism revealed no difference in the expression of the 1858C- and T-alleles in non-stimulated peripheral blood mononuclear cells (PBMCs) from non-pregnant female subjects, male subjects or pregnant female subjects in first or third trimester (P=0.70), respectively. While the transcription of PTPN22 in anti-CD3/anti-CD28 stimulated PBMCs increased fourfold (P<0.0001) and 13-fold (P<0.0001) after 48 and 72 h of activation, respectively, the expression of PTPN22 1858C- and T-alleles increased to the same extent (P=0.64). The present result essentially excludes such phenomena as a partial explanation for the female predominance in most of the autoimmune disorders that associate with the PTPN22 Trp620 variant.
Assuntos
Alelos , Substituição de Aminoácidos/genética , Doenças Autoimunes/genética , Expressão Gênica , Proteínas Tirosina Fosfatases/genética , Proteínas Tirosina Fosfatases/metabolismo , Primers do DNA , Eletroforese Capilar , Feminino , Humanos , Leucócitos Mononucleares/metabolismo , Masculino , Polimorfismo de Fragmento de Restrição , Gravidez , Proteína Tirosina Fosfatase não Receptora Tipo 22 , Fatores SexuaisRESUMO
Common variable immunodeficiency, CVID, is a primary antibody deficiency characterized by decreased levels of serum immunoglobulin G (IgG), decreased IgA and/or IgM and recurrent infections. It is assumed to be heterogeneous group of disorders caused by different genetic defects. Some patients have decreased levels of class switched memory B cells and/or decreased levels of somatic hypermutation which points to defects in the germinal centre (GC) reactions as cause of the disease in these patients. The inducible costimulator, ICOS, and its' ligand, ICOSL, are both involved in and necessary for the GC reaction and so is activation-induced cytidine deaminase, AID. Therefore, we sequenced the ICOS, ICOSL and AID genes in a cohort of 34 Danish CVID patients. We found 13 new single nucleotide polymorphisms (SNP) in the intron regions of the ICOSL gene as well as one SNP in exon 3. However, none of these polymorphisms were associated with CVID. We did not find a previously reported CVID-causing ICOS gene deletion or any other unique mutations in the ICOS or AID genes.
Assuntos
Antígenos de Diferenciação de Linfócitos T/genética , Imunodeficiência de Variável Comum/genética , Citosina Desaminase/genética , Proteínas/genética , Adolescente , Adulto , Idoso , Alelos , Antígenos CD , Criança , Estudos de Coortes , Citidina Desaminase , Dinamarca , Feminino , Predisposição Genética para Doença , Humanos , Ligante Coestimulador de Linfócitos T Induzíveis , Proteína Coestimuladora de Linfócitos T Induzíveis , Masculino , Pessoa de Meia-Idade , Polimorfismo GenéticoRESUMO
The human IGHG2 gene locus is polymorphic, encoding two known allotypes of IgG2: G2m(n-) and G2m(n+). The allele prevalence varies greatly between different ethnic groups and individual genotypes correlate with the level of plasma IgG2 and with antibody responses to certain polysaccharide antigens. In this study, we present three new alleles of IGHG2 (IGHG2*03, 04, and 05), and a complete sequence specific PCR typing system allowing discrimination between the different allotypes of IgG2. A hitherto unknown allotype, which we name G2m(ny), is encoded by IGHG2*04 and differs from G2m(n-) by asparagine rather than serine in CH1 residue 75 and by phenylalanine rather than leucine in CH1 residue 76 (EU numbering 192 and 193). The polymorphic residues are probably surface exposed near the hinge region. The same residues are also found in IgG1, IgG3, and IgG4, and G2m(ny) is therefore an isoallotype that probably arises by gene conversion within the heavy chain locus. The IGHG2*04 allele is present among Danish Caucasians with a low prevalence (2.5%), but was not found in Japanese or Mozambicans. The two other new alleles (IGHG2*03 and IGHG2*05) both encode the G2m(n-) allotype. The IGHG2*03 allele encodes most of the IgG2 of the G2m(n-) allotype in Danish Caucasians.
Assuntos
Alelos , Frequência do Gene , Imunoglobulina G/genética , Sequência de Aminoácidos , Sequência de Bases , Dinamarca , Humanos , Japão , Desequilíbrio de Ligação , Dados de Sequência Molecular , Moçambique , Polimorfismo Conformacional de Fita SimplesRESUMO
The extremely high risk reported for some types of cancer among patients with common variable immunodeficiency (CVID) is based on a limited number of investigations. Therefore, we examined the risks for cancer among 562 Danish and Swedish patients with CVID or IgA deficiency and 2071 relatives in 1958-96. The patients were identified through an Immunodeficiency Register and hospital records, while the relatives were traced through population registers. Cancer incidence was assessed by linkage to the Cancer Registries and compared with that in the general population. Among 386 patients with IgA deficiency, the incidence of cancer was not increased (standardized incidence ratio (SI) = 1.0); but two cases of stomach cancer were found, resulting in a non-significant increase in risk (SIR = 5.4; 95% CI = 0.7-19.5). Among 176 patients with common variable immunodeficiency (CVID), the incidence of cancer at all sites combined was increased (SIR = 1.8; 95% CI = 1.0-2.9), which was due mainly to significant excesses of malignant lymphoma (obs = 4; SIR = 12.1; 95% CI = 3.3-31.0) and of stomach cancer (obs = 3; SIR = 10.3; 95% CI = 2.1-30.2). Among the 626 relatives of patients with CVID, no increase in risk was found for these types of cancer or for cancer overall (obs = 53; SIR = 1.0; 95% CI = 0.8-1.3). Our data show that the risks for malignant lymphoma and stomach cancer among patients with CVID may be lower than reported previously. The absence of an increased risk among relatives suggests that the increased cancer morbidity in patients with CVID is related to the immunodeficiency per se rather than to specific genetic traits shared with their relatives.
Assuntos
Imunodeficiência de Variável Comum/complicações , Deficiência de IgG/complicações , Neoplasias/etiologia , Adulto , Idoso , Dinamarca/epidemiologia , Feminino , Humanos , Incidência , Recém-Nascido , Linfoma/epidemiologia , Linfoma/etiologia , Masculino , Pessoa de Meia-Idade , Neoplasias/epidemiologia , Pais , Risco , Distribuição por Sexo , Irmãos , Neoplasias Gástricas/epidemiologia , Neoplasias Gástricas/etiologia , Suécia/epidemiologiaRESUMO
Here we describe a hitherto unknown proline/threonine polymorphism at residue 72 of the human IgG2 CH1 domain (EU numbering 189) and show that it is linked to the known valine/methionine polymorphism at residue 52 of CH2 (EU numbering 282) defining the G2m(n+)/G2m(n-) allotypes. We sequenced the entire constant region of the heavy-chain gene for secreted IgG2 in five IGHG2*02 homozygous individuals covering CH1, hinge, CH2, and CH3 regions (approximately 2 kb). Proline 72 in CH1 of G2m(n-) is changed to threonine in the G2m(n+) [G2m(23)] allotype. Based on the crystal structure of human IgG1, this amino acid position is expected to be surface exposed in IgG2. Besides this structural difference, we identified two silent nucleotide polymorphisms in the CH1 region and seven in the introns. Finally, we developed a sequence-specific PCR typing system detecting the polymorphisms in the CH1 and CH2 regions. We typed 64 Danish Caucasians and found that the CH1 and CH2 region polymorphisms are in complete linkage disequilibrium in this population.
Assuntos
Éxons/genética , Genes de Imunoglobulinas/genética , Éxons Codificadores da Região de Dobradiça , Imunoglobulina G/genética , Cadeias Pesadas de Imunoglobulinas/genética , Desequilíbrio de Ligação , Polimorfismo Genético/genética , População Branca/genética , Alelos , Sequência de Aminoácidos , Sequência de Bases , Códon/genética , DNA Polimerase Dirigida por DNA/metabolismo , Frequência do Gene/genética , Haplótipos/genética , Humanos , Imunoglobulina G/química , Cadeias Pesadas de Imunoglobulinas/química , Modelos Moleculares , Dados de Sequência Molecular , Países Baixos , Polimorfismo de Fragmento de Restrição , Conformação Proteica , Análise de Sequência de DNA , Taq Polimerase/metabolismoRESUMO
Protective antibodies to the important childhood pathogen Haemophilus influenzae type b (Hib) are directed against the capsular polysaccharide (HibCP). Most of the antibody is encoded by a well-defined set of ("canonical") immunoglobulin genes, including the Vkappa A2 gene, and expresses an idiotypic marker (HibId-1). In comparison to noncanonical antibodies, the canonical antibody is generally of higher avidity, shows higher levels of in vitro bactericidal activity, and is more protective in infant rats. Using site-directed mutagenesis, we here characterize canonical HibCP antibodies expressed as antigen-binding fragments (Fabs) in Escherichia coli, define amino acids involved in antigen binding and idiotype expression, and propose a three-dimensional structure for the variable domains. We found that canonical Fabs, unlike a noncanonical Fab, bound effectively to HibCP in the absence of somatic mutations. Nevertheless, pronounced mutation-based affinity maturation was demonstrated in vivo. An almost perfect correlation was found between unmutated gene segments that mediated binding in vitro and those encoding canonical HibCP antibodies in vivo. Thus, the Vkappa A2a gene could be replaced by the A2c gene but not by the highly homologous sister gene, A18b, corresponding to the demonstrated usage of A2c but not of A18b in vivo. Similarly, only Jkappa1 and Jkappa3, which predominate in the response in vivo, were able to facilitate binding in vitro. These findings suggest that the restricted immunoglobulin gene usage in HibCP antibodies reflects strict structural demands ensuring relatively high affinity prior to somatic mutations-requirements met by only a limited spectrum of immunoglobulin gene combinations.
Assuntos
Anticorpos Antibacterianos/química , Haemophilus influenzae tipo b/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Antibacterianos/genética , Afinidade de Anticorpos , Sequência de Bases , Criança , Clonagem Molecular , Primers do DNA/genética , Escherichia coli/genética , Genes de Imunoglobulinas , Infecções por Haemophilus/genética , Infecções por Haemophilus/imunologia , Infecções por Haemophilus/prevenção & controle , Humanos , Fragmentos Fab das Imunoglobulinas/química , Fragmentos Fab das Imunoglobulinas/genética , Fragmentos de Imunoglobulinas/química , Fragmentos de Imunoglobulinas/genética , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Polissacarídeos Bacterianos/imunologia , Conformação Proteica , Ratos , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologiaRESUMO
The Ab response of a healthy adult to the first dose of a Haemophilus influenzae type b capsular polysaccharide (HibCP) conjugate vaccine was studied at the level of Ig gene usage by circulating Ab-secreting cells. Forty-one IgA and 17 IgG mRNA sequences were obtained. The major part of the response was confined to IgA Ab-secreting cells, and 72% of the IgA sequences were derived from the progeny of a single rearranged B cell. These sequences could be arranged in a genealogical tree showing multiple somatic mutations and at least two intraclonal isotype switches to IgA2. Fourteen somatic mutations were shared by this clonal progeny, indicating that extreme clonal selection had occurred early in the clonal development. Taking into account the frequency of somatic mutations and the clone size, it was evident that the responding cell population must have originated from a mutated, highly selected, and expanded population of cells existing before vaccination, i.e., memory B cells. The dominating heavy and light chains of the response were combined in a Fab that bound HibCP. It was shown that the shared heavy and light chain mutations increased the affinity for HibCP considerably, indicating that the clonal selection had been driven by affinity. Pre-existing memory cells in unvaccinated adults may explain several features of Ab responses to polysaccharide vaccines and may play a role in acquiring the ability to respond to pure polysaccharides during infancy.
Assuntos
Afinidade de Anticorpos , Subpopulações de Linfócitos B/imunologia , Vacinas Anti-Haemophilus/imunologia , Imunoglobulina A/biossíntese , Isotipos de Imunoglobulinas/biossíntese , Memória Imunológica/imunologia , Ativação Linfocitária/imunologia , Polissacarídeos Bacterianos/imunologia , Adulto , Sequência de Aminoácidos , Afinidade de Anticorpos/genética , Subpopulações de Linfócitos B/metabolismo , Cápsulas Bacterianas , Sequência de Bases , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Células Clonais/imunologia , Células Clonais/metabolismo , Rearranjo Gênico de Cadeia Pesada de Linfócito B , Rearranjo Gênico de Cadeia Leve de Linfócito B , Vacinas Anti-Haemophilus/administração & dosagem , Humanos , Esquemas de Imunização , Imunoglobulina A/genética , Imunoglobulina A/metabolismo , Fragmentos Fab das Imunoglobulinas/genética , Fragmentos Fab das Imunoglobulinas/metabolismo , Isotipos de Imunoglobulinas/genética , Isotipos de Imunoglobulinas/metabolismo , Região Variável de Imunoglobulina/genética , Região Variável de Imunoglobulina/metabolismo , Memória Imunológica/genética , Ativação Linfocitária/genética , Masculino , Dados de Sequência Molecular , Mutação/imunologia , Polissacarídeos Bacterianos/administração & dosagem , Vacinas Conjugadas/administração & dosagem , Vacinas Conjugadas/imunologiaRESUMO
The purpose of this study was to determine the in-vivo and in-vitro effects of insulin, at physiological and supraphysiological concentrations, on the human immune system. Ten healthy young men went through a sequential two-step hyperinsulinaemic euglycaemic clamp. Plasma insulin concentrations were increased from baseline (9.0 microU/ml) to 49.1 microU/ml after 1 h of insulin infusion (step I) and to 1281 microU/ml (step II) after 2 h of infusion. As control experiments infusions of isotonic saline were performed. The unstimulated natural killer (NK) cell activity among blood mononuclear cells (BMNC) increased in response to supraphysiological plasma insulin levels (baseline versus step II: 20.6 +/- 11.3 versus 27.8 +/- 14.4%). The percentages of the D16+ NK cells did not change, indicating an enhanced cytotoxic capability per individual NK cell. Insulin also slightly increased the activity of NK cells in vitro. A decline at step II in the concentrations of monocytes (0.29 +/- 0.09 versus 0.12 +/- 0.03 x 10(9)/L), lymphocytes (1.57 +/- 0.46 versus 1.22 +/- 0.25 x 10(9)/L), and CD16+(24.2 +/- 17.5 versus 16.7 +/- 11.2 x 10(7)/L), CD14+ (20.9 +/- 10.8 versus 8.6 +/- 3.9 x 10(7)/L), HLA-DR+ (37.2 +/- 22.1 versus 19.2 +/- 10.7 x 10(7)/L) and CD45RO+ (91.6 +/- 33.4 versus 61.7 +/- 6.4 x 10(7)/L) cells as well as in the percentages of CD14+ cells (11.2 +/- 4.7 versus 6.4 +/- 2.3%) and CD14+/HLA-DR+ monocytes (9.7 +/- 3.9 versus 4.8 +/- 2.8%) were observed. No changes were found at step I. Hyperinsulinaemia did not change the percentages of the CD3+, CD4+, CD8+, CD19+, CD56+, CD11a+, CD45RO+ and CD45RA+ cells, the numbers of circulating immunoglobulin (Ig)G-, IgA- and IgM- secreting cells, or the proliferative responses of BMNC to phytohaemagglutinin, purified derivative of tuberculin or interleukin (IL)-2. Hyperinsulinaemia did not change the in-vitro sensibility to insulin. In conclusion, supraphysiological insulin levels increased the activity of the individual NK cells, but decreased the numbers of NK cells, lymphocytes and activated monocytes. The findings are presumably of minor clinical relevance but may indicate an insulin-induced immune activation.
Assuntos
Hiperinsulinismo/imunologia , Insulina/imunologia , Adulto , Antígenos CD/imunologia , Técnica Clamp de Glucose , Humanos , Insulina/administração & dosagem , Insulina/sangue , Células Matadoras Naturais/imunologia , MasculinoRESUMO
We describe a hitherto unknown functional IGKV gene, VkLa, belonging to the IGKV1 subgroup with exon 2 having only 94% similarity to the closest known IGKV gene, 1-13/1D-13 (L4/L18a). Genomic DNA sequences spanning from 5' of the decanucleotide box to 3' of the heptamer (649 bp) were cloned and sequenced from four individuals. The new gene encodes the conserved amino acids in the exons and contains no apparent defects in known regulatory intron sequences such as pd-box, dc-box, TATA-box, CCCT-elements, splice-sequences, initiation codon, and heptamer sequence. VkLa is therefore potentially functional and, correspondingly, we found transcripts of properly rearranged VkLa with somatical hypermutations. VkLa was found in 12 of 57 (21%) healthy Caucasians by a nested polymerase chain reaction and subsequent sequencing of exon 2. This finding shows that there is more inter-individual variation in the available IGKV gene repertoire than was hitherto assumed. Finally, we describe a minor correction in the IGKV1D-43 (L23) gene sequence.
Assuntos
Imunoglobulinas/genética , Sequência de Aminoácidos , Sequência de Bases , Cromossomos Humanos Par 2 , Clonagem Molecular , DNA , Expressão Gênica , Frequência do Gene , Genes de Imunoglobulinas , Humanos , Cadeias Leves de Imunoglobulina/genética , Dados de Sequência Molecular , Família MultigênicaRESUMO
The expressed human kappa light chain gene repertoire utilized by healthy individuals was studied by two different single-sided specific PCR techniques to avoid bias for certain V genes. A total of 103 rearranged kappa sequences from peripheral blood mononuclear cells from healthy individuals were cloned from cDNA and assigned to the Vkappa and Jkappa germ-line genes with the closest overall homology. The use of cDNA rather than genomic DNA focused the analysis on activated B cells rich in mRNA. Accordingly, the sequences represented the applied repertoire and almost all were somatically mutated. V genes from the Jkappa-proximal duplication unit of the kappa locus were almost exclusively used. A total of 65% of the sequences could be assigned to four or five genes: A27 (humkv325), L6 (Vg), L2 (humkv328), and A3 and/or A19. N additions and P nucleotides were quite common and found in 32% and 21% of the sequences, respectively. Extended CDR3s more than nine residues in length were found in 18% of the sequences, and in 71% of cases this was due to insertion of an extra proline residue. This proline was usually explained from the germ-line sequences involved. These results are in good agreement with those of previous repertoire studies using potentially V-gene-biased techniques. Thus, it is clear that restricted V-gene usage, common N and P additions, and extended CDR3 regions are normal features and not, as has been claimed, characteristics of pathological autoantibodies.
Assuntos
Genes de Imunoglobulinas , Cadeias kappa de Imunoglobulina/genética , Mutação , Autoanticorpos/genética , Linfócitos B/imunologia , Linfócitos B/metabolismo , Células Cultivadas , Clonagem Molecular , DNA Complementar/genética , Rearranjo Gênico , Variação Genética , Humanos , Leucócitos Mononucleares , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Análise de Sequência de DNARESUMO
Immunoglobulin gene polymorphisms are interesting because they reflect differences in the available antibody repertoire which may affect the susceptibility to specific infections. Until recently, the human V kappa gene, A18, was known as a nonfunctional gene only. In this study, we cloned and sequenced four apparently functional alleles and determined the gene frequencies in three well-defined populations: Danish Caucasians, eastern Greenland Eskimos and Mozambican blacks. The A18b allele that was recently described in Native American Navajos by Atkinson et al. was found in all three populations with gene frequencies of 8%, 45% and 23% in Caucasians, Eskimos and blacks, respectively. Conversely, the frequencies of the nonfunctional A18a allele were 92%, 55% and 57%. Further, three new A18 alleles, c, d, and e were found exclusively in blacks, among whom they had an total frequency of 19%. These data indicate that both the A18a and A18b alleles originated before the diversification of Africans and non-Africans 90,000 years ago, whereas the A18c, A18d and A18e alleles may have a more recent origin. The functionality of the A18b allele was documented by the demonstration of properly rearranged and somatically hypermutated A18b messenger RNA present in the blood lymphocytes of individuals carrying this allele. The expression clearly exceeded that of a known functional V gene, A2, indicating that functional A18 alleles contribute significantly to the available antibody repertoire. In this context, it is surprising that the functional A18b allele apparently has been negatively selected in the Caucasian population, among whom 85% completely lack a functional gene.
Assuntos
População Negra/genética , Região Variável de Imunoglobulina/genética , Cadeias kappa de Imunoglobulina/genética , Inuíte/genética , Polimorfismo Genético , População Branca/genética , Alelos , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , DNA Complementar , Dinamarca , Expressão Gênica , Humanos , Dados de Sequência Molecular , Moçambique , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , RNA MensageiroRESUMO
Restricted V region diversity is a key feature of Abs to many haptens and simple polysaccharides. Two possible mechanisms exist: 1) selection of many clonally unrelated B cells using very similar or identical VDJ and VJ rearrangements; and 2) selection of a heavily expanded progeny of few virgin B cells. How many virgin B cells eventually give rise to the total Ab response to a simple Ag is a fundamental immunologic question. In this report, we address this question in human adults by analyzing the rearranged VkappaJkappa genes of B cells responding to a single dose of the capsular polysaccharide of Haemophilus influenzae type b coupled to tetanus toxoid. We combined affinity purification of circulating vaccine-induced Ab-secreting cells with PCR amplification of cDNA followed by cloning and sequencing. Forty-eight and 42 kappa VJ gene transcripts were analyzed from two adults, respectively. Both individuals used extremely restricted repertoires with >90% of the cells using a single Vkappa gene rearranged to a single Jkappa gene. Despite the fact that the Ab responses engaged high numbers of Ab-secreting cells, analysis of the many shared, somatically acquired mutations showed that the majority of the cells originated from a common virgin B cell. Kinetic considerations implied that an extremely selected population of hypermutated memory B cells must have existed in these individuals before the first systemic immunization with the Ag. A possible role for the mucosal immune system in the priming and selection of these cells is proposed.
Assuntos
Antígenos de Bactérias/imunologia , Linfócitos B/imunologia , Células Clonais/imunologia , Vacinas Anti-Haemophilus/imunologia , Haemophilus influenzae/imunologia , Memória Imunológica , Polissacarídeos Bacterianos/imunologia , Adulto , Antígenos de Bactérias/biossíntese , Antígenos de Bactérias/genética , Cápsulas Bacterianas , Sequência de Bases , Feminino , Rearranjo Gênico de Cadeia Leve de Linfócito B , Genes de Imunoglobulinas , Humanos , Região Variável de Imunoglobulina/genética , Cadeias kappa de Imunoglobulina/genética , Masculino , Dados de Sequência Molecular , RNA Mensageiro/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , VacinaçãoRESUMO
OBJECTIVE: To evaluate the antibody response to a Haemophilus influenzae type b capsular polysaccharide (HibCP) tetanus toxoid (TT) conjugate vaccine (HibCP-TT) in preterm infants. SUBJECTS: Thirty-five healthy preterm infants with gestational ages (GA) from 27 to 36 weeks and birth weights from 920 to 2550 g. Controls were 37 term infants. METHODS: All infants were immunized with HibCP-TT at 2, 4 and 12 months of age. Antibodies to HibCP and TT were determined at each immunization and 1 month after the second and third. RESULTS: After two doses of HibCP-TT the preterm infants with GAs < or = 30 weeks (n = 8; mean GA, 29.5 weeks) had a significantly lower HibCP antibody response than the preterm infants with GAs > 30 weeks (n = 23; mean GA, 34.2 weeks) (P = 0.004), who for their part had a response not significantly different from that of the term infants. After the third dose there were no significant differences among the groups. The response to the TT part of the vaccine showed the same pattern. CONCLUSION: Although the most immature infants may show an inadequate antibody response to the initial immunizations, many preterm infants can benefit from vaccination with HibCP-TT when starting immunization at the same chronologic age as term infants.
Assuntos
Anticorpos Antibacterianos/biossíntese , Infecções por Haemophilus/prevenção & controle , Vacinas Anti-Haemophilus/imunologia , Doenças do Prematuro/imunologia , Doenças do Prematuro/microbiologia , Polissacarídeos Bacterianos/imunologia , Anticorpos Antibacterianos/análise , Cápsulas Bacterianas/imunologia , Humanos , Lactente , Recém-Nascido , Polissacarídeos Bacterianos/administração & dosagem , Antitoxina Tetânica/análise , Toxoide Tetânico/imunologia , Vacinas Conjugadas/imunologiaRESUMO
Children with recurrent lower respiratory tract infection (RLRI) may respond poorly to polysaccharide antigens. To examine how such children respond to a polysaccharide coupled to a protein carrier, we immunized 15 children with RLRI aged 8-69 months and 15 carefully age-matched healthy controls once with a Haemophilus influenzae type b (Hib) conjugate vaccine. Total IgG subclasses, total antipolysaccharide Hib antibodies, and antipolysaccharide Hib antibodies of IgM, IgG, IgA, and IgG1-4 specificity were determined by ELISA. There were no significant differences between the two groups in any single total IgG subclass, but total IgG measured as the sum of all four subclasses was significantly lower in the children with RLRI than in the controls (P = 0.036). Before vaccination, the children with RLRI had significantly less IgG antipolysaccharide Hib antibody than the controls (P = 0.005), whereas 1 month later they had significantly more IgM antibody (P = 0.038). No other significant differences were found between the groups before or after immunization with respect to antipolysaccharide Hib antibodies. Since naturally occurring IgG antibodies are thought to be acquired partly as a consequence of antigenic stimulation on mucosal surfaces, we hypothesize that the low level of specific IgG found before immunization, as well as the low total IgG in the children with RLRI, may reflect an impaired ability to prime through mucosal surfaces. This is supported by our finding of an increased IgM response to Hib conjugate vaccine in these children, since this isotype predominates in the primary immune response, i.e., in the absence of immunologic memory.(ABSTRACT TRUNCATED AT 250 WORDS)