Influence of the polymorphism of the DUSP14 gene on the expression of immune-related genes and development of pulmonary tuberculosis.

Recently, a genome-wide screening identified a functional single-nucleotide polymorphism in dual-specificity phosphatase 14 gene (DUSP14), which was associated with pulmonary tuberculosis (TB) in a West African study. DUSP14 regulates T-cell proliferation and cytokine production in a negative way via dephosphorylation and inactivation of key signaling molecules. The aim of this study is to further explore the possible significance of the DUSP14 polymorphism. Total RNA was extracted from the whole blood of 109 healthcare workers (HCWs) in Vietnam and subjected to quantitative reverse-transcription PCR for DUSP14 and 20 immune-related genes. DUSP14 rs1051838 was genotyped in 502 new pulmonary TB patients and 506 healthy controls. Among disease-free individuals (HCWs), T-helper type-1 (Th1)-related genes, interferon-gamma receptor 2 (IFNGR2) and signal transducer and activator of transcription-1 (STAT1) mRNA levels significantly increased as the number of A alleles of rs1051838 increased, whereas the DUSP14 mRNA level tended to decrease. The AA genotype was associated with protection against active TB in younger patients (⩽45 years old, OR=0.63, 95% CI 0.44-0.90). Our results suggest that a low-expression genotype of DUSP14 accompanied by high transcript levels of Th1 immune-related genes may confer protection against early TB development.

Association of TLR polymorphisms with development of tuberculosis in Indonesian females.

Tuberculosis (TB) is caused by Mycobacterium tuberculosis and is a major cause of morbidity and mortality worldwide. Many candidate genes have been investigated for a possible association with TB. Toll-like receptors (TLRs) are known to play important roles in human innate immune systems. Polymorphisms in and functions of TLRs have been investigated to identify associations with specific infectious diseases, including TB. Here, we examined whether single-nucleotide polymorphisms (SNPs) in TLRs and genes in TLR signaling were associated with TB susceptibility in Indonesian and Vietnamese populations. A statistically significant association was observed between TB susceptibility in a classified Indonesian female group and rs352139, an SNP located in the intron of TLR9, using the genotype (P = 2.76E-04) and recessive (AA vs AG+GG, P = 2.48E-04, odds ratio = 1.827, 95% confidence interval = 1.321-2.526) models. Meta-analysis of the Indonesian and Vietnamese populations showed that rs352139 was significantly associated with TB in the recessive model. This finding indicated that a TLR9 polymorphism might have an important role in the susceptibility to M. tuberculosis in Asian populations.

Association analysis of susceptibility candidate region on chromosome 5q31 for tuberculosis.

Chromosome 5q31 spans the T helper (Th) 2-related cytokine gene cluster, which is potentially important in Th1/Th2 immune responses. The chromosome 5q23.2-31.3 has been recently identified as a region with suggestive evidence of linkage to tuberculosis in the Asian population. With the aim of fine-mapping a putative tuberculosis susceptibility locus, we investigated a family-based association test between the dense single nucleotide polymorphism (SNP) markers within chromosome 5q31 and tuberculosis in 205 Thai trio families. Of these, 75 SNPs located within candidate genes covering SLC22A4, SLC22A5, IRF1, IL5, RAD50, IL13, IL4, KIF3A and SEPT8 were genotyped using the DigiTag2 assay. Association analysis revealed the most significant association with tuberculosis in haplotypes comprising SNPs rs274559, rs274554 and rs274553 of SLC22A5 gene (P(Global)=2.02 x 10(-6)), which remained significant after multiple testing correction. In addition, two haplotypes within the SLC22A4 and KIF3A region were associated with tuberculosis. Haplotypes of SLC22A5 were significantly associated with the expression levels of RAD50 and IL13. The results show that the variants carried by the haplotypes of SLC22A4, SLC22A5 and KIF3A region potentially contribute to tuberculosis susceptibility among the Thai population.

Genome-wide SNP-based linkage analysis of tuberculosis in Thais.

Tuberculosis, a potentially fatal infectious disease, affects millions of individuals annually worldwide. Human protective immunity that contains tuberculosis after infection has not been clearly defined. To gain insight into host genetic factors, nonparametric linkage analysis was performed using high-throughput microarray-based single nucleotide polymorphism (SNP) genotyping platform, a GeneChip array comprised 59 860 bi-allelic markers, in 93 Thai families with multiple siblings, 195 individuals affected with tuberculosis. Genotyping revealed a region on chromosome 5q showing suggestive evidence of linkage with tuberculosis (Z(lr) statistics=3.01, logarithm of odds (LOD) score=2.29, empirical P-value=0.0005), and two candidate regions on chromosomes 17p and 20p by an ordered subset analysis using minimum age at onset of tuberculosis as the covariate (maximum LOD score=2.57 and 3.33, permutation P-value=0.0187 and 0.0183, respectively). These results imply a new evidence of genetic risk factors for tuberculosis in the Asian population. The significance of these ordered subset results supports a clinicopathological concept that immunological impairment in the disease differs between young and old tuberculosis patients. The linkage information from a specific ethnicity may provide unique candidate regions for the identification of the susceptibility genes and further help elucidate the immunopathogenesis of tuberculosis.

Splice acceptor site mutation of the transporter associated with antigen processing-1 gene in human bare lymphocyte syndrome.

Expression of histocompatibility leukocyte antigen (HLA) class I molecules on the cell surface depends on the heterodimer of the transporter associated with antigen processing 1 and 2 (TAP1 and TAP2), which transport peptides cleaved by proteasome to the class I molecules. Defects in the TAP2 protein have been reported in two families with HLA class I deficiency, the so-called bare lymphocyte syndrome (BLS) type I. We have, to our knowledge, identified for the first time a splice site mutation in the TAP1 gene of another BLS patient. In addition, class I heavy chains (HCs) did not form the normal complex with tapasin in the endoplasmic reticulum (ER) of the cells of our patient.

A survey of tuberculosis prevalence in Hanoi, Vietnam.

OBJECTIVE: To assess the prevalence of tuberculosis (TB) in Hanoi, Vietnam, in 2003/2004. METHODS: A random selection was carried out involving 11624 subjects from 20 communes within the city. RESULTS: On chest X-ray examination, 317 subjects (2.73%) showed abnormal lung opacity, of which 17 were sputum smear-positive, two concentrated smear-positive and three culture-positive, all with active TB. The prevalence of sputum smear-positive pulmonary TB was 146 per 100000 in persons aged >or=15 years (95%CI 65-228). CONCLUSION: This is the first large-scale assessment of the prevalence of TB in Hanoi. The prevalence rate was higher than expected, suggesting that a significant number of patients with active TB, particularly females, remain undiagnosed, thus representing a continuing potential source of transmission in the community.

Serum concentration of soluble interleukin-2 receptor as a sensitive parameter of disease activity in sarcoidosis.

We investigated the clinical value of measuring serum concentrations of soluble IL-2R in monitoring sarcoidosis. Serum concentrations of soluble IL-2R were measured in 70 patients with sarcoidosis. The mean value for active untreated sarcoidosis was 1,143 +/- 509 U/ml, while the normal range in 97 healthy control subjects was 80 to 300 U/ml. The mean value for active untreated sarcoidosis was significantly higher than that for dormant disease (353 +/- 183 U/ml) or that for corticosteroid-treated patients (380 +/- 151 U/ml). Serial changes in serum soluble IL-2R level were studied in cases of spontaneous remission or in corticosteroid-treated patients; a good correlation was noted between the changes in serum level of soluble IL-2R and clinical status. A positive correlation was noted between serum concentration of soluble IL-2R and serum ACE activity. These data confirmed that measurement of serum concentration of soluble IL-2R could be used in monitoring the disease activity in sarcoidosis.

Detection of lymphomatous involvement of the lung by bronchoalveolar lavage. Application of immunophenotypic and gene rearrangement analysis.

We report three cases of pulmonary involvement of non-Hodgkin's lymphoma in which immunophenotypic or gene rearrangement analysis of bronchoalveolar lavage (BAL) cells demonstrated monoclonality of T- or B-cell lineage. The first patient had T-cell lymphoma and developed pulmonary lesions. Surface marker analysis of the BAL cells revealed that CD8-positive lymphoid cells were dominant and Southern blot analysis of T-cell receptor gene detected gene rearrangement demonstrating monoclonality of T-cell lineage. The second patient presented with diffuse micronodular shadows on chest radiograph. Marked B-lymphocytosis in BAL fluid prompted us to analyze their clonality. The third was a case in which recurrence of primary pulmonary lymphoma was suspected. In the second and third case, the finding of marked increase in the number of CD 19-positive B lymphocytes with a single class of light chains proved a monoclonal population in BAL cells. With the review of other cases in our study and the relevant literature, we conclude that the clonal analysis of BAL cells is helpful in establishing the diagnosis of pulmonary involvement of T- or B-cell lymphoma.

Effect of adenovirus E1A on ICAM-1 promoter activity in human alveolar and bronchial epithelial cells.

In previous studies we demonstrated that the E1A DNA and proteins of group C adenovirus are present in excess in the lungs of patients with chronic obstructive pulmonary disease (COPD). Because adenovirus EIA gene products are known to regulate the expression of many genes by interacting with cellular transcription factors, we postulated that E1A enhances the production of inflammatory mediators and exacerbates the inflammatory process in smokers' lungs. We reported that LPS-induced ICAM-1 expression in A549 cells is upregulated by E1A. In the current study we investigated whether this regulation is mediated through the ICAM-1 promoter. A549 cells and primary human bronchial epithelial (HBE) cells were transiently cotransfected with a plasmid containing the ICAM-1 enhancer-promoter linked to the chloramphenicol acetyltransferase (CAT) reporter gene (pBS-CAT-P) and either a plasmid carrying the adenovirus 5 E1A gene (pE1Aneo) or a control plasmid (pneo). To compare the effect of transient versus stable E1A expression on the activity of this promoter, we also transiently transfected stable E1A-expressing A549 cells with pBS-CAT-P. Transient cotransfection of pE1Aneo and pBS-CAT-P had no effect on basal ICAM-1 promoter activity in A549 or HBE cells. After stimulation of A549 cells with TNF-alpha, IFN-gamma, or LPS, promoter activity was increased by two- to threefold in the presence of adenovirus EIA. In HBE cells, on the other hand, E1A repressed the ICAM-1 promoter after stimulation with IFN-gamma and LPS with little change after TNF-alpha stimulation. In stable E1A transfectants, ICAM-1 promoter activity was 2 to 2.5 times higher than in control transfectants with or without stimulation with TNF-alpha or LPS. These findings suggest that EIA can modulate the activity of the ICAM-1 promoter in lung epithelial cells and this modulation is different in cells of alveolar origin compared to bronchial epithelial cells.

Erythromycin promotes monocyte to macrophage differentiation.

Recent reports have suggested that long-term administration of erythromycin (EM) appears to ameliorate some of chronic inflammatory processes where macrophages and lymphocytes play important roles. Our study was initiated to examine the effect of EM on monocyte-macrophage lineage in vitro. EM (1 approximately 100 micrograms/ml) significantly increased the number of adherent monocyte-derived macrophages after 7 days of culture. The combination of EM and macrophage colony stimulating factor (M-CSF) synergistically increased the number of monocyte-derived macrophages, while the combination of EM and granulocyte-macrophage colony stimulating factor exerted an additive effect. Culture with EM induced the expression of a surface antigen CD71, one of the activation markers of macrophages as compared with control cultures. The combination of EM plus M-CSF significantly enhanced H2O2-producing capacity of those cells as compared with M-CSF alone. A differentiation process of monocytoid THP-1 cells was also augmented by EM. These results indicate that EM promotes differentiation of human monocyte-macrophage lineage, altering their functions.

Antilymphocytic activity of erythromycin distinct from that of FK506 or cyclosporin A.

Erythromycin (EM), a macrolide antibiotic has been recently reported to depress the extent of inflammation irrespective of its antimicrobial action. Our study was initiated to examine the effect of EM on T cell proliferation in vitro, since other macrolide antibiotics FK506 and rapamycin (RAP) have been well known to possess strong immunosuppressive or anti-inflammatory potential. EM had a suppressive effect on the proliferative response of human lymphocytes stimulated with mitogens and antigens, while EM had no effect on concanavalin A (Con A)-induced interleukin-2 (IL-2) production or IL-2R alpha (CD25) expression. Delayed addition of EM after the first 48 hours of mitogenic stimulation did suppress IL-2-dependent proliferation of Con A blasts, whereas pretreatment with EM for the first 48 hours of stimulation did not impede the subsequent IL-2-dependent proliferation of obtained blast cells. The results indicate that EM suppresses T cell proliferation at a late stage in the activation process by impairing their response to IL-2. This antilymphocytic action of EM was quite distinct from that of FK506 or cyclosporin A (CsA) but was similar to that of RAP. Unlike RAP, however, EM did not antagonize FK506-induced suppression but potentiated the action of FK506 and CsA. The addition of an enteric hormone motilin, a receptor of which was previously found to be occupied by EM, unaffected the lymphocyte proliferation and the subsequent EM-induced suppression. These data suggest that EM operates through an undefined mechanism probably distinct from that of FK506, CsA, RAP or motilin.

[Tuberculosis in patients with acquired immune deficiency syndrome].

HIV-1 infection is a major cause of worldwide epidemic of tuberculosis. In Japan, the cumulative number of the patients reported is 131 by the end of 1999 with 10 to 20 annual new cases. Most of Japanese cases are advanced AIDS patients with low CD4 number less than 100/microliter. The peak age of Japanese patient is 40 to 60 years old, whereas that of foreigners is 20-30 years old, suggesting that most Japanese cases are recurrent tuberculosis. There is increasing clinical evidence that coinfection with M. tuberculosis accelerates progression of AIDS. We found that, in vivo, HIV-1 load and mutation increase in involved lung segments in patients with pulmonary tuberculosis. We also reported that Mycobacterium tuberculosis stimulates HIV-1 replication by enhancing transcription on the 5' LTR in a macrophage cell line, THP-1, in vitro. In contrast, HIV-1 replication is suppressed by M. tuberculosis infection of monocytes derived macrophages (MDM) or differentiated monocytic THP-1 cells. We observed that HIV-1 5' LTR function was repressed in PMA differentiated THP-1 cells after co-infection with M. tuberculosis. Point mutations in C/EBP-beta binding domains of the HIV-1 LTR negative regulatory element (NRE) abolished promoter repression. Monocyte-derived macrophages and differentiated THP-1 cells increased expression of the 16 kDa inhibitory from of C/EBP-beta after M. tuberculosis coinfection. Bronchoalveolar lavage cells obtained from normal controls and alveolar macrophages from uninflamed lung of tuberculosis patients also expressed the 16 kDa inhibitory form of C/EBP-beta. However, alveolar macrophages from lung segments involved with pulmonary tuberculosis had markedly reduced C/EBP-beta expression. These data suggest that 16 kDa isoform of C/EBP-beta plays an important role for the control of HIV-1 replication in macrophages. We propose derepression of HIV-1 LTR mediated transcription as one mechanism for enhanced HIV-1 replication observed in pulmonary tuberculosis.

[Genetic variation of NADPH/NADH oxidase and susceptibility to diffuse panbronchiolitis (DPB) and chronic obstructive pulmonary disease (COPD)].

Diffuse panbronchiolitis (DPB) and chronic obstructive pulmonary disease (COPD) are both characterized by chronic airflow obstruction of unknown etiology. It is hypothesized that neutrophils play the major role in the pathogenesis of these diseases. Recent studies have suggested that genetic factors may be related to individual susceptibility to these diseases. We have investigated the association between the C 242 T polymorphism of p 22 phox, a critical subunit of superoxide-generating NADH/NADPH oxidase, and susceptibility to DPB and COPD. Blood samples obtained from both patients with DPB (n = 82), COPD (n = 53), and control subjects (n = 82) were used for this genotyping assay; and the polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) were performed to genotype the gene. The frequency of the C allele was 0.91, 0.92, 0.92 in the DPB, COPD, and control groups, respectively. There were no differences in the distribution of the genotype of p 22 phox among these groups, either. We concluded that there is no association between the C 242 T polymorphism of p 22 phox, and susceptibility to DPB and COPD.

HLA-A, -B, -C, -DRB1 and -DQB1 alleles and haplotypes in the Kinh population in Vietnam.

Allele and haplotype frequencies of the human leukocyte antigens (HLA) were studied in the Kinh Vietnamese population. We analyzed 170 unrelated healthy individuals. DNA-based HLA typing was performed using a microsphere-based array genotyping platform with sequence-specific oligonucleotide probes to distinguish HLA-A, -B, -C, -DRB1 and -DQB1 alleles. A total of 21 HLA-A, 37 HLA-B, 18 HLA-C, 25 HLA-DRB1, and 14 HLA-DQB1 alleles were identified. HLA-A*1101, A*2402, A*3303, B*1502, B*4601, Cw*0102, Cw*0702, Cw*0801, DRB1*1202, DQB1*0301, DQB1*0303, and DQB1*0501 were found with frequencies higher than 10%. Two representative haplotypes bearing two to five HLA loci were A*1101-B*1502 and A*3303-B*5801 for HLA-A-B; Cw*0801-B*1502 and Cw*0102-B*4601 for HLA-C-B; B*1502-DRB1*1202 and B*4601-DRB1*0901 for HLA-B-DRB1; DRB1*1202-DQB1*0301 and DRB1*0901-DQB1*0303 for HLA-DRB1-DQB1; A*1101-Cw*0801-B*1502 and A*3303-Cw*0302-B*5801 for HLA-A-C-B; A*1101-B*1502-DRB1*1202 and A*2901-B*0705-DRB1*1001 for HLA-A-B-DRB1, A*1101-Cw*0801-B*1502-DRB1*1202-DQB1*0301 and A*2901-Cw*1505-B*0705-DRB1*1001-DQB1*0501 for HLA-A-C-B-DRB1-DQB1. Allele distribution and haplotype analysis demonstrated that the Vietnamese population shares HLA patterns with southern Chinese, Thai, Javanese and Micronesians, while it also retains unique characteristics.

Pulmonary Mycobacterium avium complex infection: association with NRAMP1 polymorphisms.

The present study aimed to elucidate risk factors for nonimmunocompromised pulmonary Mycobacterium avium complex (MAC) infection. Epidemiological data and variations of candidate genes for mycobacterial diseases were analysed in 111 patients with pulmonary MAC infection. Four polymorphisms of the human natural resistance-associated macrophage protein (NRAMP)1 gene, the 5'(GT)n, 469+14 G/C, D543N and the 3'untranslated region (3'TGTG) insertion/deletion, were genotyped using PCR-based methods. Fok I and Taq I polymorphisms of the vitamin D receptor gene and -221 X/Y and codon 54 A/B polymorphisms of the mannose binding lectin gene were also evaluated. Females were more susceptible to MAC infection mainly affecting the right middle lobe or lingular segment of the lung. Patients' residence at the onset of the disease was distributed evenly irrespective of a waterfront or city water supply system. As compared with homozygotes for major alleles of the D543N and TGTG insertion/deletion polymorphism of the NRAMP1 gene, heterozygotes containing minor alleles were less often observed in MAC cases than in controls. This genetic effect was more significant in patients without comorbidity but not in patients with comorbidity. Other polymorphisms did not show any association with the MAC infection. The human natural resistance-associated macrophage protein 1 gene might be involved in susceptibility to pulmonary Mycobacterium avium complex infection.

Pulmonary Mycobacterium avium complex infection associated with the IVS8-T5 allele of the CFTR gene.

BACKGROUND: The T5 allele in intron 8 (IVS8) on specific haplotype backgrounds (e.g., long TG repeats) causes abnormal splicing in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, and is also known to be associated with chronic airway diseases. OBJECTIVE: To investigate the role of CFTR variations for susceptibility to pulmonary Mycobacterium avium complex (MAC) infection. PARTICIPANTS: Three hundred patients with pulmonary MAC infection (72 males, 228 females; mean age at onset 61.6 + or - 12.4 years) took part in this study. Diagnosis of MAC infection was based on American Thoracic Society criteria. Clinical profiles were collected and blood samples were genotyped for TG repeats, poly-T and M470V polymorphisms. RESULTS: We found significantly higher T5 frequency in MAC patients than in healthy controls from our own study (0.035 and 0.005, respectively, P = 0.023) and other reports. Homozygote for the T5 allele was found in two MAC patients. All T5 alleles were associated with longer TG repeats, the TG12 or TG13 allele. Seventeen of the 21 T5 alleles appeared to be associated with the V470 allele. Other polymorphisms did not show any significant differences in frequency. CONCLUSIONS: These findings suggest that the IVS8 5T allele might be involved in susceptibility to pulmonary MAC infection.