New molecular marker for phylogenetic reconstruction of black yeast-like fungi (Chaetothyriales) with hypothetical EIF2AK2 kinase gene.

In eukaryotes, phosphorylation of the α-subunit of eIF2 is a mechanism to adjust cellular gene expression profiles in response to specific signals. The eIF2α kinases are a group of serine-threonine kinases that perform important functions in response to infection, proteotoxicity, and nutrient scavenging. The conserved nature of eIF2α kinases among fungi makes them potential evolutionary markers, which may contribute to deeper understanding of taxonomy and evolution. To date, only few studies are available of eIF2α kinases in black yeasts, which are members of Chaetothyriales containing potential agents of a gamut of major human diseases, such as chromoblastomycosis, phaeohyphomycosis and mycetoma. To establish the phylogenetic validity of sequences of eIF2α kinases hypothetical genes, we compared these genes between members of different classes of fungi, including black yeasts and allies, aiming at evaluation of the phylogeny of this group using an alternative molecular marker, compared to standard ribosomal genes. Trees generated with eIF2α kinase sequences of fungi were compared with those generated by ribosomal internal transcribed spacers (ITS rDNA) sequences from the same species. Sequences used were obtained from the protein Non-redundant database of NCBI, were aligned using CLUSTALX v1.8 and alignments were analyzed with RAxML v8.2.9 on the CIPRES Science Gateway portal. The trees generated had similar topologies, demonstrating that eIF2α kinases hypothetical gene sequences present a coherent reflection of evolution among fungi, compared to trees reconstructed by the use of ribosomal sequences. Our preliminary findings with a limited dataset strongly suggest that the evolution of kinases among black yeasts follows a similar path as revealed by ribosomal data, which underlines the validity of current taxonomy of black yeasts and relatives.

Differential gene expression profiles may differentiate responder and nonresponder patients with rheumatoid arthritis for methotrexate (MTX) monotherapy and MTX plus tumor necrosis factor inhibitor combined therapy.

OBJECTIVE: We aimed to evaluate whether the differential gene expression profiles of patients with rheumatoid arthritis (RA) could distinguish responders from nonresponders to methotrexate (MTX) and, in the case of MTX nonresponders, responsiveness to MTX plus anti-tumor necrosis factor-α (anti-TNF) combined therapy. METHODS: We evaluated 25 patients with RA taking MTX 15-20 mg/week as a monotherapy (8 responders and 17 nonresponders). All MTX nonresponders received infliximab and were reassessed after 20 weeks to evaluate their anti-TNF responsiveness using the European League Against Rheumatism response criteria. A differential gene expression analysis from peripheral blood mononuclear cells was performed in terms of hierarchical gene clustering, and an evaluation of differentially expressed genes was performed using the significance analysis of microarrays program. RESULTS: Hierarchical gene expression clustering discriminated MTX responders from nonresponders, and MTX plus anti-TNF responders from nonresponders. The evaluation of only highly modulated genes (fold change > 1.3 or < 0.7) yielded 5 induced (4 antiapoptotic and CCL4) and 4 repressed (4 proapoptotic) genes in MTX nonresponders compared to responders. In MTX plus anti-TNF non-responders, the CCL4, CD83, and BCL2A1 genes were induced in relation to responders. CONCLUSION: Study of the gene expression profiles of RA peripheral blood cells permitted differentiation of responders from nonresponders to MTX and anti-TNF. Several candidate genes in MTX non-responders (CCL4, HTRA2, PRKCD, BCL2A1, CAV1, TNIP1, CASP8AP2, MXD1, and BTG2) and 3 genes in MTX plus anti-TNF nonresponders (CCL4, CD83, and BCL2A1) were identified for further study.

Bosentan, an endothelin receptor antagonist, ameliorates collagen-induced arthritis: the role of TNF-α in the induction of endothelin system genes.

OBJECTIVE: Endothelins (ETs) are involved in several inflammatory events. The present study investigated the efficacy of bosentan, a dual ETA/ETB receptor antagonist, in collagen-induced arthritis (CIA) in mice. TREATMENT: CIA was induced in DBA/1J mice. Arthritic mice were treated with bosentan (100 mg/kg) once a day, starting from the day when arthritis was clinically detectable. METHODS: CIA progression was assessed by measurements of visual clinical score, paw swelling and hypernociception. Histological changes, neutrophil infiltration and pro-inflammatory cytokines were evaluated in the joints. Gene expression in the lymph nodes of arthritic mice was evaluated by microarray technology. PreproET-1 mRNA expression in the lymph nodes of mice and in peripheral blood mononuclear cells (PBMCs) was evaluated by real-time PCR. The differences were evaluated by one-way ANOVA or Student's t test. RESULTS: Oral treatment with bosentan markedly ameliorated the clinical aspects of CIA (visual clinical score, paw swelling and hyperalgesia). Bosentan treatment also reduced joint damage, leukocyte infiltration and pro-inflammatory cytokine levels (IL-1ß, TNFα and IL-17) in the joint tissues. Changes in gene expression in the lymph nodes of arthritic mice returned to the levels of the control mice after bosentan treatment. PreproET mRNA expression increased in PBMCs from rheumatoid arthritis (RA) patients but returned to basal level in PBMCs from patients under anti-TNF therapy. In-vitro treatment of PBMCs with TNFα upregulated ET system genes. CONCLUSION: These findings indicate that ET receptor antagonists, such as bosentan, might be useful in controlling RA. Moreover, it seems that ET mediation of arthritis is triggered by TNFα.

Delayed effects of exposure to a moderate radiation dose on transcription profiles in human primary fibroblasts.

Ionizing radiation (IR) is used in a wide variety of medical and nonmedical applications and poses a potential threat to human health. Knowledge of changes in gene expression in irradiated cells may be helpful for the establishment of effective paradigms for radiation protection. IR-induced DNA damage triggers a complex cascade of signal transduction. Recently, genome-wide approaches have allowed the detection of alterations in gene expression across a wide range of radiation doses. However, the delayed or long-term biological effects of mild-doses of IR remain largely unknown. The main objective of the present study was to investigate the effects of a moderate dose of gamma-rays (50 cGy) on gene expression 6 days post-irradiation. Gene expression using cDNA microarrays revealed statistically significant changes in the expression of 59 genes (FDR < 0.07), whose functions are related to cell-cycle control, protein trafficking, ubiquitin cycle, Rho-GTPAse pathway, protein phosphatase signalization, oxidoreductase control, and stress response. A set of 464 genes was also selected by a less stringent approach, and we demonstrate that this broader set of genes can efficiently distinguish the irradiated samples from the unirradiated, defining a long-term IR signature in human primary fibroblasts. Our findings support the existence of persistent responses to mild doses of IR detectable by changes in gene expression profiles. These results provide insight into delayed effects observed in human primary cells as well as the role of long-term response in neoplastic transformation. Environ.

Collagen induced arthritis (CIA) in mice features regulatory transcriptional network connecting major histocompatibility complex (MHC H2) with autoantigen genes in the thymus.

Considering that imbalance of central tolerance in the thymus contributes to aggressive autoimmunity, we compared the expression of peripheral tissue autoantigens (PTA) genes, which are involved in self-representation in the thymic stroma, of two mouse strains; DBA-1/J (MHC-H2(q)) susceptible and DBA-2/J (MHC-H2(d)) resistant to collagen induced arthritis (CIA). We evaluate whether these strains differ in their thymic gene expression, allowing identification of genes that might play a role in susceptibility/resistance to CIA. Microarray profiling showed that 1093 PTA genes were differentially modulated between collagen immunized DBA-1/J and DBA-2/J mice. These genes were assigned to 17 different tissues/organs, including joints/bone, characterizing the promiscuous gene expression (PGE), which is implicated in self-representation. Hierarchical clustering of microarray data and quantitative RT-PCR analysis showed that Aire (autoimmune regulator), an important regulator of the PGE process, Aire-dependent (insulin), Aire-independent (Col2A1 and Gad67), and other 22 joint/bone autoantigen genes were down-regulated in DBA-1/J compared with DBA-2/J in the thymus. Considering the importance of MHC-H2 in peptide-self presentation and autoimmunity susceptibility, we reconstructed transcriptional networks of both strains based on actual microarray data. The networks clearly demonstrated different MHC-H2 transcriptional interactions with PTAs genes. DBA-1/J strain featured MHC-H2 as a node influencing downstream genes. Differently, in DBA-2/J strain network MHC-H2 was exclusively self-regulated and does not control other genes. These findings provide evidence that CIA susceptibility in mice may be a reflex of a cascade-like transcriptional control connecting different genes to MHC-H2 in the thymus.

Alterations in gene expression profiles correlated with cisplatin cytotoxicity in the glioma U343 cell line.

Gliomas are the most common tumors in the central nervous system, the average survival time of patients with glioblastoma multiforme being about 1 year from diagnosis, in spite of harsh therapy. Aiming to study the transcriptional profiles displayed by glioma cells undergoing cisplatin treatment, gene expression analysis was performed by the cDNA microarray method. Cell survival and apoptosis induction following treatment were also evaluated. Drug concentrations of 12.5 to 300 µM caused a pronounced reduction in cell survival rates five days after treatment, whereas concentrations higher than 25 µM were effective in reducing the survival rates to ~1%. However, the maximum apoptosis frequency was 20.4% for 25 µM cisplatin in cells analyzed at 72 h, indicating that apoptosis is not the only kind of cell death induced by cisplatin. An analysis of gene expression revealed 67 significantly (FDR < 0.05) modulated genes: 29 of which down- and 38 up-regulated. These genes belong to several classes (metabolism, protein localization, cell proliferation, apoptosis, adhesion, stress response, cell cycle and DNA repair) that may represent several affected cell processes under the influence of cisplatin treatment. The expression pattern of three genes (RHOA, LIMK2 and TIMP2) was confirmed by the real time PCR method.

Alterations in gene expression profiles correlated with cisplatin cytotoxicity in the glioma U343 cell line

Gliomas are the most common tumors in the central nervous system, the average survival time of patients with glioblastoma multiforme being about 1 year from diagnosis, in spite of harsh therapy. Aiming to study the transcriptional profiles displayed by glioma cells undergoing cisplatin treatment, gene expression analysis was performed by the cDNA microarray method. Cell survival and apoptosis induction following treatment were also evaluated. Drug concentrations of 12.5 to 300 μM caused a pronounced reduction in cell survival rates five days after treatment, whereas concentrations higher than 25 μM were effective in reducing the survival rates to ~1 percent. However, the maximum apoptosis frequency was 20.4 percent for 25 μM cisplatin in cells analyzed at 72 h, indicating that apoptosis is not the only kind of cell death induced by cisplatin. An analysis of gene expression revealed 67 significantly (FDR < 0.05) modulated genes: 29 of which down- and 38 up-regulated. These genes belong to several classes (metabolism, protein localization, cell proliferation, apoptosis, adhesion, stress response, cell cycle and DNA repair) that may represent several affected cell processes under the influence of cisplatin treatment. The expression pattern of three genes (RHOA, LIMK2 and TIMP2) was confirmed by the real time PCR method.

Evidence for a network transcriptional control of promiscuous gene expression in medullary thymic epithelial cells.

The expression of peripheral tissue antigens (PTAs) in the thymus by medullary thymic epithelial cells (mTECs) is essential for the central self-tolerance in the generation of the T cell repertoire. Due to heterogeneity of autoantigen representation, this phenomenon has been termed promiscuous gene expression (PGE), in which the autoimmune regulator (Aire) gene plays a key role as a transcription factor in part of these genes. Here we used a microarray strategy to access PGE in cultured murine CD80(+) 3.10 mTEC line. Hierarchical clustering of the data allowed observation that PTA genes were differentially expressed being possible to found their respective induced or repressed mRNAs. To further investigate the control of PGE, we tested the hypothesis that genes involved in this phenomenon might also be modulated by transcriptional network. We then reconstructed such network based on the microarray expression data, featuring the guanylate cyclase 2d (Gucy2d) gene as a main node. In such condition, we established 167 positive and negative interactions with downstream PTA genes. Silencing Aire by RNA interference, Gucy2d while down regulated established a larger number (355) of interactions with PTA genes. T- and G-boxes corresponding to AIRE protein binding sites located upstream to ATG codon of Gucy2d supports this effect. These findings provide evidence that Aire plays a role in association with Gucy2d, which is connected to several PTA genes and establishes a cascade-like transcriptional control of promiscuous gene expression in mTEC cells.

Shared and unique gene expression in systemic lupus erythematosus depending on disease activity.

Patients presenting with active systemic lupus erythematosus (SLE) manifestations may exhibit distinct pathogenetic features in relation to inactive SLE. Also, cDNA microarrays may potentially discriminate the gene expression profile of a disease or disease variant. Therefore, we evaluated the expression profile of 4500 genes in peripheral blood lymphocytes (PBL) of SLE patients. We studied 11 patients with SLE (seven with active SLE and four with inactive SLE) and eight healthy controls. Total RNA was isolated from PBL, reverse transcribed into cDNA, and postlabeled with Cy3 fluorochrome. These probes were then hybridized to a glass slide cDNA microarray containing 4500 human IMAGE cDNA target sequences. An equimolar amount of total RNA from human cell lines served as reference. The microarray images were quantified, normalized, and analyzed using the R environment (ANOVA, significant analysis of microarrays, and cluster-tree view algorithms). Disease activity was assessed by the SLE disease activity index. Compared to the healthy controls, 104 genes in active SLE patients (80 repressed and 24 induced) and 52 genes in nonactive SLE patients (31 induced and 21 repressed) were differentially expressed. The modulation of 12 genes, either induced or repressed, was found in both disease variants; however, each disease variant had differential expression of different genes. Taken together, these results indicate that the two lupus variants studied have common and unique differentially expressed genes. Although the biological significance of the differentially expressed genes discussed above has not been completely understood, they may serve as a platform to further explore the molecular basis of immune deregulation in SLE.

Genetic susceptibility loci in rheumatoid arthritis establish transcriptional regulatory networks with other genes.

Linkage studies have identified the human leukocyte antigen (HLA)-DRB1 as a putative rheumatoid arthritis (RA) susceptibility locus (SL). Nevertheless, it was estimated that its contribution was partial, suggesting that other non-HLA genes may play a role in RA susceptibility. To test this hypothesis, we conducted microarray transcription profiling of peripheral blood mononuclear cells in 15 RA patients and analyzed the data, using bioinformatics programs (significance analysis of microarrays method and GeneNetwork), which allowed us to determine the differentially expressed genes and to reconstruct transcriptional networks. The patients were grouped according to disease features or treatment with tumor necrosis factor blocker. Transcriptional networks that were reconstructed allowed us to identify the interactions occurring between RA SL and other genes, for example, HLA-DRB1 interacting with FNDC3A (fibronectin type III domain containing 3A). Given that fibronectin fragments can stimulate mediators of matrix and cartilage destruction in RA, this interaction is of special interest and may contribute to a clearer understanding of the functional role of HLA-DRB1 in RA pathogenesis.

Transcriptional response of peripheral lymphocytes to early fibrosarcoma: a model system for cancer detection based on hybridization signatures.

Since circulating leukocytes, mainly B and T cells, continuously maintain vigilant and comprehensive immune surveillance, these cells could be used as reporters for signs of infection or other pathologies, including cancer. Activated lymphocyte clones trigger a sensitive transcriptional response, which could be identified by gene expression profiling. To assess this hypothesis, we conducted microarray analysis of the gene expression profile of lymphocytes isolated from immunocompetent BALB/c mice subcutaneously injected with different numbers of tumorigenic B61 fibrosarcoma cells. Flow cytometry demonstrated that the number of circulating T (CD3(+)CD4(+) or CD3(+)CD8(+)) or B (CD19(+)) cells did not change. However, the lymphocytes isolated from tumor cell-injected animals expressed a unique transcriptional profile that was identifiable before the development of a palpable tumor mass. This finding demonstrates that the transcriptional response appears before alterations in the main lymphocyte subsets and that the gene expression profile of peripheral lymphocytes can serve as a sensitive and accurate method for the early detection of cancer.

Gene expression profiles in radiation workers occupationally exposed to ionizing radiation.

Ionizing radiation (IR) imposes risks to human health and the environment. IR at low doses and low dose rates has the potency to initiate carcinogenesis. Genotoxic environmental agents such as IR trigger a cascade of signal transduction pathways for cellular protection. In this study, using cDNA microarray technique, we monitored the gene expression profiles in lymphocytes derived from radiation-exposed individuals (radiation workers). Physical dosimetry records on these patients indicated that the absorbed dose ranged from 0.696 to 39.088 mSv. Gene expression analysis revealed statistically significant transcriptional changes in a total of 78 genes (21 up-regulated and 57 down-regulated) involved in several biological processes such as ubiquitin cycle (UHRF2 and PIAS1), DNA repair (LIG3, XPA, ERCC5, RAD52, DCLRE1C), cell cycle regulation/proliferation (RHOA, CABLES2, TGFB2, IL16), and stress response (GSTP1, PPP2R5A, DUSP22). Some of the genes that showed altered expression profiles in this study can be used as biomarkers for monitoring the chronic low level exposure in humans. Additionally, alterations in gene expression patterns observed in chronically exposed radiation workers reinforces the need for defining the effective radiation dose that causes immediate genetic damage as well as the long-term effects on genomic instability, including cancer.

Differential gene expression of peripheral blood mononuclear cells from rheumatoid arthritis patients may discriminate immunogenetic, pathogenic and treatment features.

This study aimed to evaluate the association between the differential gene expression profiling of peripheral blood mononuclear cells of rheumatoid arthritis patients with their immunogenetic (human leucocyte antigen shared-epitope, HLA-SE), autoimmune response [anti-cyclic citrullinated peptide (CCP) antibodies], disease activity score (DAS-28) and treatment (disease-modifying antirheumatic drugs and tumour necrosis factor blocker) features. Total RNA samples were copied into Cy3-labelled complementary DNA probes, hybridized onto a glass slide microarray containing 4500 human IMAGE complementary DNA target sequences. The Cy3-monocolour microarray images from patients were quantified and normalized. Analysis of the data using the significance analysis of microarrays algorithm together with a Venn diagram allowed the identification of shared and of exclusively modulated genes, according to patient features. Thirteen genes were exclusively associated with the presence of HLA-SE alleles, whose major biological function was related to signal transduction, phosphorylation and apoptosis. Ninety-one genes were associated with disease activity, being involved in signal transduction, apoptosis, response to stress and DNA damage. One hundred and one genes were associated with the presence of anti-CCP antibodies, being involved in signal transduction, cell proliferation and apoptosis. Twenty-eight genes were associated with tumour necrosis factor blocker treatment, being involved in intracellular signalling cascade, phosphorylation and protein transport. Some of these genes had been previously associated with rheumatoid arthritis pathogenesis, whereas others were unveiled for future research.

Comprehensive gene expression profiling in lungs of mice infected with Mycobacterium tuberculosis following DNAhsp65 immunotherapy.

BACKGROUND: The continued increase in tuberculosis (TB) rates and the appearance of extremely resistant Mycobacterium tuberculosis strains (XDR-TB) worldwide are some of the great problems of public health. In this context, DNA immunotherapy has been proposed as an effective alternative that could circumvent the limitations of conventional drugs. Nonetheless, the molecular events underlying these therapeutic effects are poorly understood. METHODS: We characterized the transcriptional signature of lungs from mice infected with M. tuberculosis and treated with heat shock protein 65 as a genetic vaccine (DNAhsp65) combining microarray and real-time polymerase chain reaction analysis. The gene expression data were correlated with the histopathological analysis of lungs. RESULTS: The differential modulation of a high number of genes allowed us to distinguish DNAhsp65-treated from nontreated animals (saline and vector-injected mice). Functional analysis of this group of genes suggests that DNAhsp65 therapy could not only boost the T helper (Th)1 immune response, but also could inhibit Th2 cytokines and regulate the intensity of inflammation through fine tuning of gene expression of various genes, including those of interleukin-17, lymphotoxin A, tumour necrosis factor-alpha, interleukin-6, transforming growth factor-beta, inducible nitric oxide synthase and Foxp3. In addition, a large number of genes and expressed sequence tags previously unrelated to DNA-therapy were identified. All these findings were well correlated with the histopathological lesions presented in the lungs. CONCLUSIONS: The effects of DNA therapy are reflected in gene expression modulation; therefore, the genes identified as differentially expressed could be considered as transcriptional biomarkers of DNAhsp65 immunotherapy against TB. The data have important implications for achieving a better understanding of gene-based therapies.

Microarray-based gene expression analysis of human osteoblasts in response to different biomaterials.

Several biomaterials have been widely used in bone regeneration/substitution procedures in orthopedic and oral surgery. However, how these biomaterials alter osteoblast gene expression is poorly understood. We therefore attempted to address this question by using cDNA microarray technique to identify genes that are differentially regulated in osteoblasts exposed to biomaterials comprehending the biocompatibility spectrum of bioactive (bioglass and hydroxyapatite), bioinert (Ti and stainless steel), and biotolerant (polymethylmethacrylate). By using a cDNA microarray containing 687 human IMAGE sequences, we identified in primary cultures of osteoblastic cells differentiated from the human bone marrow and exposed to these biomaterials, genes whose expression was significantly upregulated or downregulated. Among the differentially expressed genes we have found those involved with cell cycle regulation, cell differentiation and proliferation, apoptosis, cell adhesion, bone mineralization and skeletal development. These results can be relevant to a better understanding of the molecular mechanism underlying the behavior of osteoblasts in bone regenerative procedures.

The effect of TAK-778 on gene expression of osteoblastic cells is mediated through estrogen receptor.

This study evaluated the effect of TAK-778 [(2R, 4S)-(-)-N-(4-diethoxyphosphorylmethylphenyl)-1,2,4,5-tetrahydro-4-methyl-7,8-methylenedioxy-5-oxo-3-benzothiepin-2-carboxamide)] on in vitro osteogenic events and on gene expression of osteoblastic cells derived from human alveolar bone and the participation of estrogen receptors (ERs) on such effect. Osteoblastic cells were subcultured, with or without TAK-778 (10(-5) M), to evaluate cell growth and viability, total protein content, and alkaline phosphatase (ALP) activity at 7, 14, and 21 days; bone-like formation at 21 days; and gene expression, using cDNA microarray, at 7 days. Also, osteoblastic cells were exposed to TAK-778 (10(-5) M) combined to ICI182,780, a nonspecific ER antagonist (10(-6) M), and gene expression was evaluated by real-time polymerase chain reaction (PCR) at 7 days. TAK-778 induced a reduction in culture growth and an increase in cell synthesis, ALP activity, and bone-like formation. The cDNA microarray showed genes associated with cell adhesion and differentiation, skeletal development, ossification, and transforming growth factor-beta receptor signaling pathway, with a tendency to be higher expressed in cells exposed to TAK-778. The gene expression of ALP, osteocalcin, Msh homeobox 2, receptor activator of NF-kappa B ligand, and intercellular adhesion molecule 1 was increased by TAK-778 as demonstrated by real-time PCR, and this effect was antagonized by ICI182,780. The present results demonstrated that TAK-778 acts at a transcriptional level to enhance the in vitro osteogenic process and that its effect on gene expression of osteoblastic cells is mediated, at least partially, through ERs. Based on these findings, TAK-778 could be considered in the treatment of bone metabolic disorders.

Genes that code for T cell signaling proteins establish transcriptional regulatory networks during thymus ontogeny.

Gene expression profiling by cDNA microarrays during murine thymus ontogeny has contributed to dissecting the large-scale molecular genetics of T cell maturation. Gene profiling, although useful for characterizing the thymus developmental phases and identifying the differentially expressed genes, does not permit the determination of possible interactions between genes. In order to reconstruct genetic interactions, on RNA level, within thymocyte differentiation, a pair of microarrays containing a total of 1,576 cDNA sequences derived from the IMAGE MTB library was applied on samples of developing thymuses (14-17 days of gestation). The data were analyzed using the GeneNetwork program. Genes that were previously identified as differentially expressed during thymus ontogeny showed their relationships with several other genes. The present method provided the detection of gene nodes coding for proteins implicated in the calcium signaling pathway, such as Prrg2 and Stxbp3, and in protein transport toward the cell membrane, such as Gosr2. The results demonstrate the feasibility of reconstructing networks based on cDNA microarray gene expression determinations, contributing to a clearer understanding of the complex interactions between genes involved in thymus/thymocyte development.

Share epitope, citrullinated cyclic peptide antibodies and smoking in Brazilian rheumatoid arthritis patients.

The events involved in the pathogenesis of rheumatoid arthritis (RA) still remain unclear, but certainly the etiology is multifactorial. Shared epitope (SE) of HLADRbeta1 is the most important genetic risk factor. Environmental risk factors are less understood. Smoking is a candidate, associated with the rising of citrullinated cyclic peptide antibodies (anti-CCP). Anti-CCP antibodies are highly specific for RA. In this study, we investigated whether the association between anti-CCP production and smoking was influenced by carriage of SE in a highly miscegenated population of patients with RA. One hundred Brazilian patients were inquired about cigarette smoking. For all of them, DNA for HLA typing and serum to anti-CCP antibodies quantification were obtained. Forty-two were smokers and 58 were nonsmokers. The SE was present in 61 patients and the anti-CCP was positive in 71 patients. We found that, among smokers, 25 were SE-positive, 22 presented with anti-CCP and 3 without anti-CCP, and 17 were SE-negative, 9 presented with anti-CCP and 8 without anti-CCP (OR 6.5, 95% CI 1.40 to 30.20). These results suggest that environmental factors contribute to the raising of anti-CCP in individuals with HLA background to RA, smoking being a strong candidate.

Gene expression profiles stratified according to type 1 diabetes mellitus susceptibility regions.

The MHC region (6p21) aggregates the major genes that contribute to susceptibility to type 1 diabetes (T1D). Three additional relevant susceptibility regions mapped on chromosomes 1p13 (PTPN22), 2q33 (CTLA-4), and 11p15 (insulin) have also been described by linkage studies. To evaluate the contribution of these susceptibility regions and the chromosomes that house these regions, we performed a large-scale differential gene expression on lymphomononuclear cells of recently diagnosed T1D patients, pinpointing relevant modulated genes clustered in these regions and their respective chromosomes. A total of 4608 cDNAs from the IMAGE library were spotted onto glass slides using robotic technology. Statistical analysis was carried out using the SAM program, and data regarding gene location and biological function were obtained at the SOURCE, NCBI, and FATIGO programs. Three induced genes were observed spanning around the MHC region (6p21-6p23), and seven modulated genes (5 repressed and 2 repressed) were seen spanning around the 6q21-24 region. Additional modulated genes were observed in and around the 1p13, 2q33, and 11p15 regions. Overall, modulated genes in these regions were primarily associated with cellular metabolism, transcription factors and signaling transduction. The differential gene expression characterization may identify new genes potentially involved with diabetes pathogenesis.