[Show abstract][Hide abstract] ABSTRACT: Mannan-binding lectin-associated serine protease 2 (MASP-2) is a key protein of the lectin pathway of complement. MASP-2 levels have been associated with different polymorphisms within MASP2 gene as well as with the risk for inflammatory disorders and infections. Despite its clinical importance, MASP-2 remains poorly investigated in rheumatoid arthritis (RA).
In this case-control study, we measured MASP-2 serum levels in 156 RA patients, 44 patient relatives, and 100 controls from Southern Brazil, associating the results with nine MASP2 polymorphisms in all patients, 111 relatives, and 230 controls genotyped with multiplex SSP-PCR.
MASP-2 levels were lower in patients than in controls and relatives (medians 181 vs. 340 or 285 ng/ml, respectively, P<0.0001). Conversely, high MASP-2 levels were associated with lower susceptibility to RA and to articular symptoms independently of age, gender, ethnicity, smoking habit, anti-CCP and rheumatoid factor positivity (OR = 0.05 [95%CI = 0.019-0.13], P<0.0001 between patients and controls; OR = 0.12, [95%CI = 0.03-0.45], P = 0.002 between patients and relatives; OR = 0.06, [95%CI = 0.004-0.73], P = 0.03 between relatives with and without articular symptoms). MASP2 haplotypes *2A1 and *2B1-i were associated with increased susceptibility to RA (OR = 3.32 [95%CI = 1.48-7.45], P = 0.004). Deficiency-causing p.120G and p.439H substitutions were associated with five times increased susceptibility to articular symptoms in relatives (OR = 5.13 [95%CI = 1.36-20.84], P = 0.02). There was no association of MASP-2 levels or MASP2 polymorphisms with autoantibodies, Sjögren's syndrome, nodules and functional class.
In this study, we found the first evidence that MASP-2 deficiency might play an important role in the development of RA and articular symptoms among relatives of RA patients.
PLoS ONE 01/2014; 9(3):e90979. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate the association of leprosy with hepatitis B virus (HBV) infection, as yet unknown for South Brazil, we assessed hepatitis B virus coinfection in 199 South Brazilian leprosy patients (119 lepromatous, 15 tuberculoid, 30 borderline, 12 undetermined and 23 unspecified) and in 681 matched blood donors by screening for the hepatitis B virus markers HBSAg and anti-HBc, using ELISA. Positive samples were retested and anti-HBc+ only samples were tested for the hepatitis B surface antibody (anti-HBs). There was a strong association between leprosy and hepatitis B virus infection (OR=9.8, 95% CI=6.4-14.7; p=0.004·E(-30)), as well as an association between HBV infection and lepromatous leprosy, compared to other forms (OR=2.4, 95% CI=1.2-4.8; p=0.017). We also found that confinement due to leprosy was associated with hepatitis B virus infection (OR=3.9, 95% CI=2.1-7.4; p=0.015·E(-3)). Leprosy patients are susceptible to develop hepatitis B virus infection, especially lepromatous. Institutionalized patients, who probably present a stronger Th2 response, have higher risk of being exposed to hepatitis B virus. This clearly emphasizes the need for special care to leprosy patients in preventing hepatitis B virus coinfection in South Brazil.
The Brazilian journal of infectious diseases: an official publication of the Brazilian Society of Infectious Diseases 08/2013; · 1.04 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The gene MASP2 (mannan-binding lectin (MBL)-associated serine protease 2) encodes two proteins, MASP-2 and MAp19 (MBL-associated protein of 19 kDa), bound in plasma to MBL and ficolins. The binding of MBL/MASP-2 and ficolin/MASP-2 complexes to microorganisms activates the lectin pathway of complement and may increase the ingestion of intracellular pathogens such as Mycobacterium leprae.
We haplotyped 11 MASP2 polymorphisms with multiplex sequence-specific PCR in 219 Brazilian leprosy patients (131 lepromatous, 29 borderline, 21 tuberculoid, 14 undetermined, 24 unspecified), 405 healthy Brazilians and 291 Danish blood donors with previously determined MASP-2 and MAp19 levels. We also evaluated MASP-2 levels in further 46 leprosy patients and 69 Brazilian controls.
Two polymorphisms flanking exon 5 of MASP2 were associated with a dominant effect on high MASP-2 levels and an additive effect on low MAp19 levels. Patients presented lower MASP-2 levels (P = 0.0012) than controls. The frequency of the p.126L variant, associated with low MASP-2 levels (below 200 ng/mL), was higher in the patients (P = 0.0002, OR = 4.92), as was the frequency of genotypes with p.126L (P = 0.00006, OR = 5.96). The *1C2-l [AG] haplotype, which harbors p.126L and the deficiency-causing p.439H variant, has a dominant effect on the susceptibility to the disease (P = 0.007, OR = 4.15). Genotypes composed of the *2B1-i and/or *2B2A-i haplotypes, both associated with intermediate MASP-2 levels (200-600 ng/mL), were found to be protective against the disease (P = 0.0014, OR = 0.6). Low MASP-2 levels (P = 0.022), as well as corresponding genotypes with *1C2-l and/or *2A2-l but without *1B1-h or *1B2-h, were more frequent in the lepromatous than in other patients (P = 0.008, OR = 8.8).
In contrast with MBL, low MASP-2 levels increase the susceptibility to leprosy in general and to lepromatous leprosy in particular. MASP2 genotypes and MASP-2 levels might thus be of prognostic value for leprosy progression.
PLoS ONE 01/2013; 8(7):e69054. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: L-ficolin (encoded by FCN2) binds to acetylated sugar moieties of many pathogens, including Trypanosoma cruzi, promoting their phagocytosis and lysis by the complement system.
We investigated L-ficolin levels in 160 T. cruzi infected patients with chronic Chagas disease and 71 healthy individuals, and FCN2 polymorphisms (-986 G>A, -602 G>A, and -4 A>G in the promoter and A258S in exon 8) in 243 patients, being 88 indeterminate (asymptomatic), 96 with cardiac, 23 with digestive and 33 with cardiodigestive manifestations (two were unspecified) and 305 controls (135 for A258S).
Patients presented lower L-ficolin plasma levels than controls (p<0.0001). Among the different groups of cardiac commitment, individuals with moderate forms had higher L-ficolin levels than the severe forms (P = 0.039). Lower L-ficolin levels were found associated with the 258S variant in the patients (P = 0.034). We found less -4A/G heterozygotes in the cardiac patients, than in the controls (OR = 0.56 [95% CI = 0.33-0.94], P = 0.034). Heterozygote -4A/G genotypes with the 258S variant and 258SS homozygotes were nevertheless more frequent among cardiodigestive patients than in controls (OR = 14.1 [95% CI = 3.5-56.8], P = 0.0001) and in indeterminate patients (OR = 3.2 [95% CI = 1.1-9.4], P = 0.037). We also found an association of the allelic frequency of the 258S variant with cardiodigestive Chagas disease compared to controls (OR = 2.24 [95% CI = 1.1-4.5], P = 0.037). Thus, decreased patient levels of L-ficolin reflect not only protein consumption due to the disease process, but also the higher frequency of the 258S variant in patients with cardiodigestive symptoms.
The very first study on Brazilian cohort associates both L-ficolin plasma levels and FCN2 variants to Chagas disease and subsequent disease progression. The prognostic value of L-ficolin levels and the FCN2*A258S polymorphism should be further evaluated in other settings.
PLoS ONE 01/2013; 8(4):e60237. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate the results of the healing process on surgical wounds in the back of Wistar rats using nanocristaline and ionic silver dressing.
Sixty rats Wistar were submitted to surgical wounds with punch of 8mm in diameter. In 30 animals (groups PN - nanocristaline and AD - control) two surgical wounds were done diametrically opposite on the upper back side. On the right side was used nanocristaline (PN) silver dressing and on the left side, distilled water dressing (AD). On the other group of 30 rats, only one wound was made with the punch, on the right side, and was used ionic silver dressing. So, the groups were divided into three subgroups, according to the day of death (7th, 14th and 21st day). In each of these days the wounds diameter were measured to evaluate the wound contraction. Microscopic data were analyzed using the H&E staining to verify the inflammatory process and neovascularization. The Masson trichrome staining was used to verify the fibrosis.
Macroscopically only the subgroup of 21st day showed statistical significance; between the groups AD and PI inflammatory process appeared in the 7th day subgroup in 90% of the cases. In neovascularization there was statistical significance between the groups PN and AD in the subgroup of 7th day. Fibrosis did not show statistical significance in the studied groups.
In relation to wound contraction, PN and PI groups showed better results than the AD group. In regard to histological analysis, H&E staining showed that there was presence of inflammation in all groups, and at the end, the control group (AD) on 7th day, was superior to PN and PI groups. In relationship to fibrosis, no differences were obtained among groups.
Acta cirurgica brasileira / Sociedade Brasileira para Desenvolvimento Pesquisa em Cirurgia 11/2012; 27(11):761-7. · 0.48 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: PURPOSE: Mycobacterium leprae exploits complement activation and opsonophagocytosis to infect phagocytes. M-ficolin is encoded by the FCN1 gene and initiates the lectin pathway on monocyte surfaces. We investigated FCN1 promoter polymorphisms that could be responsible for the high interindividual variability of M-ficolin levels and for modulating leprosy susceptibility. METHODS: We genotyped rs2989727 (-1981 G > A), rs28909068 (-791 G > A), rs10120023 (-542 G > A), rs17039495 (-399 G > A), rs28909976 (-271IndelT), rs10117466 (-144C > A) and rs10858293 (+33 T > G) in 400 controls and 315 leprosy patients from Southern Brazil, and in 296 Danish healthy individuals with known M-ficolin levels. RESULTS: Ten haplotypes were identified with sequence-specific PCR and/or haplotype-specific sequencing. We found evidence for a protective codominant additive effect of FCN1*-542A-144C with leprosy in Euro-Brazilians (P = 0.003, PBf = 0.021, OR = 0.243 [CI95% = 0.083-0.71]), which was independent of age, ethnic group and gender effects (P = 0.029). There was a trend for a positive association of the -399A variant in Afro-Brazilians (P = 0.022, PBf = 0.154, OR = 4.151 [CI95% = 1.115-15.454], as well as for a negative association of the FCN1*3A haplotype with lepromatous leprosy, compared with less severe forms of the disease (P = 0.016, PBf = 0.112, OR = 0.324 [CI95% = 0.123-0.858]). Danish individuals with this haplotype presented M-ficolin levels higher than the population average of circa 1,000 ng/ml, and -542A-144C, which is able to modify the recognition of transcription factors in silico, occurred in individuals with levels under the 25 percentil (P = 0.031). CONCLUSIONS: Our data provide the first evidence that FCN1 polymorphisms are associated with leprosy. M-ficolin may represent a novel key to understand the immunopathogenesis of M. leprae infection.
Journal of Clinical Immunology 09/2012; · 3.38 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Due to its importance both in the clearance of pathogens that contribute as rheumatic etiological agents and in the disposal of apoptotic bodies and potential autoimmune initiators, deficiencies of the components of the lectin pathway of complement have been found to increase susceptibility and modulate the severity of most rheumatic disorders. This chapter introduces the general aspects of the structure, function, and genetics of lectin pathway components and summarizes current knowledge of the field regarding rheumatic diseases predisposition and modulation.
Advances in clinical chemistry 01/2012; 56:105-53. · 3.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Hepatitis C virus (HCV) has become a major public health issue and is prevalent in most countries. We examined several MASP2 functional polymorphisms in 104 Brazilian patients with moderate and severe chronic hepatitis C using the primers set to amplify the region encoding the first domain (CUB1), a critical region for the formation of functional mannan-binding lectin (MBL)/MBL-associated serine proteases (MASP)-2 complexes, and the fifth domain (CCP2), which is essential for C4 cleavage of the MASP2 gene. We identified five single nucleotide polymorphisms in patients and controls: p. R99Q, p. D120G, p.P126L, p.D371Y, and p.V377A. Our results show that the p.D371Y variant (c.1111 G > T) is associated with susceptibility to HCV infection (p = 0.003, odds ratio = 6.33, 95% confidence interval = 1.85-21.70). Considered as a dominant function for the T allele, this variant is associated with high plasma levels of the MASP-2 in hepatitis C patients (p < 0.001). However, further functional investigations are necessary to understand the degree of involvement between MASP2 and the HCV susceptibility.
Human immunology 07/2011; 72(10):912-5. · 2.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mannose-binding lectin (MBL) initiates complement on Trypanosoma cruzi through the MBL-associated serine protease 2 (MASP2). We haplotyped six MASP2 polymorphisms in 208 chronic chagasic patients, being 81 indeterminate and 123 symptomatic (76 with cardiac, 19 with digestive and 28 with cardiodigestive forms) and 300 healthy individuals from Southern Brazil, using PCR with sequence-specific primers. The g.1961795C, p.371D diplotype (short CD) occurred at a higher frequency among symptomatic patients, compared with the indeterminate group (P(Bf)=0.012, OR=3.11), as well as genotypes with CD, but not with the g.1945560A in the promoter in cardiac patients (P(Bf)=0.012, OR=13.54). CD haplotypes linked to the p.P126L and p.V377A variants were associated with reduced MASP-2 levels (P<0.0001) but not reduced MBL/MASP-2/C4 complexes. MASP2*CD genotypes, most of them generating low MASP-2 levels, are associated with high risk of chagasic cardiomyopathy. Rapid MASP2 genotyping might be used to predict the risk of symptomatic disease.
[Show abstract][Hide abstract] ABSTRACT: Deficiency of mannan-binding lectin-associated serine protease 2 (MASP-2) has been associated with infections, whereas high levels appear to increase the risk of inflammatory disorders. Nevertheless, MASP2 haplotypes have been poorly investigated. To overcome haplotyping cost and time consumption, we developed multiplex polymerase chain reactions with sequence-specific primers (PCR-SSP) for 8 single nucleotide polymorphisms (SNPs), reducing the number of necessary reactions from 18 to 7. SNPs were distributed from the promoter to the last exon, and a single PCR-SSP was used for p.D120G. We evaluated the phylogenetic relationships and global distribution of 10 identified haplotypes in 338 Danish individuals with known MASP-2 and MAp19 levels and 309 South Brazilians. Four haplotypes were associated with reduced MASP-2 levels in plasma (lower than 200 ng/mL). Simultaneous association with the highest MASP-2 (over 600 ng/mL) and lowest MAp19 levels (lower than 200 ng/mL) was demonstrated with the intron 9 mutation (Kruskal-Wallis p < 0.0001). Cumulative genotype frequencies predict approximately 0.4% severely deficient and 25% overproducing individuals in both populations. Rapid and low-cost screening of patients with multiplex MASP2 PCR-SSP could be used to identify clinical conditions where MASP-2 (or MAp19) levels may be disease modifying, possibly improving disease outcome through early therapeutic and preventive measures.
Human immunology 06/2011; 72(9):753-60. · 2.55 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Polymorphisms of the mannose-binding lectin gene (MBL2) affect the concentration and functional efficiency of the protein. We recently used haplotype-specific sequencing to identify 23 MBL2 haplotypes, associated with enhanced susceptibility to several diseases.
In this work, we applied the same method in 288 and 470 chromosomes from Gabonese and European adults, respectively, and found three new haplotypes in the last group. We propose a phylogenetic nomenclature to standardize MBL2 studies and found two major phylogenetic branches due to six strongly linked polymorphisms associated with high MBL production. They presented high Fst values and were imbedded in regions with high nucleotide diversity and significant Tajima's D values. Compared to others using small sample sizes and unphased genotypic data, we found differences in haplotyping, frequency estimation, Fu and Li's D* and Fst results.
Using extensive testing for selective neutrality, we confirmed that stochastic evolutionary factors have had a major role in shaping this polymorphic gene worldwide.
[Show abstract][Hide abstract] ABSTRACT: Mannose binding lectin (MBL) has an important role in the activation of the complement system and opsonization of pathogenic microorganisms. Frequent polymorphisms found in the MBL2 gene affect the concentration and functionality of the protein and are associated with enhanced susceptibility to severe malaria in African children. Most MBL2 typing strategies were designed to the analysis of selected variants, the significance of whole haplotypes is poorly known. In this work, a new typing strategy was developed and validated in an association analysis of MBL2 with adult asymptomatic infection.
MBL2 allele-specific fragments of 144 healthy Gabonese adults were amplified by using haplotype-specific sequencing (HSS), a new strategy that combines sequence-specific PCR and sequence-based typing. The Gabonese were investigated for the presence of Plasmodium falciparum parasitaemia by the amplification of parasite genes, immunochromatographic antigen detection and microscopic analysis. HSS results were also compared with a previously used real-time PCR (RT-PCR) method in 72 Euro-Brazilians.
Fourteen polymorphisms were identified beside the commonly investigated promoter (H, L; X, Y; P, Q) and exon 1 (A, O; O = B, C or D) variants. The MBL2*LYPA/LYPA genotype was associated with the absence of asymptomatic infection (P = 0.017), whereas the MBL2*LYQC haplotype and YA/YO + YO/YO genotypes were associated with positive parasite counts in asymptomatic adults (P = 0.033 and 0.018, respectively). The associations were specific to LYPA (identical to the reference sequence Y16577) and LYQC (Y16578) and would not have been revealed by standard genotyping, as there was no association with LYPA and LYQC haplotypes carrying new polymorphisms defined by sequence-based typing. In contrast, HSS and RT-PCR produced very similar results in the less diverse European-derived population.
In this work, a new typing strategy for a highly polymorphic gene was developed and validated focusing on the asymptomatic status of P. falciparum-infected adults. In populations with high nucleotide diversity, it allowed for the identification of associations with fine-scaled haplotypes that would not have been found using common typing techniques. In this preliminary study, MBL2 haplotypes or SNPs linked to them were found associated with susceptibility to infection and parasitaemia control of asymptomatic adults.
[Show abstract][Hide abstract] ABSTRACT: The CC chemokine receptor 5 (CCR5) molecule is an important co-receptor for HIV. The effect of the CCR5*D32 allele in susceptibility to HIV infection and AIDS disease is well known. Other alleles than CCR5*D32 have not been analysed before, neither in Amerindians nor in the majority of the populations all over the world. We investigated the distribution of the CCR5 coding region alleles in South Brazil and noticed a high CCR5*D32 frequency in the Euro-Brazilian population of the Paraná State (9.3%), which is the highest thus far reported for Latin America. The D32 frequency is even higher among the Euro-Brazilian Mennonites (14.2%). This allele is uncommon in Afro-Brazilians (2.0%), rare in the Guarani Amerindians (0.4%) and absent in the Kaingang Amerindians and the Oriental-Brazilians. R223Q is common in the Oriental-Brazilians (7.7%) and R60S in the Afro-Brazilians (5.0%). A29S and L55Q present an impaired response to β-chemokines and occurred in Afro- and Euro-Brazilians with cumulative frequencies of 4.4% and 2.7%, respectively. Two new non-synonymous alleles were found in Amerindians: C323F (g.3729G > T) in Guarani (1.4%) and Y68C (g.2964A > G) in Kaingang (10.3%). The functional characteristics of these alleles should be defined and considered in epidemiological investigations about HIV-1 infection and AIDS incidence in Amerindian populations.
Genetics and Molecular Biology 01/2009; 32(1):12-9. · 0.74 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Interferon-alpha (IFNalpha) is a critical mediator of immunity to hepatitis B virus (HBV) infection. Although IFN has been used in the treatment of viral hepatitis for more than a decade, the role of IFN-alpha-receptor in HBV infection has not been intensively studied. We have evaluated the impact of two variants of the IFNAR1 gene on the outcome of HBV infection. Four hundred and fifty eight HBV-infected Vietnamese patients, with well-characterised clinical profiles including all forms of hepatic disease, and 160 non-infected, healthy Vietnamese individuals were enrolled in the study. Of these patients, 54 had acute hepatitis B, 88 had chronic hepatitis B, 118 had liver cirrhosis, 146 had a hepatocellular carcinoma and 52 were asymptomatic carriers of HBV. We analysed two SNPs for unequal distribution between these groups. The first SNP, rs1012335 is situated in intron 3 of the interferon alpha receptor 1 (IFNAR1). A C at position 17470 in the IFNAR1 on both chromosomes was detected more frequently in HBV-infected patients compared to healthy controls (OR: 2.6; 95% CI: 1.46-4.72, p < 0.001). The same homozygosity is also associated with higher concentrations of AST and ALT (aspartate and alanine amino-transferase) in the plasma of the patients. The second SNP (rs2257167) is situated in exon 4, causing a change of amino acids from Val (GTT) to Leu (CTT). Subjects having GTT on both chromosomes were more frequent in the healthy control group (OR: 0.54, 95% CI: 0.35-0.84, p = 0.004) and had lower plasma ALT concentrations. The findings indicate that two variants of the IFNAR1 gene are associated with the clinical presentation of HBV infection.
[Show abstract][Hide abstract] ABSTRACT: Inhibition of T cell function is an important pathological feature in malaria. We investigated which T cell population is reduced contributing to immunosuppression. We examined protein and RNA level of various cell receptors, specific for T cells in children having Plasmodium falciparum infection and compared those to healthy children of the same age. We observe reduced levels of cluster of differentiation (CD)3 and T cell receptor (TCR)alphabeta in both RNA and protein expression level. This reduced expression was associated with a collapsed membrane asymmetry as determined by enhanced annexinV binding. Also human leukocyte antigen (HLA)-A,B,C- and HLA-DR-positive cells increasingly bound annexinV. The enhanced binding of annexinV was paralleled by a reduced expression of transcription factors such as T cell transcription factor 7 and GATA binding protein 3, which are involved in the expression of T cell specific genes. Also the expression of the transcription factors major histocompatibility complex class II transactivator and regulatory factor X 5, which are part of the HLA transcription machinery, is reduced during infection. We show that two mechanisms may lead to a suppression of the immune system during malaria: cell damage and reduction of gene expression of the CD3/TCR complex.
Parasitology Research 11/2008; 104(3):575-82. · 2.85 Impact Factor