Studies of association of polimorfismos of only base in genes candidates in the leprosy. Role of gelatinases, their inhbitors, and matrix molecules in cutaneous lesions from american cutaneous leishmaniasis

by Cardoso, Cynthia Chester

Abstract (Summary)
Leprosy is a chronic infectious disease caused by the intracellular pathogen Mycobacterium leprae, which has tropism for Schwann cells in peripheral nerves and macrophages in the skin. The spectrum with five different clinical forms is determined by host:pathogeninteractions that influences a polarized immune response represented as the tuberculoid and lepromatous forms of the disease, which are associated to Th1 x Th2 immunological patterns, respectively. The sequence analysis of M. leprae showed low genetic variability in a way that both leprosy per se and the spectrum of manifestations are explained mostly due to host responses. This observation has encouraged many studies aiming the identification of markersof susceptibility to leprosy per se or to one of the clinical forms. The aim of this study was to replicate in a new population previous published results that reported association between single nucleotide polymorphisms (SNPs) and leprosy susceptibility. It was designed a casecontrolstudy using 233 healthy unrelated household contacts and 212 patients from all clinical forms of leprosy. Samples were genotyped using PCR-RFLP or allelic discriminationmethod for SNPs in TNFA (-308), LTA (+252) and PARK2 (-2599 and rs1040079) genes. Deviations from the Hardy-Weinberg Equilibrium were detected in SNPs located at TNFlocus (-308 in patients and +252 in control group) and for the SNP ?2599 at PARK2 gene in both populations. The population-based analysis replicated the protection previously reportedfor allele TNF-308A (OR=0,59; P=0,03) and also for ?308A/+252A haplotype, although it was not significant (OR=0,47 P=0,06). Analysis with PARK2 SNPs and haplotypes did not show any association. A novel SNP (asp110ala) at NINJ1 gene was also investigated so as to determine a possible association with leprosy. There was no evidence of association of this SNP and leprosy susceptibility and severity, but the first analysis suggested an associationwith neurological damage as assessed by the disability grade. So, using an extended patients population (n=765) we confirmed the association between allele 110ala and incapacity grade (OR=1,54; P=0,007). Functional analysis detected a higher ninjurin mRNA and protein expression in response to M. leprae stimulus. Allele-specific transcript quantitation demonstrated that allele 110asp was augmented as compared to 110ala. Here, we confirm the association of -308A allele with resistance of leprosy per se among Brazilians. Ninjurin analysis showed that asp110ala could be a valuable prognostic marker, since carriers of ala110 allele have increased susceptibility to nerve damage possibly because this allele shows a decreased expression of this adhesion molecule. Leishmaniasis is an important antropozoonosis that affect the entire world. In Brazil, the main clinical form is American cutaneous leishmaniasis (ACL) caused by Leishmania (V.) braziliensis in the most of the cases. This disease is characterized by the formation of wounds at the site of the sand fly bite. The primary lesions are cutaneous, and after their healing or during their evolution, secondary lesions located at mucous tissues can appear as a result of the parasite metastatization. In general, these lesions are predisposed to be chronic, originating ulcers with an intense process of tissue destruction. Throughout the regenerative process, the synthesis and breaking of many different matrix compounds are necessary in order to allow proliferative and migratory events, which will conclude the repairing step. A family of endopeptidases known as matrix metalloproteinases (MMPs) takes part of this process. An important subfamily is the gelatinases MMPs 2 and 9, which are involved in the wound re-epithelization and resolution. MMP activity is strongly controlled by proteins called tissue inhibitors of matrix metalloproteinases (TIMPs). The loss of this control can result in critical tissue degradation. The present work aimed to evaluate the role of gelatinases A (MMP-2) and B (MMP-9), their respective inhibitors TIMPs 2 and 1, and matrix molecules which are substrates for these enzymes in cutaneous ulcers from ACL patients. To achieve this objective, it was used the following techniques: RT-PCR to evaluate the gelatinases and TIMPs mRNA expression, in situ zymography to visualize the tissue sites of gelatinases activity, and immunohistochemistry to detect fibronectin, type IV collagen, laminin, and tenascin. At last, data were correlated to important clinical variables of ACL. It was found that patients that have a low frequency of MMPs and TIMPs mRNA expression also showed a bad responsiveness to therapy, lesions greater than 5.8 cm2, and/or lesion located at the lower limbs, in addition to an intense gelatinase activity and a high matrix molecules expression. These facts suggest that the imbalance between MMPs synthesis and activity can impair the healing process. Moreover, recent lesions express high levels of gelatinases, while the old ones seem to have high expression levels of inhibitors, what can favor the healing process. Therefore, we suggest that gelatinases and their inhibitors can be important in the leishmanial lesions outcome.
This document abstract is also available in Portuguese.
Bibliographical Information:

Advisor:Milton Ozório Moraes

School:Faculdades Oswaldo Cruz

School Location:Brazil

Source Type:Master's Thesis

Keywords:Hansenáise Leprosy single nucleotide polymorphisms leishmaniose braziliensis gelatinases cutaneous leishmaniasis cellular matrix


Date of Publication:03/24/2006

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