Antibodies In Leprosy Research Articles

Epidemiological, clinical and immune factors that influence the persistence of antiphospholipid antibodies in leprosy

IntroductionAntiphospholipid antibodies (aPL) are described in individuals with leprosy without the clinical features of antiphospholipid antibody syndrome (APS), a condition involving thromboembolic phenomena. We have described the persistence of these antibodies for over 5 years in patients with leprosy after specific treatment.ObjectivesTo determine whether epidemiological, clinical and immunological factors played a role in the long-term persistence of aPL antibodies in leprosy patients after multidrug therapy (MDT) had finished.MethodsThe study sample consisted of 38 patients with a diagnosis of leprosy being followed up at the Dermatology and Venereology Outpatient Department at the Alfredo da Matta Foundation (FUAM) in Manaus, AM. ELISA was used to detect anticardiolipin (aCL) and anti-β2 glycoprotein I (anti-β2GPI) antibodies. Patients were reassessed on average of 5 years after specific treatment for the disease (MDT) had been completed.ResultsPersistence of aPL antibodies among the 38 leprosy patients was 84% (32/38), and all had the IgM isotype. Mean age was 48.1 ± 15.9 years, and 23 (72.0%) were male. The lepromatous form (LL) of leprosy was the most common (n = 16, 50%). Reactional episodes were observed in three patients (9.4%). Eighteen (47.37%) were still taking medication (prednisone and/or thalidomide). Mean IgM levels were 64 U/mL for aCL and 62 U/mL for anti-β2GPI. In the multivariate binary logistic regression the following variables showed a significant association: age (p = 0.045, OR = 0.91 and CI 95% 0.82–0.98), LL clinical presention (p = 0.034; OR = 0.02 and CI 95% = 0.0–0.76) and bacterial index (p = 0.044; OR = 2.74 and CI 95% = 1.03–7.33). We did not find association between prednisone or thalidomide doses and positivity for aPL (p = 0.504 and p = 0.670, respectively). No differences in the variables vascular thrombosis, pregnancy morbidity, diabetes, smoking and alcoholism were found between aPL-positive and aPL-negative patients.ConclusionPersistence of positivity for aPL antibodies was influenced by age, clinical presentation and bacterial index. However, further studies are needed to elucidate the reason for this persistence, the role played by aPL antibodies in the disease and the B cell lineages responsible for generation of these antibodies.

Anti-ceramide antibodies in leprosy: marker for nerve damage?

Leprosy is a chronic infectious disease primarily affecting the peripheral nervous system and skin. Multibacillary leprosy is associated with nerve damage which could contribute to myelin alteration. As ceramide is a constituent of myelin sheath, the present study aimed to compare anti-ceramide antibody titre in paucibacillary and multibacillary leprosy patients with controls. Serum levels of anti-ceramide antibody were measured using enzyme-linked immunosorbent assays (ELISA) in 50 leprosy patients (25 paucibacillary and 25 multibacillary) and 25 healthy controls. Results were reported in OD units as mean +/- SD and analyzed by Chi square test (significance at p < 0.05). Patients suffering from multibacillary leprosy had significantly higher anti-ceramide antibody serum levels compared to paucibacillary leprosy patients and healthy controls (p < 0.005). Since nerve damage is the most debilitating effect of leprosy, the search for a serum marker for assessing nerve damage is required in countries where leprosy is still widespread. In multibacillary leprosy patients, the role of anti-ceramide antibody as a marker for nerve damage should be explored.

Polimorfismo Val247Leu do gene β2-glicoproteína 1 pode justificar a gênese de anticorpos antiβ2GP1 e síndrome do anticorpo antifosfolípide na hanseníase multibacilar

BACKGROUND - Multibacillary (MB) leprosy may be manifested with antiphospholipid antibodies (aPL), among which anti-beta2GP1 (beta2-glycoprotein 1). High titers of aPL are associated with APS (Antiphospholipid Syndrome), characterized by thrombosis. The mutation Val247Leu in the domain V of beta2GP1 exposes hidden epitopes with consequent development of anti-beta2GP1 antibodies. To evaluate the Val247Leu polymorphism of beta2GP1 gene and its correlation with anti-beta2GP1 antibodies in leprosy patients. The Val247Leu polymorphism was performed by PCR-RFLP and anti-beta2GP1 antibodies were measured by ELISA. The genotypic Val/Val was more prevalent in the leprosy group, compared to controls. Regarding the 7 MB patients with APS, four presented heterozygosis and three, Val/Val homozygosis. Although higher titrations of anti-beta2GP1 IgM antibodies were seen in MB leprosy group with Val/Leu and Val/Val genotypes, there was no statistical difference when compared to Leu/Leu genotype. The prevalence of Val/Val homozygosis in leprosy group can partially justify the presence of anti-beta2GP1 IgM antibodies in MB leprosy. The description of heterozygosis and Val/Val homozygosis in 7 patients with MB leprosy and thrombosis corroborates the implication of anomalous phenotype expression of beta2GP1 and development of anti-beta2GP1 antibodies, with consequent thrombosis and APS.

Anti-neutrophil cytoplasmic antibodies in leprosy

Anti-Neutrophil Cytoplasmic Antibodies (ANCA) are auto-antibodies directed to intracellular components of neutrophils and used to be considered as present almost exclusively in granulomatous vasculitis. Recently, these auto-antibodies have been found in other autoimmune disorders as well as infectious diseases. We studied patients with leprosy confirmed by bacilloscopy and/or skin biopsy, in reaction phase from the Ambulatório de Hanseníase do Hospital Universitário Professor Edgar Santos. ANCA and Antinuclear antibodies (ANA) were determined by indirect immunofluorescence using commercially available kits. Twenty patients were enrolled in our study, nine males and 11 females. The mean age was 36.9+/-18.2 years. ANCA were present only in one patient, with a perinuclear staining pattern (p-ANCA), and no patient tested positive for ANA. Although other studies have shown the presence of ANCA in leprosy, the low frequency of these antibodies in leprosy sera demonstrated in the present study illustrates the high specificity of ANCA for the diagnosis of Wegener granulomatosis.

Immune profiling of leprosy and tuberculosis patients to 15-mer peptides of Mycobacterium leprae and M. tuberculosis GroES in a BCG vaccinated area: implications for development of vaccine and diagnostic reagents.

Mycobacterium leprae (ML) GroES has been shown to induce strong T cell responses in tuberculoid as well as in exposed healthy contacts of leprosy patients, and therefore this antigen has been the focus of study as a potential vaccine candidate. Paradoxically, we have shown that ML GroES also induces extremely high titres of IgG1 antibody in leprosy patients across the disease spectrum, a response associated with disease progression. IgG1 antibodies in leprosy also show a negative association with interferon-gamma, a critical T cell cytokine responsible for macrophage activation and intracellular killing of mycobacteria. We therefore queried if antibody and T cell responses were being evoked by different epitopes in ML GroES proteins. To address the issue of epitope recognition in mycobacterial diseases, we have analysed 16 peptides (15-mer peptides) spanning the entire ML and M. tuberculosis GroES protein in leprosy (n = 19) and tuberculosis (n = 9) patients and healthy endemic controls (n = 8). Our analysis demonstrates clearly that the dominant peptides evokingT cell and IgG subclass antibodies were different. The target of both T and B cell responses were cross-reactive epitopes in all groups. Differences in disease and healthy states related to the strength (mean intensity) of the T cell and antibody response. IgG1 and IgG3 antibodies were associated with disseminated disease and IgG 2 and IgG4 with disease limitation. Such comprehensive immune profiling of antigen-specific responses is critical to understanding the disease pathogenesis and also if these reagents are to be exploited for either diagnostic or vaccine purposes.

Effects of anti-endothelial cell antibodies in leprosy and malaria.

As a result of damaging endothelial cells (ECs), Mycobacterium leprae triggers the production of antibodies (Abs). These anti-EC Abs (AECAs) can be divided into two types. The first type nonspecifically reacts with components of the cytosol (CY) and can be detected by enzyme-linked immunosorbent assay (ELISA). The second specifically reacts with the EC membrane (MB) and requires fluorescence-activated cell sorter (FACS) analysis to be detected. The presence of both types of AECAs was determined in 68 leprosy patients. The ELISA was positive for 35 of them but also for 30 of 34 malaria patients and 17 of 50 healthy African controls. However, whereas FACS analysis showed MB reactivity in only three malaria patients and four controls, this reactivity was found in 27 leprosy patients, more of those having the lepromatous than the tuberculoid form. Specificity for MB, which we failed to absorb by incubation with CY lysates, predominated over that for CY in leprosy, unlike malaria, where the EC reactivity was restricted to the CY. Western blot analysis and two-dimensional electrophoresis revealed that calreticulin, vimentin, tubulin, and heat shock protein 70 were targeted by AECAs from leprosy patients, but other proteins remained unidentified. These auto-Abs, but not those from malaria patients, did activate ECs, as indicated by the E-selectin and intercellular adhesion molecule 1 upregulation, and/or induced them into apoptosis, as documented by four different methods. Our findings suggest that, in some but not all leprosy patients, AECAs may play a role in pathogenesis.

High prevalence of antiphospholipid antibodies in leprosy: evaluation of antigen reactivity.

Antiphospholipid antibodies (aPL) have been reported not only in autoimmune disorders but also in various infectious diseases. Accumulating evidence indicates that beta2 glycoprotein I (beta2GPI) and prothrombin are the main proteins to which autoimmune aPL bind. The aim of this study was to evaluate the prevalence of different aPL in patients with leprosy. We included 51 outpatients (42 lepromatous and 9 borderline leprosy) without any clinical feature of the antiphospholipid syndrome (APS). 35 had lupus anticoagulant and 31 had anticardiolipin antibodies (aCL). Anti-beta2GPI antibodies were highly positive in 29/51 and anti- prothrombin antibodies (anti-II) were detected in 23/51. Almost all aCL and anti-beta2GPI were of IgM isotype, while IgG isotype was more frequent among anti-II. No statistical difference was found when aPL were evaluated in patients grouped according to their bacteriological status. Furthermore, patients under treatment (n=33) had a similar frequency of positive aPL compared to patients in vigilance (n=14). Assessing the specificity of antibody binding to CL and beta2GPI in ELISA by means of inhibition studies with cardiolipin-beta2GPI liposomes, leprosy and APS sera showed a similar behaviour. Comparable results were also found in both groups of patients when inhibition experiments with lysate of Mycobacterium leprae were carried out. In summary, leprosy-related aPL resemble those found in patients with APS but the immunoglobulin isotype is different, with IgM much more prevalent in leprosy patients.

Antibodies to sulfatide in leprosy and leprosy reactions.

Antibodies to sulfatide have been reported in various demyelinating peripheral polyneuropathies. We have investigated the diagnostic value of these antibodies in leprosy. Anti-sulfatide IgM in leprosy patients was not significantly elevated. High anti-sulfatide IgG titers were observed in individuals from endemic areas, irrespective of their leprosy status, while western European controls were negative. No significant correlation was found between IgM or IgG antibody titers and leprosy classification, although multibacillary patients had higher anti-sulfatide IgM titers than paucibacillary patients. In addition, 23 patients developing leprosy reactions were followed longitudinally. Antibody titers in these patients fluctuated slightly during the follow-up period. There was no association with the occurrence of leprosy reactions or treatment. Thus, IgG titers against sulfatides are high in both leprosy patients and healthy controls in endemic areas, whereas such antibodies are not found in western European controls, suggesting that these antibodies are induced by environmental factors, such as microorganisms.

IgG Antibodies in Leprosy and Their Relation to Thl/Th2 Responses

The study of the IgG subclasses of antibodies across the clinical spectrum of leprosy id not reveal differences that could be related to the Th1/Th2 dichotomy; the predominance of antibodies of the IgG1, 2 and 3 subclasses suggest a possible Th1 response across the spectrum in Venezuelan patients. In contrast, the anergic form of American cutaneous leishmaniasis was characterized by a strong IgG4 specific antibody response, suggesting a Th2 response, while localized forms of the disease appeared to be associated with responses of the Th1 type. Persisting antibodies to phenolic glyco- lipid-I in a small percentage of clinically inactive multibacillary patients may reflect differences in the catabolism of this unusual antigen, since persistence was not associated with a unique initial IgG subclass response to a complex extract of Mycobacterium leprae antigens.

Staining pattern of seven monoclonal anti-CD26 antibodies in leprosy: implications for the use of CD26 as a surrogate marker of a human Th1-like reaction.

In a previous study using the monoclonal anti-CD26 antibody MIB-DS2/7 in leprosy and other granulomatous diseases, it was shown that CD26 may be a candidate for use as an operational marker of a human Th1-like reaction. In this follow-up study, we compared seven different monoclonal anti-CD26 antibodies with respect to their staining pattern in lepromatous and tuberculoid leprosy tissues. Three distinct staining patterns became apparent in this anti-CD26 antibody panel: staining of T-lymphocytes and of connective tissue; staining of T-lymphocytes, connective tissue and macrophages; and almost no staining of T-lymphocytes but staining of connective tissue and macrophages. The two antibodies assigned to the first staining pattern, including MIB-DS2/7, were found to be most suitable for the operational discrimination between Th1-like and Th2-like reactions in leprosy. The antibodies assigned to staining patterns 2 and 3 did not allow this discrimination. Although all seven monoclonal antibodies investigated were specific for CD26, only two were found to be useful in identifying a Th1-like immune reaction in human tissue.

Prevention and Early Detection of Leprosy in Children

India with its 4 million cases of leprosy, accounts for one-third of the world's population of leprosy patients. One-fourth of them are below 15 years of age. We report a 5-year follow-up study of healthy children who were close contacts of leprosy patients, in order to: 1. detect subclinical infection and observe the development of overt disease by using the Fluorescent Leprosy Antibody Absorption Technique (FLA-ABS) and the lepromin test which assess the humoral and cell-mediated immunity (CMI), respectively; 2. evaluate the efficacy of dapsone as a chemoprophylactic agent in the 'at risk' contacts. Four-hundred-and-fifty-five healthy contacts were studied. Majority of the contacts of multibacillary patients (303) were FLA-ABS positive (75 percent) and lepromin negative (55 percent) showing that although most of them had been infected, the lepromin status was negative (P < 0.01). On the other hand, the majority of the contacts of paucibacillary patients (152) were lepromin positive (57 percent) (P < 0.05). Furthermore, only 61 percent of contacts of paucibacillary patients were FLA-ABS positive as compared to 75 percent of contacts of multibacillary patients demonstrating that the former had been exposed to a lesser quantum of infection (P < 0.05). On the basis of results of FLA-ABS and lepromin tests, these 455 contacts were classified into four groups, viz. Group I comprising children who were FLA-ABS positive and lepromin positive; Group II, who were FLA-ABS positive and lepromin negative; Group III, who were FLA-ABS negative and lepromin positive; and Group IV who were FLA-ABS negative and lepromin negative.(ABSTRACT TRUNCATED AT 250 WORDS)

Anticardiolipin antibodies in infections are heterogenous in their dependency on beta 2-glycoprotein I: analysis of anticardiolipin antibodies in leprosy.

We studied the effect of beta 2-GPI on binding of antibodies in sera from patients with leprosy and patients with the antiphospholipid syndrome (APS) to CL in enzyme-linked immunosorbent assays (ELISAs). Increased levels of IgG aCL were detected in 59 of 61 leprosy patients' sera by the standard aCL-ELISA in the presence of bovine beta 2-GPI and in 60 of the 61 leprosy patients' sera by the modified aCL-ELISA without beta 2-GPI. When tested by both aCL-ELISAs on the same plate, 10/31 leprosy sera and 9/10 APS sera bound better in the standard aCL-ELISA, 16/31 leprosy sera bound better in the modified aCL-ELISA and in five leprosy and one APS sera the difference was not significant. A dose-dependent enhancing effect of beta 2-GPI on the leprosy and APS sera binding to CL was confirmed using purified human beta 2-GPI. Enhanced binding was seen if beta 2-GPI was added either before or together with the test serum. In 11/61 leprosy sera increased levels of IgG antibodies against beta 2-GPI were found by ELISA. Leprosy anti-beta 2-GPI antibodies appear to be a separate antibody population recognizing only beta 2-GPI adsorbed on the ELISA plate. These results demonstrate heterogeneity of leprosy aCL with respect to their beta 2-GPI requirement for binding to CL.

The role of antiperipheral nerve antibodies in nerve damage in leprosy.

The objective of this study was to determine the role of antineural antibodies in leprosy. Indirect ELISA using antigen prepared from normal human peripheral nerves was carried out on the sera from 100 leprosy patients and 18 normal controls. In total, 9% of the patients had demonstrable levels of IgG antineural antibodies and 11% had demonstrable levels of IgM antibodies. There was no correlation with the type of leprosy, bacteriological index, treatment taken, the presence of a reactional state, the presence of enlarged nerves or active neuritis.

Antimitochondrial (pyruvate dehydrogenase) antibodies in leprosy.

Sera from 69 patients with leprosy but without liver involvement were assayed for the presence of mitochondrial pyruvate dehydrogenase (PDH)-specific autoantibodies by enzyme-linked immunoabsorbent assay (ELISA), immunoblotting using PDH as an antigen and by enzymatic inhibition test. Twenty-seven of the leprosy serum samples (39.1%) were found to react with PDH by ELISA. However, unlike sera from primary biliary cirrhosis (PBC) patients, none of these were able to inhibit the PDH enzymatic activity. By immunoblotting, it was found that only 2 of the 27 positive sera recognized the 74-kD protein of the PDH complex, which is recognized by sera of most PBC patients. The antimitochondrial antibodies in lepra most probably recognize different epitopes than those in PBC. These findings may indicate that anti-PDH autoantibodies in patients with leprosy may arise by polyclonal B cell stimulation and may represent natural anti-PDH autoantibodies.

Early Detection of Leprosy in Children

Healthy children contacts of leprosy patients had their humoral and cell-mediated immunological status assessed using the Fluorescent Leprosy Antibody Absorption Technique (FLA-ABS) and the Lepromin test, respectively. Subsequently, they were followed up for 2 1/2 years to study the development of overt disease. Two-hundred children were studied and classified into four groups, viz. Group I comprised of children who were FLA-ABS positive and Lepromin positive; Group II = FLA-ABS positive and Lepromin negative; Group III = FLA-ABS negative and Lepromin positive; Group IV = FLA-ABS negative and Lepromin negative. The good cell-mediated immune (CMI) response in the 107 children in Group I prevented them from developing the disease though they had been infected. Out of the 37 children in Group II, 15 developed the disease. There were no children in Group III. None of the 56 children in Group IV developed the disease, possibly because they had not been significantly infected. All these findings were statistically significant (P less than 0.01). This study highlights the protective role of cell-mediated immunity in leprosy. It also suggests the need to carry out surveillance surveys in the endemic population to identify and follow up those at risk.

Epidemiological Immunological Studies on Leprosy in Okinawa. 6. Lepromin, Lymphocyte Transformation,

In-and out-patients with leprosy in Okinawa were surveyed by using the following im-munological tests: cutaneous reaction to Mitsuda's lepromin (40 million bacilli/ml) (LPR), lymphocyte transformation test with Dharmendara's antigen (DL-LTT), that with phyto-hemagglutinin (PHA-LTT), fluorescent leprosy antibody absorption test (FLA-ABS) and Leproagglutination test with cardiolipin-lecithin antigen (LAT). A correlation between two tests and a relationship between the test and personal or family history or clinical finding of the patients were evaluated by non-parametric statistics. Spearman's rank cor-relation coefficient was significant between LPR and DL-LTT, while a reverse correlation was found beween LPR and FLA-ABS, DL-LTT and FLA-ABS, and DL-LTT and LAT, respectively.High reactivity in PHA-LTT was found in almost all of patients, irrespective of type and stage of leprosy, suggesting general immune responsiveness of leprosy patients in Okinawa. The other tests showed a significant difference in the mean rank of reaction values according to clinical findings such as type and stage of leprosy, property of skin lesion, loss of eyebrow, and bacteriological test. The use of LPR and DL-LTT for the study of cell-mediated immunity in leprosy was therefore confirmed. FLA-ABS was found useful for early serodiagnosis, while LAT for the study of autoimmunity in leprosy.None of these immunological tests showed significant difference among the subgroups of patients classified by the following items: presence or absence of consanguineous patient with leprosy, living place, place of onset, presence or absence of neural symptoms such as the enlargement of peripheral nerve, claw hand and facial palsy, ocular and nasal involvement, ENL in lepromatous and borderline leprosy and conditions in the treatment. On the other hand, LPR showed higher reactivity in female than in male, in the patients at home than those discharged, and in the patients with drop foot than those without. Higher reactivity of DL-LTT was also found in male than in female and in the patients with plantar ulcer than those without. FLA-ABS reactivity was significantly higher in the discharged patients than the inpatients, in the period of 5 to 9 years after onset than that of 20 or more years, but the reactivity was lower in the patients with drop foot than those without. LAT reactivity was significantly higher in the 30-39 years old patients than 0-19 years old, and in the patients with physical work than those with mental. These findings were discussed by comparing with those described in the previous papers of this series and from immunological point of view on leprosy.

The sensitivity and specificity of fluorescent leprosy antibody absorption (FLA-ABS) test for detecting subclinical infection with Mycobacterium leprae

The sensitivity and specificity of fluorescent leprosy antibody absorption (FLA-ABS) test for detecting subclinical infection with Mycobacterium leprae

Subclinical infection in leprosy--its detection and control by fluorescent leprosy antibody absorption (FLA-ABS) test.

Subclinical infection in leprosy--its detection and control by fluorescent leprosy antibody absorption (FLA-ABS) test.

Clinical significance of autoimmune antibodies in leprosy

Abstract Specimens of sera from 73 leprosy patients and from 20 controls were tested for rheumatoid factor, antinuclear factor, thyroglobulin antibody and for certain organ specific antibodies (gastric parietal cell, thyroid cytoplasmic, and mitochondrial antibodies). A significant proportion of the lepromatous leprosy patients showed positive reactions to rheumatoid factor, thyroglobulin, and antinuclear factor in low titre. The presence of non-organ specific autoantibodies bears no clinical significance to the disease. Organ specific autoantibodies were not common in leprosy sera.