HLA segregation of tuberculoid leprosy: confirmation of the DR2 marker

Families with multiple cases of leprosy were tested for HLA (histocompatibility leukocyte antigen)-linked control of susceptibility to tuberculoid leprosy and association with HLA-DR2. Thirty-one non-HLA genetic markers were also examined for indications of non-HLA-linked genetic factors that might control susceptibility to tuberculoid leprosy. A significant (P = 0.002) preferential inheritance of HLA-DR2 by siblings affected with tuberculoid leprosy, but not by healthy siblings nor by siblings affected with lepromatous leprosy, was observed. In addition, combined family data showed a significant (P less than 0.0025) excess of identical HLA haplotypes inherited from healthy parents by siblings affected with tuberculoid leprosy. Segregation on non-HLA polymorphisms did not deviate significantly from what would have occured randomly. These data are compatible with a recessive inheritance of HLA-linked susceptibility to tuberculoid leprosy. The preferential segregation of DR2 observed in children with tuberculoid leprosy (P less than 0.001 for the combined data from India) indicates that the HLA-linked susceptibility gene is either DR2 or in linkage disequilibrium with it.     

HLA-linked control of predisposition to lepromatous leprosy

In a study of the relation between HLA and lepromatous leprosy, HLA haplotype segregation was analyzed in 28 families with multiple cases of different types of leprosy. The inheritance of HLA-DR2, HLA-DR3, and HLA-MT1, which had previously been shown to be associated with susceptibility to leprosy or with a leprosy type, was analyzed separately. Segregation occurred in a significantly nonrandom fashion in both polar tuberculoid leprosy and lepromatous leprosy. This finding indicated HLA-encoded control of a predisposition to both of these forms of the disease. In both cases the segregation observed among healthy siblings was random. Thus, susceptibility to leprosy per se is probably not controlled by HLA-linked genes. HLA-DR3 was inherited preferentially by children with polar tuberculoid leprosy rather than lepromatous disease (P = .02), and HLA-MT1 was inherited preferentially by children with lepromatous leprosy (P = .04). The results confirmed the association of these genetic markers with leprosy type.     

Human leukocyte antigens in forms of leprosy among Japanese patients

Human leukocyte antigens (HLA) class II alleles were analyzed among Japanese leprosy patients to ascertain whether immunogenetic differences exist among the leprosy classification forms of Ridley and Jopling. Ninety-three unrelated Japanese leprosy patients (21 lepromatous, 24 borderline lepromatous, 17 mid-borderline, 26 borderline tuberculoid, 5 tuberculoid) and 114 healthy control subjects were investigated. The frequencies of HLA-DRB1*1501, -DRB5*0101, -DQA1*0102 and DQB1*0602 were significantly increased in all of the Japanese leprosy patients. The frequencies of HLA-DRB1*0405, -DQA1*03 and -DQB1*0401 were significantly decreased in the Japanese patients after correction of the p value. Conversely, there were no significantly different distributions of the HLA-DRB1, -DRB5, -DQA1, DQB1 alleles in the five subgroups of these patients. We conclude that HLA class II alleles were not associated with the form of leprosy. Other HLA, a non-HLA gene, and/or environmental factors may play a critical role in the different manifestations of leprosy.     

Determination of circulating IgG subclasses against lipoarabinomannan in the leprosy spectrum and reactions

IgG subclasses against lipoarabinomannan of mycobacteria were analyzed in the sera of leprosy patients. Patients with active leprosy [tuberculoid and lepromatous, patients undergoing erythema nodosum leprosum (ENL) and reversal reactions] and inactive cases (tuberculoid and lepromatous who were cured after chemotherapy) were included in this study. Active lepromatous patients had higher levels of IgG subclasses, except IgG4, compared to active tuberculoid patients. Some of the inactive cases (lepromatous patients cured after chemotherapy) were positive for the IgG1, IgG2 and IgG3 subclasses. However, their levels are lower than active lepromatous cases. On the other hand, no difference in the subclass levels between the active and inactive tuberculoid groups could be observed. While a significant fall in the level of IgG3 in ENL was observed as compared to lepromatous leprosy without ENL, higher levels of IgG1 and IgG2 were found in patients with reversal reactions compared to their active counterparts without reactions.     

The Incidence of HLA Antigens in Leprosy

Abstract. HLA antigens were studied in 36 patients, with leprosy, 20 cases of lepromatous and 16 cases of tuberculoid type. Eleven out of 36 (30.55%) had BW40 as compared to 9.33% of 150 controls. The frequency of BW40 in tuberculoid patients (31.25%) was not different from that in lepromatous cases (30%).     

HLA-linked genes and leprosy: a family study in Karigiri, South India.

The evidence for a genetic determination of susceptibility to leprosy is reviewed. To test the hypothesis that an HLA (histocompatibility leukocyte antigen)-linked gene is associated with such susceptibility, the association between the distribution of leprosy within a family and the segregation of HLA haplotypes was investigated among 72 families who lived in Karigiri, Tamil Nadu State, South India. A statistically significant association was found for families in which siblings had tuberculoid leprosy and in which neither parent had leprosy. The findings from the data of this study agree with those of two previous studies carried out among smaller populations is Surinam and Wardha, Maharashtra State, India. Such an agreement suggests that a genetic determinant which is linked to the major HLA locus on chromosome 6 and which is probably recessive affects susceptibility to tuberculoid leprosy in humans.     

Histologic responses in sixty multibacillary leprosy patients inoculated with autoclaved Mycobacterium leprae and live BCG

Sixty lepromatous or borderline lepromatous patients were submitted to immunotherapy with a mixture of autoclaved Mycobacterium leprae and BCG. The histopathologic findings in skin biopsy specimens taken before and after immunotherapy were evaluated independently by six histopathologists in a workshop setting. Their pooled observations on diagnosis and classification were analyzed to assess the histopathologic changes following various periods of immunotherapy. Expressing the results as the average value of five to six independent observations, there were changes in classification of reversal or upgrading toward the tuberculoid end of the leprosy spectrum in 90.5% of the patients initially classified as lepromatous (LL), and in 83.3% of those initially classified as borderline lepromatous (BL). The histopathologic findings amply support the clinical, bacteriologic and immunological changes following immunotherapy from LL or BL, to BL, mid-borderline (BB) or even borderline tuberculoid (BT) leprosy.     

Defective blood mononuclear phagocyte function in patients with leprosy.

Patients with lepromatous leprosy possess a defective lymphocyte function in vivo and in vitro that is less evident in the tuberculoid form. Data concerning their macrophage ability to digest Mycobacterium leprae are controversial. The purpose of this study was to determine whether monocytes from patients with either tuberculoid or lepromatous leprosy were altered in their enzyme systems, that is myeloperoxidase-dependent and myeloperoxidase-independent systems. The ability of adherent blood monocytes to ingest and kill Candida pseudotropicalis after 30 and 60 min of incubation with yeast cells was tested. Mononuclear phagocytic cells from patients with either principal form of leprosy functioned similarly to normal monocytes in phagocytosis while their fungicidal activity for C. pseudotropicalis was statistically significantly altered and was more evident in the lepromatous than in the tuberculoid type. The results indicate that peripheral blood monocytes from patients with leprosy possess an impaired enzymatic candidacidal activity.     

In vivo responses to Mycobacterium leprae: antigen presentation, interleukin-2 production, and immune cell phenotypes in naturally occurring leprosy lesions

To investigate the immune defect in lepromatous leprosy we studied immune cell phenotypes, lymphocyte activation states, and interleukin-2 (IL-2) production in naturally occurring leprosy skin lesions. Mouse hybridoma monoclonal antibodies reacting with the IL-2 receptor (anti-Tac), unbound IL-2 (DMS-1), antigen-presenting Langerhans' cells (OKT6) and the OKT4-Leu3 and OKT8 T-lymphocyte subpopulations were used with indirect horseradish peroxidase and alkaline phosphatase techniques on frozen biopsy sections. The percentage of Tac+ lymphocytes and the number of OKT6+ cells in the epidermis and dermal granuloma were significantly correlated in naturally occurring lesions (correlation coefficient 0.79) and were higher in tuberculoid than in lepromatous lesions. Leu3 antigen was expressed by 70-90% of Tac+ cells in tuberculoid lesions. Although the percentage of cells producing IL-2 was low in lesions of both lepromatous and tuberculoid patients, it was about 15 times greater in tuberculoid than in lepromatous lesions (0.032 +/- 0.037 tuberculoid vs 0.0019 +/- 0.023 lepromatous). There was an association between the number of OKT6+ cells and the percentage of IL-2-producing cells, but the association was weaker than that of OKT6+ cells and the percentage of IL-2 receptor-bearing cells (r = 0.2), implying that IL-2 production is not an intervening variable in the latter association. The absolute number of OKT4-Leu3+ lymphocytes was significantly different in different clinical leprosy groups and was positively correlated with host resistance (mean OKT4-Leu3+ cells/mm2 in 6 micron sections; 1412 +/- 288 tuberculoid, 400 +/- 93 borderline lepromatous, 200 +/- 100 polar lepromatous; r = 0.95). Absolute numbers of OKT8+ cells/mm2 in lesions were not significantly different. We conclude that there is a relative paucity of OKT4-Leu3+ cells as well as IL-2-producing cells at the local level in lepromatous leprosy lesions. Possible functional relationships between these findings and the failure of macrophage activation and destruction of Mycobacterium leprae in lepromatous leprosy are discussed.     

HORMONAL CHANGES ASSOCIATED WITH TESTICULAR ATROPHY AND GYNAECOMASTIA IN PATIENTS WITH LEPROSY

Basal LH, FSH, 17 beta-oestradiol and testosterone and the gonadotrophin responses to luteinizing hormone releasing hormone (LHRH) were studied in male patients with leprosy (twenty-four with lepromatous and six with tuberculoid leprosy). The mean basal LH and FSH was significantly elevated in the lepromatous group and was associated with an excessive response of both gonadotrophins following LHRH administration. The mean basal testosterone and 17 beta-oestradiol values in the lepromatous group were significantly lower than those of the tuberculoid and control groups. The abnormal gonadotrophin and sex steroid values in the lepromatous group are in keeping with the testicular atrophy and gynaecomastia accompanying this form of leprosy. However, the lack of a significant correlation between basal FSH and testicular atrophy should be noted. In addition, no correlation between any of these hormonal values and gynaecomastia could be demonstrated. The patients with tuberculoid leprosy had essentially normal hormonal profiles (except for two who had raised 17 beta-oestradiol values). This is compatible with the lack of gonadal involvement in these patients.     

Biochemical, immunological and genetic studies in leprosy. II. Profile of immunoglobulins, complement components and C-reactive protein in sera of leprosy patients and healthy controls.

Various classes of immunoglobulins (IgA, IgM, IgG, IgD and IgE), complement components (C3 and C4) and C-reactive protein (CRP) were estimated in sera from normal healthy controls and leprosy (lepromatous and tuberculoid) patients from Ethiopia. Higher levels of IgA, IgM, IgG and IgD were found in lepromatous leprosy compared with normal healthy people while in tuberculoid leprosy only IgM, IgG and IgD levels were increased. Borderline leprosy patients showed increase in IgG level only. Although an increase in IgE was noted in lepromatous leprosy, it was not significant; the variations in IgE levels could be due to different socioeconomic background and exposure to intestinal parasites. C3 component was significantly reduced in leprosy patients compared with healthy controls while no difference in C4 component was observed. The results point towards an involvement of the "alternate pathway". A positive test against C-reactive protein antiserum was given by about 20% of the normal healthy controls while more than 60% lepromatous and tuberculoid leprosy patients were CRP positive. The results are discussed in relation to the status of immunoglobulins and complement components in leprosy and possible factors (environmental and genetic) which might affect them.     

In vitro studies on dermal leprosy granulomas: assessment of division and protein synthesis of cells

Single cell suspension from dermal leprosy granulomas (10 tuberculoid and 10 lepromatous) was prepared and an assessment of the division and protein synthesis by the cells was made. The cells of tuberculoid granulomas showed a high incorporation of 3H-thymidine and 14C-leucine. On the contrary, the cells of the lepromatous granulomas exhibited poor division but their protein synthesis remained unimpaired. These observations suggest that the epithelioid cell granuloma of tuberculoid leprosy appears to be more active and secretory than the macrophage granuloma of lepromatous leprosy.     

Salivary immunoglobulins and antibody activities in leprosy

The technics of immunodiffusion and the fluorescent leprosy antibody absorption (FLA-ABS) test were used to determine the levels of immunoglobulins and their antibody activities against Mycobacterium leprae in the serum and the saliva collected from a total of 110 patients with leprosy (50 lepromatous, 24 borderline, and 36 tuberculoid). The average levels of serum IgG, IgM, and IgA were not significantly different among these patients. In saliva, however, IgM was detected in only two cases with lepromatous leprosy and three tuberculoid cases. Salivary IgG and IgA levels and their ratios to those in the sera were not significantly different according to the classification of leprosy. The percentages of positive FLA-ABS tests in the sera and saliva were compared by using fluorescent antibodies specific for IgG, IgM, and IgA, respectively. The results indicated that M. leprae-specific antibodies in the serum were mainly found in IgG and IgM and, less frequently, in IgA. IgG antibodies were found more frequently in lepromatous and borderline patients than in tuberculoid cases. On the other hand, salivary IgA antibodies against M. leprae were found in a significant number of specimens; whereas IgG and IgM antibodies were scarcely found. However, the percentage of positive FLA-ABS tests caused by salivary IgA antibodies was higher in the patients with tuberculoid or borderline leprosy than in those with lepromatous leprosy. A significant number of patients with tuberculoid or borderline leprosy secreted M. leprae-specific IgA antibodies into saliva without detection of circulating IgA antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serum calcium and magnesium in different types of leprosy

Serum calcium and magnesium were studied in 200 leprosy patients and 25 apparently healthy individuals. Serum calcium was found to be significantly decreased in all types of leprosy except tuberculoid. The decrease in serum magnesium was highly significant in tuberculoid, lepromatous and borderline lepromatous cases.     

A preliminary study of correlation of immuno-histological and ultrastructural characteristics of neural granuloma in leprosy patients.

With an aim to better understand the pathogenesis of nerve damage in leprosy, peripheral nerve biopsies from six untreated leprosy cases (3 BT/TT and 3 BL/LL) were studied by electronmicroscopy and immuno-histology. In addition to routine histopathology for diagnosis, infiltrating cells of granuloma were characterized after preparation of single cell suspension. The lymphocytes in the lesion were characterized by E and EAC rosetting and macrophage phagocytic system (MPS) cells were studied using histochemical markers like esterase and peroxidase. The results indicate that the lymphocyte content was significantly greater in tuberculoid neural granuloma compared to lepromatous nerves and these formed rosettes with sheep erythrocytes (E) and expressed HLA-DR antigen suggesting that they are activated T cells. Infiltrating macrophages in both the tuberculoid and lepromatous neural granuloma were esterase positive, peroxidase negative and did not form rosettes with sheep erythrocytes or EAC. Ultrathin sections of tuberculoid granuloma showed lymphocytes clearly associated to epithelioid macrophages having well developed Golgi apparatus and rough endoplasmic reticulum. Correlation of these immunological and ultrastructural characters suggests that hypersensitivity mechanisms are possibly responsible for nerve damage in tuberculoid leprosy. Ultrastructural examination of lepromatous nerves, on the other hand, showed the predominance of macrophages with large nucleus, heavily bacillated Schwann cells, and a few lymphocytes. The correlation of immuno-histological and ultrastructural characters indicates that the mechanism(s) of nerve damage in lepromatous leprosy are basically different wherein hypersensitivity appears to play a very limited role.     

Serum immunoglobulins in leprosy.

Serum immunoglobulins were quantitated by radial immunodiffusion in 25 cases each of tuberculoid and lepromatous leprosy. Immunoglobulins estimated from 50 normal healthy adults were the control. Serum IgG was markedly raised in both tuberculoid (mean 2420 mg/dl) and lepromatous leprosy (mean 2493 mg/dl) when compared with the controls (mean 1288 mg/dl) and the difference was significant (p less than 0.01). However the difference in serum IgM and IgA levels in cases as compared to controls were not statistically significant. Serum IgM was slightly raised, the mean values obtained being 222 mg/dl in tuberculoid leprosy, 221 mg/dl in lepromatous leprosy and 202 mg/dl in control. Serum IgA was reduced in lepromatous leprosy (mean 129 mg/dl) as compared to the controls (mean 168 mg/dl) and the cases of tuberculoid leprosy (mean 165 mg/dl). The range of values obtained in both groups of patients showed greater scatter than the controls and a few cases of both forms of leprosy showed very low values of both serum IgA and IgM.     

HLA antigens and neural reversal reactions in Ethiopian borderline tuberculoid leprosy patients

Reversal reactions (RR) or acute neuritis episodes are frequently observed in borderline tuberculoid (BT) leprosy patients during the first year of treatment, and are associated with a rapid increase in cell-mediated immunity. Because HLA-linked genes have been shown to be an important factor in determining the type of leprosy that develops in susceptible individuals and because HLA molecules regulate cellular interactions in the immune system, we have investigated whether RR are associated with HLA antigens in Ethiopian patients. The data reported here indicate that this is not the case: no significant differences in the distribution of HLA class I and class II antigens were observed among three groups: 28 BT patients with a history of RR, 27 BT patients with no history of RR, and 33 healthy individuals. In contrast to these negative results, we observed that HLA-DR3 was associated with high skin-test responsiveness against Mycobacterium leprae antigens among RR patients. Since DR3 was not associated with RR per se, the observed DR3-associated high responsiveness to M. leprae may not be primarily related to the development of RR.     

Blastogenic response with autologous plasma and Mitsuda's reaction in leprosy patients and their unaffected sibs

Many investigators have evaluated the influence of genetic constitution on the susceptibility to leprosy in studies linked to different types of research. To determine the possible existence of a family trait linked to the lymphoproliferation and to lepromin reactivity we studied the blastogenic response to phytohaemagglutinin, lepromin and M. leprae and the Mitsuda's reaction in leprosy patients and their unaffected sibs. Sixty-eight individuals were studied, 34 were leprosy patients (17 lepromatous and 17 tuberculoid leprosy) and the remaining were their sibs previously matched by sex and age. The indices of blastogenesis and lepromin reactivity were lower in lepromatous than in tuberculoid patients, that confirmed the immunological polarity of the two types of leprosy. Both the lymphoproliferation and Mitsuda's reaction results suggest different cell immune responses in leprosy patients and their unaffected sibs, so that the hypothesis of a family trait favouring the similarity of responses to these tests among sibs becomes unlikely.     

Cured tuberculoid patients have a greater life expectancy than cured lepromatous patients in Japan

Leprosy patients lack specific cellular immunity against Mycobacterium leprae, but other immunological functions are thought to be preserved. However, in a leprosy sanatorium in South Japan between 1982 and 2000, we found that the average age at death of cured lepromatous leprosy patients was about 5 yrs younger than that of cured tuberculoid patients; [male/lepromatous, 76.0 +/- 10.0 yrs old vs. male/tuberculoid, 79.7 +/- 9.4 yrs old, p = 0.026], and [female/lepromatous, 78.0 +/- 10.5 vs. female/tuberculoid, 85.3 +/- 9.8, p = 0.0001]. This trend was also observed in autopsy records of two other leprosy sanatoria in Japan. In a prospective study based on their age in 1982, among females in the age group between 60 and 69, lepromatous patients (75.3 +/- 6.0 yrs) died earlier than tuberculoid patients (81.0 +/- 5.1 yrs) (p < 0.01). These findings suggest that lepromatous patients have higher risk of death even in a post-chemotherapy era.     

Biochemical, immunological and genetic studies in leprosy. I. Changes in serum lactate dehydrogenase isoenzymes, creatine phosphokinase and aldolase activity in different forms of leprosy.

Serum lactate dehydrogenase isoenzymes, creatine phosphokinase and aldolase activity were determined in healthy control subjects and in lepromatous and tuberculoid leprosy patients from Ethiopia. Sera from lepromatous patients showed a higher total LDH activity compared with control subject. The values for tuberculoid leprosy patients were similar to those of controls. Sera from normal healthy controls showed a higher proportion of LDH-H form (72%) while lepromatous leprosy patient's sera exhibited a higher proportion of LDH-M form (55%). Tuberculoid leprosy patients showed a pattern similar to that of healthy controls. A possible significance of these observations is discussed. No significant variations were observed in fructose-1,6-diphosphate aldolase activity within the different types of disease and controls. Although creatine phosphokinase levels in different types of leprosy decreased significantly from those of normal healthy, it falls within the reported variation of the activity in normal sera.