[Immunohistochemical analysis of cell surface markers in leprosy patients].

Nine biopsy specimens from 3 tuberculoid (TT), 1 borderline tuberculoid (BT), 2 borderline (BB), 2 borderline lepromatous (BL), and 1 lepromatous (LL) patients were studied using immunoperoxidase procedures with monoclonal antibodies. In TT patients 4B4+, TCR alpha beta+, CD4+ T cells were dominant in dermal granulomas. In LL patient, dermal granulomas with 4B4+, TCR alpha beta+, CD4+ T cells and 2H4+, TCR alpha beta+, CD4+ T cells also had CD8+ T cells. The proportion of 4B4+ T cells increased in granulomas from LL to TT and 2H4+ T cells proportionately increased from TT to LL. TCR gamma delta+ T cells were detected only in BB and BL patients. However, there is no difference of staining pattern of ICAM-1 or LFA-1 antigen in any type of leprosy. These results indicate that the immunohistochemical pattern of T cells (CD4, CD8, 4B4, 2H4, TCR) may be useful in the diagnosis of the spectrum of leprosy.

Selective expansion of V delta 1 + T cells from leprosy skin lesions.

T cells bearing gamma delta T-cell receptors (TCRs) are prominent residents of murine epidermis and appear to be important participants in the immune response to infection in human skin. The Mitsuda reaction in leprosy, induced by intradermal challenge with Mycobacterium leprae, provides an opportunity to study the cellular events that mediate a form of delayed-type hypersensitivity (DTH) in skin. T cells bearing gamma delta TCRs comprise a significant proportion of the T-cell population in these DTH reactions. Presently we have generated T-cell lines from Mitsuda reactions in vitro and compared their TCR repertoire to that found in situ. gamma delta T cells comprised 20-40% of lines derived from these skin lesions, but < 10% of lines derived from the peripheral blood of the same individuals. Flow-cytometric analysis of variable (V) chain usage in T-cell lines derived from skin lesions indicated that V delta 1 was predominant. Evaluation of the TCR repertoire using PCR indicated that V delta 1-J delta 1 and V gamma 2-J gamma P gene rearrangements were prevalent. In comparison, V delta 2-J delta 1 gene rearrangements predominated in situ. Furthermore, nucleotide sequence analysis of the V-J junction of one T-cell line revealed limited genetic diversity of the gamma delta TCR. These findings suggest that the V delta 1 subpopulation of gamma delta T cells in Mitsuda skin reactions selectively outgrows from leprosy skin lesions in vitro. Such V delta 1 + T-cell lines should be useful for determining the relevant antigens and restriction elements in this response to a pathogen in skin.

Suppression of T-cell proliferation by Mycobacterium leprae and its products: the role of lipopolysaccharide.

Addition of soluble molecules obtained from sonicated Mycobacterium leprae markedly suppressed the proliferative response to the mitogen anti-CD3 of peripheral blood mononuclear cells and isolated T cells. Suppression was nonspecific and occurred with cells from lepromatous and tuberculoid leprosy patients as well as control donors. The purified lipoarabinomannans from M. leprae and Mycobacterium tuberculosis had a similar spectrum of inhibition whereas their deacylated derivatives were without effect. All mycobacterial preparations of either a crude or purified state, which suppressed cellular responses, contained appreciable quantities of bacterial lipopolysaccharide by the Limulus amebocyte assay. Contamination with lipopolysaccharide could account for the extent and nonselectivity of the T-cell suppression. Suppression was also monocyte-dependent and in part due to the release of arachidonate metabolites of the cyclooxygenase pathway.

Immunohistological analysis of nerve granulomas in neuritic leprosy.

Immunohistological analysis of infiltrates of nerves in patients with neuritic leprosy was carried out using monoclonal antibodies defining T cell subsets, Langerhans cells, HLA DR antigens, and indirect immunofluorescence. In all, eight nerves were analyzed. 2 of the 8 nerves showed epithelioid cell granulomas surrounded by large numbers of lymphocytes. The predominant lymphocytes in these granulomas were activated T cells expressing CD3 and HLA DR antigens. The proportion of CD3+ and CD4+ cells was higher than that of CD8+ cells. The ratio of CD4+/CD8+ cells in these two biopsy specimens was 5.6 and 1.5, respectively. In these nerves CD4+ cells were diffusely scattered into epithelioid cell granulomas, while CD8+ cells were localized at the periphery of the granuloma. The remaining six nerves showed macrophages containing numerous bacilli, and a few lymphocytes and plasma cells diffusely distributed into the granuloma. In these nerves, only occasional lymphocytes expressing CD3 or CD4 or CD8 and HLA DR antigens were noticed. In two fo the biopsy specimens, a small proportion of CD8+ cells were visualized. Macrophages and Schwann cells were HLA DR+ in all nerves. CD1+ cells were not seen in the infiltrates of any of these nerves. A similar pattern and distribution of cells was noticed in the nerve granulomas of tuberculoid and lepromatous leprosy. These findings suggest that the mechanisms of nerve damage in the patients with neuritic leprosy could be either immunological or non-immunological, depending on the nature and characteristics of the infiltrates.

Defective intralesional interferon-gamma activity in patients with lepromatous leprosy.

Cryostat sections of full-thickness skin biopsies from 21 patients along the whole spectrum of leprosy were subjected to immunohistological examination with special regard to defective lymphokine production. There was an inverse relationship between intra-lesional IL-1 reactivity and IL-2R expression, in that the latter was markedly observed in tuberculoid lesions. Whenever epithelioid cell containing granulomas were present in paucibacillary forms, significant reactivity within the central phagocytic cells with the monoclonal antibody directed against interferon-gamma was detectable. The keratinocytes covering tuberculoid lesions abundantly expressed class II alloantigens (HLA-DR antigens), indicating high intra-lesional interferon-gamma activity. In contrast, multibacillary forms revealed significant anti-IL-1 reactivity within the cellular infiltrate. IL-2R bearing cells were virtually absent as was anti-HLA-DR reactivity of the keratinocytes, underlining a defective intra-lesional interferon-gamma activity.

Quantitative analysis of contrasuppressor T lymphocytes in leprosy; induction of Ia antigens with gamma interferon.

Lepromatous leprosy is characterized by immune anergy and abnormal suppressor T-cell function. Contrasuppressor cells are a subset of CD8+, vicia villosa-adherent T lymphocytes. T-contrasuppressor (Tcs) cells act on T-helper cells to cause them to become unresponsive to the action of T-suppressor cells. In 8 lepromatous (LL) and 7 tuberculoid (TT) patients, and 6 healthy contacts we studied the percent of the following lymphocyte subsets: CD3+, CD4+, CD8+, Ia+, vicia villosa+ (VV+), CD8, VV+, VV, Ia+, and Ia, Tac+. This was done in baseline status as well as post-stimulation with recombinant gamma interferon (rIFN-gamma). We found that peripheral blood mononuclear cells from LL and TT patients and controls exhibit a similar number of putative contrasuppressor lymphocytes (CD8, VV+ cells). However, in the contrasuppressor subset from LL patients we found a low percent of Ia+ (p less than 0.05 compared to controls or TT). In the three groups studied, the rIFN-gamma enhanced the percent of Ia+ lymphocytes in the CD8, VV+ cell subpopulation. However, the CD8, VV+ lymphocytes from LL patients, despite the effect of rIFN-gamma, continue to have a low percent of Ia+ cells (p less than 0.05 compared to controls or TT). These findings suggest that LL patients might have abnormalities in the contrasuppressor immune circuit. Future functional studies on the role of Tcs cells in the anergy seen in LL will be required in order to define the apparent dysfunction occurring in this disease.