Identification of an immunostimulating protein from Mycobacterium leprae.

Despite the recent identification of a number of Mycobacterium leprae proteins, the major immunogenic determinants of this organism remain obscure. We isolated from M. leprae a potent immunostimulatory preparation, designated the MLP fraction, which contains a major protein of 35 kilodaltons (kDa). This protein was precipitated by monoclonal antibody ML03-A1, which recognizes a 35-kDa protein of M. leprae, and by sera obtained from patients with lepromatous leprosy. Neither sera from healthy controls nor sera from patients with pulmonary tuberculosis recognized the 35-kDa protein, and only one of four serum samples from patients with borderline tuberculoid leprosy reacted with this protein. The MLP fraction stimulated T-cell proliferation in patients with leprosy whose T cells proliferate in response to whole M. leprae cells. Apparently, the T-cell epitope associated with MLP is also expressed on M. tuberculosis and M. bovis BCG, since patients with pulmonary tuberculosis and BCG-vaccinated individuals demonstrated significant responses to the MLP fraction. The 35-kDa M. leprae protein, purified to homogeneity in the laboratory of P. J. Brennan, stimulated T-cell proliferative responses in all MLP-responsive subjects. These findings suggest that the 35-kDa protein present in MLP is an immunostimulatory component of M. leprae. In addition to serving as a useful probe for study of the T-cell anergy associated with lepromatous disease, this protein may ultimately be useful as a component of a vaccine designed to provide protection against infection with M. leprae.

Effective vaccination of mice against leprosy bacilli with subunits of Mycobacterium leprae.

Model vaccines against leprosy bacilli have consisted of nonvirulent, live, attenuated Mycobacterium bovis BCG and irradiated, heat-killed, or autoclaved intact M. leprae. We report that immunization with various cell wall fractions of M. leprae, progressively depleted of lipids, carbohydrates, and soluble proteins, as well as a partially purified protein(s) derived from a pellet fraction of sonicated M. leprae, conferred significant protection against subsequent infection with live leprosy bacilli. Moreover, lymphocytes from regional lymph nodes and spleens of mice immunized with these M. leprae-derived subunits responded by proliferation when stimulated with M. leprae in vitro. Our results provide the first evidence that vaccination with M. leprae-derived fractions protects mice against leprosy bacilli.

T cell defect in lepromatous leprosy is reversible in vitro in the absence of exogenous growth factors.

T lymphocytes from patients with lepromatous leprosy (LL) characteristically fail to respond to Mycobacterium leprae. This specific immunologic defect is thought to contribute to the aggressive clinical course that typifies patients with LL. We report that although fresh CD4+ (helper) T cells from most LL patients are specifically unresponsive to M. leprae, after culture in medium alone for 48 hr the same cells respond to M. leprae antigens. The recovery of T cell function is specific for M. leprae, occurs at the level of responder CD4+ T cells, and is not affected by monocytes or CD8+ (suppressor) T cells. Recovery of T cell reactivity is blocked by the presence of M. leprae bacilli in the preculture medium. These findings indicate that despite the apparent specific anergy seen in patients with LL, the T cells of most LL patients can respond to M. leprae. Their failure to do so, in vivo, may be due to the persistence of antigen, which renders antigen-reactive T cells nonresponsive either directly or via activation of CD4+ suppressor cells.

Defective cell-mediated immunity in leprosy: failure of T cells from lepromatous leprosy patients to respond to Mycobacterium leprae is associated with defective expression of interleukin 2 receptors and is not reconstituted by interleukin 2.

Patients with lepromatous leprosy (LL) but not borderline tuberculoid leprosy (BT) have defective cell-mediated immune responses to Mycobacterium leprae, despite normal responses to other stimuli, as judged by in vivo skin testing and in vitro lymphocyte transformation. To investigate the basis of the immune defect in LL patients, we studied the ability of patient mononuclear leukocytes to produce interleukin 1 (IL 1) and interleukin 2 (IL 2) upon stimulation with M. leprae, and determined the ability of exogenous IL 1 and IL 2 to reconstitute the LL patient response to this antigen in vitro. Equal numbers of adherent non-T cells from LL and BT patients produced similar amounts of IL 1 upon challenge with M. leprae, and addition of IL 1 to the culture medium failed to reconstitute the response of lymphocytes from LL patients to M. leprae. On the other hand, T cells of LL patients failed to express receptors for IL 2 or to produce IL 2 in response to M. leprae, whereas similarly treated T cells of BT patients both expressed IL 2 receptors and produced IL 2. Finally, recombinant human IL 2 purified to homogeneity as well as crude supernatants of mitogen-activated lymphocytes failed to reconstitute the response of LL patients to M. leprae. These results suggest that T cells of LL patients fail to respond to M. leprae despite an ability to produce IL 1 and that their failure to express receptors for IL 2 may explain both defective proliferation and the failure of exogenous IL 2 to reconstitute the response.