Jacinto Convit

Determination of the cytokine profile in American cutaneous leishmaniasis using the polymerase chain reaction.

The lymphokine profiles were determined in the skin lesions of the three distinct clinical forms of American cutaneous leishmaniasis (ACL), using a reverse transcriptase polymerase chain reaction (RT‐PCR) and primers for various lymphokines. The message for interferon‐gamma (IFN‐y). tumour necrosis factor‐beta (TNF–β) and IL‐8 was expressed in the three clinical forms of ACL. IL‐lβ inRNA was expressed in most localized (LCL) and mucoeutaneous (MCL) leishmaniasis. but in only few of the diffuse cutaneous leishmaniasis (DCL). IL‐2 mRNA was detected in about half of the lesions, with more prominent values for MCL. IL‐4 mRNA was present in most lesions from the three clinical forms, but markedly increased in DCL. IL‐5 and IL‐10 mRNAs were expressed in all MCL and in half of the DCL lesions and weakly expressed in LCL lesions. IL‐10 mRNA was more abundant in MCL lesions. In contrast. IL‐6 and TNF‐s mRNAs were expressed in a Iarge number of LCL. In MCL, IL‐6 mRNA was expressed in most cases and TNF–α mRNA in all the cases. In DCL. 11,–6 mRNA was absent and TNF–α mRNA was weakly expressed. These results suggest that most T cells present in the MCL and DCL lesions secrete a mixture of type 1 and type 2 cytokine patterns, but in DCL granulomas type 2 cytokines predominate. In LCL the cytokine patterns show a mixture of type l and type o with a preponderance of IFN–γ over IL‐4, and low levels of IL‐5 and IL‐10. The Iack of IL‐6 and TNF–α mRNAs, and the low expression of IL‐lβ in DCL lesions suggest a defect in the anligcn‐processing cells that may account for the state of unrcsponsiveness in these patients.

Diffuse cutaneous leishmaniasis: A disease due to an immunological defect of the host

Abstract Diffuse cutaneous leishmaniasis, with its characteristic diffusion of lesions, great abundance of parasites, anergy to skin tests with the specific antigen and resistance to treatment, has been described as a disease produced by a special strain of Leishmania, L. pifanoi. Our concept is that this form of leishmaniasis is due, not to a different type of parasite, but to an immunological defect of the human host, which makes him respond with these special clinical and parasitological manifestations. The basis for our belief is: (1) epidemiologically, the disease appears as isolated cases in endemic areas; (2) accidental inoculation of a laboratory technician with a strain taken from an animal with diffuse cutaneous leishmaniasis lesions produced a nodule with the clinical, pathological and parasitological characteristics of an American cutaneous lesion, with a strongly positive leishmanin reaction; (3) the coexistence, in a leishmaniasis focus, of two patients living in the same house, one of whom had diffuse cutaneous leishmaniasis and the other American cutaneous leishmaniasis; (4) clinical and pathological characteristics of an experimental inoculation in human volunteers with material obtained from diffuse cutaneous leishmaniasis lesions. This produced, in all the hosts, a typical American cutaneous leishmaniasis type of response.

The clinical and immunological spectrum of American cutaneous leishmaniasis

American cutaneous leishmaniasis is characterized by a spectrum of clinical manifestations. These include localized, often self-healing single lesions, intermediate forms which frequently produce mucosal lesions and often show exaggerated delayed-type hypersensitivity (DTH), and the rare diffuse cutaneous leishmaniasis in which no reaction of protective cell-mediated immunity or DTH can be demonstrated. Clinical, pathological and immunological studies have begun to unravel some of the mechanisms associated with different disease manifestations, dependent on complex interactions between the host immune response, measured in terms of indices including lymphocyte subsets and lymphokines in vitro and within active lesions, and different species of Leishmania.

IMMUNOTHERAPY VERSUS CHEMOTHERAPY IN LOCALISED CUTANEOUS LEISHMANIASIS

In a randomised trial a combination vaccine consisting of live BCG together with killed leishmania promastigotes was compared with a standard antimonial regimen in 94 patients with localised cutaneous leishmaniasis. Three vaccinations over 32 weeks gave a similar cure rate (94%) to three 20-day courses of meglumine antimonate. In the immunotherapy group side-effects were few (5.8%) and slight whereas in the chemotherapy group they were frequent (52.4%) and often serious. Immunotherapy is a low-cost, low-risk alternative to chemotherapy in localised cutaneous leishmaniasis, applicable by primary health services in rural areas.

Diffuse cutaneous leishmaniasis responds to miltefosine but then relapses

Summary  Background  Diffuse cutaneous leishmaniasis (DCL), although rare, is profoundly incapacitating. At present there is no successful treatment for this progressive protozoan infection, which is associated with the absence of specific cell‐mediated immunity (CMI) to Leishmania. This disease shares features with visceral leishmaniasis (VL), including specific CMI inactivity during active disease and a heavy parasitic burden, but VL responds well to treatment. Miltefosine is the first orally administered drug which has shown efficacy in the treatment of VL; it has not been adequately evaluated in the treatment of DCL.

Immunoprophylactic trial with combined Mycobacterium Leprae/BCG vaccine against leprosy: preliminary results

In an attempt to find a vaccine that gives greater and more consistent protection against leprosy than BCG vaccine, we compared BCG with and without killed Mycobacterium leprae in Venezuela. Close contacts of prevalent leprosy cases were selected as the trial population since they are at greatest risk of leprosy. Since 1983, 29,113 contacts have been randomly allocated vaccination with BCG alone or BCG plus 6 x 10(8) irradiated, autoclaved M leprae purified from the tissues of infected armadillos. We excluded contacts with signs of leprosy at screening and a proportion of those whose skin-test responses to M leprae soluble antigen (MLSA) were 10 mm or more (positive reactions). By July, 1991, 59 postvaccination cases of leprosy had been confirmed in 150,026 person-years of follow-up through annual clinical examinations of the trial population (31 BCG, 28 BCG/M leprae). In the subgroup for which we thought an effect of vaccination was most likely (onset more than a year after vaccination, negative MLSA skin-test response before vaccination), leprosy developed in 11 BCG recipients and 9 BCG/M leprae recipients; there were 18% fewer cases (upper 95% confidence limit [CL] 70%) in the BCG/M leprae than in the BCG alone group. For all cases with onset more than a year after vaccination irrespective of MLSA reaction the relative efficacy was 0% (upper 95% CL 54%; 15 cases in each vaccine group). Retrospective analysis of data on the number of BCG scars found on each contact screened suggested that BCG alone confers substantial protection against leprosy (vaccine efficacy 56%, 95% CL 27-74%) and there was a suggestion that several doses of BCG offered additional protection. There is no evidence in the first 5 years of follow-up of this trial that BCG plus M leprae offers substantially better protection against leprosy than does BCG alone, but the confidence interval on the relative efficacy estimate is wide.

Immunotherapy of American cutaneous leishmaniasis in Venezuela during the period 1990–1999

Of a total of 11532 Venezuelan patients with American cutaneous leishmaniasis (ACL) receiving immunotherapy with a combined vaccine containing heat-killed Leishmania promastigotes and bacille Calmette-Guerin (BCG) during the period 1990-99, we evaluated 5341 from 4 widely separated geographical states. Clinical healing varied from 91.2 to 98.7%, with an average of 95.7%. Adverse reactions were mild and limited to those associated with BCG vaccination alone. Immunotherapy failures in 143 patients included 54.5% with typical localized ulcers and 45.5% with non-mucosal intermediate cutaneous leishmaniasis (ICL). Less than 2% of the patients in this study had lesions suggestive of ICL. The disproportionately large number of immunotherapy failures in the ICL group suggests that it should not be used as monotherapy in this group. Weaker reactivity to purified protein derivative in immunotherapy failures, while not statistically significant in the small group reported here, suggests the possibility that these patients develop a relatively torpid immune response. The high percentage of clinical cures achieved with immunotherapy, associated with few secondary effects and low cost, support the use of immunotherapy in the routine treatment of localized ACL.

Studies on human polymorphonuclear leukocyte enzymes. I. Assay of acid hydrolases and other enzymes.

Abstract Eight acid hydrolases, peroxidase (EC 1.11.1.7), cyanide-insensitive NADH oxidase and alkaline phosphatase (EC 3.1.3.1.) were demonstrated in homogenates of human polymorphonuclear leukocytes. The main kinetic characters of these enzymes have been studied and methods for their quantitative assay have been developed. The activities present in human leukocytes are given and compared with those found in rabbit heterophil leukocytes. Homogenates were separated by differential centrifugation into two particulate fractions and a soluble fraction. The influence of the homogenization medium on the distribution of protein, DNA, cyanide-insensitive NADII oxidase and acid β-glycerophosphatase in these fractions was studied.

Therapy of Venezuelan patients with severe mucocutaneous or early lesions of diffuse cutaneous leishmaniasis with a vaccine containing pasteurized Leishmania promastigotes and bacillus Calmette-Guerin: preliminary report

Severe mucocutaneous (MCL) and diffuse (DCL) forms of American cutaneous leishmaniasis (ACL) are infrequent in Venezuela. Chemotherapy produces only transitory remission in DCL, and occasional treatment failures are observed in MCL. We have evaluated therapy with an experimental vaccine in patients with severe leishmaniasis. Four patients with MCL and 3 with early DCL were treated with monthly intradermal injections of a vaccine containing promastigotes of Leishmania (Viannia) braziliensis killed by pasteurization and viable Bacillus Calmette- Guerin. Clinical and immunological responses were evaluated. Integrity of protein constituents in extracts of pasteurized promastigotes was evaluated by gel electrophoresis. Complete remission of lesions occurred after 5-9 injections in patients with MCL or 7-10 injections in patients with early DCL. DCL patients developed positive skin reactions, average size 18.7 mm. All have been free of active lesions for at least 10 months. Adverse effects of the vaccine were limited to local reactivity to BCG at the injection sites and fever in 2 patients. Extracts of pasteurized and fresh promastigotes did not reveal differences in the integrity of protein components detectable by gel electrophoresis. Immunotherapy with this modified vaccine offers an effective, safe option for the treatment of patients who do not respond to immunotherapy with vaccine containing autoclaved parasites or to chemotherapy.

Immune response in healthy volunteers vaccinated with killed leishmanial promastigotes plus BCG. I: Skin-test reactivity, T-cell proliferation and interferon-γ production

This study reports the results of a vaccine trial established to study the cellular immune responses in vivo (skin-test reactivity) and in vitro (T-cell proliferation and interferon-gamma production) to both leishmanial and mycobacterial antigens following vaccination of healthy volunteers from a leishmaniasis-endemic area with killed leishmanial promastigotes, with or without BCG (Bacille Calmètte-Guerin). Skin tests were performed using purified protein derivative of tuberculin (PPD) and leishmanial antigen in 692 volunteers, and 208 doubly negative subjects (< or = 7 mm induration) were selected to participate in the trial. The study subjects were divided into four vaccine groups: (A) killed promastigotes plus BCG, (B) BCG alone, (C) killed promastigotes alone, and (D) placebo. Three vaccine doses were administered at 6-10-week intervals. The skin-test responses to PPD and leishmanial antigen were reassessed at 4-6- and 12-18-month follow-ups. The results of this trial demonstrated that the combined vaccine, i.e. killed promastigotes of Leishmania plus BCG, results in the stimulation of an immune response to both leishmania and mycobacterial antigens in a high percentage of vaccines (> 85%), manifested either by skin-test conversion, lymphocyte proliferation and/or interferon-gamma production. This was evident after the first dose of vaccine for lymphocyte proliferation and interferon-gamma production and was maintained for a year after the three doses of vaccine. Group B (which received BCG alone), responded as well as group A to PPD but not as well to leishmanial antigen. The reverse was true for group C which received promastigotes alone. Group A attained a 38% leishmanin skin-test conversion at the 4-6-month follow-up, which was associated with double PPD/leishmanial antigen responder status. In contrast, a 35% skin-test conversion was found at the 12-18-month follow-up in group C (promastigotes alone), but this was not associated with responses to PPD. A significant percentage of conversion was observed in the placebo group at the 12-18-month follow-up, both to PPD (58%) and leishmanial (21%) antigens, which suggests either environmental exposure to mycobacterial or leishmanial antigens during the vaccine trial or, more probably, a response to the repeated leishmanial skin tests. Further studies are required to determine whether the presence of proliferative and/or interferon-gamma responses in the absence of a skin-test response are sufficient indicators of potential vaccine success.