Apoptosis and frequency of total and effector CD8+ T lymphocytes from cutaneous leishmaniasis patients during antimonial therapy.

BACKGROUND: Leishmaniasis is an important parasitic disease affecting millions worldwide. Human cutaneous leishmaniasis (CL) is endemic in Rio de Janeiro, Brazil, where is caused by Leishmania braziliensis. The adaptive immune response is accountable for the healing of CL and despite of key role of CD8+ T cells in this immune response little is known about the CD8+ T lymphocytes frequencies, apoptosis and antigen-responsive CD8+ T lymphocytes of CL patients during antimonial therapy. METHODS: Using flow cytometry, we examined total and effector CD8+ T cells from CL patients before (PBT), during (PDT) and after (PAT) treatment for apoptosis and frequencies upon isolation and after in vitro L. braziliensis antigens (LbAg)-stimulation culture. Besides, a correlation study between immunological findings and lesion size was done. RESULTS: PDT showed lower frequencies of total CD8+ T lymphocytes and higher levels of apoptosis of these cells, which were also observed following LbAg-stimulation culture. Regarding effector CD8+ T cells, high frequencies were observed in PDT, while lower frequencies were observed in PAT. Interestingly, PDT showed higher frequencies of apoptotic-effector CD8+ T lymphocytes. Similar results were seen after in vitro antigenic-stimulation assays. Correlation analysis showed that the greater the size of lesion, the smaller the frequency of effector CD8+ T lymphocytes in PDT and PAT, as well as a positive correlation between apoptotic-effector CD8+ T cells frequency and lesion size of PDT. CONCLUSIONS: Changes in effector CD8+ T-lymphocyte frequencies, during and after treatment, seem to represent a critical stage to generate an efficient immune response and suggest that these cells would be evolved in the triggering or in the resolution of lesion, under the influence of therapy. This hypothesis opens new perspectives to clarify controversial statements about the protective or deleterious role of CD8+ T cells in the cure or aggravation of CL and the new approach of evaluating patients during treatment proved to be of utmost importance for understanding the immune response in the healing process of human CL.

Kinetoplastid membrane protein-11 is present in promastigotes and amastigotes of Leishmania amazonensis and its surface expression increases during metacyclogenesis

Kinetoplastid membrane protein-11 (KMP-11), a protein present in all kinetoplastid protozoa, is considered a potential candidate for a leishmaniasis vaccine. A suitable leishmaniasis vaccine candidate molecule must be expressed in amastigotes, the infective stage for mammals. However, the expression of KMP-11 in Leishmania amastigotes has been a subject of controversy. We evaluated the expression of this molecule in logarithmic and stationary growth phase promastigotes, as well as in amastigotes, of Leishmania amazonensis by immunoblotting, flow cytometry and immunocytochemistry, using a monoclonal antibody against KMP-11. We found that KMP-11 is present in promastigotes and amastigotes. In both stages, the protein was found in association with membrane structures (at the cell surface, flagellar pocket and intracellular vesicles). More importantly, its surface expression is higher in amastigotes than in promastigotes and increases during metacyclogenesis. The increased expression of KMP-11 in metacyclic promastigotes, and especially in amastigotes, indicates a role for this molecule in the parasite relationship with the mammalian host. The presence of this molecule in amastigotes is consistent with the previously demonstrated immunoprotective capacity of vaccine prototypes based on the KMP-11-coding gene and the presence of humoral and cellular immune responses to KMP-11 in Leishmania-infected humans and animals.