Apoptose na infecção experimental com Ehrlichia canis em cães domésticos

Abstract

The Canine MonocyticEhrlichiosis is a zoonotic disease caused by Ehrlichiacanis, a Gram negative bacteria, with a worldwide distribution. Some dogs with ehrlichiosis become asymptomaticcarriers while others develop an acute disease with rapid death. Apoptosis can be important in the elimination of intracellular pathogens in infections caused by Ehrlichia sp. and Anaplasma sp. occur modulation of apoptosis to prolong the survival of these organisms allowing their proliferation and spread, acting as a Trojan horse by inhibition the immune response. In this context apoptosis may contribute to the progression or regression of the lesions. To evaluate the role of apoptosis in Canine MonocyticEhrlichiosis, seven mongrel dogs, clinically healthy and free of ticks, aged among six and eight months, were randomly assigned into two groups: GI, Inoculated - with four dogs and GII, Control -with three dogs. Each dog was inoculated intravenously with 3 ml of either infected blood (strain Jaboticabal) in GI or sterile PBS in GII. Weekly and 35 days post-inoculation, blood samples were collected and subjected to nested reaction chain polymerase (nPCR) and immunofluorescence assay (IFA) for confirmation of infection. On day 36 post-inoculation, animals were euthanized and samples of spleen, lymph node, kidney and liver were collected and processed for TUNEL and immunohistochemical techniques (Bcl-2, Bax). Another set of the same samples, with bone marrow and heart were processed for gene expression of apoptotic molecules (Bax, Bcl-2, caspase-3, caspase 8 and caspase 9) by RTPCR. As verified by nPCR inoculated animals became positive for E. canis from the second week after inoculation. IFAT showed seropositive animals from the third week after inoculation. In the control group, all tests were negative for E. canis. Both pro and anti-apoptotic molecules showed higher expression of genes in tissues of inoculated animals than in the control group. TUNEL assay showed increased incidence of apoptosis in the group inoculated, with more intense labeling in spleen and lymph nodes, however the results of the immunohistochemistry for Bcl-2 and Bax indicated that the intrinsic pathway of apoptosis was not important. In conclusion we found that in the acute phase of the experimental disease,Ehrlichiacanis modulates apoptosis in different ways in different tissues and that apoptosis plays a role in disease progression.