Mice with Genetic Deletion of Group VIA Phospholipase A2β Exhibit Impaired Macrophage Function and Increased Parasite Load in Trypanosoma cruzi-Induced Myocarditis

Abstract

Trypanosoma cruzi infection, which is the etiological agent of Chagas disease, is associated with intense inflammation during the acute and chronic phases. The pathological progression of Chagas disease is influenced by the infiltration and transmigration of inflammatory cells across the endothelium to infected tissues, which are carefully regulated processes involving several molecular mediators, including adhesion molecules and platelet-activating factor (PAF). We have shown that PAF production is dependent upon calcium-independent group VIA phospholipase A2β (iPLA2β) following infection of human coronary artery endothelial cells (HCAECs) with T. cruzi, suggesting that the absence of iPLA2β may decrease the recruitment of inflammatory cells to the heart to manage parasite accumulation. Cardiac endothelial cells isolated from iPLA2β-knockout (iPLA2β-KO) mice infected withT. cruzi demonstrated decreased PAF production compared to that by cells isolated from wild-type (WT) mice but demonstrated increases in adhesion molecule expression similar to those seen in WT mice. Myocardial inflammation in iPLA2β-KO mice infected with T. cruzi was similar in severity to that in WT mice, but the iPLA2β-KO mouse myocardium contained more parasite pseudocysts. Upon activation, macrophages from iPLA2β-KO mice produced significantly less nitric oxide (NO) and caused lessT. cruzi inhibition than macrophages from wild-type mice. Thus, the absence of iPLA2β activity does not influence myocardial inflammation, but iPLA2β is essential forT. cruzi clearance.

FIG 1

Expression of ICAM-1 and VCAM-1 and PAF production in endothelial cells isolated from the hearts of WT and iPLA₂β-KO mice infected with T. cruzi (MOI, 0.2) for up to 48 h. Values are normalized to those observed in the absence of infection. The values shown are means + SEMs for 4 separate cell cultures. *, P < 0.05 compared to untreated controls; **, P < 0.01 compared to untreated controls.

FIG 2

(A) Myocardial inflammation score for WT and iPLA₂β-KO mice following T. cruzi infection for up to 12 weeks. (B) Number of parasites in the myocardium per field for WT and iPLA₂β-KO mice following T. cruzi infection for 4 weeks. **, P < 0.01 compared to WT. The results in panels A and B are means + SEMs for at least 6 separate samples. (C) Hematoxylin-and-eosin-stained sections of hearts from WT and iPLA₂β-KO mice following 4 weeks of T. cruzi infection. Arrows, parasite pseudocysts in the myocardium.

FIG 3

NO release from cells of the murine macrophage cell line RAW 264.7 following infection with T. cruzi (MOI, 0.2) with or without IFN-γ (100 units) for 24 h. **, P < 0.01 compared to untreated cells; +, P < 0.05 compared to treated or infected cells in the presence or absence of BEL (0.5 μM); ++, P < 0.01 compared to treated or infected cells in the presence or absence of BEL (0.5 μM). The values shown are means + SEMs for 4 separate cell cultures.

FIG 4

NO release from murine BMDMs and inhibition of T. cruzi intracellular parasites. (A) NO release from WT mouse BMDMs incubated with T. cruzi (MOI, 0.2) with or without IFN-γ (100 units) for 24 h. **, P < 0.01 compared to untreated cells; +, P < 0.05 compared to treated or infected cells in the presence or absence of BEL (10 μM); ++, P < 0.01 compared to treated or infected cells in the presence or absence of BEL (10 μM). (B) BMDMs isolated from WT or iPLA₂β-KO mice were treated with T. cruzi (MOI, 0.2) or IFN-γ (100 units) for 24 h. **, P < 0.01 compared to untreated cells; ++, P < 0.01 compared to NO release between WT and iPLA₂β-KO mice. (C) T. cruzi inhibition in BMDMs isolated from WT or iPLA₂β-KO mice infected with T. cruzi (MOI, 5). **, P < 0.01 compared to data between WT and iPLA₂β-KO mice. The values shown are means + SEMs for at least 4 separate cell cultures.