Leishmania infantum-specific production of IFN-γ and IL-10 in stimulated blood from dogs with clinical leishmaniosis

Abstract

BackgroundThere is limited information available on cytokine profiles in dogs with different degrees of disease severity due to natural infection of Leishmania infantum. The aim of this study was to investigate L. infantum-specific IFN-γ and IL-10 production in blood from dogs with leishmaniosis at diagnosis and correlate these findings with disease severity, humoral immune response and blood parasitemia.MethodsSixty dogs were diagnosed based on physical examination, routine laboratory tests, L. infantum-specific antibody levels measured by quantitative ELISA and blood parasitemia by real-time PCR. Heparin whole blood was stimulated with L. infantum soluble antigen (LSA) and concanavalin A (ConA) and incubated for 5 days. IFN-γ and IL-10 concentrations were measured in supernatants with sandwich ELISAs.ResultsThe majority of dogs (n = 36) were classified as LeishVet stage II (moderate disease). The rest of the dogs were classified as stage I (n = 10), III (n = 10) and IV (n = 4). Dogs classified with stage I and IIa presented significantly higher (P = 0.02) LSA IFN-γ concentrations, lower (P <0.0001) antibody levels and a tendency for lower blood parasitemia (P = 0.1) than dogs classified with stages IIb, III or IV while no differences in ConA IFN-γ or IL-10 concentrations were observed among groups. Thirty-five dogs produced significantly higher LSA IFN-γ (mean ± SD: 2320 ± 3960 pg/ml) and ConA IFN-γ (mean ± SD: 7887 ± 7273 pg/ml) when compared with 25 dogs that did not produce detectable LSA IFN-γ but produced ConA IFN-γ (mean ± SD: 4917 ± 5233 pg/ml). IFN-γ producer dogs presented lower (mean ± SD: 5750 ± 14,082 ELISA units (EU), P = 0.001) antibody levels and blood parasitemia (mean ± SD: 5 ± 10 parasites/ml, P = 0.001) when compared with IFN-γ non-producers (mean ± SD: 19,638 ± 28,596 EU and 1100 ± 5112 parasites/ml), respectively. LSA IL-10 was not detectable in 34 dogs while 49 dogs secreted ConA IL-10 (mean ± SD of 90 ± 103 pg/ml). LSA IFN-γ concentration was negatively correlated with blood parasitemia and antibody levels and positively correlated with ConA IFN-γ and LSA IL-10 concentrations.ConclusionsThe results of this study demonstrate that sick dogs lacking L. infantum specific IFN-γ production in stimulated whole blood produce a strong humoral response, have a high blood parasitemia and severe clinical disease. IL-10 does not appear to be a marker of disease severity.

The inflammatory cytokine effect of Pam3CSK4 TLR2 agonist alone or in combination with Leishmania infantum antigen on ex-vivo whole blood from sick and resistant dogs

“…Slight modifications were done for the IFN-γ ELISA as described elsewhere [27]. Standard curve for TNF-α started with 1000 pg/ml and two-fold dilutions were made until 7.8 pg/ml concentration.…”

The effects of increased heme oxygenase-1 on the lymphoproliferative response in dogs with visceral leishmaniasis

“…It has been accepted that susceptibility is associated with an exacerbated humoral immune response (Th2) and a reduced cellular immune response (Th1) (Alvar et al, 2004;Baneth et al, 2008). Increased production of IL-10, a major Th2 cytokine, is associated with progression of the canine infection (Boggiatto et al, 2010), while higher expression of IFN-gamma, a Th1 cytokine, is correlated with a lower blood parasite load (Solano-Gallego et al, 2016).…”

Induction of haem oxygenase‐1 increases infection of dog macrophages by L. infantum

“…We attempted to ascertain the involvement of the Th1 cytokine TNF‐alpha and the Th2 cytokine IL‐10 and observed that infection of macrophages from healthy dogs with promastigotes of L. infantum elicited IL‐10 production in the supernatant of 3‐day‐cultured macrophages. Higher IL‐10 correlated with increased parasite load and severity of visceral leishmaniasis in dogs caused by L. infantum , although some studies have suggested that IL‐10 is not associated with disease susceptibility in dogs . However, the association between increased HO‐1 levels and increased IL‐10 levels in infected dogs has been previously addressed, and this could also be a mechanism responsible for the susceptibility of this species.…”

“…Higher IL-10 correlated with increased parasite load and severity of visceral leishmaniasis in dogs caused by L. infantum, 26 although some studies have suggested that IL-10 is not associated with disease susceptibility in dogs. 33 However, the association between increased HO-1 levels and increased IL-10 levels in infected dogs has been previously addressed, 10 and this could also be a mechanism responsible for the susceptibility of this species. Additionally, it is already known that IL-10 induces expression of HO-1 via a p38 mitogenactivated protein kinase-dependent pathway and that inhibition of HO-1 protein synthesis or activity significantly reverses the inhibitory effect of IL-10.…”