DH82 Canine and RAW264.7 Murine Macrophage Cell Lines Display Distinct Activation Profiles Upon Interaction With Leishmania infantum and Leishmania amazonensis.

Leishmaniasis is an anthropozoonotic disease, and dogs are considered the main urban reservoir of the parasite. Macrophages, the target cells of Leishmania sp., play an important role during infection. Although dogs have a major importance in the epidemiology of the disease, the majority of the current knowledge about Leishmania-macrophage interaction comes from murine experimental models. To assess whether the canine macrophage strain DH82 is an accurate model for the study of Leishmania interaction, we compared its infection by two species of Leishmania (Leishmania infantum and L. amazonensis) with the murine macrophage cell line (RAW264.7). Our results demonstrated that L. amazonensis survival was around 40% at 24 h of infection inside both macrophage cell lines; however, a reduction of 4.3 times in L. amazonensis infection at 48 h post-infection in RAW 264.7 macrophages was observed. The survival index of L. infantum in DH82 canine macrophages was around 3 times higher than that in RAW264.7 murine cells at 24 and 48 h post-infection; however, at 48 h a reduction in both macrophages was observed. Surprisingly at 24 h post-infection, NO and ROS production by DH82 canine cells stimulated with LPS or menadione or during Leishmania infection was minor compared to murine RAW264.7. However, basal arginase activity was higher in DH82 cells when compared to murine RAW264.7 cells. Analysis of the cytokines showed that these macrophages present a different response profile. L. infantum induced IL-12, and L. amazonensis induced IL-10 in both cell lines. However, L. infantum and L. amazonensis also induced TGF-ß in RAW 264.7. CD86 and MHC expression showed that L. amazonensis modulated them in both cell lines. Conversely, the parasite load profile did not show significant difference between both macrophage cell lines after 48 h of infection, which suggests that other mechanisms of Leishmania control could be involved in DH82 cells.

Immunomodulatory Role of Capsular Polysaccharides Constituents of Cryptococcus neoformans.

Cryptococcosis is a systemic fungal infection caused by Cryptococcus neoformans. In immunocompetent patients, cryptococcal infection is often confined to the lungs. In immunocompromised individuals, C. neoformans may cause life-threatening illness, either from novel exposure or through reactivation of a previously acquired latent infection. For example, cryptococcal meningitis is a severe clinical disease that can manifest in people that are immunocompromised due to AIDS. The major constituents of the Cryptococcus polysaccharide capsule, glucuronoxylomannan (GXM), and galactoxylomannan (GalXM), also known as glucuronoxylomanogalactan (GXMGal), are considered the primary virulence factors of Cryptococcus. Despite the predominance of GXM in the polysaccharide capsule, GalXM has more robust immunomodulatory effects on host cellular immunity. This review summarizes current knowledge regarding host-Crytococcus neoformans interactions and the role of capsular polysaccharides in host immunomodulation. Future studies will likely facilitate a better understanding of the mechanisms involved in antigenic recognition and host immune response to C. neoformans and lead to the development of new therapeutic pathways for cryptococcal infection.

B-1 Cells May Drive Macrophages Susceptibility to Trypanosoma cruzi Infection.

B-1 cells can directly and indirectly influence the immune response. These cells are known to be excellent producers of natural antibodies and can secrete a variety of immunomodulatory molecules. They are also able to differentiate into B-1 cell-derived phagocytes (B-1CDP). B-1 cells can modulate macrophages to become less effective, and B-1CDP cells are more susceptible in infection models. In this work, we investigated the microbicidal ability of these cells in Trypanosoma cruzi infection in vitro. The results show that macrophages from BALB/c mice are more susceptible to infection than macrophages from XID mice. The resistance observed in macrophages from XID mice was abolished in the presence of B-1 cells, and this event seems to be associated with IL-10 production by B-1 cells, which may have contributed to the decrease of NO production. Additionally, B-1CDP cells were more permissive to intracellular T. cruzi infection than peritoneal macrophages. These findings strongly suggest that B-1 cells and B-1CDP cells have a potential role in the persistence of the parasite in host cells.

Involvement of the capsular GalXM-induced IL-17 cytokine in the control of Cryptococcus neoformans infection.

Cryptococcus neoformans is an opportunistic fungus that can cause lethal brain infections in immunosuppressed individuals. Infection usually occurs via the inhalation of a spore or desiccated yeast which can then disseminate from the lung to the brain and other tissues. Dissemination and disease is largely influence by the production of copious amounts of cryptococcal polysaccharides, both which are secreted to the extracellular environment or assembled into a thick capsule surrounding the cell body. There are two important polysaccharides: glucuronoxylomannan (GXM) and galactoxylomannan, also called as glucuronoxylomanogalactan (GXMGal or GalXM). Although GXM is more abundant, GalXM has a more potent modulatory effect. In the present study, we show that GalXM is a potent activator of murine dendritic cells, and when co-cultured with T cells, induces a Th17 cytokine response. We also demonstrated that treating mice with GalXM prior to infection with C. neoformans protects from infection, and this phenomenon is dependent on IL-6 and IL-17. These findings help us understand the immune biology of capsular polysaccharides in fungal pathogenesis.

Canine Macrophage DH82 Cell Line As a Model to Study Susceptibility to Trypanosoma cruzi Infection.

Trypanosoma cruzi is an obligatory intracellular protozoan parasite, and it is the etiological agent of Chagas' disease that is endemic in the Americas. In addition to humans, a wide spectrum of mammals can be infected by T. cruzi, including dogs. Dogs develop acute and chronic disease, similar to human infection. T. cruzi can infect almost all cell types and after cell invasion, the metacyclics trypomastigotes localize in the cytoplasm, where they transform into amastigotes, the replicative form of T. cruzi in mammals. After amastigote multiplication and differentiation, parasites lyse host cells and spread through the body by blood circulation. In this work, we evaluated the in vitro ability of T. cruzi to infect a canine macrophage cell line DH82 compared with RAW264.7, a murine tissue culture macrophage. Our results have shown that the T. cruzi is able to infect, replicate and differentiate in DH82 cell line. We observed that following treatment with LPS and IFN-γ DH82 cells were more resistant to infection and that resistance was not related reactive oxygen species production in our system. In this study, we also found that DH82 cells became more susceptible to T. cruzi infection when cocultured with apoptotic cells. The analysis of cytokine production has showed elevated levels of the TGF-ß, IL-10, and TNF-α produced by T. cruzi-infected canine macrophages. Additionally, we demonstrated a reduced expression of the MHC class II and CD80 by infected DH82 cell line.

Neutrophils increase or reduce parasite burden in Trypanosoma cruzi-infected macrophages, depending on host strain: role of neutrophil elastase.

Neutrophils are involved in the initial steps of most responses to pathogens and are essential components of the innate immune response. Due to the ability to produce and release various soluble mediators, neutrophils may participate in the regulation of the inflammatory response. Little is known about the role of neutrophils during protozoan infections including infection by Trypanosoma cruzi. In the present study we investigated the importance of inflammatory neutrophils on macrophage activation and T. cruzi replication in vitro, in cells obtained from BALB/c mice and C57Bl/6 mice. Co-cultures of BALB/c apoptotic or live neutrophils with infected peritoneal macrophages resulted in increased replication of the parasites and in the production of TGF-ß and PGE2. The treatment with anti-TGF-ß neutralizing antibody and COX inhibitor blocked the parasite replication in vitro. On the other hand, co-cultures of T. cruzi infected macrophages with live neutrophils isolated from C57BL/6 mice resulted in decreased number of trypomastigotes in culture and increased production of TNF-α and NO. The addition of anti-TNF-α neutralizing antibody or elastase inhibitor resulted in the abolishment of macrophage microbicidal effect and increased parasite replication. Addition of elastase to infected macrophages reduced the replication of the parasites, and on the other hand, addition of a selective inhibitor of iNOS increased parasite growth, suggesting the role of NO in this system. Our findings reveal that neutrophils may regulate T. cruzi experimental infection and determine susceptibility and resistance to infection.