investigating-the-phagocytosis-of-leishmania-using-confocal-microscopy

Investigating the Phagocytosis of Leishmania using Confocal Microscopy

Phagocytosis is an orchestrated process that involves distinct steps: recognition, binding, and internalization. Professional phagocytes take up ...

in-vitro-differentiation-human-dendritic-cells-their-markers

In Vitro Differentiation of Human Dendritic Cells and their Markers in Leishmania Infection

Leishmaniasis comprises a collection of clinical manifestations associated with the infection of obligate intracellular protozoans, Leishmania. The life ...

Gonçalo Moniz Institute

CBA mouse macrophages control  infection yet are permissive to . Few studies have been conducted to assess the role played by autophagy in  infection. Therefore, we assessed whether the autophagic response of infected macrophages may account for the differential behavior of these two parasite strains. After 24 h of infection, the LC3-II/Act ratio increased in both - and -infected macrophages compared to uninfected controls, but less than in chloroquine-treated cells. This suggests that  and  activate autophagy in infected macrophages, without altering the autophagic flux. Furthermore, infected cells exhibited higher percentages of DQ-BSA-labeled parasitophorous vacuoles (50%) than those infected by  (25%). However, - and -induced parasitophorous vacuoles accumulated LysoTracker similarly, indicating that the acidity in both compartment was equivalent. At as early as 30 min, endogenous LC3 was recruited to both - and -induced parasitophorous vacuoles, while after 24 h a greater percentage of LC3 positive vacuoles was observed in -infected cells (42.36%) compared to those infected by  (18.10%). Noteworthy, principal component analysis (PCA) and an hierarchical cluster analysis completely discriminated -infected macrophages from -infected cells accordingly to infection intensity and autophagic features of parasite-induced vacuoles. Then, we evaluated whether the modulation of autophagy exerted an influence on parasite infection in macrophages. No significant changes were observed in both infection rate or parasite load in macrophages treated with the autophagic inhibitors wortmannin, chloroquine or VPS34-IN1, as well as with the autophagic inducers rapamycin or physiological starvation, in comparison to untreated control cells. Interestingly, both autophagic inducers enhanced intracellular  and  viability, while the pharmacological inhibition of autophagy exerted no effects on intracellular parasite viability. We also demonstrated that autophagy induction reduced NO production by - and -infected macrophages but not alters arginase activity. These findings provide evidence that although -induced parasitophorous vacuoles recruit LC3 more markedly,  and  similarly activate the autophagic pathway in CBA macrophages. Interestingly, the exogenous induction of autophagy favors  intracellular viability to a greater extent than  related to a reduction in the levels of NO.

Autophagic Induction Greatly Enhances  Intracellular Survival Compared to  in CBA/j-Infected Macrophages.

Sand flies bite mammalian hosts to obtain a blood meal, driving changes in the host inflammatory response that support the establishment of  infection. This effect is partially attributed to components of sand fly saliva, which are able to recruit and activate leukocytes. Our group has shown that heme oxygenase-1 (HO-1) favors  survival in infected cells by reducing inflammatory responses. Here, we show that exposure to sand fly bites is associated with induction of HO-1 . Histopathological analyses of skin specimens from human volunteers experimentally exposed to sand fly bites revealed that HO-1 and Nrf2 are produced at bite sites in the skin. These results were recapitulated in mice ears injected with a salivary gland sonicate (SGS) or exposed to sand fly bites, indicating that vector saliva may be a key factor in triggering HO-1 expression. Resident skin macrophages were the main source HO-1 at 24-48 h after bites. Additionally, assays  after bites and  after stimulation with saliva both demonstrated that HO-1 production by macrophages was Nrf2-dependent. Collectively, our data demonstrates that vector saliva induces early HO-1 production at the bite sites, representing a major event associated with establishment of naturally-transmitted  infections.

 Saliva Induces Heme Oxygenase-1 Expression at Bite Sites.

Poorly controlled diabetes mellitus leads to several comorbidities, including susceptibility to infections. Hyperglycemia increases phagocyte responsiveness, however immune cells from people with diabetes show inadequate antimicrobial functions. We and others have shown that aberrant production of leukotriene B (LTB) is detrimental to host defense in models of bacterial infection. Here, we will unveil the consequences of high glucose in the outcome of  skin infection in people with diabetes and determine the role of LTB in human phagocytes. We show that diabetes leads to higher systemic levels of LTB, IL-6 and TNF-α in cutaneous leishmaniasis. Only LTB correlated with blood glucose levels and healing time in diabetes comorbidity. Skin lesions of people with leishmaniasis and diabetes exhibit increased neutrophil and amastigote numbers. Monocyte-derived macrophages from these individuals showed higher  loads, reduced production of Reactive Oxygen Species and unbalanced LTB/PGE ratio. Our data reveal a systemic inflammation driven by diabetes comorbidity in opposition to a local reduced capacity to resolve  infection and a worse disease outcome.

Unbalanced production of LTB/PGE driven by diabetes increases susceptibility to cutaneous leishmaniasis.

Inflammasome Activation by CD8 T Cells from Patients with Cutaneous Leishmaniasis Caused by Leishmania braziliensis in the Immunopathogenesis of the Disease.

, an intracellular parasite species, causes lesions on the skin and in the mucosa and internal organs. The dissemination of infected host cells containing  is crucial to parasite survival and the establishment of infection. Migratory phenomena and the mechanisms underlying the dissemination of -infected human dendritic cells (hDCs) remain poorly understood. The present study aimed to investigate differences among factors involved in hDC migration by comparing infection with visceral leishmaniasis (VL) induced by  with diverse clinical forms of tegumentary leishmaniasis (TL) induced by  or . Following the infection of hDCs by isolates obtained from patients with different clinical forms of , the formation of adhesion complexes, actin polymerization, and CCR7 expression were evaluated. We observed increased hDC migration following infection with isolates of  (VL), as well as disseminated (DL) and diffuse (DCL) forms of cutaneous leishmaniasis (CL) caused by  and , respectively. Increased expression of proteins involved in adhesion complex formation and actin polymerization, as well as higher CCR7 expression, were seen in hDCs infected with , DL and DCL isolates. Together, our results suggest that hDCs play an important role in the dissemination of  parasites in the vertebrate host.

ArticleTotal : 2	Year
Publication title Cited by 3	2021
Publication title	2022

Article	Year
Autophagic Induction Greatly Enhances Intracellular Survival Compared to in CBA/j-Infected Macrophages. Frontiers in microbiology\| PubMed ID: 30158914	2018
Saliva Induces Heme Oxygenase-1 Expression at Bite Sites. Frontiers in immunology\| PubMed ID: 30546363	2018
Unbalanced production of LTB/PGE driven by diabetes increases susceptibility to cutaneous leishmaniasis. Emerging microbes & infections\| PubMed ID: 32525457	2020
Inflammasome Activation by CD8 T Cells from Patients with Cutaneous Leishmaniasis Caused by Leishmania braziliensis in the Immunopathogenesis of the Disease. The Journal of investigative dermatology\| PubMed ID: 32544477	2021
-Induced Dendritic Cell Migration and Its Potential Contribution to Parasite Dissemination. Microorganisms\| PubMed ID: 34207943	2021

Cláudia I. Brodskyn

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