Karen Henriette Pinke

Proinflammatory profile of in vitro monocytes in the ageing is affected by lymphocytes presence

BackgroundAging is associated with complex and constant remodeling of the immune function, resulting in an increasing susceptibility to infection and others diseases. The infections caused by Gram-negative microorganisms, present in nursing homes and hospitals, constitute one of the most common infections in the elderly, and are mainly combated by innate immune cells. Although the functions of innate immunity seem more preserved during aging than of adaptive immune mechanisms, two systems operate in an integrated way in the body, so that injury in one part of the immune system inevitably affects the other as they are part of a defensive network. The aim of this study was to investigate the in vitro production of proinflammatory (TNF-α, IL-6, IL-1β, CXCL-8 and MCP-1) and anti-inflammatory (TGF-β and IL-10) cytokines by monocytes, stimulated or not (basal) with lipopolysaccharide, from healthy young and elderly subjects. By means of PBMCs, we also studied if cytokine profile is altered in these different patient groups, in the presence of lymphocytes, under the same experimental conditions.ResultsThe monocytes from elderly presented higher basal production of TNF-α, MCP-1 and lower of TGF-β than young monocytes. PBMC showed similar cytokines production, irrespective age or stimulation presence. In the presence of lymphocytes, the spontaneous production of IL-10 was higher and of TGF-β was lower than monocytes, regardless of age. After LPS-stimulation, the presence of lymphocytes resulted in increased IL-6, IL-1β, MCP-1 and IL-10 and decreased CXCL-8 and TGF-β in comparison to pure culture of monocytes from young patients. With age, the same differences were observed, except for CXCL-8 and TGF-β which production was the same between monocytes and PBMC stimulated with LPS.ConclusionThese findings reinforce the systemic state of inflamm-aging frequently reported in elderly and considered a factor of susceptibility to numerous diseases. Still, the cytokine production from just monocytes of the elderly showed alterations, while in the lymphocyte presence not, suggesting an immunomodulator role of lymphocytes on monocytes. In addition, the differences between the production patterns by LPS-stimulated PBMC between young and elderly volunteers can be related with an imbalance in response against Gram-negative bacteria in throughout life.

Mast cells phagocyte Candida albicans and produce nitric oxide by mechanisms involving TLR2 and Dectin-1

Candida albicans (C. albicans) is a fungus commonly found in the human mucosa, which may cause superficial and systemic infections, especially in immunosuppression. Until now, the main actors in the defense against this fungus are the epithelial cells, neutrophils, macrophages/monocytes and dendritic cells. However, mast cells are strategically located to play a first line of anti-Candida defense and it has appropriate mechanisms to do it. As with other cells, the recognition of C. albicans occurs meanly via TLR2 and Dectin-1. We assess the TLR2/Dectin-1 involvement in phagocytosis and production of nitric oxide (NO) and reactive oxygen species (ROS) by mast cells challenged with C. albicans. Bone marrow-derived mast cells (MC) from wild type (Wt) or knockout (TLR2-/-) mice C57BL/6 were subjected to in vitro Dectin-1 blockade. After challenged with FITC-labeled C. albicans or zymosan, phagocytosis was analyzed by microscopy. The intracellular production of NO and ROS was measured by DAF-FM diacetate and CellROX Deep/Red Reagent kits. The nitrite formation and hydrogen peroxide release were analyzed by Griess reaction and Amplex Red Hydrogen Peroxide/Peroxidase Assay Kit. Wt/MC phagocytose C. albicans with production of intracellular NO, but not ROS. Moreover, increased levels of nitrite were also observed. The absence and/or blockade of TLR2/Dectin-1 caused significant decreased in C. albicans phagocytosis and NO production. Our results showed that mast cells are able to phagocytose and produce NO against C. albicans via TLR2/Dectin-1. Therefore, mast cells could be important during the course of Candida infection and as a therapeutic target.

Aging does not affect the ability of human monocyte-derived dendritic cells to phagocytose Candida albicans

BackgroundDendritic cells (DCs) are the most potent antigen-presenting cells, playing a key role in induction of both innate and adaptive immunity. Immunosenescence refers to age-associated changes in the immune system, which may be associated with susceptibility to infections and their clinical complications. The precise effects of aging on DCs in immunity to infections are not well understood. Among the common pathogenic microorganisms, the fungus Candida albicans is an important pathogen for the development of invasive infections, especially in immunocompromised individuals, as well as during aging.AimsTo make a comparative in vitro evaluation of the immunomodulatory function of DCs challenged with C. albicans, by phagocytosis of the fungal cells, and determine the involvement of TLR2 and TLR4 receptors. For this purpose, DCs were generated with the use of peripheral blood monocytes from healthy young and aged subjects.ResultsThe phagocytosis of C. albicans is developed by DCs in TLR2- and TLR4-dependent way. This mechanism is not affected by aging.ConclusionGiven the important role of the DCs in responses against the fungus, it is evident that if changes in phagocytosis occurred with aging, impairment in the elderly could develop. However, the evidence that phagocytosis of this fungus by DCs is not impaired with aging, brings us to the question of which are the mechanisms truly associated with the prevalence of certain diseases in the elderly.

Mast Cells Act as Phagocytes Against the Periodontopathogen Aggregatibacter Actinomycetemcomitans

BACKGROUND Evidence to date shows that mast cells play a critical role in immune defenses against infectious agents, but there have been no reports about involvement of these cells in eliminating periodontopathogens. In this study, the phagocytic ability of mast cells against Aggregatibacter actinomycetemcomitans compared with macrophages is evaluated. METHODS In vitro phagocytic assays were conducted using murine mast cells and macrophages, incubated with A. actinomycetemcomitans, either opsonized or not, with different bacterial load ratios. After 1 hour, cells were stained with acridine orange and assessed by confocal laser-scanning electron microscopy. RESULTS Phagocytic ability of murine mast cells against A. actinomycetemcomitans was confirmed. In addition, the percentage of mast cells with internalized bacteria was higher in the absence of opsonization than in the presence of opsonization. Both cell types showed significant phagocytic activity against A. actinomycetemcomitans. However, the percentage of mast cells with non-opsonized bacteria was higher than that of macrophages with opsonized bacteria in one of the ratios (1:10). CONCLUSIONS This is the first report about the participation of murine mast cells as phagocytes against A. actinomycetemcomitans, mainly in the absence of opsonization with human serum. Our results may indicate that mast cells act as professional phagocytes in the pathogenesis of biofilm-associated periodontal disease.

Decreased production of proinflammatory cytokines by monocytes from individuals presenting Candida-associated denture stomatitis

Candida-associated denture stomatitis (DS) is the most frequent lesion among denture wearers, especially the elderly. DS is strongly associated with Candida albicans, as well as local and systemic factors, such as impaired immune response. Monocytes are important in the protective immune response against the fungus by the production of cytokines that recruit and activate leukocytes. There are functional changes in these cells with age, and individual alterations involving monocyte response may predispose the host to developing infections by Candida spp. In this study, our aim was to evaluate the production of TNF-α, IL-6, CXCL8, IL-1β, MCP-1 and IL-10 by monocytes from elderly denture wearers with/without DS and elderly or young non-denture wearers. We detected that monocytes from elderly denture wearers with Candida-related denture stomatitis produced lower levels of CXCL-8, IL-6 and MCP-1. This imbalance in cytokine levels was observed in spontaneous or LPS-stimulated production. Therefore, our data suggested that inherent aspects of the host, such as changes in cytokine production by monocytes, might be associated with the development and the persistence of DS irrespective of aging.

Antimicrobial activity of denture adhesive associated with Equisetum giganteum- and Punica granatum-enriched fractions against Candida albicans biofilms on acrylic resin surfaces

Abstract Candida biofilms adhere to the internal surface of removable dentures, which is an etiological factor in the pathogenesis of denture stomatitis (DS). Adhesive materials are used at the base of maxillary complete dentures to improve their retention and chewing qualities. This article reports the antimicrobial activity of the enriched fractions of Equisetum giganteum and Punica granatum incorporated into a denture adhesive against C. albicans biofilm. The biofilms were induced on the surface of heat-cured acrylic resin specimens that were previously treated with a mixture of adhesive/herb extracts. The antimicrobial activity was evaluated by CFU counts, XTT reduction, and SEM and CLSM analysis. Both herb extracts amplified the anti-biofilm action of the adhesive on the acrylic resin by up to 12 h. Therefore, when these extracts were combined with COREGA®, they played a collaborative and innovative role in biofilm control and can be considered alternatives for temporary use in the treatment and/or prevention of DS.

Phagocytosis and nitric oxide production by peritoneal adherent cells in response to Candida albicans in aging: a collaboration to elucidate the pathogenesis of denture stomatitis

Abstract Elderly denture wearers are commonly affected by Candida-associated denture stomatitis (DS), an inflammatory process of the oral mucosa strongly associated with Candida spp and other microorganisms, as well as local and systemic factors. The impaired immune response against pathogens is among the inherent host factors that have been also associated with the pathogenesis of DS. Mononuclear phagocytes respond to the pathogens through phagocytosis followed by the production of several substances inside the phagosomes, among them are the reactive nitrogen species (RNS). A failure in these mechanisms may contribute to the DS development. Objective The aim of this study was to investigate the influence of aging on the internalization and the production of nitric oxide (NO) by peritoneal adherent cells (PAC), in response to Candida albicans (C. albicans). Material and methods PAC obtained from young and aged mice were challenged with dead or viable C. albicans by using predetermined proportions (cells:yeast) for 30 and 120 minutes. Phagocytosis was analyzed by acridine orange dye, and NO production by the Griess reaction. Results C. albicans phagocytosis by PAC from aged mice was similar to that of young mice, although the cells from older mice cells present more internalized fungi compared with matched control. In addition, a tendency towards impaired NO production by peritoneal mononuclear phagocytes from aged mice was observed. Conclusions PAC from aged mice may capture and store many fungi, which in turn may mean that these cells are effectively unable to eliminate fungi, probably due to impaired NO production. Therefore, considering the important role of C. albicans overgrowth in the pathogenesis of DS and the aspects observed in this study, aging may favor the onset and severity of local candidosis such as DS and its systemic forms.