Claudia E. Sotomayor

Evidence of a role for galectin‐1 in acute inflammation

Galectin‐1 (Gal‐1), a member of a family of β‐galactoside‐binding proteins, has been suggested to play key roles in immunological and inflammatory processes. The present study deals with the concept of an in vivo role for Gal‐1 in acute inflammation by using the rat hind paw edema test. Local administration of Gal‐1 (0.5, 2, 4 and 8 μg / ml) inhibited acute inflammation induced by bee venom phospholipase A2 (PLA2) when it was injected 30 min before the enzyme or co‐injected together with PLA2. The anti‐inflammatory effect was prevented by a specific antibody, but independent of its carbohydrate‐binding properties. In contrast, Gal‐1 failed to inhibit histamine‐induced edema. Histopathological studies showed a clear reduction of the inflammatory process when Gal‐1 was injected before PLA2, evidenced by a diminished number of infiltrated polymorphonuclear neutrophils and scarce degranulated mast cells. The anti‐inflammatory effect was also assessed in vitro, showing that Gal‐1 treatment reduced prostaglandin E2 secretion and arachidonic acid release from stimulated peritoneal macrophages. Results presented here provide the first evidence for a role of Gal‐1 in acute inflammation and suggest that the anti‐inflammatory effect involves the inhibition of both soluble and cellular mediators of the inflammatory response.

Regulated expression of a 16-kd galectin-like protein in activated rat macrophages.

We investigated the presence of a galectin‐like protein in rat mononuclear cells using a polyclonal antibody raised against a soluble lactose‐binding lectin purified from adult chicken liver that immunoreacted strongly with a broad protein band of about 16 kd in Western blot assays. Immunochemical studies revealed a constitutive expression of this protein in mononuclear cells mainly in the macrophage (Mφ) population. Subcellular localization was assessed by Western blot assays of the cytosolic and membrane fractions of different cell populations studied: (1) spleen mononuclear cells, (2) T cell‐enriched, (3) B cell‐ and Mφ‐enriched populations, and (4) peritoneal cells, processed in the presence of lactose. In broad agreement with immunocytochemical studies of nonpermeabilized and permeabilized cells, Western blot assays suggest that this protein is localized mainly in the cytoplasmic compartment but also associated with the cell surface. By flow cytometric analyses we detected about a 14% of ED1 double‐positive cells corresponding to Mφs that constitutively express this galectin‐like protein associated with their cell surface. The cytosolic fraction obtained from the Mφ‐enriched cell population showed hemagglutinating activity specifically inhibited by β‐galactoside–related sugars. Moreover, this galectin‐like protein was retained in a lactosyl‐Sepharose matrix and specifically eluted with lactose. In this work, evidence is also provided to show that different stimuli are able to modulate the expression of the galectin‐like protein. Expression was upregulated in inflammatory and activated Mφs, revealing a significant increase in phorbol ester– and formylmethionine oligopeptide–treated cells. Both stimuli involving protein kinase C activation pathway have been able not only to up‐regulate the total expression of this protein but also to modulate its subcellular localization.

Oxidative and nitrosative stress in Staphylococcus aureus biofilm.

Diverse chemical and physical agents can alter cellular functions associated with oxidative metabolism, thus stimulating the production of reactive oxygen species (ROS) and reactive nitrogen intermediates (RNI) in planktonic bacterial physiology. However, more research is necessary to determine the precise role of cellular stress in biofilm. The present study was designed to address the issues of Staphylococcus aureus biofilm formation with respect to the generation of oxidative and nitrosative stress. We studied three pathogenic S. aureus clinical strains and an ATCC strain exposed to a different range of culture conditions (time, temperature, pH, reduction and atmospheric conditions) using quantitative methods of biofilm detection. We observed that cellular stress could be produced inside biofilms, thereby affecting their growth, resulting in an increase of ROS and RNI production, and a decrease of the extracellular matrix under unfavorable conditions. These radical oxidizers could then accumulate in an extracellular medium and thus affect the matrix. These results contribute to a better understanding of the processes that enable adherent biofilms to grow on inert surfaces and lead to an improved knowledge of ROS and RNI regulation, which may help to clarify the relevance of biofilm formation in medical devices.

Galectins: a key intersection between glycobiology and immunology

Galectins are a family of evolutionarily conserved animal lectins, widely distributed from lower invertebrates to mammals. They share sequence and structure similarities in the carbohydrate recognition domain and specificity for polylactosamine-enriched glycoconjugates. In the last few years significant experimental data have been accumulated concerning their participation in different biological processes requiring carbohydrate recognition such as cell adhesion, cell growth regulation, inflammation, immunomodulation, apoptosis and metastasis. In the present review we will discuss some exciting questions and advances in galectin research, highlighting the significance of these proteins in immunological processes and their implications in biomedical research, disease diagnosis and clinical intervention. Designing novel therapeutic strategies based on carbohydrate recognition will provide answers for the treatment of autoimmune disorders, inflammatory processes, allergic reactions and tumor spreading.

Impaired Activity of Phagocytic Cells in Candida albicans Infection after Exposure to Chronic Varied Stress

Objective: Candidiasis is a prototypic opportunistic fungal disease that may follow severe modulations of the immune system of the host. The purpose of this study was to evaluate which innate immune mechanisms involved in the protection against fungal invasion are impaired under stress conditions. Methods: Wistar rats were infected intraperitoneally with Candida albicans and immediately exposed to chronic varied stress (CVS) over 10 days (CVS; Ca-S); the fungal burden (CFU), histopathological lesion and ACTH levels were evaluated. Additionally, functional assessment of peritoneal cells (PC) included the phagocytic and anticandidacidal activities and the production of H2O2 and NO. Results: In the only infected animals (Ca), C. albicans colonization stimulated an efficient inflammatory response, while in Ca-S rats poor tissue reactions were associated with increased CFU in livers and kidneys (p < 0.05, Ca vs. Ca-S). Whereas the phagocytic process was not modified, the candidacidal activity of PC was significantly decreased after the application of CVS (p < 0.001, Ca vs. Ca-S). The H2O2 production by macrophages and neutrophils was downregulated by the infection, and while at early intervals these cells possessed a residual oxidative capacity, by day 10, the production of this metabolite was blocked. Spontaneous NO production by macrophages was significantly increased in both Ca and Ca-S animals (p < 0.001), but in stressed rats, this reactive nitrogen intermediate was noticeably downregulated (p < 0.05, Ca vs. Ca-S). The hyperactivity of hypothalamus-pituitary-adrenal axis after exposure to stress was confirmed by an increase in baseline plasma ACTH levels. Conclusion: These results show that during infection with C. albicans, the exposure to CVS contributes to the spread of the fungus and downregulates critical functions of phagocytic cells involved in the control of this opportunistic pathogen.

Evaluation of Syngonanthus nitens (Bong.) Ruhl. extract as antifungal and in treatment of vulvovaginal candidiasis

The purpose of this study was to evaluate the in vitro anticandidal activity of a methanolic extract of Syngonanthus nitens scapes against different Candida species and clinical isolates from patients with vulvovaginal candidiasis (VVC), and its effect in vivo in the treatment of vaginal infection. Chemical characterization of the extract was performed by HPLC-UV analyses and showed the presence of flavones derivatives. The extract was effective against several Candida strains from our collection and species recovered from VVC patients, and was able to inhibit the yeast-hyphal transition. No cytotoxic activity against human female reproductive tract epithelial cells and no hemolytic activity against human red blood cells were observed. In the in vivo model of VVC, we evaluated the efficacy of the intravaginal treatment with a cream containing the extract at doses of 0.5, 1.0 and 2.0%. The treatment eradicated the vaginal fungal burden in infected rats after 8 days of treatment. S. nitens extract could be considered as an effective and non-toxic natural antifungal agent in the treatment of vulvovaginal candidiasis.

Immunosuppression in experimental cryptococcosis in rats. Induction of afferent T suppressor cells to a non-related antigen

To demonstrate the nature of the suppressor cells elicited in rats infected with Cryptococcus neoformans and immunized with human serum albumin (HSA), spleen mononuclear (SpM) cells were fractionated through a nylon wool column. The adherent and non-adherent populations were collected and transferred to syngeneic rats. In all cases, the non-adherent or T-enriched cells adoptively transferred suppression to HSA, however, the suppressive effects of the non-adherent cells were never as great as those of the unpassed population of SpM cells. The fractions adherent to nylon wool also diminished the delayed-type hypersensitivity response to HSA although this was not significant, but glass-adherent cells did exhibit significant suppressor activity. Immunized, non-infected rats were used as donor controls. Furthermore, we showed that the T-enriched-cells are sensitive to treatment with low doses of cyclophosphamide and that they bind HSA. These data indicate that immune suppression of the induction of the delayed-type hypersensitivity response to HSA in cryptococcosis can occur as a result of infection with C. neoformans, and that at least one mechanism involved is the induction of adherent and non-adherent suppressor cells. Characterization of the non-adherent cells indicates that they are Ts1 cells.

Immunosuppression in experimental cryptococcosis in rats: Modification of macrophage functions by T suppressor cells

The influence of lymphocytes on the modulation of macrophage functions in altered immune states induced by Cryptococcus neoformans infection in rats has been investigated. In this report we observed a decrease of ‘in vitro’ phagocytic activity by peritoneal cells (PC) from rats that received T suppressor cells induced by cryptococcal infection, against both the same microorganism that stimulated this suppressor population (p<0.05) and another non-pathogenic primary yeast (Candida tropicalis), (p<0.02). The microbicide function of the PC from these animals present a significant decrease in challenge by C. tropicalis (p<0.002) when compared with PC from animals transferred with T normal cells. The transference of T suppressor cells induced by cryptococcal infection in animals immunized with human serum albumin-complete Freunds adjuvant (HSA-CFA) produces a significant alteration of the phagocytosis to HSA-human red cells (HSA-HRC) when compared with the phagocytosis observed in animals that received T normal cells or the phagocytosis of normal animals (p<0.001). We could also observe that the DTH to HSA studied during 30 days was negative in rats transferred with PC sensitizated with HSA and treated with suppressor T cells, when compared with the DTH response of animals transferred with PC-HSA cocultured with normal cells (p<0.05 21st day). The data presented in this paper illustrated that following infection of rats with C. neoformans there is a change in some population of accessory cells behavior reflected by the modification of several functions, such as phagocytosis, lytic activity and antigen presentation.

Non-specific immunosuppression in experimental cryptococcosis in rats

The delayed type hypersensitivity response to human serum albumin (HSA) of rats infected intraperitoneally with 107 viable C. neoformans cells, and 7 days after, immunized with human serum albumin was significantly diminished (p<0.05) when compared with the response observed in rats immunized with human serum albumin and non infected. The spleen mononuclear cells from suppressed rats transferred to normal syngeneic recipients of the same sex suppress the afferent phase of the response (p<0.02) suggesting that cells present in the spleen might be one of the responsible mechanisms for the suppression to nonrelated antigens in infected animals.

Correlation among Antioxidant, Antimicrobial, Hemolytic, and Antiproliferative Properties of Leiothrix spiralis Leaves Extract

The biological activities of a plant extract depend on a complex sum of individual properties including the antioxidant activity. Several biological activities protect against the harmful action of reactive oxygen species (ROS), and here we focused our attention on the relationship between the biological activities tested and the antioxidant properties. In this study, the total flavonoid content as well as the antioxidant, antimicrobial, hemolytic and cytotoxicity activities of the methanolic extract of Leitothrix spiralis leaves were evaluated. The extract showed a total flavonoid content of 19.26% and the chemical characterization by HPLC-PAD confirmed the presence of flavonoids as the major secondary metabolite compounds. Significant antioxidant activity (IC50 = 1.743 μg/mL ± 0.063) was demonstrated and was effective against Gram-negative organisms and all Candida strains tested, and showed an ability to inhibit hyphal formation. Non-hemolytic and antiproliferative activity could be demonstrated.