Chemiluminescence activity in whole blood phagocytes of dogs naturally infected with Leishmania infantum.

Dogs are the domestic reservoir of Leishmania infantum, a vector-borne intracellular protozoan agent of human visceral leishmaniasis. The role of polymorphonuclear leukocytes (PMNs) in the immune defence against this parasite has been poorly studied. We have investigated the function of peripheral blood PMNs in naive beagle dogs that have been naturally exposed to phlebotomine vectors in an area highly endemic for canine leishmaniasis, and found infected by Leishmania at the end of the transmission season. Whole blood phagocyte oxidative metabolism was assessed by a rapid method that determines a luminol-amplified chemiluminescence (CL) emission. This was evaluated using either a soluble stimulant, phorbol mirystate acetate (PMA), or phagocytic stimuli, such as zymosan unopsonized (ZYM) or opsonized with autologous serum (OPZ). In blood samples taken 2 months after exposure to Leishmania transmission, data on CL emission revealed a significant decrease of reactive oxygen intermediates (ROI) production in the presence of both PMA and ZYM, compared with blood samples obtained from dogs before exposure. On the contrary, no variations in CL emission were detected in presence of OPZ. Our data indicate that immunological changes occur early in canine leishmaniasis and confirm that the role of PMNs and their products need to be clarified.

Immunopharmacological properties of flavonoids.

The flavonoids are a heterogeneous group of ubiquitous plant polyphenols that abound in the human diet and are endowed with several biological activities, including immunomodulating and antioxidant activities. The bioactivity of flavonoids is tightly correlated with their chemical structure and action mechanisms, mostly inhibitory, on enzymatic systems involved in cellular activation. The aim of this study was to evaluate the effects of acetonic extracts from Lunularia cruciata gametophytes and Feijoa sellowiana fruits on human phagocyte functions. A purified native flavonoid, a catechin, and two of its derivatives were also tested. The extract, and its flavonoid activity, was evaluated by analyzing luminol-dependent chemiluminescence emitted by human circulating phagocytes, at rest or activated by phorbol miristate acetate (PMA). The results demonstrate that both the raw extracts and flavonoids significantly inhibit CL emission by leukocytes, especially when these cells are activated by PMA. The antioxidant activity of flavonoids can be increased by changing the chemical structure of the native molecule.

Activation of peripheral phagocytes in BCG-vaccinated subjects.

The role of polymorphonuclear leukocytes (PMNs) in the immune defence against intracellular bacteria has long been neglected. Only recently have studies begun to address this issue. In this study the behavior of peripheral PMNs in Bacillus Calmette-Guerin (BCG) vaccinated subjects was investigated. Twenty healthy and purified protein derivative-negative adults were studied before, and two and four months after, BCG administration. Luminol-amplified chemiluminescence (CL) emission was evaluated in whole blood phagocytes using a soluble stimulus, such as phorbol mirystate acetate, or particulates such as zymosan opsonized with homologous (OZH) or autologous (OZA) serum. Specific IgG, IgA and IgM against antigen -60 by ELISA, total immunoglobulin, C3 and C4 components of complement, were assessed by immunochemical tests. The results revealed a late heightened production of reactive oxygen intermediates in vaccinated subjects in presence of OZA and OZH. Our findings confirm that the role of PMNs and their mediators in immunoregulation of intracellular diseases needs to be re-evaluated.

Antimicrobial and antioxidant activities of Feijoa sellowiana fruit.

The present study was designed to evaluate the antibacterial and antioxidant activities of an aqueous extract from the tropical Feijoa sellowiana Berg. fruit which is widely used for human food. The extract was tested against gram-positive and gram-negative bacteria by a broth dilution test and on human whole blood leukocytes, as well as isolated neutrophils using a chemiluminescence (CL) assay. The extract inhibited bacterial growth; Pseudomonas aeruginosa, Enterobacter aerogenes and Enterobacter cloacae were the most sensitive. The fruit extract significantly decreased CL emission from human whole blood phagocytes and isolated polymorphonuclear leukocytes whether they were activated or not by soluble or phagocytic stimuli. F. sellowiana showed both antibacterial and antioxidant properties and therefore its extract might be used as a new multifaceted drug.

Early suppression of lymphoproliferative response in dogs with natural infection by Leishmania infantum.

Dogs are the domestic reservoirs of zoonotic visceral leishmaniasis caused by Leishmania infantum. Early detection of canine infections evolving to clinically patent disease may be important to leishmaniasis control. In this study we firstly investigated the peripheral blood mononuclear cell (PBMC) response to leishmanial antigens and to polyclonal activators concanavalin A, phytohemagglutinin and pokeweed mitogen, of mixed-breed dogs with natural L. infantum infection, either in presymptomatic or in patent disease condition, compared to healthy animals. Leishmania antigens did not induce a clear proliferative response in any of the animals examined. Furthermore, mitogen-induced lymphocyte proliferation was found strongly reduced not only in symptomatic, but also in presymptomatic dogs suggesting that the cell-mediated immunity is suppressed in progressive canine leishmaniasis. To test this finding, naive Beagle dogs were exposed to natural L. infantum infection in a highly endemic area of southern Italy. Two to 10 months after exposure all dogs were found to be infected by Leishmania, and on month 2 of exposure they all showed a significant reduction in PBMC activation by mitogens. Our results indicate that suppression of the lymphoproliferative response is a common occurrence in dogs already at the beginning of an established leishmanial infection.

Antioxidant properties of Lunularia cruciata (Bryophyta) extract.

The effects of Lunularia cruciata (L.) Dum (Bryophyta) acetonic extract was studied in vitro by means of luminol-dependent chemiluminescence (CL) emission from human peripheral whole blood phagocytes and isolated polymorphonuclear leukocytes (PMNs). L. cruciata adult thalli underwent extraction with acetone. CL emission was evaluated in an automated luminometer, measuring the oxygen free-radical production by phagocytes incubated with opsonized zymosan (OZ) or phorbol myristate acetate (PMA), in absence or in presence of various concentrations of L. cruciata extract. The CL results indicated that L. cruciata induced significant changes in light emission from whole blood phagocytes, as well as isolated PMNs. Its inhibitory activity was more evident when resting isolated PMNs were studied. When the cells were activated, the greatest inhibitory effect was observed with PMA. The L. cruciata activity could be caused by several compounds, such as flavonoids and/or sesquiterpenes, present in the acetonic extract.