Superoxide Anion Generation in Human Milk Macrophages: Opsonin-Dependent Versus Opsonin-Independent Stimulation Compared with Blood Monocytes

Abstract

Macrophages are believed to play an important role within the immunoprotective effects of human breast milk. It was the purpose of this study to evaluate the capability of human milk macrophages (MM⌽) to generate superoxide anions (O 2 -) in comparison with peripheral blood monocytes (BMo) after stimulation with opsonized and unopsonized zymosan. Potential inhibitors of attachment and phagocytosis such as mannose and cytochalasin B were used. Expression of the mannose receptor on MM⌽ was demonstrated by staining with MAb. BMo generated more O 2 -than MM⌽ (417 Ϯ 79 versus 216 Ϯ 15 nmol O 2 -/mg protein, p Ͻ 0.05) after stimulation with opsonized zymosan. When unopsonized zymosan was used as a serum-independent stimulus, BMo generated slightly less O 2 -in comparison with MM⌽ (150 Ϯ 34 versus 176 Ϯ 18 nmol O 2 -/mg protein, p Ͻ 0.05). These findings imply a higher proportion of opsoninindependent phagocytosis in MM⌽ than in BMo (82 versus 36%). Preincubation with mannose resulted in a significantly higher reduction of O 2 -generation in MM⌽ compared with BMo stimulated with opsonized zymosan, whereas no difference was found when unopsonized zymosan was used. After addition of cytochalasin B, equal inhibition of O 2 -generation was observed regardless of the cell type or stimulus used. Thus, MM⌽ are stimulated to a greater extent by serum-independent mechanisms than BMo. As opsonins like complement or IgG are rare in the colostrum and the neonatal intestinal environment, such a differentiation toward serum-independent phagocytic abilities could play an important role for protective functions of human MM⌽. Besides well-known nutritional benefits, transfer of antimicrobial activity is of great importance in infant breast-feeding. A great variety of humoral defense factors (e.g. secretory Ig, especially sIgA, lactoferrin, lysozyme, oligosaccharides, mucins, and others) contribute to the beneficial effects. In addition, large numbers of viable cells are present in human colostrum and breast milk with a high proportion of macrophages likely being responsible for antiinfectious properties.These human MM⌽ with a diameter of 18 to 40 m contain a high amount of phagocytosed lipids, "foamy cells." Their morphologic and cytochemical properties are similar to differentiated macrophages. For example, they bear Fc receptors for different subclasses of IgG, IgA, and C3b receptors and synthesize various humoral defensive factors such as complement factors, lactoferrin, and lysozyme (1-5).Because human MM⌽ remain viable in conditions similar to those in the small intestine (6), show relative resistance to an environment with a pH Ͻ3, and resist trypsinization (7), it seems very likely that MM⌽ can develop their immunoprotective functions within the gastrointestinal tract of the breast-fed baby.Interaction of macrophage membrane receptors with complementary coating substances on a pathogen's surface is well described as opsonophagocytosis. These coatings (opsonins), derived from various sites of the hosts immune system, consist of b...

Immuno-physiological effects of dietary beta-glucan on Rohu, Labeo rohita challenged with Aeromonas veronii

“…As the rst line of defence against invaders, neutrophils and monocytes generate huge amounts of superoxide anion, as reported by Anderson and Siwicki 20 . A number of different sh species have shown that beta-glucan actually increases superoxide anion generation by macrophages, which is contrary to the current study [29][30][31] . Nevertheless, the experimental designs used in each of these studies were distinctive in form of betaglucan, dosage, and administration time.…”

Effect of fetal bovine serum and heat-inactivated fetal bovine serum on microbial cell wall-induced expression of procoagulant activity by equine and canine mononuclear cells in vitro

“…For example, serum-free conditions fail to provide sources of either LPS-binding protein or opsonin. Results of in vitro studies 6,14 performed with cells from other species have indicated that the presence of LPS-binding protein and opsonin markedly increase the sensitivity of the cells to endotoxin and zymosan, thereby reflecting one of the roles of these proteins in vivo. We are unaware of any studies regarding expression of tissue factor by canine monocytes.…”

“…Procoagulant activity was determined by assessing the ability of cellular lysates to shorten the recalcification time of pooled equine plasma in a 1-stage clotting time assay. 14 Clotting times were determined in triplicate photoelectronically. c Procoagulant activities of samples were determined by reference to a standard curve generated by known dilutions (0.09% to 100%) of equine brain thromboplastin prepared in our laboratory.…”

Host Factors in Amniotic Fluid and Breast Milk that Contribute to Gut Maturation

“…Chemokinetic agents such as TNFα in human milk may aid in recruiting macrophages to mucosal sites of the recipient's alimentary tract [63]. The human milk macrophage appears lipid laden and has diminished respiratory burst patterns compared with blood monocytes after opsonized phagocytosis; however, when stimulated by opsonin-independent mechanisms, milk macrophages have greater respiratory burst than blood monocytes [147][148][149][150]. It is postulated that as the intestinal environment of the neonate offers only a small amount of opsonins (complement and immunoglobulin G [IgG]), such differentiation of lectinophagocytic properties of milk macrophages could render a greater advantage to the neonate [147].…”

“…These macrophages survive in the neonatal gut environment during at least the first postnatal week and appear ideally suited to secrete various growth factors and cytokines to the recipient newborn gut. As a source of growth factors, macrophages that are present in human milk may "prime" the gut for continued epithelial maturation and closure [31, 41,44,61,81,147,150,[152][153][154][155]. Cell-to-cell interactions appear to play an integral part in the maturation of the GI tract of newborns [156].…”