Modulation by bovine lactoferrin of parameters associated with the IgA response in the proximal and distal small intestine of BALB/c mice.

BACKGROUND: Secretory IgA (SIgA) and the polymeric immunoglobulin receptor (pIgR) have a pivotal role in gut homeostasis. Bovine lactoferrin (bLf) has been shown to modulate intestinal immunity and endogenous corticosterone. Considering the regionalization of the intestinal immune response, the aim of this work was to compare the impact of bLf on the IgA response in the proximal versus distal small intestine under physiological conditions. METHODS: Groups of healthy male BALB/c mice were orally treated with one daily dose of bLf (50, 500, or 5000 µg) or untreated (control) for 7 d, and then sacrificed. From plasma samples, corticosterone levels were determined by an enzyme-linked immunosorbent assay (ELISA) kit. From distal and proximal segments of the small intestine, the following material was obtained: intestinal secretions to evaluate IgA levels by ELISA; epithelial cell extracts for protein-analysis of α-chain and pIgR by Western blot; mucosa samples for mRNA analysis of α-/J-chain, pIgR, and interleukin (IL)-2, -4, -5, and -6 by real-time quantitative reverse transcription polymerase chain reaction (RT-PCR). RESULTS: With 5000 µg of bLf, there were greater modulatory effects in the distal (versus proximal) segment, evidenced by an increase in the (i) level of total and specific IgA, (ii) protein expression of α-chain and pIgR, (iii) mRNA transcripts of α-chain, IL-2 and IL-5, and (iv) level of plasma corticosterone. CONCLUSIONS: Endogenous corticosterone elicited by bLf may have allowed for an IL profile that favored the IgA antibody response. The latter has a key role in maintaining intestinal homeostasis.

Effect of Bovine Lactoferrin Treatment Followed by Acute Stress on the IgA-Response in Small Intestine of BALB/c Mice.

Secretory IgA (SIgA) has a pivotal role in gut homeostasis, which can be disturbed by stress. SIgA is formed by IgA-dimers (associated by the J-chain) and the secretory component, a protein derivative of polymeric immunoglobulin receptor (pIgR). Given the gut immuno-modulatory properties of bovine lactoferrin (bLf), the aim of this study was to compare, after bLf treatment followed by acute stress, the IgA response and IgA-associated parameters in proximal versus distal small intestine. Male BALB/c mice (n = 6) were orally treated with bLf (50, 500, and 5000 µg) for 7 days, then stressed by immobilization for 1 h, and sacrificed. In proximal and distal segments, levels were determined of IgA in gut secretions (enzyme-linked immunosorbent assay [ELISA]), the α-/J-chain and pIgR proteins in epithelial cells (Western-blot), and mRNA expression of the α-/J-chain, pIgR, and interleukins (ILs) in mucosa (RT-PCR). Data were compared by two-way analysis of variance (ANOVA) (significance at P < 0.05). Under acute stress, bLf triggered higher levels of IgA, SIgA, and anti-bLf-IgA as well as greater mRNA expression of pIgR, IL-4, and IL-6 (500 µg) in proximal intestine, while inducing higher levels of the total IgA, α-/J-chain, and pIgR proteins as well as greater mRNA expression of the α-chain and IL-4 (5000 µg) in distal intestine. Compared to unstressed/bLf-untreated mice, plasma corticosterone (a stress biomarker, measured by ELISA) increased in stressed/bLf-treated (0, 50 and 500 µg) and unstressed/bLf-treated (5000 µg) mice. The interplay of corticosterone with gut neuroendocrine factors may have elicited signals creating anti-inflammatory conditions for an IgA-response profile in each intestinal region, according to the bLf concentration administered.

Intermittent fasting favored the resolution of Salmonella typhimurium infection in middle-aged BALB/c mice.

Intermittent fasting (IF) reportedly increases resistance and intestinal IgA response to Salmonella typhimurium infection in mature mice. The aim of this study was to explore the effect of aging on the aforementioned improved immune response found with IF. Middle-aged male BALB/c mice were submitted to IF or ad libitum (AL) feeding for 40 weeks and then orally infected with S. typhimurium. Thereafter, infected animals were all fed AL (to maximize their viability) until sacrifice on day 7 or 14 post-infection. We evaluated body weight, bacterial load (in feces, Peyer's patches, spleen and liver), total and specific intestinal IgA, lamina propria IgA+ plasma cells, plasma corticosterone, and messenger RNA (mRNA) expression of α-chain, J-chain, and the polymeric immunoglobulin receptor (pIgR) in liver and intestinal mucosa. In comparison with the infected AL counterpart, the infected IF group (long-term IF followed by post-infection AL feeding) generally had lower intestinal and systemic bacterial loads as well as higher total IgA on both post-infection days. Both infected groups showed no differences in corticosterone levels, body weight, or food and caloric intake. The increase in intestinal IgA was associated with enhanced pIgR mRNA expression in the intestine (day 7) and liver. Thus, to maintain body weight and caloric intake, IF elicited metabolic signals that possibly induced the increased hepatic and intestinal pIgR mRNA expression found. The increase in IgA probably resulted from intestinal IgA transcytosis via pIgR. This IgA response along with phagocyte-induced killing of bacteria in systemic organs (not measured) may explain the resolution of the S. typhimurium infection.

Intermittent fasting modulates IgA levels in the small intestine under intense stress: a mouse model.

Intermittent fasting prolongs the lifespan and unlike intense stress provides health benefits. Given the role of the immunoglobulin A (IgA) in the intestinal homeostasis, the aim of this study was to assess the impact of intermittent fasting plus intense stress on secretory IgA (SIgA) production and other mucosal parameters in the duodenum and ileum. Two groups of six mice, with intermittent fasting or fed ad libitum for 12weeks, were submitted to a session of intense stress by a bout of forced swimming. Unstressed ad libitum fed or intermittently fasted groups were included as controls. After sacrifice, we evaluated intestinal SIgA and plasma adrenal hormones, lamina propria IgA+ plasma-cells, mRNA expression of polymeric immunoglobulin receptor, α- and J-chains in the liver and intestinal mucosa, as well as pro- (tumor necrosis factor-α, interleukin-6 and Interferon-γ) and anti- (interleukin-2, -4, -10 and transforming growth factor-ß) inflammatory cytokines in mucosal samples. Under intense stress, intermittent fasting down- or up-modulated the levels of most parameters in the duodenum and ileum, respectively while up-regulated corticosterone levels without affecting epinephrine. Our data suggest intermittent fasting plus intense stress elicited neuroendocrine pathways that differentially controlled IgA and pIgR expression in duodenum and ileum. These findings provide experimental foundations for a presumable impact of intermittent fasting under intense stress on the intestinal homeostasis or inflammation by triggering or reducing the IgA production in ileum or duodenum respectively.

Stress modulates intestinal secretory immunoglobulin A.

Stress is a response of the central nervous system to environmental stimuli perceived as a threat to homeostasis. The stress response triggers the generation of neurotransmitters and hormones from the hypothalamus pituitary adrenal axis, sympathetic axis and brain gut axis, and in this way modulates the intestinal immune system. The effects of psychological stress on intestinal immunity have been investigated mostly with the restraint/immobilization rodent model, resulting in an up or down modulation of SIgA levels depending on the intensity and time of exposure to stress. SIgA is a protein complex formed by dimeric (dIgA) or polymeric IgA (pIgA) and the secretory component (SC), a peptide derived from the polymeric immunoglobulin receptor (pIgR). The latter receptor is a transmembrane protein expressed on the basolateral side of gut epithelial cells, where it uptakes dIgA or pIgA released by plasma cells in the lamina propria. As a result, the IgA-pIgR complex is formed and transported by vesicles to the apical side of epithelial cells. pIgR is then cleaved to release SIgA into the luminal secretions of gut. Down modulation of SIgA associated with stress can have negative repercussions on intestinal function and integrity. This can take the form of increased adhesion of pathogenic agents to the intestinal epithelium and/or an altered balance of inflammation leading to greater intestinal permeability. Most studies on the molecular and biochemical mechanisms involved in the stress response have focused on systemic immunity. The present review analyzes the impact of stress (mostly by restraint/immobilization, but also with mention of other models) on the generation of SIgA, pIgR and other humoral and cellular components involved in the intestinal immune response. Insights into these mechanisms could lead to better therapies for protecting against pathogenic agents and avoiding epithelial tissue damage by modulating intestinal inflammation.

Effects on secretory IgA levels in small intestine of mice that underwent moderate exercise training followed by a bout of strenuous swimming exercise.

Intestinal homeostasis effectors, secretory IgA (SIgA) and polymeric immunoglobulin receptor (pIgR), have been evaluated in proximal and distal small intestine with moderate-exercise training but not with strenuous exercise or a combination of these two protocols. Therefore, two groups of mice (n=6-8) were submitted to strenuous exercise, one with and one without previous training. The control group had no exercise protocol. Assessment was made of intestinal SIgA and plasma adrenal hormones (by immunoenzymatic assay), alpha-chain and pIgR proteins in intestinal mucosa (by Western blot), lamina propria IgA plasma-cells (by cytofluorometry), mRNA expression (by real-time PCR) for pIgR, alpha- and J-chains in liver and intestinal mucosa, and pro- and anti-inflammatory cytokines in mucosa samples. Compared to other exercise protocols, training plus strenuous exercise elicited: (1) higher levels of SIgA and pIgR in the proximal intestine (probably by hepatobiliary contribution); (2) higher levels of SIgA in the distal segment; (3) lower mRNA expression of some SIgA- and most pro-inflammatory pIgR-producing cytokines. SIgA and pIgR in both segments were derived from an existing pool of their corresponding producing cells. The apparent decreased translation of mRNA transcripts underlies lower levels of SIgA and pIgR in distal than proximal small intestine. There was no significant difference in the relatively high adrenal hormone levels found in both exercised groups. Further study is required about the effects of training plus strenuous exercise on pool-derived SIgA levels and mRNA expression of pro-inflammatory pIgR-producing cytokines. These results could have important implications for intestinal disorders involving inflammation and infection.

Caloric restriction modifies both innate and adaptive immunity in the mouse small intestine

Although caloric restriction (CR) apparently has beneficial effects on the immune system, its effects on the immunological function of the intestinal mucosa are little known. The present study explored the effect of CR on the innate and adaptive intestinal immunity of mice. Balb/c mice were either fed ad libitum (control) or on alternate days fed ad libitum and fasted (caloric restriction). After 4 months, an evaluation was made of IgA levels in the ileum, the gene expression for IgA and its receptor (pIgR), as well as the expression of two antimicrobial enzymes (lysozyme and phospholipase A2) and several cytokines of the intestinal mucosa. CR increased the gene expression of lysozyme and phospholipase A2. The levels of IgA were diminished in the ileum, which apparently was a consequence of the reduced transport of IgA by pIgR. In ileum, CR increased the gene expression for most cytokines, both pro- and anti-inflammatory. Hence, CR differentially modified the expression of innate and adaptive immunity mediators in the intestine (AU)

Caloric restriction modifies both innate and adaptive immunity in the mouse small intestine.

Although caloric restriction (CR) apparently has beneficial effects on the immune system, its effects on the immunological function of the intestinal mucosa are little known. The present study explored the effect of CR on the innate and adaptive intestinal immunity of mice. Balb/c mice were either fed ad libitum (control) or on alternate days fed ad libitum and fasted (caloric restriction). After 4 months, an evaluation was made of IgA levels in the ileum, the gene expression for IgA and its receptor (pIgR), as well as the expression of two antimicrobial enzymes (lysozyme and phospholipase A2) and several cytokines of the intestinal mucosa. CR increased the gene expression of lysozyme and phospholipase A2. The levels of IgA were diminished in the ileum, which apparently was a consequence of the reduced transport of IgA by pIgR. In ileum, CR increased the gene expression for most cytokines, both pro- and anti-inflammatory. Hence, CR differentially modified the expression of innate and adaptive immunity mediators in the intestine.

Effect of moderate exercise on IgA levels and lymphocyte count in mouse intestine.

The aim of the present study was to determine the effect of moderate exercise on the production and secretion of IgA in mouse duodenum, on lymphocyte levels in the lamina propria, and on gene expression encoding for cytokines that regulate the synthesis of α-chain of IgA and the expression of pIgR in the lamina propria. Two groups of young Balb/c mice were fed ad libitum, one sedentary and the other with an exercise program (swimming) for 16 weeks. IgA levels in the duodenum were quantified by ELISA; the number of IgA containing cells as well as B cells, CD4(+) and CD8(+) T cells in the duodenal mucosa was determined by immunohistochemistry; gene expression was analyzed by real-time PCR, and the expression of proteins by Western blotting. Because of physical training, in the duodenum there was a decrease in the number of IgA producing cells, but an increase in the levels of IgA. Additionally, exercise increased the expression of the genes encoding for IL-4, IL-6, IL-10, TNF-α and TGF ß, cytokines that regulate the synthesis of IgA and pIgR, the inflammatory response, and the immune response in the intestine. Thus, the increased IgA found in the duodenal lumen is probably due to the increased production of IgA in the LP and the increased transport of the pIgA-pIgR complex across epithelial cells. Possibly the increased S-IgA levels in the bile also contribute to the change in IgA levels.

Caloric restriction reduces IgA levels and modifies cytokine mRNA expression in mouse small intestine.

The aim of this study was to determine the effect of caloric restriction (CR) in mouse small intestine on the production and secretion of immunoglobulin (Ig) A, the population of lymphocytes in the lamina propria, and the expression of cytokines that mediate and regulate innate and adaptive immunity. One group of young Balb/c mice was fed ad libitum, while the CR group was fed ad libitum and fasted on alternate days. When mice were six months old, IgA levels in the proximal small intestine were quantified by enzyme-linked immunosorbent assay, while the number of IgA containing cells, CD4(+) T cells and CD8(+) T cells in the duodenal mucosa was determined by immunohistochemistry. Furthermore, the expression of several intestinal cytokines, the genes for α-chain IgA, and the polymeric Ig receptor (pIgR) were analyzed by real-time polymerase chain reaction. CR decreased the levels of IgA in the intestine, apparently a consequence of a reduced number of IgA(+) cells in the lamina propria that decrease the production and secretion of this Ig, and a reduced secretion of S-IgA into the bile, which in turn discharges into the proximal intestine. Contrarily, CR increased the expression of genes for α-chain IgA, and the pIgR, indicating that transport of IgA was not a key factor in the decrease of this Ig. Additionally, CR modified the expression of genes for tumor necrosis factor-α, interferon-γ, tumor growth factor-ß, interleukin (IL)-2 and IL-10, all of which regulate the synthesis of IgA and pIgR, the inflammatory response, and the immune response in the intestine.

Caloric restriction increases free radicals and inducible nitric oxide synthase expression in mice infected with Salmonella Typhimurium.

It is well known that CR (caloric restriction) reduces oxidative damage to proteins, lipids and DNA, although the underlying mechanism is unclear. However, information concerning the effect of CR on the host response to infection is sparse. In this study, 6-month-old mice that were fed AL (ad libitum) or with a CR diet were infected with Salmonella serovar Typhimurium. EPR (electron paramagnetic resonance; also known as ESR (electron spin resonance)) was used to identify FRs (free radicals). These results were subsequently correlated with SOD (superoxide dismutase) catalytic activity, iNOS [inducible NOS (nitric oxide synthase) or NOSII] expression and NO (nitric oxide) content. EPR analysis of liver samples demonstrated that there was a higher quantity of FRs and iron-nitrosyl complex in infected mice provided with a CR diet as compared with those on an AL diet, indicating that CR was beneficial by increasing the host response to Salmonella Typhimurium. Furthermore, in infected mice on the CR diet, NOSII expression was higher, NO content was greater and spleen colonization was lower, compared with mice on the AL diet. No changes in SOD activity were detected, indicating that the NO produced participated more in the formation of iron-nitrosyl complexes than peroxynitrite. These results suggest that CR exerts a protective effect against Salmonella Typhimurium infection by increasing NO production.

Repeated restraint stress increases IgA concentration in rat small intestine.

The most abundant intestinal immunoglobulin and first line of specific immunological defense against environmental antigens is secretory immunoglobulin A. To better understand the effect of repeated stress on the secretion of intestinal IgA, the effects of restraint stress on IgA concentration and mRNA expression of the gene for the alpha-chain of IgA was assessed in both the duodenum and ileum of the rats. Restraint stress induced an increase in intestinal IgA, which was blocked by an adrenalectomy, suggesting a role of catecholamines and glucocorticoids. Whereas the blocking of glucocorticoid receptors by RU-486 did not affect the increased IgA concentration, it did reduce IgA alpha-chain mRNA expression in both segments, indicating a possible mediation on the part of glucocorticoids in IgA secretion by individual cells. Treatment with corticosterone significantly increased both the IgA concentration and IgA alpha-chain mRNA expression in ileum but not in duodenum, suggesting that glucocorticoids may act directly on IgA-antibody forming cells to increase IgA secretion in the former segment. A probable role by catecholamines was evidenced by the reduction in IgA concentration and IgA alpha-chain mRNA expression in both segments after a chemical sympathectomy with 6-hydroxydopamine (6-OHDA). Additionally, norepinephrine significantly reduced IgA alpha-chain mRNA levels but increased pIgR mRNA expression and IgA concentration in both intestinal segments. We propose that the increased intestinal IgA levels caused by repeated restraint stress is likely due to the effects of catecholamines on the transport of plgA across the epithelium.

Role of the striatum in the humoral immune response to thymus-independent and thymus-dependent antigens in rats.

Since the role of striatal GABAergic medium-sized spiny (MSP) neurons in the modulation of the immune responses is largely unknown, we evaluated the humoral immune response in rats with bilateral lesion of the striatum caused by quinolinic acid, which destroys MSP neurons. Sham-operated rats and those with striatal lesions were immunized either with TNP-LPS, a T-independent antigen type 1, or one of several T-dependent antigens: ovoalbumin, bovine serum albumin, lysozyme, sheep red blood cells (SRBC) or outer membrane proteins (OMP) of Salmonella enterica serovar Typhimurium. The specific levels of serum IgM and IgG, as well as intestinal IgA antibodies were determined either by enzyme-linked immunosorbent assay (ELISA) or a haemagglutination assay 5 or 7 days after immunization. Our results show that the lesion of striatal MSP neurons attenuated the primary antibody response to the T-independent antigen type 1 (TNP-LPS), but increased the antibody response to T-dependent antigens (proteins, SRBC and OMP), indicating that the striatal neurons modulate the humoral immune response in rats. The mechanisms involved are probably related to a reduction in both the number of B cells and the expression of caveolin-1 in the spleen, as well as an increase in the number of CD4(+) T cells and in corticosterone levels of the serum.