Anna Pérez-Bosque

Dietary Plasma Protein Supplements Prevent the Release of Mucosal Proinflammatory Mediators in Intestinal Inflammation in Rats

Spray-dried plasma (SDP) is a complex mixture of active proteins that modulates the immune response of gut-associated lymphoid tissue. We examined whether SDP and Ig concentrate (IC) supplementation could modulate cytokine expression and inflammatory mediators in rats challenged with Staphylococcus aureus enterotoxin B (SEB). Wistar-Lewis rats were fed diets supplemented with SDP (8% wt:wt), IC (1.5% wt:wt), or milk proteins (control diet) from weaning (d 21) to d 34 after birth. On d 32 and 35, the rats were given SEB (0.5 mg/kg; intraperitoneal). Six hours after the second SEB dose, jejunal mucosa and Peyers patches (PP) from the small intestine were collected. The cytokines interferon-gamma (IFNgamma), tumor necrosis factor-alpha (TNFalpha), interleukin (IL)-6, IL-10, transforming growth factor-beta (TGFbeta), and leukotrienne B(4) (LTB(4)) were analyzed using commercial kits. SEB increased the release of proinflammatory mediators (IFNgamma, TNFalpha, IL-6, and LTB(4)) in PP (P < 0.05) and in the mucosa (P < 0.05). In both tissues, SDP prevented the increase in IFNgamma, IL-6, and LTB(4) induced by SEB (P < 0.05). IC reduced the expression of TNFalpha and LTB(4) in PP and mucosa (P < 0.05). SDP supplementation increased IL-10 and mature TGFbeta concentrations in intestinal mucosa from both inflamed and noninflamed rats. Both SDP and IC increased the mature:total TGFbeta ratio (all P < 0.05). Both supplements were effective at preventing the SEB-induced increase in proinflammatory:antiinflammatory cytokine ratios in PP and mucosa and in serum. The preventive effects of plasma supplements on intestinal inflammation involve modulation of intestinal cytokines, characterized by an increased expression of antiinflammatory cytokines.

Dietary plasma proteins attenuate the innate immunity response in a mouse model of acute lung injury

We examined whether oral plasma protein supplements affect the innate immune response in a model of acute lung inflammation. Mice were fed diets supplemented with 8 % spray-dried plasma (SDP) or 2 % plasma Ig concentrate (IC) from day 19 (weaning) until day 34. The mice were challenged with intranasal lipopolysaccharide (LPS) at day 33 (and killed 24 h later for cytokine and leucocyte analyses) or at day 34 (and killed 6 h later for cytokine determinations). In bronchoalveolar lavage fluid (BALF), LPS increased the number of leucocytes by twenty-sevenfold, an effect that was partly prevented by both SDP and IC, and by twentyfold the percentage of activated monocytes, which was partly prevented by SDP. In the lung tissue, LPS increased the infiltrated leucocytes, and this effect was prevented in part by SDP. In unchallenged mice, both SDP and IC diets reduced the percentage of resident neutrophils and monocytes (P < 0·05). In the blood, both SDP and IC completely prevented LPS-dependent monocyte activation (CD14⁺; P < 0·05). LPS dramatically increased the concentration of cytokines (TNF-α, IL-1α, IL-6, granulocyte-macrophage colony-stimulating factor and granulocyte colony-stimulating factor) and chemokines (CXCL1, CCL2, CCL3 and CCL4) in BALF. The acute response of cytokine production was reduced by 20-80 % by both SDP and IC. For chemokines, plasma supplements had no effect on LPS-induced CXCL1 expression but significantly reduced CCL2, CCL3 and CCL4 production (P < 0·05). The results support the view that dietary plasma proteins can be used to attenuate endotoxin-associated lung inflammation.

Aldosterone Reduces Crypt Colon Permeability during Low-Sodium Adaptation

Fluid and electrolyte absorption by colonic crypts depends on the transport properties of crypt cellular and paracellular routes and of the pericryptal sheath. As a low-Na+ diet increases aldosterone and angiotensin II secretion, either hormone could affect absorption. Control and adrenalectomized (ADX) Sprague-Dawley rats were kept at a high-NaCl (HS) diet and then switched to low-NaCl (LS) diet for 3 days. Aldosterone or angiotensin II plasma concentrations were maintained using implanted osmotic mini-pumps. The extracellular Na+ concentration in isolated rat distal colonic mucosa was determined by confocal microscopy using a low-affinity Na+-sensitive fluorescent dye (Sodium red, and Na+-insensitive BODIPY) bound to polystyrene beads. Crypt permeability to FITC-labelled dextran (10 kDa) was monitored by its rate of escape from the crypt lumen into the pericryptal space. Mucosal ion permeability was estimated by transepithelial electrical resistance (TER) and amiloride-sensitive short-circuit current (SCC). The epithelial Na+ channel, ENaC, was determined by immunolocalization. LS diet decreased crypt wall permeability to dextran by 10-fold and doubled TER. Following ADX, aldosterone decreased crypt wall dextran permeability, increased TER, increased Na+ accumulation in the pericryptal sheath and ENaC expression even in HS. Infusion of angiotensin II to ADX rats did not reverse the effects of aldosterone deprivation. These findings indicate that aldosterone alone is responsible for both the increase in Na+ absorption and the decreased paracellular and pericryptal sheath permeability.

A rat model of mild intestinal inflammation induced by Staphylococcus aureus enterotoxin B.

The epithelial barrier of the intestine and the gut-associated lymphoid tissue (GALT) protects the host against luminal pathogenic micro-organisms. This is important at weaning, when animals are exposed to infectious agents and stresses. We have developed a rat model of intestinal inflammation post weaning, based on the systemic administration of Staphylococcus aureus enterotoxin B (SEB). Since the inflammatory response obtained is mild, the food intake pattern is not affected, which makes this model useful for studies of nutritional therapies for intestinal inflammatory disease. SEB increased T-lymphocytes in Peyers patches and the number of activated T-lymphocytes in mesenteric lymph nodes (organized GALT). In the lamina propria, SEB increased activated T-lymphocytes as well as cytotoxic and natural killer-cell populations of the diffuse GALT. It also increased pro-inflammatory cytokines and inflammatory mediators in both Peyers patches and mucosa. Rats given SEB had higher paracellular permeability to macromolecules, which was associated with a reduction in epithelial tightness. This model was used to examine whether dietary supplementation with spray-dried animal plasma proteins affects intestinal inflammation. Results showed that dietary plasma proteins can attenuate the mucosal immune response in both organized and diffuse GALT and that these effects are mediated by a reduction in the production of pro-inflammatory cytokines.

Dietary Plasma Proteins Modulate the Adaptive Immune Response in Mice with Acute Lung Inflammation

We examined the effects of oral plasma protein supplements on the pulmonary adaptive immune response in mice challenged with intranasal LPS. C57BL/6 mice were fed a control diet or a diet supplemented with plasma proteins [spray-dried plasma (SDP) 80 g/kg] or with an Ig concentrate [(IC) 20 g/kg] from postnatal d 19 (weaning) until d 34. Mice were challenged with PBS or LPS from Escherichia coli at d 33 and killed 24 h later for leukocyte analyses or at d 34 and killed 6 h later for cytokine determination. LPS induced the activation of T helper (Th) lymphocytes in lung and blood and this response was reduced by SDP and IC (P < 0.05). In both tissues, LPS increased the Th1 and Th2 subpopulations and this effect was inhibited by the two plasma protein supplements (P < 0.05). The LPS challenge increased the expression of all the cytokines studied (P < 0.01). SDP and IC reduced the expression of IFNγ, IL-5, IL-12p40, IL-12p70, IL-13, and IL-17 in both tissues, whereas they increased the percentage of regulatory Th lymphocytes in lung, even in PBS-treated mice (P < 0.05). LPS reduced the concentration of mature TGFβ1 (P < 0.05) in the lung but did not modify the expression of IL-10. Mice exposed to LPS and supplemented with SDP or IC showed an increased expression of the anti-inflammatory cytokine IL-10 (P < 0.05). Moreover, the two supplements increased the concentration of IL-10 in intestinal mucosa (P < 0.05). Our results show that plasma supplementation reduces the immune response that characterizes the acute lung inflammation syndrome.

Pericryptal myofibroblast growth in rat descending colon induced by low-sodium diets is mediated by aldosterone and not by angiotensin II.

Pericryptal myofibroblast growth in descending colonic crypts correlates with the activation of the renin-angiotensin-aldosterone system. Earlier work showed that during the transition from a high-Na+ (HS) to low-Na+ (LS) diet there are changes in the colonic crypt wall and pericryptal sheath. As LS diet increases both aldosterone and angiotensin II, the aim here was to determine their individual contributions to the trophic changes in colonic crypts. Experiments were conducted on control and adrenalectomized Sprague-Dawley rats fed an HS diet and then switched to LS diet for 3 days and supplemented with aldosterone or angiotensin II. The actions of the angiotensin-converting enzyme inhibitor captopril, the angiotensin receptor antagonist losartan and the aldosterone antagonist spironolactone on extracellular matrix proteins, claudin 4 and E-cadherin myofibroblast proteins, α-smooth muscle actin (α-SMA) and OB-cadherin (cadherin 11), angiotensin type 1 and TGFβr1 membrane receptors were determined by immunolocalization in fixed distal colonic mucosa. The LS diet or aldosterone supplementation following ADX in HS or LS increased extracellular matrix, membrane receptors and myofibroblast proteins, but angiotensin alone had no trophic effect on α-SMA. These results show that aldosterone stimulates myofibroblast growth in the distal colon independently of dietary Na+ intake and of angiotensin levels. This stimulus could be a genomic response or secondary to stretch of the pericryptal sheath myofibroblasts accompanying enhanced rates of crypt fluid absorption.

W1801 Dietary Supplementation With Porcine Plasma Proteins Reduce Lymphocyte Recruitment and Cytokine and Chemokine Expression in a Mouse Model of Spontaneous Colitis

between CD and controls for either phagocytosis or RORs. Conclusion: The ability of monocytes fromCD patients to engulf E.coli appears normal. However, there is good evidence of an impaired capacity to kill these bacteria as measured by reduced levels of RORs. These results points to either an inability to complete the process of phagolysosome formation or defects in the enzyme pathways involved in formation of reactive oxygen products that are induced by phagocytosis and support the concept of impaired innate immunity in CD.

Aldosterone induces myofibroblast EGF secretion to regulate epithelial colonic permeability

In vivo studies show that raised aldosterone (Aldo) during low-Na adaptation regulates the growth of pericryptal myofibroblasts and reduces the permeability of the colonic epithelium. The aim of this study was to reproduce in vitro the in vivo condition of increased Aldo using human CCD-18Co myofibroblasts and T84 colonic epithelial cells to measure myofibroblast and epithelial proliferation and the expression of intercellular junction proteins. Proliferation was quantified by measuring 5-bromo-2-deoxyuridine incorporation. The myofibroblast expression of EGF, VEGFa, and transforming growth factor-β1 (TGF-β1) was measured by real-time PCR and the expression of junctional complex proteins by Western blot. Aldo stimulated the proliferation of myofibroblasts by 70% (P < 0.05) and increased EGF mRNA expression by 30% (P < 0.05) without affecting VEGFa and TGF-β1. EGF concentration in the incubation medium increased by 30% (P < 0.05) 24 h after Aldo addition, and these effects were prevented by the addition of spironolactone. Myofibroblast proliferation in response to Aldo was mediated by EGF receptor (EGFR) and involved both MAPKK and phosphatidylinositol 3-kinase pathways. When T84 cells were incubated with medium from myofibroblasts stimulated with Aldo (conditioned medium), the expression of β-catenin and claudin IV was increased by 30% (P < 0.05) and proliferation by 40% (P < 0.05). T84 proliferation decreased when α-EGF, or the EGFR antagonist AG1478, was present. Results in vivo indicate that rats fed a low-salt diet showed an increased expression of EGF and EGFR in the colonic mucosa. These results support the view that changes in colonic permeability during low-Na adaptation are mediated by the EGF secreted by myofibroblasts in response to raised Aldo.

Type I Interferon Response Is Delayed in Human Astrovirus Infections

Type I interferon (IFN) activation and its subsequent effects are important in the response to viral infections. Here we show that human astroviruses (HAstVs), which are important agents of acute gastroenteritis in children, induce a mild and delayed IFN response upon infecting CaCo-2 cells. Although IFN-β mRNA is detected within infected cells and supernatant from infected cells show antiviral activity against the replication of other well-known IFN-sensitive viruses, these responses occur at late stages of infection once genome replication has taken place. On the other hand, HAstV replication can be partially reduced by the addition of exogenous IFN, and inhibition of IFN activation by BX795 enhances viral replication, indicating that HAstVs are IFN-sensitive viruses. Finally, different levels of IFN response were observed in cells infected with different HAstV mutants with changes in the hypervariable region of nsP1a/4, suggesting that nsP1a/4 genotype may potentially have clinical implications due to its correlation with the viral replication phenotype and the antiviral responses induced within infected cells.

Spray dried plasma as an alternative to antibiotics in piglet feeds, mode of action and biosafety

The use of growth promoting and therapeutic antibiotics in piglet feed has been a concerning subject over the last few decades because of the risk of generating antimicrobial resistance that could be transferred to humans. As a result, many products have been proposed as potential alternatives to the use of antibiotics, and among these, spray dried plasma is considered one of the most promising. However, there have been concerns about its biosafety, particularly during periods of emergence or re-emergence of swine diseases in different regions of the world, such as the recent porcine epidemic diarrhea virus outbreak in North America. The objectives of this paper are to review recent publications about the use of spray dried plasma as an alternative to antibiotics in weaned pig diets, the possible mechanisms of action of spray dried plasma, and the existing evidence related to the biosafety of spray dried animal plasma. Particular attention is given to studies in which spray dried plasma has been directly compared to antibiotics or other alternative antimicrobial products. Several studies on the possible modes of action for spray dried plasma, such as preservation of gut barrier function or modulation of the immune response, are also reviewed. Finally, the paper focuses on the review of the existing studies on the risks of disease transmission with the use of spray dried plasma from porcine origin. Overall, spray dried plasma is a promising alternative to in-feed antimicrobials for piglets, particularly during the early stages of the post-weaning phase. Additionally, there is enough evidence to support that commercial spray dried porcine plasma is a safe product for pigs.