Enteral feeding preserves gut Th-2 cytokines despite mucosal cellular adhesion molecule-1 blockade

BACKGROUND: Parenteral nutrition (PN) decreases gut-associated lymphoid tissue (GALT), the intestinal IgA stimulating cytokines IL-4 and IL-10 in gut homogenates, intestinal IgA levels and the expression of Peyer patch (PP) mucosal cellular adhesion molecule-1 (MAdCAM-1), an adhesion molecule found on the high endothelial venules of PP and other tissues. IL-4 in PP stimulates MAdCAM expression in vitro. MAdCAM-1 blockade with MECA-367 reduces GALT cell populations to PN levels but maintains intestinal IgA levels if the animals are chow fed. This study compares IL-4 levels in PP of chow and PN fed mice and measures the effects of MAdCAM blockade on IL-4 and IL-10 levels in gut homogenates of chow fed mice. We hypothesized that in vivo IL-4 levels drop in PP of PN fed mice and IL-4 and IL-10 levels are maintained after MAdCAM-1 blockade in chow fed mice. METHODS: Exp 1: 18 mice received chow or PN for 5 days to determine PP IL-4 levels. Exp 2: 44 mice were randomized to chow + control monoclonal antibody (mAb), chow + MECA-367 (anti-MAdCAM-1 mAb) or PN for 4 days before measurement of IL-4 and IL-10 levels in gut homogenates. RESULTS: Exp 1: IL-4 levels in vivo were lower in PP of PN-fed mice than chow fed mice (92.0 +/- 15.1 pg/mL vs 251.1 +/- 14.8, p = .0003). Exp 2: IL-4 levels were significantly higher in chow + control mAb (187.1 +/- 44.1 pg/mL) and chow + MECA-367 (110.9 +/- 19.1 pg/mL) groups than PN mice (21.8 +/- 30.6 pg/mL, p < .02 vs chow + control or chow + MECA-367). IL-10 levels were significantly lower with PN (23.1 +/- 40.9 pg/mL) with chow+control (174.0 +/- 22.2 pg/mL p < .01), or chow + MECA-367 (181.7 +/- 23.1 pg/mL, p < .02 vs PN). CONCLUSIONS: PN-feeding reduces in vivo IL-4 levels in PP (consistent with lowered MAdCAM-1 expression) and IL-4 and IL-10 levels in gut homogenates compared with chow. Despite MAdCAM-1 blockade, enteral feeding preserved gut IL-4 levels and increased IL-10 levels consistent with preserved IgA levels.     

Interleukin-7 dose-dependently restores parenteral nutrition-induced gut-associated lymphoid tissue cell loss but does not improve intestinal immunoglobulin a levels

BACKGROUND: Without enteral nutrition, the mass and function of gut-associated lymphoid tissue (GALT), a center of systemic mucosal immunity, are reduced. Therefore, new therapeutic methods, designed to preserve mucosal immunity during parenteral nutrition (PN), are needed. Our recent study revealed that exogenous interleukin-7 (IL-7; 1 microg/kg twice a day) restores the GALT cell mass lost during intravenous (IV) PN but does not improve secretory immunoglobulin A (IgA) levels. Herein, we studied the IL-7 dose response to determine the optimal IL-7 dose for recovery of GALT mass and function during IV PN. We hypothesized that a high dose of IL-7 would increase intestinal IgA levels, as well as GALT cell numbers. METHODS: Male mice (n = 42) were randomized to chow, IL-7-0, IL-7-0.1, IL-7-0.33, IL-7-1 and IL-7-3.3 groups and underwent jugular vein catheter insertion. The IL-7 groups were fed a standard PN solution and received IV injections of normal saline (IL-7-0), 0.1, 0.33, 1, or 3.3 microg/kg of IL-7 twice a day. The chow group was fed chow ad libitum. After 5 days of treatment, the entire small intestine was harvested and lymphocytes were isolated from Peyer's patches (PPs), intraepithelial (IE) spaces, and the lamina propria (LP). The lymphocytes were counted and phenotypes determined by flow cytometry (alphabetaTCR, gammadeltaTCR, CD4, CD8, B cell). IgA levels of small intestinal washings were also examined using ELISA (enzyme-linked immunoabsorbent assay). RESULTS: IL-7 dose-dependently increased total lymphocyte numbers in PPs and the LP. The number of lymphocytes harvested from IE spaces reached a plateau at 1 microg/kg of IL-7. There were no significant differences in any phenotype percentages at any GALT sites among the groups. IgA levels of intestinal washings were significantly higher in the chow group than in any of the IL-7 groups, with similar levels in all IL-7 groups. CONCLUSIONS: Exogenous IL-7 dose-dependently reverses PN-induced GALT cell loss, with no major changes in small intestinal IgA levels. IL-7 treatment during PN appears to have beneficial effects on gut immunity, but other therapeutic methods are needed to restore secretory IgA levels.     

Parenteral nutrition and fasting reduces mucosal addressin cellular adhesion molecule-1 (MAdCAM-1) mRNA in Peyer's patches of mice

BACKGROUND: Mucosal addressin cellular adhesion molecule-1 (MAdCAM-1) in Peyer's patches (PP) is the gateway molecule for cellular migration into the mucosal immune system. Lack of enteral feeding during parenteral nutrition (PN) rapidly decreases PP MAdCAM-1, leading to drops in mucosal T and B cells and intestinal and respiratory IgA. We determined the molecular events associated with MAdCAM-1 mRNA and protein during PN (short and long term) and fasting (1 and 2 days). METHODS: Experiment 1: Cannulated mice received PN for 8 hours (short-term PN, n = 6) or chow + saline (chow, n = 6). Experiment 2: Cannulated mice received PN (long-term PN, n = 4) or chow (n = 3) for 5 days. Experiment 3: Noncannulated chow mice were fasted for 1 and 2 days (n = 2/time). Total cellular RNA from the PP was quantified for MAdCAM-1 mRNA by real-time polymerase chain reaction (PCR). MAdCAM-1 protein was measured by Western blot. RESULTS: PN rapidly down-regulated MAdCAM-1 gene expression. After 8 hours of PN with lack of enteral feeding, MAdCAM-1 mRNA levels dropped 20% (0.8-fold vs chow, p > .05); 5 days of PN reduced MAd-CAM-1 levels 64% (0.34-fold vs chow, p < .05). PN reduced MAdCAM-1 protein levels by 30% (chow: 329 +/- 14 vs PN: 230 +/- 35, p < .05) after 5 days. Fasting of uncannulated mice decreased MAdCAM-1 mRNA levels by 16% (0.84-fold, p < .05) at day 1 and 30% (0.7-fold, p < .05) by day 2 compared with chow. CONCLUSIONS: Both PN with lack of enteral feeding and fasting down-regulate MAdCAM-1 mRNA and protein levels in PP. The MAdCAM-1 changes are due to lack of enteral stimulation rather than toxic effects of PN.     

Influence of adding pyrroloquinoline quinone to parenteral nutrition on gut-associated lymphoid tissue

Background: Experimental intravenous (IV) parenteral nutrition (PN) diminishes gut‐associated lymphoid tissue (GALT) cell number and function. PN solution cannot maintain GALT at the same level as a normal diet, even when delivered intragastrically (IG). Previous studies demonstrated pyrroloquinoline quinone (PQQ)–deficient mice to be less immunologically responsive. Because standard (STD) PN solution lacks PQQ, PQQ supplementation may prevent PN‐induced GALT changes. This study was designed to determine the influence of adding PQQ to PN on GALT. Methods: In experiment 1, mice (n = 32) were randomized to chow, IV‐STD‐PN, and IV‐PQQ‐PN groups. The chow group was fed chow with the same caloric content as PN. The IV‐STD‐PN group received STD‐PN solution, whereas the IV‐PQQ‐PN group was given PQQ (3 mcg/d)–enriched PN by the IV route. After 5 days of feeding, lymphocytes were isolated from the Peyer's patch (PPs), intraepithelial space (IE), and lamina propria (LP) of the small intestine. GALT lymphocyte number and phenotype (αβTCR+, γδTCR+, CD4+, CD8+, B220+ cells) and intestinal immunoglobulin A (IgA) level were determined. In experiment 2, mice (n = 28) were randomized to IG‐STD‐PN or IG‐PQQ‐PN group. After IG nutrition supports, GALT mass and function were determined as in experiment 1. Results: The IV‐PQQ‐PN group showed increased PP lymphocyte number and PP CD8+ cell number compared with the IV‐STD PN group. The IG‐PQQ‐PN group had significantly greater PP lymphocyte number and PP CD4+ cell numbers than the IG‐STD‐PN group. Neither IV nor IG PQQ treatment raised IgA level. Conclusions: PQQ added to PN partly restores GALT mass, although its effects on GALT function remain unclear.     

Mucosal immunity preservation with bombesin or glutamine is not dependent on mucosal addressin cell adhesion molecule-1 expression

BACKGROUND: Mucosal addressin cell adhesion molecule-1 (MAdCAM-1) is an adhesion molecule that directs naive T and B cells into Peyer's patches for sensitization and distribution to intestinal and extraintestinal sites. With no enteral stimulation, its expression drops rapidly in association with reduced Peyer's patch cell populations and increases rapidly with reinstitution of enteral feeding. Because both glutamine (GLN) and bombesin (BBS) preserve mucosal immunity, this study examined whether they preserve MAdCAM-1 expression. METHODS: In 2 separate experiments, animals were randomized to IV cannulation with chow, total parenteral nutrition (TPN), and (experiment 1) 15 microg/kg BBS 3 times per day or (experiment 2) an isocaloric, isonitrogenous 2% GLN-supplemented solution. After 5 days of feeding, MAdCAM-1 expression in Peyer's patches, spleen, and intestine was measured using a dual radiolabeled monoclonal antibody technique. RESULTS: MAdCAM-1 expression was not significantly improved from TPN levels either with BBS or GLN supplementation. Levels of MAdCAM-1 expression remained unchanged in non-Peyer's patch sites. CONCLUSIONS: Although MAdCAM-1 is considered the gateway molecule for cell entry into mucosal immunity, this does not seem to be the mechanism for mucosal immunity preservation in nonenterally fed mice receiving bombesin or glutamine.     

Influence of adding fish oil to parenteral nutrition on gut-associated lymphoid tissue

BACKGROUND: Lack of enteral nutrition reduces gut-associated lymphoid tissue (GALT) mass and function, a mechanism underlying the increased morbidity of infectious complications in severely injured or critically ill patients. Strategies to restore parenteral nutrition (PN)-induced changes of GALT mass and function have been pursued. However, the influences of adding fish oil to PN on gut immunity remain to be clarified. METHODS: Male Institute of Cancer Research (ICR) mice (n = 50) were randomized to 4 groups: ad libitum chow (chow), fat free PN (fat (-)-PN), PN + fish oil (FO-PN), and PN + safflower oil (SO-PN). The PN groups were given isocaloric and isonitrogenous PN solutions. The FO- and SO-PN groups received 20% of total calories from fat emulsions. After 5 days of feeding, lymphocytes from Peyer's patches (PPs), the intraepithelial space (IE), and the lamina propria (LP) of the entire small intestine were isolated. GALT lymphocyte numbers and phenotypes (CD4+, CD8+, alphabetaTCR+, gammadeltaTCR+, B220+ cells) were determined. Immunoglobulin A (IgA) levels of small intestinal washings were also measured by enzyme-linked immunosorbent assay. Another set of mice (n = 24) was used to determine plasma fatty acid compositions after feeding. RESULTS: Lymphocyte numbers from PPs and the LP and intestinal IgA levels were significantly lower in the PN groups than in the chow group, with no significant differences between any 2 PN groups. The FO- and SO-PN groups showed moderate recovery of IE cell numbers compared with the fat (-)-PN group. Omega-3 and omega-6 fatty acid levels were increased with fish and safflower oil additions, respectively, compared with the fat (-)-PN group. CONCLUSIONS: Adding fish oil to PN does not exacerbate PN-induced GALT changes but rather partially reverses these changes, with increased plasma omega-3 fatty acid levels.     

Alterations in hepatic mitochondrial function during total parenteral nutrition in immature rats

To evaluate the effects of total parenteral nutrition (TPN) on hepatic mitochondrial function in immature rats, changes in hepatic energy charge levels and oxidative phosphorylation rates of hepatic mitochondria were studied along with the examination of serum chemical test. Male Wistar rats weighing 30 to 45 g were used and randomized into TPN (n = 8), enteral (n = 7), and control groups (n = 8). Parenteral and enteral groups were fed with TPN solution containing 19.3% dextrose, 3.19% amino acids, 1.05% fat emulsion, minerals and vitamins, and the control group with rat chow. The number of calories per kilogram per day was 550 x 1/4 on the 1st day, 550 x 1/2 on the 2nd, 550 x 3/4 on the 3rd, and 550 x 1 on the 4th day, based on the body weight on the 1st day. After the 5th day, 550 Kcal/kg/day was given, based on the body weight of the respective day. After 13-day feeding, hepatic energy charge (EC), phosphorylation rate (PR) of hepatic mitochondria and serum chemical examination were carried out. EC was 0.871 +/- 0.016 in the control group, 0.830 +/- 0.019 in the enteral, and 0.785 +/- 0.011 in TPN group (p less than 0.001, compared with control group). PR was 138.9 +/- 1.9, 133.0 +/- 6.7, 111.0 +/- 4.3, respectively, (p less than 0.05, compared with control and enteral groups). There was no difference between the three groups on SGOT, SGPT, and total bilirubin. TPN group showed a deterioration of hepatic phosphorylation rate and energy charge in spite of normal serum transaminase levels.(ABSTRACT TRUNCATED AT 250 WORDS)     

L-arginine-enriched parenteral nutrition affects lymphocyte phenotypes of gut-associated lymphoid tissue

BACKGROUND: Experimentally, total parenteral nutrition (TPN) diminishes gut-associated lymphoid tissue (GALT) cell numbers and function. Although glutamine supplementation is known to reverse TPN-induced changes in GALT, effects of another conditionally essential amino acid, L-arginine (ARG), on GALT remain unclear. METHODS: Twenty-two male Institute of Cancer Research mice were randomized to standard TPN (0.3% arginine, STD-total parenteral nutrition) or 1% ARG-enriched TPN (ARG-total parenteral nutrition). After 5 days of feeding, lymphocytes were harvested from Peyer's patches (PP), the lamina propria, and intraepithelial (IE) spaces of the small intestine to determine cell yields. Lymphocyte phenotypes (alphabetaTCR, gammadeltaTCR, CD4, CD8, and B220 as a B cell marker) were determined using flow cytometry. IgA levels in washings of the small intestine, upper respiratory tract, and lungs were measured with ELISA. RESULTS: ARG-total parenteral nutrition did not affect lymphocyte yields. The percentages of CD4+ cells in PP and IE, and alphabetaTCR+ cells in PP, were significantly higher in the ARG-total parenteral nutrition than in the STD-total parenteral nutrition mice, without marked differences in other phenotypes examined. There were no significant differences in intestinal and respiratory tract IgA levels between the 2 groups of mice. CONCLUSIONS: One percent ARG supplementation of TPN does not improve GALT cell number or mucosal IgA level but benefits to increase CD4+ cell percentages in GALT.     

Individual neuropeptides regulate gut-associated lymphoid tissue integrity, intestinal immunoglobulin A levels, and respiratory antibacterial immunity

BACKGROUND: Total parenteral nutrition (TPN) leads to atrophy of the gut-associated lymphoid tissue (GALT) and a significant decrease in intestinal immunoglobulin A (IgA) levels, a major constituent of mucosal immunity. Bombesin (BBS) prevents TPN-induced GALT atrophy and maintains intestinal IgA levels. BBS, a neuropeptide analogous to gastrin-releasing peptide in humans, stimulates the release of other gut neuropeptides including cholecystokinin (CCK), gastrin, and neurotensin (NT). This study investigates the ability of CCK, gastrin, or NT to individually prevent TPN-induced GALT atrophy and preserve respiratory immunity. METHODS: Experiment 1: Male mice were randomly assigned to receive chow, TPN, TPN plus CCK, TPN plus gastrin, or TPN plus NT. After 5 days of feeding, Peyer's patches (PP) from the proximal and distal small bowel were harvested and analyzed for cell yields. PP cells were also analyzed for GALT cell type. Small bowel IgA levels were measured by enzyme-linked immunosorbent assay (ELISA). Experiment 2: Mice were randomly assigned to receive either liposomes containing Pseudomonas antigen or liposomes without antigen. After 10 days, mice were randomly assigned to the same five treatment groups, fed for 5 days, and then given intratracheal Pseudomonas. Mortality was assessed after 48 hours. RESULTS: Experiment 1: GALT cell reductions due to IV-TPN were greater in the distal than proximal small bowel. All three neuropeptides prevented most TPN-induced GALT atrophy due mainly to the maintenance of the B-cell and T-cell populations in the PP of the distal bowel. Intestinal IgA levels were significantly higher in the animals treated with neuropeptides than animals treated with TPN only; however, these IgA levels were not maintained at levels observed in chow-fed animals. Experiment 2: Immunization resulted in significantly lower mortality in animals fed chow, TPN plus CCK, and TPN plus gastrin. TPN alone and TPN plus NT resulted in loss of immunity and mortality rate at comparable levels to unimmunized animals. CONCLUSIONS: Supplementation of IV-TPN with CCK, gastrin, and NT prevents GALT atrophy, primarily in the distal bowel. Intestinal IgA levels improve but not to normal levels. CCK and gastrin reversed IV-TPN-induced effects on antibacterial pneumonia in immunized animals while NT did not.     

Influence of enteral diets supplemented with key nutrients on lymphocyte subpopulations in Peyer's patches of endotoxin-boostered mice

BACKGROUND AND AIMS: This study was undertaken to compare the effect of different key nutrients on lymphocyte subsets of Peyer's patches (PP) and spleen in endotoxemic mice. METHODS: Female Balb/c mice were fed over a period of 10 days either with an isocaloric and isonitrogenous control diet (Control), a glutamine enriched diet (Diet I) or a diet containing glutamine, arginine, glycine, and n-3 fatty acids (Diet II). On day 7 the mice were challenged intraperitoneally with 25 microg LPS. The lymphocyte subpopulations (B cells, T cells, CD4+ and CD8+) of PP and spleen were analysed by flow cytometry. Glutathione content of small intestinal mucosa and spleen was determined by HPLC and luminal small intestinal IgA by ELISA. RESULTS: Both experimental diets increased the number of B and T cells in the PP and that of T cells in the spleen (P<0.01). Glutathione content in PP and spleen was higher under administration of key nutrients (P<0.05). Diet II reduced luminal small intestinal IgA content in comparison to the two other groups. CONCLUSION: The addition of arginine, glycine and n-3 fatty acids to a glutamine supplemented diet does not enhance lymphocyte numbers in PP and spleen, but reduces intestinal IgA content.     

Urogastrone reduces gut atrophy during parenteral alimentation

Urogastrone (UG) exerts trophic effects on the intestine and may play a role in maintaining normal intestinal structure and function. Since administration of nutrients parenterally results in intestinal hypoplasia and hypofunction, the aim of this study was to determine the effects of UG on intestinal structure and function in parenterally fed rats. Central venous catheters were placed into 28 Sprague-Dawley rats. Group I (n = 10) received TPN alone. Group II (n = 8) received TPN and 15 micrograms/day of UG and group III (n = 10) received rat chow ad libitum. The animals that received urogastrone had significantly greater (p less than 0.05) intestinal weight (25.6 +/- 2.5 mg/cm vs 22.6 +/- 3.0 mg/cm), mucosal weight (8.4 +/- 1.4 mg/cm vs 6.2 +/- 0.9 mg/cm), mucosal protein content (6.2 +/- 1.7 mg/cm vs 2.7 +/- 0.6 mg/cm), villous height (427 +/- 27 microns vs 293 +/- 75 microns), crypt cell production rate (14.5 +/- 1.4 metaphases/hr vs 12.3 +/- 0.7 metaphases/hr) and sucrase specific activity (6.5 +/- 2.6 vs 3.7 +/- 2.0) than animals receiving only TPN. However, these parameters remained less than in chow-fed animals. Thus, simultaneous infusion of UG prevents, in part, intestinal hypofunction and hypoplasia which occurs during TPN. This may be due to maintenance of mucosal proliferative activity and brush border enzyme activity.     

Glutamine-enriched total parenteral nutrition maintains intestinal interleukin-4 and mucosal immunoglobulin A levels

BACKGROUND: Total parenteral nutrition (TPN) prevents progressive malnutrition but fails to maintain intestinal gut-associated lymphoid tissue (GALT) or established respiratory antiviral or antibacterial mucosal immunity. Our previous work demonstrated that decreases in intestinal immunoglobulin A (IgA) were associated with decreases in Th2-type IgA-stimulating cytokines, interleukin (IL)-4 and IL-10. Because glutamine supplementation of TPN partially preserves respiratory defenses and normalizes GALT, we investigated the ability of parenteral glutamine to normalize respiratory and intestinal IgA levels and measured Th2 cytokines in intestinal homogenates. METHODS: Animals were cannulated and randomly assigned to receive chow (n = 17), TPN (n = 18), or an isonitrogenous, isocaloric TPN solution formulated by removing the appropriate amount of amino acids and replacing them with 2% glutamine (n = 18) for 5 days. Respiratory tract and intestinal washings were obtained for IgA and the intestine homogenized and analyzed for IL-4 and IL-10. RESULTS: TPN decreased intestinal and respiratory IgA in association with decreases in intestinal IL-4 and IL-10 compared with chow-fed animals. Glutamine significantly improved respiratory and intestinal IgA levels, significantly improved IL-4 compared with TPN animals, and maintained IL-10 levels midway between chow-fed and TPN animals. CONCLUSIONS: Glutamine-enriched TPN preserved both extraintestinal and intestinal IgA levels and had a normalizing effect on Th2-type IgA-stimulating cytokines.     

Metabolic response of simultaneous versus sequential intravenous administration of amino acids and energy substrates to rats

Three groups of rats were maintained on total intravenous nutrition for ten days. Group SA and SB were infused sequentially (2 X 12 h periods per day), SA received amino acids (AA) during the night and carbohydrates (CHO) + FAT during the day. The SB group received nutrients in the opposite order. A control group received a mixed solution simultaneously for 24 h/day. The sequentially fed groups showed a lower weight gain (2.4 +/- 0.4, 2.6 +/- 0.2 vs 4.9 +/- 0.3 g/day), nitrogen balance (95 +/- 7, 95 +/- 6 vs 139 +/- 7 mg/day) and nitrogen utilization (69 +/- 3, 67 +/- 3 vs 87 +/- 3%) compared with the control group. Administration of energy substrate in the SA and SB was a stronger denominator for O2 consumption and changes in RQ than the periods of physical activity. Control animals did not show any diurnal variations in O2 and RQ. Glucose, FFA and insulin were higher with CHO + FAT administration compared to AA infusion or simultaneous AA/CHO/FAT administration. In conclusion, the results suggest that simultaneous administration of a mixture of AA/CHO/FAT is preferable for whole body nitrogen economy during TPN.     

Influence of short-term protein malnutrition of mice on the phenotype and costimulatory signals of lymphocytes from spleen and Peyer's patches

The objective of this study was to investigate the impact of short-term protein malnutrition (PM) on immunoglobulin A (IgA) production and on the number and phenotype of lymphocytes in Peyer's patches (PP) and in the spleen. Balb/c mice were fed for 4, 7, or 10 d with a protein-deficient diet (0.1% protein). We determined B lymphocytes (CD40(+)), T lymphocytes (CD3(+)), T-helper (CD4(+)), and T-suppressor (CD8(+)) cells and the expression of costimulatory signals B7.1 (CD80) and B7.2 (CD86) on B cells and their counter receptors CD28 and CTLA-4 on T cells by fluorescence-activated cell-sorting analysis. Luminal IgA concentration in the small intestine was determined by an enzyme-linked immunosorbent assay. Four days of PM caused a significant reduction in the number of mononuclear cells in the spleen (5.6 x 10(7) +/- 1 x 10(7) versus 2. 4 x 10(7) +/- 0.5 x 10(7), P < 0.001) and the PP (13 x 10(6) +/- 3 x 10(6) versus 8.6 x 10(6) +/- 2 x 10(6), P < 0.01). There was a relative increase of T cells in the spleen and a relative increase of B cells in the PP. Luminal IgA content of small intestine was significantly reduced after 4 d of PM (242 +/- 55 microg versus 173 +/- 39 microg, P < 0.05) and remained at about this level until day 10 of PM. Four days after PM, the costimulatory signals B7.1 and B7. 2 on B cells were upregulated in the PP but markedly downregulated in the spleen, which was inversely related to the expression of the counter receptor CD28 on T-helper cells. We conclude that short-term PM increases the activation of B cells in the PP but reduces the relative number and activation state of splenic B cells. Only 4 d of PM caused a systemic and intestinal immunodepression, as indicated by a markedly decreased content of mononuclear cells in the PP and the spleen.     

Innate Mucosal Immune System Response of BALB/c vs C57BL/6 Mice to Injury in the Setting of Enteral and Parenteral Feeding

Background: Outbred mice exhibit increased airway and intestinal immunoglobulin A (IgA) following injury when fed normal chow, consistent with humans. Parenteral nutrition (PN) eliminates IgA increases at both sites. Inbred mice are needed for detailed immunological studies; however, specific strains have not been evaluated for this purpose. BALB/c and C57BL/6 are common inbred mouse strains but demonstrate divergent immune responses to analogous stress. This study addressed which inbred mouse strain best replicates the outbred mouse and human immune response to injury. Methods: Intravenously cannulated mice received chow or PN for 5 days and then underwent sacrifice at 0 or 8 hours following controlled surgical injury (BALB/c: n = 16–21/group; C57BL/6: n = 12–15/group). Bronchoalveolar lavage (BAL) was analyzed by enzyme‐linked immunosorbent assay for IgA, tumor necrosis factor–α (TNF‐α), interleukin (IL)–1β, and IL‐6, while small intestinal wash fluid (SIWF) was analyzed for IgA. Results: No significant increase in BAL IgA occurred following injury in chow‐ or PN‐fed BALB/c mice (chow: P = .1; PN: P = .7) despite significant increases in BAL TNF‐α and SIWF IgA (chow: 264 ± 28 vs 548 ± 37, P < .0001; PN: 150 ± 12 vs 301 ± 17, P < .0001). Injury significantly increased mucosal IgA in chow‐fed C57BL/6 mice (BAL: 149 ± 33 vs 342 ± 87, P = .01; SIWF: 236 ± 28 vs 335 ± 32, P = .006) and BAL cytokines. After injury, PN‐fed C57BL/6 mice exhibited no difference in BAL IgA (P = .9), BAL cytokines, or SIWF IgA (P = .1). Conclusions: C57BL/6 mice exhibit similar airway responses to injury as outbred mice and humans, providing an appropriate model for studying mucosal responses to injury. The BALB/c mucosal immune system responds differently to injury and does not replicate the human injury response.     

Intestinal polymeric immunoglobulin receptor is affected by type and route of nutrition

BACKGROUND: Secretory immunoglobulin A (SIgA) prevents adherence of pathogens at mucosal surfaces to prevent invasive infection. Polymeric immunoglobulin receptor (pIgR) is located on the basolateral surface of epithelial cells and binds dimeric immunoglobulin A (IgA) produced by plasma cells in the lamina propria. This IgA-pIgR complex is transported apically, where IgA is exocytosed as SIgA to the mucosal surface. Our prior work shows that mice fed intragastric (IG, an elemental diet model) and IV parenteral nutrition (PN) solution have reduced intestinal T and B cells, SIgA, and interleukin-4 (IL-4) compared with mice fed chow or a complex enteral diet (CED). Prior work also demonstrates a reduction in IgA transport to mucosal surfaces in IV PN-fed mice. Because IL-4 up-regulates pIgR production, this work studies the effects of these diets on intestinal pIgR. METHODS: Male Institute of Cancer Research (ICR) mice were randomized to chow (n = 11) with IV catheter, CED (n = 10) or IG PN (n = 11) via gastrostomy and IV PN (n = 12) for 5 days. CED and PN were isocaloric and isonitrogenous. Small intestine was harvested for pIgR and IL-4 assays after mucosal washing for IgA. IgA and IL-4 levels were analyzed by enzyme-linked immunosorbent assay and pIgR by Western blot. RESULTS: Small intestinal pIgR expression, IgA levels, and IL-4 levels decreased significantly in IV PN and IG PN groups. CONCLUSIONS: Lack of enteral stimulation affects multiple mechanisms responsible for decreased intestinal SIgA levels, including reduced T and B cells in the lamina propria, reduced Th-2 IgA-stimulating cytokines, and impaired expression of the IgA transport protein, pIgR.     

GLP-2 Prevents Intestinal Mucosal Atrophy and Improves Tissue Antioxidant Capacity in a Mouse Model of Total Parenteral Nutrition

We investigated the effects of exogenous glucagon-like peptide-2 (GLP-2) on mucosal atrophy and intestinal antioxidant capacity in a mouse model of total parenteral nutrition (TPN). Male mice (6–8 weeks old) were divided into three groups (n = 8 for each group): a control group fed a standard laboratory chow diet, and experimental TPN (received standard TPN solution) and TPN + GLP-2 groups (received TPN supplemented with 60 µg/day of GLP-2 for 5 days). Mice in the TPN group had lower body weight and reduced intestinal length, villus height, and crypt depth compared to the control group (all p < 0.05). GLP-2 supplementation increased all parameters compared to TPN only (all p < 0.05). Intestinal total superoxide dismutase activity and reduced-glutathione level in the TPN + GLP-2 group were also higher relative to the TPN group (all p < 0.05). GLP-2 administration significantly upregulated proliferating cell nuclear antigen expression and increased glucose-regulated protein (GRP78) abundance. Compared with the control and TPN + GLP-2 groups, intestinal cleaved caspase-3 was increased in the TPN group (all p < 0.05). This study shows GLP-2 reduces TPN-associated intestinal atrophy and improves tissue antioxidant capacity. This effect may be dependent on enhanced epithelial cell proliferation, reduced apoptosis, and upregulated GRP78 expression.     

Parenteral Feeding Depletes Pulmonary Lymphocyte Populations

Background: The effect of parenteral nutrition (PN) on lymphocyte mass in the lung is unknown, but reduced mucosal lymphocytes are hypothesized to play a role in the reduced immunoglobulin A–mediated immunity in both gut and lung. The ability to transfer and track cells between mice may allow study of diet‐induced mucosal immune function. The objectives of this study are to characterize lung T‐cell populations following parenteral feeding and to study distribution patterns of transferred donor lung T cells in recipient mice. Methods: In experiment 1, cannulated male Balb/c mice are randomized to receive chow or PN for 5 days. Lung lymphocytes are obtained via collagenase digestion, and flow cytometric analysis is used to identify total T (CD3+) and B (CD45/B220+) cells. In experiment 2, isolated lung T cells from chow‐fed male Balb/c mice are pooled and labeled in vitro with a fluorescent dye (carboxyfluorescein diacetate succinimidyl ester [CFSE]), and 1.1 × 10⁸ CFSE+ cells (3.1 × 10⁶ T cells) are transferred to chow‐fed Balb/c recipients. Cells recovered from recipient lungs and intestinal lamina propria (LP) are analyzed by flow cytometry to determine CFSE/CD3+ T cells at 1, 2, and 7 days. In experiment 3, cells are transferred to PN‐fed recipients. Results: In experiment 1, PN significantly decreases lung T‐ and B‐cell populations compared with chow feeding. In experiment 2, CFSE+ T‐cell retention is highest on day 1 in lung and LP, and decreases on day 2. Cells are gone by day 7; 98.1% of retained donor lung T cells migrate to recipient lungs and 1.9% to the intestine on day 1. Similar results are seen in experiment 3 after transfer of cells to PN‐fed recipients. Conclusions: PN reduces pulmonary lymphocyte populations consistent with impaired respiratory immunity. Transferred lung T cells preferentially localize to recipient lungs rather than intestine with maximal accumulation at 24 hours. Limited cross‐talk of transferred lung T cells to the intestine indicates that mucosal lymphocyte traffic might be programmed to localize to specific effector sites.     

Bombesin Improves Adaptive Immunity of the Salivary Gland During Parenteral Nutrition

Background: The parotid and submandibular salivary glands are gut‐associated lymphoid tissues (GALTs) that secrete immune compounds into the oral cavity. Parenteral nutrition (PN) without enteral stimulation decreases GALT function, including intestinal lymphocyte counts and secretory immunoglobulin A (sIgA) levels. Since the neuropeptide bombesin (BBS), a gastrin‐releasing peptide analogue, stimulates intestinal function and restores GALT parameters, we hypothesized that PN + BBS would stimulate parotid and salivary gland IgA levels, T lymphocytes, and IgA plasma cell counts compared with PN alone. Methods: Male (Institute of Cancer Research) ICR mice received intravenous catheters and were randomized to chow with saline, PN, or PN + BBS (15 µg/tid/mouse) for 5 days (8/group), 2 days after cannulation. Salivary glands were weighed and either frozen for IgA and amylase analysis or fixed for histological analysis of acinar cells, IgA+ plasma cells, and T lymphocytes. Small intestinal wash fluid was collected for IgA regression analysis with salivary glands. Results: PN reduced organ weight, acinar cell size, and amylase activity compared with chow; BBS had no significant effects on these parameters. Compared with chow, PN significantly reduced salivary gland IgA levels, IgA+ plasma cells, and T lymphocytes. PN + BBS significantly elevated IgA and restored cellularity compared with PN. Salivary gland tissue homogenate IgA levels significantly correlated with intestinal fluid IgA levels. Conclusions: Compared with chow, PN results in atrophy of the salivary glands characterized by reduced amylase, IgA, and immune cellularity. BBS has no effect on acinar cells or amylase activity compared with PN but maintains tissue IgA and plasma cells and T‐lymphocyte numbers compared with chow.