L-Arginine Pretreatment Reduces Intestinal Mucositis as Induced by 5-FU in Mice

Beneficial effects of L-arginine on immune responses and bowel function have been reported. Mucositis is a side effect of chemotherapy treatment that affects approximately 40% of patients. This complication is characterized by inflammation that affects the gastrointestinal tract, increasing permeability and causing abdominal pain, nausea, vomiting, and diarrhea, which worsen the patient's nutritional status and increases morbimortality. The aim of this study was to evaluate the effect of pretreating with 2% L-arginine supplementation in water on mucositis as induced by 5-fluorouracil (5-FU; a single dose of 200 mg/kg body weight) in Swiss male mice. The effect of L-arginine on weight, intestinal permeability, morphology, and the histopathological score of the small intestine (from 0 to 12), oxidative stress, myeloperoxidase (MPO), and N-acetylglucosaminidase (NAG) activities were evaluated. Intestinal length improvement was observed, in addition to the partial recovery of the mucosal architecture. L-arginine attenuated the histopathological score and MPO activity. There was also an improvement in intestinal permeability, despite weight loss after 5-FU administration. In conclusion, L-arginine can positively impact intestinal mucositis by promoting partial mucosal recovery, reducing inflammation and improving intestinal permeability.     

Effects of adding butyric acid to PN on gut-associated lymphoid tissue and mucosal immunoglobulin A levels

Background: Parenteral nutrition (PN) causes intestinal mucosal atrophy, gut‐associated lymphoid tissue (GALT) atrophy and dysfunction, leading to impaired mucosal immunity and increased susceptibility to infectious complications. Therefore, new PN formulations are needed to maintain mucosal immunity. Short‐chain fatty acids have been demonstrated to exert beneficial effects on the intestinal mucosa. We examined the effects of adding butyric acid to PN on GALT lymphocyte numbers, phenotypes, mucosal immunoglobulin A (IgA) levels, and intestinal morphology in mice. Methods: Male Institute of Cancer Research mice (n = 103) were randomized to receive either standard PN (S‐PN), butyric acid–supplemented PN (Bu‐PN), or ad libitum chow (control) groups. The mice were fed these respective diets for 5 days. In experiment 1, cells were isolated from Peyer's patches (PPs) to determine lymphocyte numbers and phenotypes (αβTCR⁺, γδTCR⁺, CD4⁺, CD8⁺, B220⁺ cells). IgA levels in small intestinal washings were also measured. In experiment 2, IgA levels in respiratory tract (bronchoalveolar and nasal) washings were measured. In experiment 3, small intestinal morphology was evaluated. Results: Lymphocyte yields from PPs and small intestinal, bronchoalveolar, and nasal washing IgA levels were all significantly lower in the S‐PN group than in the control group. Bu‐PN moderately, but significantly, restored PP lymphocyte numbers, as well as intestinal and bronchoalveolar IgA levels, as compared with S‐PN. Villous height and crypt depth in the small intestine were significantly decreased in the S‐PN group vs the control group, however Bu‐PN restored intestinal morphology. Conclusions: A new PN formula containing butyric acid is feasible and would ameliorate PN‐induced impairment of mucosal immunity.     

Pretreatment and Treatment With L‐Arginine Attenuate Weight Loss and Bacterial Translocation in Dextran Sulfate Sodium Colitis

Background: Imbalances in a variety of factors, including genetics, intestinal flora, and mucosal immunity, can contribute to the development of ulcerative colitis and its side effects. This study evaluated the effects of pretreatment or treatment with arginine by oral administration on intestinal permeability, bacterial translocation (BT), and mucosal intestinal damage due to colitis. Methods: C57BL/6 mice were distributed into 4 groups: standard diet and water (C: control group), standard diet and dextran sodium sulfate (DSS) solution (Col: colitis group), 2% L ‐arginine supplementation for 7 days prior to DSS administration and during disease induction (PT: pretreated group), and 2% L ‐arginine supplementation during disease induction (T: treated group). Colitis was induced by administration of 1.5% DSS for 7 days. After 14 days, intestinal permeability and BT were evaluated; colons were collected for histologic analysis and determination of cytokines; feces were collected for measurement of immunoglobulin A (IgA). Results: The Col group showed increased intestinal permeability (C vs Col: P < .05) and BT (C vs Col: P < .05). In the arginine‐supplemented groups (PT and T), this amino acid tended to decrease intestinal permeability. Arginine decreased BT to liver during PT (P < .05) and to blood, liver, spleen, and lung during T (P < .05). Histologic analysis showed that arginine preserved the intestinal mucosa and tended to decreased inflammation. Conclusions: Arginine attenuates weight loss and BT in mice with colitis.     

Intestinal Microbial Diversity and Perioperative Complications

Background and Aims: Enteral nutrient deprivation via parenteral nutrition (PN) in a mouse model leads to a local mucosal inflammatory response. This proinflammatory response leads to a loss of epithelial barrier function and atrophy of the intestine. Although the underlying mechanisms are unknown, a potential contributing factor is the impact PN has on the intestinal microbiome. We recently identified a shift in the intestinal microbial community in mice given PN; however, it is unknown whether such changes occur in humans. We hypothesized that similar microbial changes occur in humans during periods of enteral nutrient deprivation. Methods: A series of small bowel specimens were obtained from pediatric and adult patients undergoing small intestinal resection. Mucosally associated bacteria were harvested and analyzed using 454 pyrosequencing techniques. Statistical analysis of microbial diversity and differences in microbial characteristics were assessed between enterally fed and enterally deprived portions of the intestine. Occurrence of postoperative infectious and anastomotic complications was also examined. Results: Pyrosequencing demonstrated a wide variability in microbial diversity within all groups. Principal coordinate analysis demonstrated only a partial stratification of microbial communities between fed and enterally deprived groups. Interestingly, a tight correlation was identified in patients who had a low level of enteric microbial diversity and those who developed postoperative enteric‐derived infections or intestinal anastomotic disruption. Conclusions: Loss of enteral nutrients and systemic antibiotic therapy in humans is associated with a significant loss of microbial biodiversity within the small bowel mucosa. These changes were associated with a number of enteric‐derived intestinal infections and intestinal anastomotic disruptions.     

Glutamine Supplementation Decreases Intestinal Permeability and Preserves Gut Mucosa Integrity in an Experimental Mouse Model

Background: Glutamine (GLN) is the preferred fuel for enterocytes, and GLN supplementation is critical during stressful conditions. The aim of this study was to evaluate the effect of GLN on intestinal barrier permeability and bacterial translocation in a murine experimental model. Methods: Swiss male mice (25–30 g) were randomized into 3 groups: (1) sham group; (2) intestinal obstruction (IO) group; (3) IO and GLN (500 mg/kg/d) group. Two different experiments were carried out to assess intestinal permeability and bacterial translocation. In the first experiment, the animals were divided into the 3 groups described above and received diethylenetriamine pentaacetate radiolabeled with technetium (^99mTc) on the eighth day. At different time points after intestinal obstruction, blood was collected to determine radioactivity. The animals were killed, and the small intestine was removed for histological analyses. In the bacterial translocation study, on the eighth day all groups received Escherichia coli labeled with ^99mTc. After 90 minutes, the animals underwent intestinal obstruction and were killed 18 hours later. Blood, mesenteric lymph nodes, liver, spleen, and lungs were removed to determine radioactivity. Statistical significance was considered when P ≤ .05. Results: The levels of intestinal permeability and bacterial translocation were higher in the IO group than in the sham and GLN groups (P < .05). GLN decreased intestinal permeability and bacterial translocation to physiologic levels in the treated animals and preserved intestinal barrier integrity. Conclusions: GLN had a positive impact on the intestinal barrier by reducing permeability and bacterial translocation to physiologic levels and preserving mucosal integrity.     

Pretreatment with citrulline improves gut barrier after intestinal obstruction in mice

Background: Citrulline has been shown to be an important marker of gut function, regulator of protein metabolism, and precursor of arginine. The authors assessed the effects of citrulline on gut barrier integrity and bacterial translocation (BT) in mice undergoing intestinal obstruction. Methods: Mice were divided into 3 groups: sham, intestinal obstruction (IO), and citrulline (CIT). The CIT group received a diet containing 0.6% citrulline; the IO and sham groups were fed a standard chow diet. On the eighth day of treatment, all animals received a diethylenetriamine pentaacetic acid (DTPA) solution labeled with ^99mTechnetium (^99mTc‐DTPA) by gavage for the intestinal permeability study. Terminal ileum was ligated except the sham group, which only underwent laparotomy. After 4, 8, and 18 hours, blood was collected to determine radioactivity. Samples of ileum were removed 18 hours after intestinal obstruction for histological analysis. In another set of animals, BT was evaluated. Animals received 10⁸ CFU/mL of ^99mTc–Escherichia coli by gavage; 90 minutes later, they underwent ileum ligation. Intestinal fluid and serum were collected to measure sIgA and cytokines. Results: The CIT group presented decreased intestinal permeability and BT when compared with the IO group (P < .05). Histopathology showed that citrulline preserved the ileum mucosa. The sIgA concentration was higher in the CIT group (P < .05). The IO group presented the highest levels of interferon‐γ (P < .05). Conclusions: Pretreatment with citrulline was able to preserve barrier integrity and also modulated the immune response that might have affected BT decrease.     

Effect of Intestinal Atrophy and Hepatic Impairment Induced by Parenteral Nutrition on Drug Absorption and Disposition in Rats

Background: Long‐term parenteral nutrition (PN) has a high risk of hepatic dysfunction and intestinal atrophy. The present study investigated the effect of PN‐induced intestinal atrophy and hepatic impairment on drug pharmacokinetics by using 2 contrasting compounds: phenolsulfonphthalein (PSP) and cyclosporin A (CyA). Materials and Methods: PSP or CyA was administered to 7‐day PN‐fed Rats (PN rats) and sham operated rats (control rats) via intravenous (IV) or intraloop administration of the intestine. Pharmacokinetic parameters with 2‐compartment analysis including area under the concentration vs time curve (AUC) and the permeability after in situ intraloop administration (P_loop) were obtained from both concentration profiles after different administration routes. Results: After IV administration of PSP to control and PN rats, there was no notable difference in any of the pharmacokinetic parameters. In contrast, after intraloop administration, AUC and P_loop in PN rats were approximately 2.6‐ and 2.0‐fold higher than that in control rats, respectively. On the other hand, after IV administration of CyA, the terminal half‐life and total body clearance were prolonged and decreased in PN rats, respectively, resulting in 2.0‐fold increase in AUC. After intraloop administration, the AUC of PN rats was increased to approximately 1.3‐fold that of control rats, whereas no notable difference was observed in P_loop. Conclusion: The intestinal permeability of PSP was enhanced by intestinal atrophy induced by PN, while the metabolism of CyA was diminished by hepatic impairment by PN. These results revealed the physicochemical property‐based pharmacokinetic alterations during PN; for a more detailed understanding, however, further studies are needed.     

The Role of L‐Arginine and Inducible Nitric Oxide Synthase in Intestinal Permeability and Bacterial Translocation

Background: Arginine has been shown to have several immunological and trophic properties in stressful diseases. Its metabolites, nitric oxide (NO) and polyamines, are related to arginine's effects. Thus, the aim of this study was to determine the effects of the NO donor L‐arginine and the role of inducible NO synthase (iNOS) on intestinal permeability and bacterial translocation in a model of intestinal obstruction (IO) induced by a simple knot in the terminal ileum. Material and Methods: Male C57BL6/J wild‐type (WT) and iNOS knockout (iNOS–/–) mice were randomized into 6 groups: Sham and Sham–/– (standard chow), IO and IO–/– (standard chow +IO), and Arg and Arg–/– (standard chow supplemented with arginine + IO). After 7 days of treatment with standard or supplemented chows, IO was induced and intestinal permeability and bacterial translocation were evaluated. The small intestine and its contents were harvested for histopathological and morphometric analysis and the determination of polyamine concentration. Results: Pretreatment with arginine maintained intestinal permeability (P > .05; Arg and Arg–/– groups vs Sham and Sham–/– groups), increased polyamine concentration in intestinal content (P < .05; Arg vs IO group), and decreased bacterial translocation in WT animals (Arg group vs IO and IO–/– groups). Absence of iNOS also presented a protective effect on permeability but not on bacterial translocation. Conclusion: Arginine supplementation and synthesis of NO by iNOS are important factors in decreasing bacterial translocation. However, when intestinal permeability was considered, NO had a detrimental role.     

Effects of Supernatants from Escherichia coli Nissle 1917 and Faecalibacterium prausnitzii on Intestinal Epithelial Cells and a Rat Model of 5-Fluorouracil-Induced Mucositis

Faecalibacterium prausnitzii (Fp) and Escherichia coli Nissle 1917 (EcN) are probiotics, which have been reported to ameliorate certain gastrointestinal disorders. We evaluated the effects of supernatants (SN) derived from Fp and EcN on 5-fluorouracil (5-FU)-treated intestinal cells and in a rat model of mucositis. In vitro: IEC-6, Caco-2, and T-84 cells were analyzed for viability and monolayer permeability. In vivo: Female dark agouti rats were gavaged with Fp or EcN SN and injected intraperitoneally with saline (control) or 5-FU to induce mucositis. Rats were euthanized and intestinal tissues collected for myeloperoxidase assay and histological analyses. In vitro: Caco-2 cell viability was further reduced when treated with Fp SN + 5-FU compared to 5-FU controls. In both Caco-2 and T-84 cells, Fp SN partially prevented the decrease in transepithelial electrical resistance (TER) caused by 5-FU administration. In vivo: 5-FU-injected rats administered Fp SN or EcN SN partly prevented body weight loss and normalized water intake compared to 5-FU controls. These results suggest a growth inhibitory mechanism of Fp SN action on transformed epithelial cells that could be mediated by effects on tight junctions. Factors derived from Fp SN and EcN SN could have a role in reducing the severity of intestinal mucositis.     

Protective effects of bovine colostrum on non-steroidal anti-inflammatory drug induced intestinal damage in rats

The aim of this study was to examine whether bovine colostrum was able to prevent the NSAID induced small intestinal damage in animals. The animal model population of the study consisted of 4 groups: control group, diclofenac group, diclofenac with 10% low fat milk group and diclofenac with 5% colostrum group. The animals with milk or colostrum were fed with 10% low fat milk or 5% colostral solution for 5 days before the administration of diclofenac. Gut injuries were induced by administration of a single dose of diclofenac (100 mg/kg orally). Epithelial permeability values (24 hour urinary excretion of 51Cr-ethylenediaminetetraacetic acid [51Cr-EDTA]), enteric aerobic bacterial counts, serum biochemical profiles and pathologic findings of distal ileum were measured. Diclofenac caused a marked increase in the intestinal permeability, enteric bacterial numbers and intestinal villous damage, and enteric protein and albumin loss. Combined administration of bovine colostrum reduced the increase in intestinal permeability, enteric bacterial overgrowth, protein losing enteropathy and mucosal villous damage of the small intestine induced by diclofenac. Bovine colostrum may have a beneficial effect in prevention of NSAID induced small intestinal injuries.     

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.     

Glutamine Improves Innate Immunity and Prevents Bacterial Enteroinvasion During Parenteral Nutrition

Background: Patients receiving parenteral nutrition (PN) are at increased risk of infectious complications compared with enteral feeding, which is in part explained by impaired mucosal immune function during PN. Adding glutamine (GLN) to PN has improved outcome in some clinical patient groups. Although GLN improves acquired mucosal immunity, its effect on innate mucosal immunity (defensins, mucus, lysozymes) has not been investigated. Methods: Forty‐eight hours following venous cannulation, male Institute of Cancer Research mice were randomized to chow (n = 10), PN (n = 12), or PN + GLN (n = 13) for 5 days. Small intestine tissue and luminal fluid were collected for mucin 2 (MUC2), lysozyme, cryptdin 4 analysis, and luminal interleukin (IL)–4, IL‐10, and IL‐13 level measurement. Tissue was also harvested for ex vivo intestinal segment culture to assess tissue susceptibility to enteroinvasive Escherichia coli. Results: In both luminal and tissue samples, PN reduced MUC2 and lysozyme (P < .0001, respectively) compared with chow, whereas GLN addition increased MUC2 and lysozyme (luminal, P < .05; tissue, P < .0001, respectively) compared with PN alone. PN significantly suppressed cryptdin 4 expression, while GLN supplementation significantly enhanced expression. IL‐4, IL‐10, and IL‐13 decreased significantly with PN compared with chow, whereas GLN significantly increased these cytokines compared with PN. Functionally, bacterial invasion increased with PN compared with chow (P < .05), while GLN significantly decreased enteroinvasion to chow levels (P < .05). Conclusions: GLN‐supplemented PN improves innate immunity and resistance to bacterial mucosal invasion lost with PN alone. This work confirms a clinical rationale for providing glutamine for the protection of the intestinal mucosa.     

Starvation‐Induced Proximal Gut Mucosal Atrophy Diminished With Aging

Background: Starvation induces small bowel atrophy with increased intestinal epithelial apoptosis and decreased proliferation. The authors examined these parameters after starvation in aged animals. Methods: Sixty‐four 6‐week‐old and 26‐month‐old C57BL/6 mice were randomly assigned to either an ad libitum fed or fasted group. The small bowel was harvested at 12, 48, and 72 hours following starvation. Proximal gut mucosal height was measured and epithelial cells counted. Apoptosis was identified by terminal deoxynucleotidyl transferase‐mediated dUTP nick end labeling (TUNEL) staining. Proliferation was determined by immunohistochemical staining for proliferating cell nuclear antigen. Comparison of fed vs fasted and adult vs old groups was done by one‐way ANOVA with Tukey's test and unpaired Student's t test. Significance was accepted at P < .05. Results: Aged mice had higher proximal gut weights, mucosal heights, and cell numbers at baseline compared with the adult group (P < .05). The rate of apoptosis was lower in the aged (P < .05), but proliferation was not different between groups before starvation. After starvation, proximal gut wet weight decreased only in adult mice (P < .05). Gut mucosal height and mucosal cell number decreased more in adult than in aged mice (P < .05). This was related to decreased proliferation only in the adult group (P < .05). The fold of epithelial apoptosis that increased was higher in the aged group than in the adult group after starvation (P < .05). Conclusions: Gut mucosal kinetics change with age and have lower rates of apoptosis and greater mucosal mass; the character of starvation‐induced atrophy is diminished with aging.     

The dependence of the rate of BP metabolism in the rat small intestinal mucosa on the composition of the dietary fat

We studied the effects that dietary fat has on the capacity of preparations of rat small intestinal mucosal cells to metabolize benzo[a]pyrene (BP) in vitro and on the composition of fatty acids in the endoplasmic reticulum of the intestinal mucosa. When rats were fed diets containing different types of fat, there were significant changes in the incorporation of fatty acids into the endoplasmic reticulum of the mucosal cells of the small intestine: the proportions of polyunsaturated fatty acids in the endoplasmic reticulum reflected the amounts of these fatty acids in the dietary fat. The rate of BP oxidation in the intestinal mucosa was dependent on the amount and composition of the dietary fat, but the range and proportions of the metabolites produced were not affected. Dietary C_18:2 was particularly important in elevating the rate of BP oxidation, but dietary C_20:5 and C_22:6 also effectively increased the rate of BP oxidation. The rate of BP oxidation in the small intestine of rats fed different diets was positively correlated with the proportion of polyunsaturated fatty acids in the endoplasmic reticulum of the mucosal cells.     

Enteral Refeeding Rapidly Restores PN‐Induced Reduction of Hepatic Mononuclear Cell Number Through Recovery of Small Intestine and Portal Vein Blood Flows

Background: Absence of enteral nutrition (EN) reduces hepatic mononuclear cell (MNC) numbers and impairs their functions. However, enteral refeeding (ER) for as little as 12 hours following parenteral nutrition (PN) rapidly restores hepatic MNC numbers. We hypothesized that changes in small intestine and portal vein blood flows related to feeding route might be responsible for this phenomenon. Methods: In experiment 1, mice (n = 19) were randomized to Chow (n = 5), PN (n = 7) or ER (n = 7) groups. The Chow group was given chow ad libitum with intravenous (IV) saline for 5 days. The PN group was fed parenterally for 5 days, while the ER group was re‐fed with chow for 12 hours following 5 days of PN. Then, small intestine and portal vein blood flows were monitored and hepatic MNCs were isolated and counted. In experiment 2, the effects of intravenous administration of prostaglandin E₁ (PGE₁) on hepatic MNC numbers were examined in fasted mice for 12 hours. Mice (n = 28) were randomized to Control (n = 8), PG0 (n = 10), or PG1 (n = 10) groups. The Control group was fed chow ad libitum with IV saline, while the PG0 and PG1 groups were fasted for 12 hours with infusions, respectively, of saline and PGE₁ at 1μ g/kg/minute. Blood flows and hepatic MNC numbers were examined. Results: Experiment 1: ER restored PN‐induced reductions in small intestine and portal vein blood flows and hepatic MNC number to the levels in the Chow group. Small intestine and portal vein blood flows correlated positively with hepatic MNC number. Experiment 2: Fasting decreased small intestine and portal vein blood flows and hepatic MNC number. However, PGE₁ restored portal vein blood flow to the level of the Control group, and moderately increased hepatic MNC number. There was a positive correlation between portal blood flow and hepatic MNC number. Conclusions: Reduced small intestine and portal vein blood flows may contribute to impaired hepatic immunity in the absence of EN. ER quickly restores hepatic MNC number through recovery of blood flow in both the small intestine and the portal vein.     

Plasmodium chabaudi Infection Alters Intestinal Morphology and Mucosal Innate Immunity in Moderately Malnourished Mice

Plasmodium falciparum is a protozoan parasite which causes malarial disease in humans. Infections commonly occur in sub-Saharan Africa, a region with high rates of inadequate nutrient consumption resulting in malnutrition. The complex relationship between malaria and malnutrition and their effects on gut immunity and physiology are poorly understood. Here, we investigated the effect of malaria infection in the guts of moderately malnourished mice. We utilized a well-established low protein diet that is deficient in zinc and iron to induce moderate malnutrition and investigated mucosal tissue phenotype, permeability, and innate immune response in the gut. We observed that the infected moderately malnourished mice had lower parasite burden at the peak of infection, but damaged mucosal epithelial cells and high levels of FITC-Dextran concentration in the blood serum, indicating increased intestinal permeability. The small intestine in the moderately malnourished mice were also shorter after infection with malaria. This was accompanied with lower numbers of CD11b⁺ macrophages, CD11b⁺CD11c⁺ myeloid cells, and CD11c⁺ dendritic cells in large intestine. Despite the lower number of innate immune cells, macrophages in the moderately malnourished mice were highly activated as determined by MHCII expression and increased IFNγ production in the small intestine. Thus, our data suggest that malaria infection may exacerbate some of the abnormalities in the gut induced by moderate malnutrition.     

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.     

Influences of Short‐Term Fasting and Carbohydrate Supplementation on Gut Immunity and Mucosal Morphology in Mice

Background: Preoperative carbohydrate (CHO) supplementation has been recommended in enhanced recovery after surgery protocols. However, the effects of CHO supplementation on gut and systemic immunity are not well understood. Methods: Mice (n = 60) were randomized to 1 of the following 5 groups: control (ad lib feeding), 12‐hour fasting without CHO administration (fasting), and 12 hours of fasting with CHO administration at 2, 4, and 8 hours before sacrifice. Then, lymphocytes were isolated from gut‐associated lymphoid tissue, that is, Peyer's patches, the intraepithelial space, and the lamina propria of the small intestine. These lymphocyte numbers and phenotypes were evaluated. IgA levels in respiratory and small‐intestinal washings were determined by ELISA. Morphology, proliferation, and apoptosis of the intestinal epithelium were also evaluated histologically. Results: Although there were no significant differences in IgA levels among the 5 groups, fasting decreased intraepithelial and lamina propria, but not Peyer's patches lymphocyte numbers. CHO at 2 hours prevented lymphocyte loss in intraepithelial, whereas CHO at 4 hours reversed lamina propria lymphocytes numbers. Percentages of lymphocyte phenotypes were similar in each site among the 5 groups. Fasting caused villous atrophy; however, CHO at 2 hours restored villous structure along with maintenance of epithelial cell proliferation rate. Conclusions: Only 12 hours of fasting causes marked gut‐associated lymphoid tissue cell loss along with gut atrophy. However, CHO at 2 hours preserves gut immunity and morphology not completely but moderately.     

Pyruvate Is Superior to Citrate in Oral Rehydration Solution in the Protection of Intestine via Hypoxia‐Inducible Factor‐1 Activation in Rats With Burn Injury

Background: Recent studies have suggested that pyruvate‐enriched oral rehydration solution (Pyr‐ORS) may be superior to the standard bicarbonate‐based ORS in the protection of intestine from ischemic injury. The aim of this study was to compare the effects of Pyr‐ORS with citrate‐enriched ORS (Cit‐ORS) on the intestinal hypoxia‐inducible factor‐1 (HIF‐1)–erythropoietin (EPO) signaling pathway for enteral rehydration in a rat model of burn injury. Methods: Rats were randomly assigned to 4 groups (N = 20, 2 subgroups each: n = 10): scald sham (group SS), scald with no fluid resuscitation (group SN), scald and resuscitation with enteral Cit‐ORS (group SC), and scald and resuscitation with enteral Pyr‐ORS (group SP). At 2.5 and 4.5 hours after a 35% total body surface area (TBSA) scald, intestinal mucosal blood flow (IMBF), contents of HIF‐1, EPO, endothelial nitric oxide synthase (eNOS), nitric oxide (NO), barrier protein (ZO‐1), levels of serum diamine oxidase (DAO), and intestinal mucosal histology injury score were determined. Results: Serum DAO activities in the scalded groups were significantly elevated, but less raised in group SP than in group SC, at 2.5 hours and at 4.5 hours after the scald. Further, group SP more profoundly preserved intestinal HIF‐1 expression compared with group SC at the 2 time points. Compared with group SC, group SP had markedly elevated intestinal EPO, eNOS, and NO levels at the same time points, respectively (P < .05). Similarly, IMBF and ZO‐1 levels were significantly higher in group SP than in group SC. Intestinal mucosal histopathological scores were statistically higher at 2.5 hours and 4.5 hours after scalding but were more attenuated in group SP than in group SC (P < .05). Immunofluorescence expression of intestinal mucosal ZO‐1 was consistent with the above changes. The above parameters were also significantly different between groups SC and SN (all P < .05). Conclusion: Pyr‐ORS provides a superior option to Cit‐ORS for the preservation of intestinal blood flow and barrier function and the attenuation of histopathological alterations in enteral resuscitation of rats with burn injury. Its underlying mechanism may be closely related to the pyruvate in activation of intestinal HIF‐1‐EPO signaling cascades.     

Influences of long-term antibiotic administration on Peyer's patch lymphocytes and mucosal immunoglobulin A levels in a mouse model

BACKGROUND: Long-term antibiotic administration is sometimes necessary to control bacterial infections during the perioperative period. However, antibiotic administration may alter gut bacterial flora, possibly impairing gut mucosal immunity. We hypothesized that 1 week of subcutaneous (SC) antibiotic injections would affect Peyer's patch (PP) lymphocyte numbers and phenotypes, as well as mucosal immunoglobulin A (IgA) levels. METHODS: Sixty-one male Institute of Cancer Research mice were randomized to CMZ (cefmetazole 100 mg/kg, administered SC twice a day), IPM (imipenem/cilastatin 50 mg/kg x 2), and control (saline 0.1 mL x 2) groups. After 7 days of treatment, the mice were killed and their small intestines removed. Bacterial numbers in the small intestine were determined using sheep blood agar plates under aerobic conditions (n = 21). PP lymphocytes were isolated to determine cell numbers and phenotypes (CD4, CD8, alphabetaTCR, gammadeltaTCR, B220; n = 40). IgA levels in the small intestinal and bronchoalveolar washings were also measured with ELISA. RESULTS: Antibiotic administration decreased both bacterial number and the PP cell yield compared with the control group. There were no significant differences in either phenotype percentages or IgA levels at any mucosal sites among the 3 groups. CONCLUSIONS: Long-term antibiotic treatment reduces PP cell numbers while decreasing bacterial numbers in the small intestine. It may be important to recognize changes in gut mucosal immunity during long-term antibiotic administration.