Sustained Dysbiosis and Decreased Fecal Short-Chain Fatty Acids after Traumatic Brain Injury and Impact on Neurologic Outcome.

Traumatic brain injury (TBI) alters microbial populations present in the gut, which may impact healing and tissue recovery. However, the duration and impact of these changes on outcome from TBI are unknown. Short-chain fatty acids (SCFAs), produced by bacterial fermentation of dietary fiber, are important signaling molecules in the microbiota gut-brain axis. We hypothesized that TBI would lead to a sustained reduction in SCFA producing bacteria, fecal SCFAs concentration, and administration of soluble SCFAs would improve functional outcome after TBI. Adult mice (n = 10) had the controlled cortical impact (CCI) model of TBI performed (6 m/sec, 2-mm depth, 50-msec dwell). Stool samples were collected serially until 28 days after CCI and analyzed for SCFA concentration by high-performance liquid chromatography-mass spectrometry/mass spectrometry and microbiome analyzed by 16S gene sequencing. In a separate experiment, mice (n = 10/group) were randomized 2 weeks before CCI to standard drinking water or water supplemented with the SCFAs acetate (67.5 mM), propionate (25.9 mM), and butyrate (40 mM). Morris water maze performance was assessed on post-injury Days 14-19. Alpha diversity remained stable until 72 h, at which point a decline in diversity was observed without recovery out to 28 days. The taxonomic composition of post-TBI fecal samples demonstrated depletion of bacteria from Lachnospiraceae, Ruminococcaceae, and Bacteroidaceae families, and enrichment of bacteria from the Verrucomicrobiaceae family. Analysis from paired fecal samples revealed a reduction in total SCFAs at 24 h and 28 days after TBI. Acetate, the most abundant SCFA detected in the fecal samples, was reduced at 7 days and 28 days after TBI. SCFA administration improved spatial learning after TBI versus standard drinking water. In conclusion, TBI is associated with reduced richness and diversity of commensal microbiota in the gut and a reduction in SCFAs detected in stool. Supplementation of soluble SCFAs improves spatial learning after TBI.

Effect of dietary cellulose supplementation on gut barrier function and apoptosis in a murine model of endotoxemia.

The gut plays a vital role in critical illness, and alterations in the gut structure and function have been reported in endotoxemia and sepsis models. Previously, we have demonstrated a novel link between the diet-induced alteration of the gut microbiome with cellulose and improved outcomes in sepsis. As compared to mice receiving basal fiber (BF) diet, mice that were fed a non-fermentable high fiber (HF) diet demonstrated significant improvement in survival and decreased organ injury in both cecal-ligation and puncture (CLP) and endotoxin sepsis models. To understand if the benefit conferred by HF diet extends to the gut structure and function, we hypothesized that HF diet would be associated with a reduction in sepsis-induced gut epithelial loss and permeability in mice. We demonstrate that the use of dietary cellulose decreased LPS-mediated intestinal hyperpermeability and protected the gut from apoptosis. Furthermore, we noted a significant increase in epithelial cell proliferation, as evidenced by an increase in the percentage of bromodeoxyuridine-positive cells in HF fed mice as compared to BF fed mice. Thus, the use of HF diet is a simple and effective tool that confers benefit in a murine model of sepsis, and understanding the intricate relationship between the epithelial barrier, gut microbiota, and diet will open-up additional therapeutic avenues for the treatment of gut dysfunction in critical illness.

Dietary Cellulose Supplementation Modulates the Immune Response in a Murine Endotoxemia Model.

The role of dietary fiber in chronic inflammatory disorders has been explored, but very little is known about its benefits in acute inflammation. Previously, we have demonstrated that dietary cellulose supplementation confers protection in a murine model of sepsis by promoting the growth of the gut microbiota that are linked to metabolic health. The survival benefit is associated with a decrease in serum concentration of proinflammatory cytokines, reduced neutrophil infiltration in the lungs, and diminished hepatic inflammation. Here, we aim to understand if the benefit of manipulating the gut microbiome exerts a broader "systemic" influence on the immune system in a lethal murine endotoxemia model. We hypothesize that mice-fed high-fiber cellulose (HF) diet will demonstrate a reduction in activated macrophages and dendritic cells (DCs) and a concomitant increase in the suppressive capacity of T-regulatory cells (Tregs) toward T cells responsiveness. We characterized the immunological profile and activation status of macrophages, DCs, and T cells in mice on HF diet that were then subjected to endotoxemia. Supplementation with HF diet decreased the number and activation of splenic macrophages and DCs in mice after LPS administration. Similarly, HF diet amplified the suppressive function of Tregs and induced anergy in T cells as compared with mice on a regular diet. Our data suggest that the use of HF diet can be a simple, yet effective tool that decreases the hepatic DNA-binding activity of NF-κB leading to a reduction in proinflammatory cytokine response in a murine endotoxemia model.

Dietary Supplementation With Nonfermentable Fiber Alters the Gut Microbiota and Confers Protection in Murine Models of Sepsis.

OBJECTIVES: Links between microbial alterations and systemic inflammation have been demonstrated in chronic disease, but little is known about these interactions during acute inflammation. This study investigates the effect of dietary supplementation with cellulose, a nonfermentable fiber, on the gut microbiota, inflammatory markers, and survival in two murine models of sepsis. DESIGN: Prospective experimental study. SETTING: University laboratory. SUBJECTS: Six-week-old male C57BL/6 wild-type mice. INTERVENTIONS: Mice were assigned to low-fiber, normal-fiber, or high-fiber diets with or without antibiotics for 2 weeks and then subjected to sepsis by cecal ligation and puncture or endotoxin injection. Fecal samples were collected for microbiota analyses before and after dietary interventions. MEASUREMENTS AND MAIN RESULTS: Mice that received a high-fiber diet demonstrated increased survival after cecal ligation and puncture relative to mice receiving low-fiber or normal-fiber diets. The survival benefit was associated with decreased serum concentration of pro-inflammatory cytokines, reduced neutrophil infiltration in the lungs, and diminished hepatic inflammation. The high-fiber diet also increased survival after endotoxin injection. Bacterial 16S ribosomal RNA gene sequences from each sample were amplified, sequenced, and analyzed. Fiber supplementation yielded an increase in relative abundance of the genera Akkermansia and Lachnospiraceae, taxa commonly associated with metabolic health. Administration of antibiotics to mice on the high-fiber diet negated the enrichment of Akkermansia species and the survival benefit after cecal ligation and puncture. CONCLUSION: Dietary supplementation with cellulose offers a microbe-mediated survival advantage in murine models of sepsis. Improved understanding of the link between diet, the microbiota, and systemic illness may yield new therapeutic strategies for patients with sepsis.

Pinhole imaging of 131I-metaiodobenzylguanidine (131I-MIBG) in an animal model of neuroblastoma.

PURPOSE: To evaluate (131)I-MIBG scintigraphic localization of xenotransplanted and spontaneously arising neuroblastomas in murine models of high-risk neuroblastoma. METHODS: Neuroblastoma xenografts were created by inoculation of human neuroblastoma cell suspensions into the subcutaneous flanks of athymic nude mice. In addition, spontaneous paraspinal neuroblastomas were detected by direct palpation in MYCN transgenic mice. After measured tumor volumes exceeded 200 mm(3), each mouse received an intraperitoneal injection of 18 muCi/g (131)I-metaiodobenzylguanidine ((131)I-MIBG). Pinhole scintigraphy was performed to evaluate the MIBG biodistribution and to attempt to visualize the tumors. Each mouse was imaged on a gamma camera equipped with a 3-mm pinhole on one head and an HEGP collimator on the other. RESULTS: Images demonstrated absorption of radiolabeled MIBG and visualization of tumors. Analysis of the images allowed for quantification of relative MIBG uptake and for determination of linear and area measurements of the tumors. CONCLUSION: High-energy pinhole imaging effectively demonstrates uptake of radiolabeled MIBG by human neuroblastoma tumors in murine laboratory models. This technique allows for in vivo assessment of tumor burden. In the future, we plan to use this method to evaluate sensitivity for detecting metastatic spread as well as investigating the therapeutic efficacy of high-dose (131)I-MIBG in combination with radiosensitizing agents.