The in vitro assessment of the bioavailability of iron in New Zealand beef : a thesis presented in partial fulfillment of the requirements for the degree of Master of Science in Physiology at Massey University, Palmerston North, New Zealand /

The bioavailability of iron in New Zealand beef either alone or as part of a 'typical' New Zealand meal was investigated. The solubility of iron and its in vitro absorption by mouse intestinal tissue were used to evaluate iron bioavailability. The solubility of haem and/or non-haem iron in meat (beef longissimus muscle), vegetables and meat-plus-vegetables was investigated. Samples were cooked and then subjected to in vitro gastrointestinal digestion with pepsin followed by a combination of pancreatic enzymes and bile. Cooking at 65°C for 90 minutes reduced the soluble iron concentration in meat by 81% and reduced the haem iron concentration by 27%, which coincided with a 175% increase in non-haem iron concentrations. However, gastrointestinal digestion increased the solubility of iron in cooked meat (333%), vegetables (367%) and meat-plus-vegetables (167%). A proportion (35%) of the haem iron in the meat was broken down by the action of pancreatic enzymes leading to a 46% increase in non-haem iron concentrations, although this was not the case for the meat-plus-vegetables. Validation studies showed that mouse intestinal segments mounted in Ussing chambers maintained integrity and viability, and were responsive to glucose, theophylline and carbachol. Intestinal tissue from iron deficient mice was then used in the Ussing chambers to investigate the absorption of iron from ferrous gluconate and the soluble fractions of meat, vegetables and meat-plus-vegetables after gastrointestinal digestion. Results indicated a trend towards a higher absorption of iron from meat and ferrous gluconate, compared to vegetables and meat-plus-vegetables. However, iron absorption results were difficult to interpret due to the wide variation in the data. This variation was possibly due to errors associated with the sample processing and the analysis of iron, which was by inductively coupled-mass spectroscopy. Overall, the present study showed that before estimations can be made on the bioavailability of food iron, the effects of the cooking and gastrointestinal digestion processes must be considered. Further, the use of in vitro gastrointestinal digestion followed by the use of Ussing chambers to assess intestinal absorption is a potentially valuable system for assessing mineral bioavailability

Oral serum-derived bovine immunoglobulin improves duodenal immune reconstitution and absorption function in patients with HIV enteropathy.

ObjectivesTo examine the impact of serum-derived bovine immunoglobulin, an oral medical food known to neutralize bacterial antigen and reduce intestinal inflammation, on restoration of mucosal immunity and gastrointestinal function in individuals with HIV enteropathy.DesignOpen-label trial with intensive 8-week phase of bovine serum immunoglobulin (SBI) 2.5 g twice daily with a 4-week washout period and an optional 9-month extension study.MethodsHIV enteropathy was defined as chronic gastrointestinal symptoms including frequent loose or watery stools despite no identifiable, reversible cause. Upper endoscopy for tissue immunofluorescent antibody assay and disaccharide gut permeability/absorption studies were performed before and after 8 weeks of SBI to test mucosal immunity and gastrointestinal function. Blood was collected for markers of microbial translocation, inflammation, and collagen kinetics. A validated gastrointestinal questionnaire assessed changes in symptoms.ResultsAll eight participants experienced profound improvement in symptoms with reduced bowel movements/day (P = 0.008) and improvements in stool consistency (P = 0.008). Gut permeability was normal before and after the intervention, but D-xylose absorption increased in seven of eight participants. Mucosal CD4 lymphocyte densities increased by a median of 139.5 cells/mm2 from 213 to 322 cells/mm2 (P = 0.016). Intestinal-fatty acid binding protein (I-FABP), a marker of enterocyte damage, initially rose in seven of eight participants after 8 weeks (P = 0.039), and then fell below baseline in four of five who continued receiving SBI (P = 0.12). Baseline serum I-FABP levels were negatively correlated with subsequent rise in mucosal CD4 lymphocyte densities (r = -0.74, P = 0.046).ConclusionSBI significantly increases intestinal mucosal CD4 lymphocyte counts, improves duodenal function, and showed evidence of promoting intestinal repair in the setting of HIV enteropathy

Review article: pathogenesis and clinical manifestations of gastrointestinal involvement in systemic sclerosis.

BACKGROUND: Gastrointestinal tract (GIT) involvement is a common cause of debilitating symptoms in patients with systemic sclerosis (SSc). There are no disease modifying therapies for this condition and the treatment remains symptomatic, largely owing to the lack of a clear understanding of its pathogenesis. AIMS: To investigate novel aspects of the pathogenesis of gastrointestinal involvement in SSc. To summarise existing knowledge regarding the cardinal clinical gastrointestinal manifestations of SSc and its pathogenesis, emphasising recent investigations that may be valuable in identifying potentially novel therapeutic targets. METHODS: Electronic (PubMed/Medline) and manual Google search. RESULTS: The GIT is the most common internal organ involved in SSc. Any part of the GIT from the mouth to the anus can be affected. There is substantial variability in clinical manifestations and disease course and symptoms are nonspecific and overlapping for a particular anatomical site. Gastrointestinal involvement can occur in the absence of cutaneous disease. Up to 8% of SSc patients develop severe GIT symptoms. This subset of patients display increased mortality with only 15% survival at 9 years. Dysmotiity of the GIT causes the majority of symptoms. Recent investigations have identified a novel mechanism in the pathogenesis of GIT dysmotility mediated by functional anti-muscarinic receptor autoantibodies. CONCLUSIONS: Despite extensive investigation, the pathogenesis of gastrointestinal involvement in systemic sclerosis remains elusive. Although treatment currently remains symptomatic, an improved understanding of novel pathogenic mechanisms may allow the development of potentially highly effective approaches including intravenous immunoglobulin and microRNA based therapeutic interventions

Unique Contractile and Structural Protein Expression in Dog Ileal Inner Circular Smooth Muscle

This study was designed to test the hypothesis that there is heterogeneous expression of contractile and structural proteins between the smooth muscle cells (SMCs) in the inner and outer circular muscle (ICM and OCM) layers of the ileum. Immunohistochemical staining and quantitation of fresh frozen sections of the dog ileum was performed using protein specific antibodies. Smooth muscle (SM) SMA myosin heavy chain (MHC), α- and γ-SM actin, and vinculin all show greater expression in the ICM relative to the OCM. SMB MHC and fibronectin show the opposite pattern, with greater expression in the OCM relative to the ICM. Differences in expression of these proteins are consistent with proposed differences in function of these muscle layers. Hypotheses regarding muscle tone and the coordination and regulation of peristalsis via these different muscle layers based on this data can now be made and tested

Mesenteric Vascular Dysregulation and Intestinal Inflammation Accompanies Experimental Spinal Cord Injury

Cervical and high thoracic spinal cord injury (SCI) drastically impairs autonomic nervous system function. Individuals with SCI at thoracic spinal-level 5 (T5) or higher often present cardiovascular disorders that include resting systemic arterial hypotension. Gastrointestinal (GI) tissues are critically dependent upon adequate blood flow and even brief periods of visceral hypoxia triggers GI dysmotility. The aim of this study was to test the hypothesis that T3-SCI induces visceral hypoperfusion, diminished postprandial vascular reflexes and concomitant visceral inflammation. We measured in vivo systemic arterial blood pressure and superior mesenteric artery (SMA) and duodenal blood flow in anesthetized T3-SCI rats at 3 days and 3 weeks post-injury either fasted or following enteral feeding of a liquid mixed-nutrient meal (Ensure™). In separate cohorts of fasted T3-SCI rats, markers of intestinal inflammation were assayed by qRT-PCR. Our results show that T3-SCI rats displayed significantly reduced SMA blood flow under all experimental conditions (p\u3c0.05). Specifically, the anticipated elevation of SMA blood flow in response to duodenal nutrient infusion (postprandial hyperemia) was either delayed or absent after T3-SCI. The dysregulated SMA blood flow in acutely-injured T3-SCI rats coincides with abnormal intestinal morphology and elevation of inflammatory markers, all of which resolve after 3 weeks. Specifically, Icam1, Ccl2 (MCP-1) and Ccl3 (MIP-1α) were acutely elevated following T3-SCI. Our data suggest that arterial hypotension diminishes mesenteric blood flow necessary to meet mucosal demands at rest and during digestion. The resulting GI ischemia and low-grade inflammation may be an underlying pathology leading to GI dysfunction seen following acute T3-SCI

Feed interventions and skatole deposition

Skatole produced in the large intestine of the pig and the testicular steroid androstenone are the main substances contributing to boar tainted meat from entire male pigs. Boar taint decreases the quality of the meat and is not accepted by consumers. Until now boar taint has been avoided by castrating male pigs. Surgical castration reduces lean meat percentage, growth rate and feed efficiency, and it causes pain to the animal. This constitutes a problem in relation to productivity and welfare. Different attempts on avoiding surgical castration were either not fully effective, not accepted by the market, or they have a long time horizon for implementation. However, when focusing on the effect of feed interventions on boar taint, previous studies have showed a reducing effect through reduced skatole production in the large intestine after a one week application period. Skatole is produced from the microbial fermentation of L-tryptophan in the large intestine. In the literature it is well documented that skatole production in the large intestine is positively correlated with skatole deposition in adipose tissue. Moreover skatole production can be decreased by adding non-digestible and easy fermentable carbohydrates to the feed. However, little is known about skatole producing bacteria from the large intestine of pigs, and how these bacteria can be affected. This thesis reviews skatole production and metabolism in the pig, and how skatole production can be reduced by affecting the microbial production of skatole in the large intestine. A skatole producing bacterium SK9 K4 was isolated from the gastrointestinal tract of pigs. No such bacterium has previously been described. The bacterium was characterised by 16S RNA sequencing, gram stain, analysis of DNA G-C content, cellular fatty acids composition and DNA hybridisation with closely related bacteria. The fermentation of different carbohydrate sources, the growth pattern, the production of organic acids and the skatole production were studied in vitro. The production of skatole in the large intestine was correlated with skatole deposition in adipose tissue. Skatole production could be reduced when adding a minimum of 20 % raw potato starch or 9 % inulin to the feed. The problem concerning deposition of skatole in adipose tissue seems to be solved through the introduction of feed interventions. However high concentrations of androstenone deposited in adipose tissue remains a challenge. Thus, the feed interventions were not fully effective against boar taint. SK9 K4 was described as cells being strictly anaerobic, occurred singly or in pairs and were gram positive. It was identical with an Olsenella sp. strain isolated from the rumen, an uncultured Olsenella sp. clone isolated from sludge and an uncultured bacterium colon isolated from the oral cavity. Moreover SK9 K4 was closely related to Olsenella uli, Olsenella profusa, Olsenella umbonata and Atopobium parvulum. SK9 K4 and O. uli produced skatole from idole-3-acetic acid but not from L-tryptophan. The major fermentation products were lactic acid together with acetic acid and formic acid. SK9 K4 was not able to ferment raw potato starch, inulin and raw corn starch. Thus, when feeding resistant starch or inulin, the growth of skatole producing bacteria might be reduced followed by a reduced ability to produce skatole. The characterisation of a skatole producing bacterium isolated from the gastrointestinal tract of pigs gives the opportunity to further study the bacterium in vivo. Studies should be conducted to investigate the effect of a control diet compared to a diet added a non-digestible and easy fermentable carbohydrate on the growth of SK9 K4 in the large intestine of the pig

Predominance of weakly cytotoxic, T-betLowEomesNeg CD8+ T-cells in human gastrointestinal mucosa: implications for HIV infection.

The gastrointestinal mucosa is an important site of HIV acquisition, viral replication, and pathogenesis. Immune cells in mucosal tissues frequently differ in phenotype and function from their non-mucosal counterparts. Although perforin-mediated cytotoxicity as measured in blood is a recognized correlate of HIV immune control, its role in gastrointestinal tissues is unknown. We sought to elucidate the cytotoxic features of rectal mucosal CD8+ T-cells in HIV infected and uninfected subjects. Perforin expression and lytic capacity were significantly reduced in rectal CD8+ T-cells compared with their blood counterparts, regardless of HIV clinical status; granzyme B (GrzB) was reduced to a lesser extent. Mucosal perforin and GrzB expression were higher in participants not on antiretroviral therapy compared with those on therapy and controls. Reduction in perforin and GrzB was not explained by differences in memory/effector subsets. Expression of T-bet and Eomesodermin was significantly lower in gut CD8+ T-cells compared with blood, and in vitro neutralization of TGF-β partially restored perforin expression in gut CD8+ T-cells. These findings suggest that rectal CD8+ T-cells are primarily non-cytotoxic, and phenotypically shaped by the tissue microenvironment. Further elucidation of rectal immune responses to HIV will inform the development of vaccines and immunotherapies targeted to mucosal tissues