Ketogenic Diet Protects from Experimental Colitis in a Mouse Model Regardless of Dietary Fat Source.

While ketogenic diets (KDs) may have potential as adjunct treatments for gastrointestinal diseases, there is little knowledge on how the fat source of these diets impacts intestinal health. The objective of this study was to investigate how the source of dietary fat of KD influences experimental colitis. We fed nine-week-old male C57BL/6J mice (n = 36) with a low-fat control diet or KD high either in saturated fatty acids (SFA-KD) or polyunsaturated linoleic acid (LA-KD) for four weeks and then induced colitis with dextran sodium sulfate (DSS). To compare the diets, we analyzed macroscopic and histological changes in the colon, intestinal permeability to fluorescein isothiocyanate-dextran (FITC-dextran), and the colonic expression of tight junction proteins and inflammatory markers. While the effects were more pronounced with LA-KD, both KDs markedly alleviated DSS-induced histological lesions. LA-KD prevented inflammation-related weight loss and the shortening of the colon, as well as preserved Il1b and Tnf expression at a healthy level. Despite no significant between-group differences in permeability to FITC-dextran, LA-KD mitigated changes in tight junction protein expression. Thus, KDs may have preventive potential against intestinal inflammation, with the level of the effect being dependent on the dietary fat source.

Randomized controlled pilot study: effect of carrageenan emulsifier on inflammation and gastrointestinal symptoms in quiescent ulcerative colitis.

Background: Animal models have provided some evidence of the pro-inflammatory effects of the commonly used emulsifier carrageenan. However, the effects of food-grade carrageenan among people with ulcerative colitis (UC) are largely unknown. Methods: A randomized, placebo-controlled cross-over study comparing high molecular carrageenan and oat-based beta-glucan preparation (placebo) among patients (n = 7) with quiescent UC was performed. Primary endpoint was Simple Clinical Colitis Activity Index (SCCAI) at the end of the treatment (7th day). Secondary analyses included biochemical biomarkers of inflammation, intestinal permeability, detoxification of intestinal lipopolysaccharide (LPS), and gastrointestinal symptoms measured by visual analog scale. Results: There were no statistically significant differences in SCCAI or any biochemical markers between carrageenan and placebo periods, nor were there any significant differences when comparing either period to baseline. Gastrointestinal symptoms were higher during the placebo period; the sum of all symptoms and borborygmi was statistically significantly higher at the end of the placebo period than at the end of the carrageenan period (20.8 ± 18.6 vs. 13.3 ± 16.4; P = 0.031, and 29.7 ± 28.6 vs. 17.9 ± 23.6; P = 0.016). Conclusions: Our study suggests that at least short-term usage of food-grade carrageenan is safe among people with UC, but given the limitations of the current study, robust human studies are still urgently needed.

Adverse effects of an aldosterone synthase (CYP11B2) inhibitor, fadrozole (FAD286), on inflamed rat colon.

Recently, we described local aldosterone production in the murine large intestine. Upregulated local aldosterone synthesis in different tissues has been linked with inflammatory conditions, which have been attenuated by the aldosterone synthase (CYP11B2) inhibitor, fadrozole (FAD286). Therefore, we investigated the effect of inhibition of intestinal aldosterone synthesis on the development of intestinal inflammation. Sprague-Dawley rats were administered 5% (v/w) dextran sodium sulphate (DSS) for 7 days with or without daily FAD286 (30 mg/kg/d) subcutaneous injections on 3 days before, during and one day after DSS. Tissue aldosterone concentrations were evaluated by ELISA, CYP11B2 by Western blot and RT-qPCR. FAD286 halved adrenal aldosterone production but, intriguingly, increased the colonic aldosterone concentration. The lack of inhibitory effect of FAD286 in the colon might have been affected by the smaller size of colonic vs. adrenal CYP11B2, as seen in Western blot. When combined with DSS, FAD286 aggravated the macroscopic and histological signs of intestinal inflammation, lowered the animals' body weight gain and increased the incidence of gastrointestinal bleeding and the permeability to iohexol in comparison to DSS-animals. To conclude, FAD286 exerted harmful effects during intestinal inflammation. Local intestinal aldosterone did not seem to play any role in the inflammatory pathogenesis occurring in the intestine.

Long-chain polyphosphates inhibit type I interferon signaling and augment LPS-induced cytokine secretion in human leukocytes.

Inorganic polyphosphates are evolutionarily conserved bioactive phosphate polymers found as various chain lengths in all living organisms. In mammals, polyphosphates play a vital role in the regulation of cellular metabolism, coagulation, and inflammation. Long-chain polyphosphates are found along with endotoxins in pathogenic gram-negative bacteria and can participate in bacterial virulence. We aimed to investigate whether exogenously administered polyphosphates modulate human leukocyte function in vitro by treating the cells with 3 different chain lengths of polyphosphates (P14, P100, and P700). The long-chain polyphosphates, P700, had a remarkable capacity to downregulate type I interferon signaling dose dependently in THP1-Dual cells while only a slight elevation could be observed in the NF-κB pathway with the highest dose of P700. P700 treatment decreased lipopolysaccharide-induced IFNß transcription and secretion, reduced STAT1 phosphorylation, and downregulated subsequent interferon-stimulated gene expression in primary human peripheral blood mononuclear cells. P700 also augmented lipopolysaccharide-induced secretion of IL-1α, IL-1ß, IL-4, IL-5, IL-10, and IFNγ. Furthermore, P700 has previously been reported to increase the phosphorylation of several intracellular signaling mediators, such as AKT, mTOR, ERK, p38, GSK3α/ß, HSP27, and JNK pathway components, which was supported by our findings. Taken together, these observations demonstrate the extensive modulatory effects P700 has on cytokine signaling and the inhibitory effects specifically targeted to type I interferon signaling in human leukocytes.

Ketogenic Diet High in Saturated Fat Promotes Colonic Claudin Expression without Changes in Intestinal Permeability to Iohexol in Healthy Mice.

Ketogenic diets (KDs) have been studied in preclinical models of intestinal diseases. However, little is known of how the fat source of these diets influences the intestinal barrier. Herein, we studied the impact of four-week feeding with KD high either in saturated fatty acids (SFA-KD) or polyunsaturated linoleic acid (LA-KD) on paracellular permeability of the intestine to iohexol in healthy male C57BL/6J mice. We investigated jejunal and colonic tight junction protein expression, histological changes, and inflammatory markers (Il1b, Il6, Tnf, and Lcn2), as well as the activity and expression of intestinal alkaline phosphatase (IAP) in feces and jejunal tissue, respectively, and plasma lipopolysaccharide. KDs did not change intestinal permeability to iohexol after two or twenty-six days of feeding regardless of fat quality. SFA-KD, but not LA-KD, upregulated the colonic expression of tight junction proteins claudin-1 and -4, as well as the activity of IAP. Both KDs resulted in increased epithelial vacuolation in jejunum, and this was pronounced in SFA-KD. Jejunal Il1ß expression was lower and colonic Il6 expression higher in LA-KD compared to SFA-KD. In colon, Tnf mRNA was increased in LA-KD when compared to controls. Overall, the results suggest that KDs do not influence intestinal permeability to iohexol but elicit changes in colonic tight junction proteins and inflammatory markers in both jejunum and colon. Future research will show whether these changes become of importance upon proinflammatory insults.

Effects of Butyrate Supplementation on Inflammation and Kidney Parameters in Type 1 Diabetes: A Randomized, Double-Blind, Placebo-Controlled Trial.

Type 1 diabetes is associated with increased intestinal inflammation and decreased abundance of butyrate-producing bacteria. We investigated the effect of butyrate on inflammation, kidney parameters, HbA1c, serum metabolites and gastrointestinal symptoms in persons with type 1 diabetes, albuminuria and intestinal inflammation. We conducted a randomized placebo-controlled, double-blind, parallel clinical study involving 53 participants randomized to 3.6 g sodium butyrate daily or placebo for 12 weeks. The primary endpoint was the change in fecal calprotectin. Additional endpoints were the change in fecal short chain fatty acids, intestinal alkaline phosphatase activity and immunoglobulins, serum lipopolysaccharide, CRP, albuminuria, kidney function, HbA1c, metabolites and gastrointestinal symptoms. The mean age was 54 ± 13 years, and the median [Q1:Q3] urinary albumin excretion was 46 [14:121] mg/g. The median fecal calprotectin in the butyrate group was 48 [26:100] µg/g at baseline, and the change was -1.0 [-20:10] µg/g; the median in the placebo group was 61 [25:139] µg/g at baseline, and the change was -12 [-95:1] µg/g. The difference between the groups was not significant (p = 0.24); neither did we find an effect of butyrate compared to placebo on the other inflammatory markers, kidney parameters, HbA1c, metabolites nor gastrointestinal symptoms. Twelve weeks of butyrate supplementation did not reduce intestinal inflammation in persons with type 1 diabetes, albuminuria and intestinal inflammation.

Renin-angiotensin system in intestinal inflammation-Angiotensin inhibitors to treat inflammatory bowel diseases?

Inflammatory bowel diseases (IBDs) are chronic disorders of the gastrointestinal tract, which manifest in recurring gastrointestinal inflammation. The current treatment options of IBD are not curative and are lacking in aspects like prevention of fibrosis. New treatment options are needed to fulfil the unmet needs and provide alternatives to drugs with resistances and side effects. Drugs targeting the renin-angiotensin system (RAS), besides being antihypertensive, also possess anti-inflammatory and antifibrotic properties and could offer an inexpensive alternative to control inflammation and fibrosis in the gut. RAS inhibitors have been effective in preventing and alleviating colitis in preclinical studies, but available human data are still sparse. This review outlines the pathophysiological functions of RAS in the gut and summarizes preclinical studies utilizing pharmacological RAS inhibitors in the treatment of experimental colitis. We discuss the alterations in intestinal RAS and the available evidence of the benefits of RAS inhibitors for IBD patients. Retrospective studies comparing IBD patients using ACE inhibitors or angiotensin II receptor blockers have provided optimistic results regarding a milder disease course and fewer hospitalizations and corticosteroid use in patients using RAS inhibitors. Prospective studies are needed to evaluate the effectiveness of these promising medications in the treatment of IBD.

Adiponectin receptor agonist AdipoRon ameliorates renal inflammation in diet-induced obese mice and endotoxin-treated human glomeruli ex vivo.

AIMS/HYPOTHESIS: Chronic low-grade inflammation with local upregulation of proinflammatory molecules plays a role in the progression of obesity-related renal injury. Reduced serum concentration of anti-inflammatory adiponectin may promote chronic inflammation. Here, we investigated the potential anti-inflammatory and renoprotective effects and mechanisms of action of AdipoRon, an adiponectin receptor agonist. METHODS: Wild-type DBA/2J mice were fed with high-fat diet (HFD) supplemented or not with AdipoRon to model obesity-induced metabolic endotoxaemia and chronic low-grade inflammation and we assessed changes in the glomerular morphology and expression of proinflammatory markers. We also treated human glomeruli ex vivo and human podocytes in vitro with AdipoRon and bacterial lipopolysaccharide (LPS), an endotoxin upregulated in obesity and diabetes, and analysed the secretion of inflammatory cytokines, activation of inflammatory signal transduction pathways, apoptosis and migration. RESULTS: In HFD-fed mice, AdipoRon attenuated renal inflammation, as demonstrated by reduced expression of glomerular activated NF-κB p65 subunit (NF-κB-p65) (70%, p < 0.001), TNFα (48%, p < 0.01), IL-1ß (51%, p < 0.001) and TGFß (46%, p < 0.001), renal IL-6 and IL-4 (21% and 20%, p < 0.05), and lowered glomerular F4/80-positive macrophage infiltration (31%, p < 0.001). In addition, AdipoRon ameliorated HFD-induced glomerular hypertrophy (12%, p < 0.001), fibronectin accumulation (50%, p < 0.01) and podocyte loss (12%, p < 0.001), and reduced podocyte foot process effacement (15%, p < 0.001) and thickening of the glomerular basement membrane (18%, p < 0.001). In cultured podocytes, AdipoRon attenuated the LPS-induced activation of the central inflammatory signalling pathways NF-κB-p65, c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38-MAPK) (30%, 36% and 22%, respectively, p < 0.001), reduced the secretion of TNFα (32%, p < 0.01), and protected against podocyte apoptosis and migration. In human glomeruli ex vivo, AdipoRon reduced the LPS-induced secretion of inflammatory cytokines IL-1ß, IL-18, IL-6 and IL-10. CONCLUSIONS/INTERPRETATION: AdipoRon attenuated the renal expression of proinflammatory cytokines in HFD-fed mice and LPS-stimulated human glomeruli, which apparently contributed to the amelioration of glomerular inflammation and injury. Mechanistically, based on assays on cultured podocytes, AdipoRon reduced LPS-induced activation of the NF-κB-p65, JNK and p38-MAPK pathways, thereby impelling the decrease in apoptosis, migration and secretion of TNFα. We conclude that the activation of the adiponectin receptor by AdipoRon is a potent strategy to attenuate endotoxaemia-associated renal inflammation.

Randomised Controlled Trial: Partial Hydrolysation of Casein Protein in Milk Decreases Gastrointestinal Symptoms in Subjects with Functional Gastrointestinal Disorders.

Unspecific gastrointestinal symptoms associated with milk consumption are common. In addition to lactose, also other components of milk may be involved. We studied whether the partial hydrolysation of milk proteins would affect gastrointestinal symptoms in subjects with functional gastrointestinal disorders. In a randomised, placebo-controlled crossover intervention, subjects (n = 41) were given ordinary or hydrolysed high-protein, lactose-free milkshakes (500 mL, 50 g protein) to be consumed daily for ten days. After a washout period of ten days, the other product was consumed for another ten days. Gastrointestinal symptoms were recorded daily during the study periods, and a validated irritable bowel syndrome-symptom severity scale (IBS-SSS) questionnaire was completed at the beginning of the study and at the end of both study periods. Blood and urine samples were analysed for markers of inflammation, intestinal permeability and immune activation. Both the IBS-SSS score (p = 0.001) and total symptom score reported daily (p = 0.002) were significantly reduced when participants consumed the hydrolysed product. Less bloating was reported during both study periods when compared with the baseline (p < 0.01 for both groups). Flatulence (p = 0.01) and heartburn (p = 0.03) decreased when consuming the hydrolysed product but not when drinking the control product. No significant differences in the levels of inflammatory markers (tumor necrosis factor alpha, TNF-α and interleukin 6, IL-6), intestinal permeability (fatty acid binding protein 2, FABP2) or immune activation (1-methylhistamine) were detected between the treatment periods. The results suggest that the partial hydrolysation of milk proteins (mainly casein) reduces subjective symptoms to some extent in subjects with functional gastrointestinal disorders. The mechanism remains to be resolved.

The use of unlicensed bone marrow-derived platelet lysate-expanded mesenchymal stromal cells in colitis: a pre-clinical study.

BACKGROUND: Mesenchymal stromal cells (MSCs) are a promising candidate for treatment of inflammatory disorders, but their efficacy in human inflammatory bowel diseases (IBDs) has been inconsistent. Comparing the results from various pre-clinical and clinical IBD studies is also challenging due to a large variation in study designs. METHODS: In this comparative pre-clinical study, we compared two administration routes and investigated the safety and feasibility of both fresh and cryopreserved platelet-lysate-expanded human bone marrow-derived MSCs without additional licensing in a dextran sodium sulfate (DSS) colitis mouse model both in the acute and regenerative phases of colitis. Body weight, macroscopic score for inflammation and colonic interleukin (IL)-1ß and tumor necrosis factor (TNF)α concentrations were determined in both phases of colitis. Additionally, histopathology was assessed and Il-1ß and Agtr1a messenger RNA (mRNA) levels and angiotensin-converting enzyme (ACE) protein levels were measured in the colon in the regenerative phase of colitis. RESULTS: Intravenously administered MSCs exhibited modest anti-inflammatory capacity in the acute phase of colitis by reducing IL-1ß protein levels in the inflamed colon. There were no clear improvements in mice treated with fresh or cryopreserved unlicensed MSCs according to weight monitoring results, histopathology and macroscopic score results. Pro-inflammatory ACE protein expression and shedding were reduced by cryopreserved MSCs in the colon. CONCLUSIONS: In conclusion, we observed a good safety profile for bone marrow-derived platelet lysate-expanded MSCs in a mouse pre-clinical colitis model, but the therapeutic effect of MSCs prepared without additional licensing (i.e. such as MSCs are administered in graft-versus-host disease) was modest in the chosen in vivo model system and limited to biochemical improvements in cytokines without a clear benefit in histopathology or body weight development.

Pilot Study: Comparison of Sourdough Wheat Bread and Yeast-Fermented Wheat Bread in Individuals with Wheat Sensitivity and Irritable Bowel Syndrome.

Many patients suspect wheat as being a major trigger of their irritable bowel syndrome (IBS) symptoms. Our aim was to evaluate whether sourdough wheat bread baked without baking improvers and using a long dough fermentation time (>12 h), would result in lower quantities of alpha-amylase/trypsin inhibitors (ATIs) and Fermentable, Oligo-, Di-, Mono-saccharides and Polyols (FODMAPs), and would be better tolerated than yeast-fermented wheat bread for subjects with IBS who have a poor subjective tolerance to wheat. The study was conducted as a randomised double-blind controlled 7-day study (n = 26). Tetrameric ATI structures were unravelled in both breads vs. baking flour, but the overall reduction in ATIs to their monomeric form was higher in the sourdough bread group. Sourdough bread was also lower in FODMAPs. However, no significant differences in gastrointestinal symptoms and markers of low-grade inflammation were found between the study breads. There were significantly more feelings of tiredness, joint symptoms, and decreased alertness when the participants ate the sourdough bread (p ≤ 0.03), but these results should be interpreted with caution. Our novel finding was that sourdough baking reduces the quantities of both ATIs and FODMAPs found in wheat. Nonetheless, the sourdough bread was not tolerated better than the yeast-fermented bread.

Local corticosterone production and angiotensin-I converting enzyme shedding in a mouse model of intestinal inflammation.

AIM: To investigate local corticosterone production and angiotensin-I converting enzyme (ACE) protein expression and their interaction in healthy and inflamed intestine. METHODS: Acute intestinal inflammation was induced to six weeks old male Balb/c mice by administration of either 3% or 5% dextran sodium sulfate (DSS) in drinking water for 7 d (n = 12 in each group). Healthy controls (n = 12) were given tap water. Corticosterone production and ACE protein shedding were measured from ex vivo incubates of the small and large intestine using EIA and ELISA, respectively. Morphological changes of the intestinal wall were assessed in hematoxylin-eosin stained tissue preparations of jejunum and distal colon. Effects of angiotensin II, captopril and metyrapone on corticosterone production was assessed by incubating pieces of small intestine of healthy mice in the presence of 0.1, 1 or 10 µmol/L angiotensin II, 1, 10 or 100 µmol/L captopril or 1, 10 or 100 µmol/L metyrapone solutions and measuring corticosterone released to the incubation buffer after 90 min (n = 5 in each group). RESULTS: Both concentrations of DSS induced inflammation and morphological changes in large intestines but not in small intestines. Changes were observed as distortions of the crypt structure, mucosal erosion, immune cell infiltration to the mucosa and submucosal edema. Ex vivo corticosterone production (2.9 ± 1.0 ng/mL vs 2.0 ± 0.8 ng/mL, P = 0.034) and ACE shedding (269.2 ± 97.1 ng/mL vs 175.7 ± 52.2 ng/mL, P = 0.016) were increased in small intestines in 3% DSS group compared to the controls. In large intestine, corticosterone production was increased compared to the controls in both 3% DSS (229 ± 81 pg/mL vs 158 ± 30 pg/mL, P = 0.017) and 5% DSS groups (366 ± 163 pg/mL vs 158 ± 30 pg/mL, P = 0.002). Large intestine ACE shedding was increased in 5% DSS group (41.5 ± 9.0 ng/mL vs 20.9 ± 5.2 ng/mL, P = 0.034). Angiotensin II treatment augmented corticosterone production in small intestine at concentration of 10 µmol/L (0.97 ± 0.21 ng/mg protein vs 0.40 ± 0.09 ng/mg protein, P = 0.036). CONCLUSION: Intestinal ACE shedding is increased by DSS-induced intestinal inflammation and parallels local corticosterone production. ACE product angiotensin II stimulates corticosterone formation in healthy intestine.