Introduction

Ulcerative colitis (UC) is a chronic intestinal inflammatory disease mainly involving the mucosa and submucosa. The pathogenesis of UC has not been clearly elucidated and there are limitations of existing treatment measures. Dietary factors have been implicated closely related to the pathogenesis and clinical outcome of UC. Our team in the early clinical trials observed that UC patients had aggravation or recurrence of their disease after taking in lamb. However, the relationship between them is unclear. Does lamb aggravate the intestinal inflammation of UC patients? What is the mechanism if lamb has a negative impact on intestinal inflammation? It is urgent for us to study further.

Aims & Methods

We aimed to clarify the effect of lamb on intestinal inflammation in UC mice and to reveal the mechanism, so as to provide scientific basis for clinical dietary guidance for UC patients.
Mice were given 2.5% DSS (w/v) solution for 5 days and deionized water for 7 days for 1 cycle, repeated 3 cycles to establish the chronic UC model. Take the start of giving DSS solution as day1, corresponding measures of lamb suspensions at a dose of 0.25, 0.50, 1.00 g/mL or sterile water were administered for 31 days. Then the general condition and the severity of colitis were monitored. Colon tissues were obtained to assess the degree of pathological damage by HE staining and detect the levels of key factors of intestinal barrier by RT-qPCR. Spleen tissues were collected to detect the levels of M1/M2 macrophages and related factors by flow cytometry and RT-qPCR.

Results

Our results showed that lamb aggravated intestinal inflammation of UC mice, as indicated by higher weight loss and DAI, shorter colon and more severe pathological damage. Also, the colonic mechanical barrier-associated factors including ZO-1, Claudin-1, Occludin, E-cadherin mRNA and colonic mucosal barrier associated-factors such as Muc1, Muc2, Muc3 mRNA showed significant decrease in lamb-treated colitis mice. Moreover, colitis mice with lamb treatment showed significant increased M1 macrophages and reduced M2 macrophages in the spleen, leading to an increased the ratio of M1/M2 macrophage. Meanwhile, the M1 macrophage-associated inflammatory factors gene TNF-α, IL-6 mRNA increased and M2 macrophage-associated inflammatory factors gene IL-10, TGF-β mRNA decreased significantly in lamb-treated colitis mice.

Conclusion

Our results show that lamb aggravates the intestinal inflammation and damages the mucosal barrier of colitis mice, whose mechanism is related to the promotion of macrophages to M1 cells and inhibition M2 cells differentiation, thereby aggravating the imbalance of M1/M2 macrophages.

Introduction

Mucosa protective effects can be mediated by physical as well as by anti-inflammatory mechanisms of action. For the root extract of marshmallow (Althaea officinalis L., REAo), known to cure mild irritation of stomach/gut mucosa [1], only a physical action had been considered. Recent studies have shown that it also induces regeneration of irritated mucosa [2,3]. Mucosal wound healing depends on the recruitment of several cell types, e.g., endothelial cells, fibroblasts, and macrophages (MΦ). Reactive oxygen species (ROS)-induced damage is associated with mucosal infiltration of activated leukocytes, such as MΦ, which produce excessive ROS and pro-inflammatory cytokines that may overwhelm the antioxidant and immune defenses of the cells and exacerbate mucosal inflammation. Additionally, fibroblasts exhibit a metabolic adaptation to control inflammation-related oxidative stress.

Aims & Methods

We aimed to investigate whether REAo has, on top of its physical effects, also anti-inflammatory/-oxidative properties in human dermal fibroblasts (NHDF), human umbilical vein endothelial cells (HUVEC), and human acute monocytic leukemia (THP-1) differentiated MΦ, critical cellular components of the gastrointestinal and oral mucosa.

The cells were pre-treated with REAo or its final formulation STW 42, containing REAo and selected excipients, followed by incubation with H₂O (pro-oxidative) or lipopolysaccharide (LPS, pro-inflammatory). Intracellular ROS were quantified with DCFDA. The releases of IL-6 and TNF-α were determined by ELISA. THP-1 cells were differentiated into MΦ with 0.1 μg/ml phorbol-12-myristate-13-acetatefor 3–5 days.

Results

Pre-treatment (24 h) of HUVEC, NHDF or THP-1-MΦ with STW 42 or REAo (100-1000 μg/ml) significantly inhibited the H₂O-induced intracellular ROS production by 30.0% to 58.7%. Whereas 24h(HUVEC, NHDF) or 48 h (THP-1-MΦ) pre-treatment with STW 42 or REAo (100-1000 μg/ml) inhibited the LPS-activated IL6 release by 25.0-67.0%. Additionally, 48 h pre-treatment of THP-1-MΦ with 50-500 μg/ml STW 42 or REAo significantly inhibited LPS-induced TNF-α release by 21.5% (50 μg/ml) to 52.0% (500 μg/ml), in comparison with LPS-treated MΦ. The observed effects of STW 42 or REAo were similar to 25-100 μM Diclofenac, which was used as an anti-inflammatory control.

Conclusion

These anti-inflammatory and antioxidant properties may support the benefit of STW 42 and its constituent REAo in patients with mucosal irritation with subsequent irritative cough or gastric complaints due to its protective/reparative effects on the mucosa.

References

1. HMPC of EMA 2016. European Union herbal monograph on Althaea officinalis L., radix. EMA/HMPC/436679/2015
2. Deters A, Zippel J, Hellenbrand N, Pappai D, Possemeyer C, Hensel A. 2010. Aqueous extracts and polysaccharides from Marshmallow roots (Althea officinalis L.): Cellular internalisation and stimulation of cell physiology of human epithelial cells in vitro. J Ethnopharmacol. 127: 62-9. doi: 10.1016/j.jep.2009.09.050.
3. Bonaterra GA, Schmitt J, Schneider K, Schwarzbach H, Aziz-Kalbhenn H, Kelber O, Müller J, Kinscherf R. Phytohustil^® and root extract of Althaea officinalis L. exert anti-inflammatory and anti-oxidative properties and improve the migratory capacity of endothelial cells in vitro. Front Pharmacol. 2022 Dec 8;13:948248.
   

Disclosure

JS and NS have nothing to disclose. GAB and RK received project support from Steigerwald Arzneimittelwerk GmbH, Darmstadt, Germany. OK, HAK and KB are employees of Steigerwald Arzneimittelwerk GmbH, Darmstadt, Germany.

Introduction

Maintaining intestinal homeostasis relies on the interplay between individuals and their unique gut microbiota. Disruptions may result in chronic inflammation and Inflammatory Bowel Disease (IBD) [1]. This study delves into this interplay, emphasizing the characterization of diet-derived microbiome-associated metabolites and prioritizing them based on their effects on epithelial barrier integrity and inflammatory status.

Aims & Methods

Diet-derived microbiome-associated metabolites – epigallocatechin-gallate, gallic acid (GA), protocatechuic acid (PCA), ellagic acid (EA), and urolithins (Uro-A, B, C, D) – were selected through extensive literature research. In the commercial colon cell model (Caco-2) and 3D epithelial organoids from primary adult intestinal-epithelial stem cells, inflammation was induced with TNF-alpha and IFN-gamma (25ng/ml and each) to mimic IBD conditions. The expression of inflammatory cytokines genes (TNF-alpha, CXCL-9, MCP-1, IL-8) and epithelial barrier integrity markers (ZO-1 and OCLN) were evaluated by qRT-PCR. Epithelial barrier integrity was visualized using FITC-dextran assay.

Results

The induction of inflammation significantly increased in key inflammatory markers by an average 7.5-fold change (±1.8SD) in Caco-2 and 3D IBD organoid models. Metabolite analysis in Caco-2 cells showed that GA exhibited a 1.24-fold increase in TNF-alpha (p=0.036) and a -7.22-fold decrease in MCP-1 (p=0.049) gene expressions. EA decreased the expression of TNF-alpha and MCP-1 genes by -1.59-fold (p=0.045) and -3.59-fold (p=0.036) respectively, whereas IL-8 was reduced by -7.62-fold (p=0.01). Uro-A uniquely decreased MCP-1 gene expression by -7.16-fold (p=0.004) but increased IL-8 by 8-fold (p=0.01). PCA demonstrated a statistically significant decrease in all tested inflammatory markers (p<0.05). Analysis of 3D IBD organoids highlighted unique effects in restoring the epithelial barrier and expression of inflammatory marker genes.

Conclusion

Our findings demonstrate the anti-inflammatory properties of gallic acid, ellagic acid, protocatechuic acid, and urolithin-A in both Caco-2 and 3D organoid models. This research elucidates the complex interrelationships within the nutrition-microbiome-metabolome axis and its consequential impact on gut health.

References

1. Zheng, D., Liwinski, T. & Elinav, E. Interaction between microbiota and immunity in health and disease. Cell Res 30, 492–506 (2020). https://doi.org/10.1038/s41422-020-0332-7

Disclosure

No conflict of interest to declare.

Introduction

Type 1 diabetes is a disease of the endocrine pancreas, but it also affects the exocrine part, especially the ductal cells. Several studies have shown that secretin-induced ductal HCO₃^- secretion is reduced in diabetes, however, the underlying mechanism and the effect of diabetes on basal secretion is less known.

Aims & Methods

Our aim is to investigate the effect of diabetes on pancreatic exocrine function. Diabetes was induced in wild type and cystic fibrosis transmembrane conductance regulator (CFTR) knock out mice by i.p. administration of streptozotocin and disease development was confirmed by fasting blood glucose level measurement. Pancreatic ductal fluid and HCO₃^- secretion were measured by fluid secretion measurements and fluorescence microscopy, respectively. Expression of ion transporters were investigated by real-time PCR and immunohistochemistry, whereas TEM was used for the morphological characterization of the pancreas.

Results

​Basal fluid and HCO₃^- secretion are significantly elevated in diabetes. Acute or chronic glucose treatment did not affect HCO₃^- secretion, but inhibition of CFTR significantly reduced it in both normal and diabetic mice. Cl^- efflux and the expression of CFTR, ANO1, NHE-1 and AQP1 increased in diabetes. Secretin-stimulated fluid secretion was also significantly higher in diabetic mice.

Conclusion

Our results show that diabetes increases fluid and HCO₃^- secretion in ductal cells both under basal and stimulated conditions in which the increased function of ion and water transporters, especially CFTR, plays central role.
This study was supported by the National Research, Development and Innovation Office Grant (SNN134497 to VV) and the CFRD-SRC Grant (No.: SRC 007).

Introduction

Current IBD therapy regimens are still frequently inefficient, can exert severe side effects, and loss of treatment response is common. Thus, the development of novel therapeutic approaches is an unmet clinical need – especially for patients featuring a primary non-response, refractory disease and/or severe side effects. Here we assessed novel small molecule inhibitors targeting a4b7 integrin or IRAK4 regarding their efficacy to relieve colitis in a model of spontaneous intestinal inflammation either as a stand-alone treatment or in combination.

Aims & Methods

IL-10 knockout (IL-10KO) mice, which develop spontaneous immune-driven intestinal inflammation resembling human IBD, were used for all studies. Mice with established disease were fed for three weeks ad libitum with control chow or chow containing the inhibitors at two previously defined concentrations: a higher concentration intended to induce optimal treatment response or a lower, sub-optimal concentration that did not induce therapeutic effect as stand-alone therapy.

Results

When administered as stand-alone treatment at the higher dose, a4b7 and IRAK4 inhibitors both significantly reduced colitis severity (reduced endoscopic and histological colitis scores [p<0.001 and p<0.01], reduced disease activity index scores [p<0.001]). This effect was accompanied by a normalization of immune cell populations in the colonic lamina propria. Particularly, we found reduced numbers of granulocytes, Th1, Th17, and cytotoxic T cell sin a4b7 and IRAK4 treated mice. Furthermore, mRNA and protein levels of inflammatory cytokines (IFNg, IL17, TNFa and IL-1b) were normalized in intestinal tissue of a4b7 and IRAK4 inhibitor-treated mice (p<0.01, each). When administered in lower concentrations, however, the a4b7 and IRAK4 inhibitors were not able to reduce disease activity as a stand-alone treatment. Of note, a combination of the low doses of the a4b7 inhibitor and the IRAK4 inhibitor significantly reduced the disease severity as evidenced by reduced endoscopic and histologic colitis scores [p<0.001, each], reduced disease activity scores [p<0.01] and a normalization of immune cell infiltrates (reduced levels of granulocytes [p<0.001], Th1 cells [p<0.001], Th17 cells [p<0.001] and less IFNg+ CD8 T cells [p<0.001]). Furthermore, while low doses of anti-IL12/23 p40 antibody were not able to reduce disease severity as a stand-alone treatment, combination of low-dose anti-IL12/23 p40 with the low dose of the a4b7 inhibitor resulted in amelioration of colitis (normalized endoscopic [p<0.001] and histologic [p<0.01] colitis scores, reduced infiltration of granulocytes, Th1, Th17 and IFNg+ CD8 T cells [p<0.01, each], normalization of IFNg, TNFa and IL-1b protein and mRNA levels in the intestine).

Conclusion

Our results demonstrate that small molecule inhibitors for a4b7 and IRAK4 were efficient in reducing colitis in a model of spontaneous immune-cell driven intestinal inflammation. Combination of the a4b7 inhibitor with low doses of the IRAK4 inhibitor or sub-optimal doses of an anti-IL12/IL23 antibody was effective to relieve the disease in the IL-10-KO model. This indicates that combination treatment of a4b7 inhibition with other immune-modulatory drugs, such as IL12/23 neutralization or IRAK4 inhibition might be a promising approach for patients that do not respond to single therapies. Furthermore, our results indicate that combination therapy is able to induce significant disease-relieving effects when the drugs are administered in low concentrations what could minimize side effects without interfering with treatment response.

Clinical Case Summary

Crohn’s disease (CD) is a multi-factorial immune-mediated inflammatory condition of the gastrointestinal tract (GIT). While CD-related granulomas typically occur within the GIT, they can appear in other areas such as lymph nodes, mesentery, peritoneum, liver, and lungs. This case highlights a rare instance of renal granuloma secondary to CD and discusses an effective treatment approach. An 18-year-old male presented to a GP with a one-year history of abdominal pain. There was no significant medical or family history. Physical examination was unremarkable. Laboratory findings showed raised white cell count, elevated erythrocyte sediment rate, and renal dysfunction. He was referred to a nephrologist who noted additional symptoms of diarrhoea and occasional per-rectal bleeding with no significant improvement in symptoms or renal function after complete cessation of non-steroidal anti-inflammatory drugs. Renal biopsy confirmed acute interstitial nephritis and granuloma, therefore he was referred to a gastroenterologist to rule out CD or sarcoid. Colonoscopy showed pancolitis with rectal sparing and segmental biopsies revealed chronic colitis with granulomas confirming CD. Fecal calprotectin was raised. Further investigations ruled out autoimmune disease, tuberculosis and sarcoidosis as causes for renal impairment. Treatment involved steroid tapering followed by intravenous infliximab induction and maintenance therapy with subcutaneous injections fortnightly. Renal function and GIT symptoms improved with infliximab monotherapy, though dose escalation to weekly infliximab injections was required at seven months due to loss of response from subcutaneous infliximab. At one year, colonoscopy showed endoscopic remission, with ongoing improvement in renal function (creatinine 194 to 111 umol/L) and resolution of pyuria. In conclusion, we present a rare case of CD-associated interstitial nephritis, showing a positive clinical response to infliximab monotherapy.

References

1. Lamb CA, Kennedy NA, Raine T, et al. British Society of Gastroenterology consensus guidelines on the management of inflammatory bowel disease in adults. Gut. 2019;68:s1-s106.
2. Eade MN, Cooke WT, Williams JA. Liver disease in Crohn's disease. Scand J Gastroenterol. 1971;6:199-204.
3. Whitehead R. Pathology of Crohn's disease. In: Kirsner JB, Shorter RG, eds. Inflammatory bowel disease. 2nd ed. Philadelphia: Lean and Febiger; 1980:296-310.

Disclosure

Nil