Dextran Sulfate Sodium Colitis Research Articles

Effects of repeated intragastric administrations with heat-inactivated Mycobacterium aurum DSM 33539 on the stress-induced aggravation of dextran sulfate sodium (DSS) colitis in C57BL/6N mice

Stress-protective effects have been reported for M. vaccae NCTC 11659 and M. vaccae ATCC 15483T. However, it remains to be investigated whether also closely related rapidly growing environmental saprophytic non-tuberculous mycobacteria (NTM) species have protective effects against the negative consequences of chronic psychosocial stress. Therefore, the aim of the current study was to assess whether repeated i.g. administrations of a heat-inactivated preparation of Mycobacterium aurum DSM 33539 prior to 19 days of chronic subordinate colony housing (CSC) are able to ameliorate the negative effects of this preclinically validated mouse model for chronic psychosocial stress on subsequent dextran sulfate sodium (DSS) colitis in male C57BL/6N mice. The results of the present study show that repeated i.g. administrations of M. aurum DSM 33539 have stabilizing effects on the composition of the gut microbiome, indicated by the findings that M. aurum DSM 33539 prevented CSC-induced increases in the relative abundances of the colitogenic phyla Desulfobacterota and Deferribacterota. Indeed, the relative abundance of Deferribacterota on day 19 was strongly correlated with histological damage to the colon. In line with the latter, M. aurum DSM 33539 was further protective against the aggravating effects of stress on subsequent DSS colitis. Collectively, our findings confirm and extend previous findings from our group and suggest that the stress-protective effects reported for M. vaccae NCTC 11659 and M. vaccae ATCC 15483T are generalizable also to other NTM species.

P0852 Influence of a gluten-free dietary intervention on the gut microbiome and inflammation in inflammatory bowel disease: results from murine experiments and protocol of a prospective proof-of-concept human study as part of the miGut-Health project

Abstract Background A significant number of inflammatory bowel disease (IBD) patients report an improvement of gastrointestinal symptoms under a gluten-free diet (GFD), despite not suffering from celiac disease. In a recent prospective pilot study, we detected a significant downregulation of proinflammatory cytokines and chemokines on the mucosal level, as well as slight compositional changes of the luminal microbiome after implementation of an eight-week GFD intervention in PSC-IBD patients (1). For a further characterization, we established murine models of IBD, investigating the influence of a GFD on colitis susceptibility, gut microbiota, and intestinal inflammation (Part A). Subsequently, a proof-of-concept human study was implemented as part of the miGut Health project for clinical correlation (Part B). Methods A) C57BL/6 mice are either bred and kept under standard diet (SD, containing wheat) or GFD followed by induction of dextran sodium sulfate (DSS) colitis, a murine model for ulcerative colitis. Relative loss of bodyweight and mean endoscopic colitis score are measured. For characterization of the microbiota impact, murine stool transplantation into germ free mice and 16s RNA analyzes are performed. B) An investigator-initiated prospective clinical trial with one interventional arm in IBD patients will be performed at the University Medical Center Hamburg-Eppendorf to investigate a 1.5-year long wheat protein-free diet in patients with inflammatory bowel disease (IBD) including ulcerative colitis (UC), Crohn’s disease (CD) and PSC-associated IBD (PSC-IBD), n=20 each. Effects of the GFD on intestinal cell composition and state, intestinal fecal and mucosal microbiota composition will be assessed as well as the dietary impact on IBD activity using colonoscopy and biomarkers of intestinal inflammation and on symptoms and health-related quality of life. Results A) Gluten aggravates murine DSS-colitis and long-term GFD (LT-GFD) protects mice from gluten aggravated colitis susceptibility. More importantly, preliminary data in germ-free settings support that this effect is mediated through microbiota alterations during the LT-GFD. 16s RNA sequencing data from murine stool showed different relative bacterial abundance. B) The study protocol received formal ethical approval by the ethics committee of the Hamburg Medical Association, Hamburg, Germany. Patient recruitment has started and is ongoing. Conclusion These primary results underline a potential role of gut microbiota modulation for the positive effect of a GFD on gastrointestinal symptoms in IBD. Long-term tolerability and efficacy of the dietary intervention in IBD patients remain to be seen.

P0200 Evaluating the efficacy of tonsil-derived mesenchymal stem cells and intestinal stem cells-like cells in modulating fibrosis and inflammation in murine colitis models

Abstract Background Current treatments in inflammatory bowel disease (IBD) primarily target inflammation, but effective anti-fibrotic treatments are limited. This study evaluates the therapeutic potential of tonsil derived mesenchymal stem cell (TMSCs) and TMSC induced intestinal stem cell-like cell (T-ISCs) in treating intestinal fibrosis and inflammation using two murine colitis models. Methods Two chronic colitis models were established using dinitrobenzene sulfonic acid (DNBS) and dextran sulfate sodium (DSS) in mice. TMSCs and T-ISCs were administered intraperitoneally, with mice divided into seven groups, including control and treatment groups. Colitis symptoms were assessed using disease activity index (DAI) and colon length. After euthanasia on day 30, colon tissues were collected for analysis. Fibrosis markers and inflammatory cytokines were assessed using real-time PCR, while fibrosis severity was evaluated through Masson’s Trichrome staining. In addition, immunofluorescence and western blotting was performed to confirm the fibrosis-related protein expression. Results The therapeutic effects of TMSCs and T-ISCs were confirmed in both DNBS and DSS colitis models. In the DNBS colitis model, both TMSCs and T-ISCs treatments significantly reduced DAI and restored colon length compared to the PBS group (colitis control group) (all p < 0.05). TMSCs treatment markedly decreased three fibrosis markers, including COL1A1, Vimentin, and TGF-β, with statistical significance (p < 0.01, p < 0.01, and p < 0.05, respectively), while T-ISCs significantly reduced only COL1A1 expression (p < 0.05). Histological analyses using Masson’s Trichrome staining showed decreased fibrosis scores in both TMSCs- and T-ISCs-treated groups, and protein expression analysis via immunofluorescence and western blot also confirmed reductions in a-SMA, Vimentin, COL1A1, and TGF-β in both TMSCs- and T-ISCs-treated groups, with a particularly pronounced reduction observed in the TMSCs-treated group. In the DSS colitis model, both TMSCs and T-ISCs treatments significantly decreased the colon shortening compared to the PBS group (all p < 0.05). Although fibrosis markers and scores showed no significant changes, inflammatory cytokine levels were markedly reduced in the DSS colitis model. Notably, T-ISCs demonstrated significant reductions in both IL-1β (p < 0.01) and IL-6 (p < 0.05). Conclusion TMSCs and T-ISCs demonstrate therapeutic effects in murine colitis models mimicking IBD, with fibrosis reduction in the DNBS model and inflammation reduction in the DSS model. These findings suggest that TMSC-based stem cell therapies are promising strategies for treating fibrosis and inflammation in IBD.

P0202 Modeling Inflammatory Bowel Disease in mice with “Humanized” immune system

Abstract Background Inflammatory bowel disease (IBD) is a chronic recurrent inflammatory disease with unknown etiology. Dextran sulfate sodium (DSS) induced colitis is a common preclinical mouse model in IBD research. DSS colitis involves activation of the submucosal immune system and can be used to study Ulcerative Colitis characteristics in different phases (acute, chronic and remission) and predict compound efficacy. However, optimizing translation to human is still needed and the use of mice with Human Immune System (HIS) provide a better model to study human immune system responses in vivo. HIS mice are created by transplanting human immune cells or their progenitor cells into highly immunodeficient recipient mouse hosts, thereby "humanizing" their immune systems. In this study we discussed colitis feature and the use of humanized mice model for testing therapeutics in preclinical studies Methods 15-weeks-old female HIS mice were treated with 2.5% dextran sulfate sodium (DSS) in drinking water (ad.lib.) for 5 days to induce acute colitis. Colonic and systemic human cytokines were assessed by MSD plex assay. Human immune cell infiltration into the colon was measured by flow cytometry. Colonic histological damage was assessed on colon Swiss roll after H&E staining Results HIS mice challenged with DSS exhibited body weight loss and increase disease activity index (loose stool or diarrhea, rectal bleeding) compared to naive HIS mice. HIS-DSS mice show a strong human pro-inflammatory response, including elevated hIL-1B, hTNF, hIL-6 in plasma and colon as well as specific human cytokines (MCP-4, IL-8, IL-26, IL-37…). Colon microscopic analysis show loss of goblet cells, loss of crypt structure and increase of inflammatory cell infiltration at the level of mucosa and submucosa Conclusion These data show a significant increase of human pro-inflammatory cytokines after DSS intake as well as histologic score characteristic of Ulcerative Colitis. They also confirm that HIS mice represent a relevant model for the preclinical assessment of therapeutics in IBD.

Exploring the efficacy of Transcutaneous Auricular Vagus nerve stimulation (taVNS) in modulating local and systemic inflammation in experimental models of colitis

BackgroundCurrent inflammatory bowel disease (IBD) treatments often fail to achieve lasting remission and have adverse effects. Vagus nerve stimulation (VNS) offers a promising therapy due to its anti-inflammatory effects. Its invasive nature, however, has led to the development of non-invasive methods like transcutaneous auricular VNS (taVNS). This study assesses taVNS’s impact on acute colitis progression, inflammatory, anti-inflammatory, and apoptosis-related markers.MethodsMale C57BL/6 mice (11–12 weeks) were used for dextran sulfate sodium (DSS)- and dinitrobenzene sulfonic acid (DNBS)-induced colitis studies. The administration of taVNS or no stimulation (anesthesia without stimulation) for 10 min per mouse began one day before colitis induction and continued daily until sacrifice. Ulcerative colitis (UC)-like colitis was induced by administering 5% DSS in drinking water for 5 days, after which the mice were sacrificed. Crohn’s disease (CD)-like colitis was induced through a single intrarectal injection of DNBS/ethanol, with the mice sacrificed after 3 days. Disease activity index (DAI), macroscopic evaluations, and histological damage were assessed. Colon, spleen, and blood samples were analyzed via qRT-PCR and ELISA. One-way or two-way ANOVA with Bonferroni and Šídák tests were applied.ResultstaVNS improved DAI, macroscopic, and histological scores in DSS colitis mice, but only partially mitigated weight loss and DAI in DNBS colitis mice. In DSS colitis, taVNS locally decreased colonic inflammation by downregulating pro-inflammatory markers (IL-1β, TNF-α, Mip1β, MMP 9, MMP 2, and Nos2) at the mRNA level and upregulating anti-inflammatory TGF-β in non-colitic conditions at both mRNA and protein levels and IL-10 mRNA levels in both non-colitic and colitic conditions. Systemically, taVNS decreased splenic TNF-α in non-colitic mice and increased serum levels of TGF-β in colitic mice and splenic levels in non-colitic and colitic mice. Effects were absent in DNBS-induced colitis. Additionally, taVNS decreased pro-apoptotic markers (Bax, Bak1, and caspase 8) in non-colitic and colitic conditions and increased the pro-survival molecule Bad in non-colitic mice.ConclusionsThis study demonstrates that taVNS has model-dependent local and systemic effects, reducing inflammation and apoptosis in UC-like colitis while offering protective benefits in non-colitic conditions. These findings encourage further research into underlying mechanisms and developing adjunct therapies for UC.

CREEPING FAT-DERIVED FREE FATTY ACIDS INDUCE HYPERPLASIA OF INTESTINAL MUSCULARIS PROPRIA MUSCLE CELLS – A NOVEL LINK BETWEEN FAT AND INTESTINAL STRICTURE FORMATION IN CROHN’S DISEASE

In Crohn's disease, wrapping of mesenteric fat around the bowel wall, so-called "creeping fat," is highly associated with strictures. The strongest contributor to luminal narrowing in strictures is a thickening of the human intestinal muscularis propria (MP). We investigated creeping fat-derived factors and their effect on mechanisms of human intestinal MP smooth muscle cell (HIMC) hyperplasia. Free fatty acids (FFAs) in creeping fat or noncreeping mesenteric fat organ cultures were measured via lipidomic mass spectrometry. Primary HIMCs were exposed to FFAs and cell proliferation was assessed. Intracellular FFA metabolism pathways and reactive oxygen species were functionally evaluated. Muscle thickness was investigated in dextran sodium sulfate colitis with small molecule inhibition of FFA transport and a novel fat deletion mouse model. Subserosal creeping fat is associated with a markedly thickened MP. Experimental deletion of mesenteric fat (FAT-ATTAC [fat apoptosis through targeted activation of caspase 8] mouse) reduced MP thickness. Human creeping fat-conditioned medium strongly up-regulated HIMC proliferation. Creeping fat released higher amounts of 5 long-chain FFAs, including palmitate. Inhibition of HIMC long-chain FFA metabolism or FFA uptake into mitochondria through carnitine palmitoyltransferase-1 reduced the palmitate-induced HIMC proliferation. Blockade of conversion of palmitate into phospholipids reduced HIMC proliferation. Prophylactic inhibition of carnitine palmitoyltransferase-1 in experimental dextran sodium sulfate colitis did not ameliorate inflammation, but reduced MP thickness. Creeping fat-released long-chain FFAs induce a selective proliferative response by HIMC. These results point to creeping fat as a novel contributor to stricture formation in Crohn's disease.

Leucine-Enriched Diet Reduces Fecal MPO but Does Not Protect Against DSS Colitis in a Mouse Model of Crohn's Disease-like Ileitis.

Understanding the complex link between inflammation, gut health, and dietary amino acids is becoming increasingly important in the pathophysiology of inflammatory bowel disease (IBD). This study tested the hypothesis that a leucine-rich diet could attenuate inflammation and improve gut health in a mouse model of IBD. Specifically, we investigated the effects of a leucine-rich diet on dextran sulfate sodium (DSS)-induced colitis in germ-free (GF) SAMP1/YitFC (SAMP) mice colonized with human gut microbiota (hGF-SAMP). hGF-SAMP mice were fed one of four different diets: standard mouse diet (CHOW), American diet (AD), leucine-rich AD (AD + AA), or leucine-rich CHOW diet (CH + AA). Body weight, myeloperoxidase (MPO) activity, gut permeability, colonoscopy scores, and histological analysis were measured. Mice on a leucine-rich CHOW diet showed a decrease in fecal MPO prior to DSS treatment as compared to those on a regular diet (p > 0.05); however, after week five, prior to DSS, this effect had diminished. Following DSS treatment, there was no significant difference in gut permeability, fecal MPO activity, or body weight changes between the leucine-supplemented and control groups. These findings suggest that while a leucine-rich diet may transiently affect fecal MPO levels in hGF-SAMP mice, it does not confer protection against DSS-induced colitis symptoms or mitigate inflammation in the long term.

The therapeutic efficacy of adipose mesenchymal stem cell-derived microvesicles versus infliximab in a dextran sodium sulfate induced ulcerative colitis rat model

ABSTRACT Ulcerative colitis (UC) is a chronic relapsing intestinal inflammation that is becoming of increasing incidence worldwide and has insufficient treatment. Therefore, finding effective therapies remains a priority. A dextran sodium sulfate colitis model was established to elucidate colonic layers alterations and compare adipose mesenchymal stem cell-derived microvesicles (MSC-MVs) versus infliximab (IFX) efficacy through biochemical, light, and electron microscope studies. Fifty-four rats were allocated to 4 groups: Control (Con), UC, UC+IFX, and UC+MSC-MVs groups. End body weights (BW) and serum malondialdehyde (MDA) levels were recorded. Colitis severity was estimated by disease activity index (DAI). Colonic specimens were processed to evaluate the histological structure, collagen content, surface mucous and goblet cells, CD44, TNF-α, and GFAP immune expression. Morphometric and statistical analyses were performed. The UC group revealed congested, stenosed colons, a significant decline in end BW, and a significant increase in serum MDA and DAI. Furthermore, disturbed histoarchitecture, inflammatory infiltration, depletion of surface mucous and goblet cells, increased collagen, and TNF-α expression and decreased GFAP expression were observed. Alterations were partially attenuated by IFX therapy, whereas MSC-MVs significantly improved all parameters. In conclusion, MSC-MVs were a superior therapeutic option, via attenuating oxidative stress and inflammatory infiltration, in addition to restoring intestinal epithelial integrity and mucosal barrier.

A pH-sensitive opioid does not exhibit analgesic tolerance in a mouse model of colonic inflammation.

Tolerance to the analgesic effects of opioids and resultant dose escalation is associated with worsening of side effects and greater addiction risk. Here, we compare the development of tolerance to the conventional opioid fentanyl with a novel pH-sensitive μ-opioid receptor (MOR) agonist, (±)-N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide (NFEPP) that is active only in acidic inflammatory microenvironments. An opioid tolerance model was developed in male C57BL/6 mice, with and without dextran sulphate sodium colitis, using increasing doses of either fentanyl or NFEPP over 5 days. Visceral nociception was assessed in vivo by measuring visceromotor responses (VMRs) to noxious colorectal distensions and in vitro measuring colonic afferent nerve activity of mesenteric nerves and performing patch-clamp recordings from isolated dorsal root ganglia neurons. Somatic thermal nociception was tested using a tail immersion assay. Cardiorespiratory effects were analysed by pulse oximeter experiments. VMRs and tail immersion tests demonstrated tolerance to fentanyl, but not to NFEPP in colitis mice. Cross-tolerance also occurred to fentanyl, but not to NFEPP. The MOR agonist DAMGO inhibited colonic afferent nerve activity in colitis mice exposed to chronic NFEPP, but not those from fentanyl-treated mice. Similarly, in patch-clamp recordings from isolated dorsal root ganglia neurons, DAMGO inhibited neurons from NFEPP-, but not fentanyl-treated mice. NFEPP did not exhibit tolerance in an inflammatory pain model, unlike fentanyl. Consequently, dose escalation to maintain analgesia during an evolving inflammation could be avoided, mitigating the potential risk of side effects.

Toll-like receptor 7 protects against intestinal inflammation and restricts the development of colonic tissue-resident memory CD8+ T cells.

The maintenance of intestinal homeostasis depends on a complex interaction between the immune system, intestinal epithelial barrier, and microbiota. Alteration in one of these components could lead to the development of inflammatory bowel diseases (IBD). Variants within the autophagy gene ATG16L1 have been implicated in susceptibility and severity of Crohn's disease (CD). Individuals carrying the risk ATG16L1 T300A variant have higher caspase 3-dependent degradation of ATG16L1 resulting in impaired autophagy and increased cellular stress. ATG16L1-deficiency induces enhanced IL-1β secretion in dendritic cells in response to bacterial infection. Infection of ATG16L1-deficient mice with a persistent strain of murine norovirus renders these mice highly susceptible to dextran sulfate sodium colitis. Moreover, persistent norovirus infection leads to intestinal virus specific CD8+ T cells responses. Both Toll-like receptor 7 (TLR7), which recognizes single-stranded RNA viruses, and ATG16L1, which facilitates the delivery of viral nucleic acids to the autolysosome endosome, are required for anti-viral immune responses. However, the role of the enteric virome in IBD is still poorly understood. Here, we investigate the role of TLR7 and ATG16L1 in intestinal homeostasis and inflammation. At steady state, Tlr7-/- mice have a significant increase in large intestinal lamina propria (LP) granzyme B+ tissue-resident memory CD8+ T (TRM) cells compared to WT mice, reminiscent of persistent norovirus infection. Deletion of Atg16l1 in myeloid (Atg16l1ΔLyz2 ) or dendritic cells (Atg16l1ΔCd11c ) leads to a similar increase of LP TRM. Furthermore, Tlr7-/- and Atg16l1ΔCd11c mice were more susceptible to dextran sulfate sodium colitis with an increase in disease activity index, histoscore, and increased secretion of IFN-γ and TNF-α. Treatment of Atg16l1ΔCd11c mice with the TLR7 agonist Imiquimod attenuated colonic inflammation in these mice. Our data demonstrate that ATG16L1-deficiency in myeloid and dendritic cells leads to an increase in LP TRM and consequently to increased susceptibility to colitis by impairing the recognition of enteric viruses by TLR7. In conclusion, the convergence of ATG16L1 and TLR7 signaling pathways plays an important role in the immune response to intestinal viruses. Our data suggest that activation of the TLR7 signaling pathway could be an attractive therapeutic target for CD patients with ATG16L1 risk variants.

Trp53 Deletion Promotes Exacerbated Colitis, Facilitates Lgr5+ Cancer Stem Cell Expansion, and Fuels Tumorigenesis in AOM/DSS-Induced Colorectal Cancer.

Colorectal cancer CRC remains one of the leading causes of cancer-related deaths worldwide, with chronic intestinal inflammation identified as a major risk factor. Notably, the tumor suppressor TP53 undergoes mutation at higher rates and earlier stages during human inflammation-driven colon tumorigenesis than in sporadic cases. We investigated whether deleting Trp53 affects inflammation-induced tumor growth and the expression of Lgr5+ cancer stem cells in mice. We examined azoxymethane (AOM)/dextran sodium sulfate (DSS)-induced colon tumorigenesis in wild-type Trp53 (+/+), heterozygous (+/-), and knockout (-/-) mice. Trp53-/- mice showed increased sensitivity to DSS colitis and earlier accelerated tumorigenesis with 100% incidence. All groups could develop invasive tumors, but knockouts displayed the most aggressive features. Unlike wild-type CRC, knockouts selectively showed increased populations of Lgr5+ colon cancer stem-like cells. Trp53 loss also boosted laminin, possibly facilitating the disruption of the tumor border. This study highlights how Trp53 deletion promotes the perfect storm of inflammation and stemness, driving colon cancer progression. Trp53 deletion dramatically shortened AOM/DSS latency and improved tumor induction efficiency, offering an excellent inflammation-driven CRC model.

Thalidomide mitigates Crohn's disease colitis by modulating gut microbiota, metabolites, and regulatory T cell immunity

Thalidomide (THA) is renowned for its potent anti-inflammatory properties. This study aimed to elucidate its underlying mechanisms in the context of Crohn’s disease (CD) development. Mouse colitis models were established by dextran sulfate sodium (DSS) treatment. Fecal microbiota and metabolites were analyzed by metagenomic sequencing and mass spectrometry, respectively. Antibiotic-treated mice served as models for microbiota depletion and transplantation. The expression of Forkhead box P3+ (FOXP3+) Regulatory T cells (Tregs) was measured by flow cytometry and immunohistochemical assay in colitis model and patient cohort. THA inhibited colitis in DSS–treated mice by altering the gut microbiota profile, with an increased abundance of probiotics Bacteroides fragilis (Bf), while pathogenic bacteria were depleted. In addition, THA increased beneficial metabolites bile acids and significantly restored gut barrier function. Transcriptomic profiling revealed that THA inhibited interleukin-17 (IL17), IL1β and cell cycle signaling. Fecal microbiota transplantation from THA-treated mice to microbiota-depleted mice partly recapitulated the effects of THA. Specifically, increased level of gut commensal Bf was observed, correlated with elevated levels of the microbial metabolite 7-Ketolithocholic acid (7-KA) following THA treatment. This microbial metabolite may stable FOXP3 expression by targeting the bile acid receptor farnesol X receptor 1(FXR1) to inhibit autophagy. An interaction between FOXP3 and FXR1 was identified, with binding regions localized to the FOXP3 domain (aa 238-335) and the FXR1 domain (aa 82-222), respectively. Conclusively, THA modulates the gut microbiota and metabolite profiles towards a more beneficial composition, enhances gut barrier function, promotes the differentiation of FOXP3+ Tregs and curbs pro-inflammatory pathways.

YOD1 sustains NOD2-mediated protective signaling in colitis by stabilizing RIPK2

Inflammatory bowel disease (IBD) is a disorder causing chronic inflammation in the gastrointestinal tract, and its pathophysiological mechanisms are still under investigation. Here, we find that mice deficient of YOD1, a deubiquitinating enzyme, are highly susceptible to dextran sulfate sodium (DSS)-induced colitis. The bone marrow transplantation experiment reveals that YOD1 derived from hematopoietic cells inhibits DSS colitis. Moreover, YOD1 exerts its protective role by promoting nucleotide-binding oligomerization domain 2 (NOD2)-mediated physiological inflammation in macrophages. Mechanistically, YOD1 inhibits the proteasomal degradation of receptor-interacting serine/threonine kinase 2 (RIPK2) by reducing its K48 polyubiquitination, thereby increasing RIPK2 abundance to enhance NOD2 signaling. Consistently, the protective function of muramyldipeptide, a NOD2 ligand, in experimental colitis is abolished in mice deficient of YOD1. Importantly, YOD1 is upregulated in colon-infiltrating macrophages in patients with colitis. Collectively, this study identifies YOD1 as a novel regulator of colitis.

Pericentriolar material 1 promotes intestinal inflammation in ulcerative colitis by activating NLRP3/gasdermin D‐mediated macrophage pyroptosis

AbstractExcessive proinflammatory cytokine release induced by pyroptosis plays a vital role in intestinal mucosal inflammation in ulcerative colitis (UC). Several pyroptosis‐related factors are regulated by the centrosome. Pericentriolar material 1 (PCM1) is a primary component of centriolar satellites that is present as cytoplasmic granules around the centrosome. Our previous study revealed that PCM1 was highly expressed in UC patients, but the role of PCM1 in UC remains unknown. This study aimed to elucidate the role of PCM1 in the development of UC, especially the mechanism in pyroptosis process of UC. Clinical mucosal sample and dextran sulfate sodium (DSS)‐induced colitis mouse were used to reveal the association between PCM1 and intestinal inflammation. Intestinal epithelial cell‐specific PCM1‐knockout mice were constructed to determine the role of PCM1 in colitis. Finally, PCM1 RNA interference and overexpression assays in THP1 cells were employed to study the molecular mechanisms of PCM1 in inflammatory responses and pyroptosis. We found that PCM1 expression was upregulated in the colonic mucosa of UC patients and positively correlated with inflammatory indicators. PCM1 expression was elevated in DSS‐induced colitis mice and was reduced after methylprednisolone treatment. In the DSS colitis model, intestinal‐specific PCM1‐knockout mice exhibited milder intestinal inflammation and lower pyroptosis levels than wild‐type mice. In cell level, PCM1 exerted a proinflammatory effect by activating the NLRP3 inflammasome and triggering subsequent gasdermin D‐mediated pyroptosis to release IL‐1β and IL‐18. In conclusion, PCM1 mediates activation of the NLRP3 inflammasome and gasdermin D‐dependent pyroptosis, ultimately accelerating intestinal inflammation in UC. These findings revealed a previously unknown role of PCM1 in initiating intestinal mucosal inflammation and pyroptosis in UC, and this factor is expected to be a regulator in the complex inflammatory network of UC.

Microbiome- and Host Inflammasome-Targeting Inhibitor Nanoligomers Are Therapeutic in the Murine Colitis Model.

Autoimmune and autoinflammatory diseases account for more than 80 chronic conditions affecting more than 24 million people in the US. Among these autoinflammatory diseases, noninfectious chronic inflammation of the gastrointestinal (GI) tract causes inflammatory bowel diseases (IBDs), primarily Crohn's and ulcerative colitis (UC). IBD is a complex disease, and one hypothesis is that these are either caused or worsened by compounds produced by bacteria in the gut. While traditional approaches have focused on pan immunosuppressive techniques (e.g., steroids), low remission rates, prolonged illnesses, and an increased frequency of surgical procedures have prompted the search for more targeted and precision therapeutic approaches. IBD is a complex disease resulting from both genetic and environmental factors, but several recent studies have highlighted the potential pivotal contribution of gut microbiota dysbiosis. Gut microbiota are known to modulate the immune status of the gut by producing metabolites that are encoded in biosynthetic gene clusters (BGCs) of the bacterial genome. Here, we show a targeted and high-throughput screening of more than 90 biosynthetic genes in 41 gut anaerobes, through downselection using available bioinformatics tools, targeted gene manipulation in these genetically intractable organisms using the Nanoligomer platform, and identification and synthesis of top microbiome targets as a Nanoligomer BGC cocktail (SB_BGC_CK1, abbreviated as CK1) as a feasible precision therapeutic approach. Further, we used a host-directed immune target screening to identify the NF-κB and NLRP3 cocktail SB_NI_112 (or NI112 for short) as a targeted inflammasome inhibitor. We used these top two microbe- and host-targeted Nanoligomer cocktails in acute and chronic dextran sulfate sodium (DSS) mouse colitis and in TNFΔARE/+ transgenic mice that develop spontaneous Crohn's like ileitis. The mouse microbiome was humanized to replicate that in human IBD through antibiotic treatment, followed by mixed fecal gavage from 10 human donors and spiked with IBD-inducing microbial species. Following colonization, colitis was induced in mice using 1 week of 3% DSS (acute) or 6 weeks of 3 rounds of 2.5% DSS induction for a week followed by 1 week of no DSS (chronic colitis model). Both Nanoligomer cocktails (CK1 and NI112) showed a strong reduction in disease severity, significant improvement in disease histopathology, and profound downregulation of disease biomarkers in colon tissue, as assessed by multiplexed ELISA. Further, we used two different formulations of intraperitoneal injections (IP) and Nanoligomer pills in the chronic DSS colitis model. Although both formulations were highly effective, the oral pill formulation demonstrated a greater reduction in biochemical markers compared to IP. A similar therapeutic effect was observed in the TNFΔARE/+ model. Overall, these results point to the potential for further development and testing of this inflammasome-targeting host-directed therapy (NI112) and more personalized microbiome cocktails (CK1) for patients with recalcitrant IBD.

Intestinal and hepatic benefits of BBR-EVO on DSS-induced experimental colitis in mice.

Ulcerative colitis (UC), characterized by disrupted intestinal barrier integrity and chronic inflammation, was modeled in mice via dextran sulfate sodium (DSS) induction. This study explored the therapeutic potential of berberine-evodiamine (BBR-EVO), bioactive components of the traditional Chinese medicine Yulian decoction, in DSS colitis. BBR-EVO intervention ameliorated weight loss, diarrhea, colonic shortening, and histopathological damage in colitic mice. The substance increased antioxidant activity while reducing high levels of pro-inflammatory cytokines in the colon, including as TNF-α, IL-1β, and IL-6. BBR-EVO inhibited the DSS-induced decrease in the tight junction proteins ZO-1 and occludin, according to immunohistochemistry. 16S rRNA sequencing demonstrated BBR-EVO partially attenuated DSS-elicited intestinal dysbiosis, reducing opportunistic pathogens and restoring diminished beneficial taxa. Critically, BBR-EVO alleviated secondary hepatic injury in colitic mice, mitigating immune cell infiltration, oxidative stress, cytokine production, and ultrastructural damage, likely by beneficially modulating gut-liver crosstalk. This study reveals BBR-EVO, derived from a traditional Chinese medicine, confers multi-target protective effects in experimental colitis and associated hepatic pathology, warranting further evaluation as a potential therapy for inflammatory bowel diseases like UC. The mechanisms may involve simultaneous augmentation of intestinal barrier integrity, inhibition of inflammation, microbiota regulation, and gut-liver axis optimization.

PARP1 inactivation increases regulatory T / Th17 cell proportion in intestinal inflammation. Role of HMGB1

Inflammatory bowel diseases (IBD) are chronic relapsing disorders with increasing prevalence. Knowledge gaps still limit the possibility to develop more specific and effective therapies. Using a dextran sodium sulfate colitis mouse model, we found that inflammation increased the total number and altered the frequencies of leukocytes within colon mesenteric lymph nodes (cMLNs). Although the inflammation reduced the frequency of regulatory T (Treg) cells, their absolute numbers were increased. Increased frequency of colitogenic Th17 cells was also observed. Noteworthy, untreated mice lacking Poly(ADP-ribose)-Polimerase-1 functional gene (PARP-1KO) displayed higher frequency of Treg cells and lower percentage of Th17 cells in cMLNs. In colitic PARP-1KO mice the inflammation driven expansion of the Foxp3 Treg population was more pronounced than in WT mice. Conversely, colitis increased Th17 cells to a lower extent in PARP-1KO mice compared with WT mice, resulting in a more protective Treg/Th17 cell ratio. Consequently PARP-1KO mice developed less severe colitis with reduced expression of inflammatory cytokines. In ex vivo experiments PARP-1KO and WT CD11c dendritic cells (DCs) promoted naïve CD4 T cell differentiation differently, the former sustaining more efficiently the generation of Treg cells, the latter that of Th17 cells. Addition of HMGB1 B box or of dipotassium glycyrrhizate, which sequesters extracellular HMGB1, revealed a role for this alarmin in the regulation exerted by PARP-1 on the stimulating vs. tolerogenic function of DCs during colitis. Moreover, a higher percentage of CD11c DC from PARP-1KO mice expressed CD103, a marker associated with the ability of DC to induce Treg cells, compared with WT DC. Conversely, PARP-1KO DC were including a reduced percentage of CX3CR1+ DC, described to induce Th17 cells. These findings were observed in both splenic and colon lamina propria DC.

Impact of Nanoplastic Particles on Macrophage Inflammation and Intestinal Health in a Mouse Model of Inflammatory Bowel Disease.

The increasing presence of plastics in the human diet is raising public concern about the potential risks posed by nanoplastic (NP) particles, which can emerge from the degradation of plastic debris. NP ingestion poses particular risks to individuals with inflammatory bowel disease (IBD), as compromised epithelial barriers may facilitate NP translocation. In vitro, bone-marrow-derived macrophages (BMDMs) were exposed to 25 nm polymethacrylate (PMMA) or 50 nm polystyrene (PS) particles to assess morphological changes and alterations in pro- and anti-inflammatory gene expression. In vivo, mice received PMMA NP particles for 6 months before acute dextran sodium sulfate (DSS) colitis was induced to investigate NP impacts on intestinal health and inflammation. PMMA and PS NP exposure in BMDMs induced morphological changes indicative of a proinflammatory phenotype characterized by enlarged amoeboid cell shapes. It also triggered an inflammatory response, indicated by increased expression of proinflammatory cytokines such as Tnfa and Il6. Unexpectedly, long-term PMMA NP administration did not affect the intestinal epithelial barrier or exacerbate acute DSS-induced colitis in mice. Colonoscopy and histological analysis revealed no NP-related changes, suggesting adverse effects on intestinal health or inflammation. Our findings from animal models offer some reassurance to IBD patients regarding the effects of NP ingestion. However, variations in lifestyle and dietary habits may lead to significantly higher plastic intake in certain individuals, raising concerns about potential long-term gastrointestinal effects of lifelong plastic consumption.

The beneficial impact of ketogenic diets on chemically-induced colitis in mice depends on the diet's lipid composition

Previously, we showed that restrictive diets, including ketogenic diet (KD), have an anti-inflammatory impact on the healthy gastrointestinal tract of mice. Afterward, we found that energy-restricting diets mitigate inflammation in the dextran sodium sulfate (DSS) colitis mouse model. The current study aimed to verify the impact of KD on DSS colitis and assess if the diet's fat composition influences the outcomes of the intervention.Mice with mild chronic colitis were fed control chow, KD composed of long-chain triglycerides (KD LCT) or a KD containing a mix of LCT and medium-chain triglycerides (KD LCT/MCT).KDs did not reverse DSS-enhanced gut permeability and shortening of the colon. Both KDs had a similar impact on liver, cecum, and spleen weight, villi and colon length, the thickness of muscularis externa, and expression of ZO-1 and occludin. On the contrary, body weight, glutathione (GSH) and taurine-GSH levels, GSH-S transferase (GST), and myeloperoxidase (MPO) activity, as well as an abundance of several fecal bacteria, all were differentially affected by the two types of KDs. When compared to the DSS control diet, reduction in colon mucosa cytokines expression was stronger in KD LCT than in the KD LCT/MCT group.We conclude that the outcomes of the KD interventions in terms of potential therapeutical applications depend on lipid composition. KD LCT showed a strong positive impact on gut inflammation. A potential contribution of GSH to KD outcomes and a correlation between MPO activity and microbiota composition was identified.

The Impact of Peptidyl Arginine Deiminase 4-Dependent Neutrophil Extracellular Trap Formation on the Early Development of Intestinal Fibrosis in Crohn's Disease.

During early phases of inflammation, activated neutrophils extrude neutrophil extracellular traps (NETs) in a peptidyl arginine deiminase 4 (PAD4)-dependent manner, aggravating tissue injury and remodeling. In this study, we investigated the potential pro-fibrotic properties and signaling of NETs in Crohn's disease (CD). NETs and activated fibroblasts were labeled on resected ileum from CD patients by multiplex immunofluorescence staining. NETs-treated human primary intestinal fibroblasts were analyzed by bulk RNA sequencing to uncover cell signaling pathways, and by high-throughput imaging to assess collagen production and migratory activity. Consequentially, TLR2/NF-κB pathway was evaluated by transfection of CCD-18Co fibroblasts with an NF-κB-luciferase reporter plasmid, incorporating C29 to block TLR2 signaling. A chronic dextran sulfate sodium (DSS) mouse model was used to define the specific role of PAD4 deletion in neutrophils (MRP8-Cre, Pad4fl/fl). Immunofluorescence showed spatial colocalization of NETs and activated fibroblasts in ileal ulcerations of CD patients. Transcriptomic analysis revealed upregulation of pro-fibrotic genes and activation of Toll-like receptor signaling pathways in NETs-treated fibroblasts. NETs treatment induced fibroblast proliferation, diminished migratory capability, and increased collagen release. Transfection experiments indicated a substantial increase in an NF-κB expression with NETs, whereas C29 led to decreased expression and release of collagen. In line, a significant reduction in collagen content was observed in the colon of MRP8-Cre, Pad4fl/fl mice subjected to chronic DSS colitis. NETs potentially serve as an initial stimulus for pathological activation of fibroblasts within the intestine via the TLR2/NF-κB pathway. Given their early involvement in inflammation, inhibition of PAD4 might offer a strategy to modulate both inflammation and fibrogenesis in CD.