Abstract
Background
People with HIV (PWH) are at high risk for metabolic dysfunction-associated fatty liver disease (MAFLD). In the general population, sex differences seem to exist in frequency and ...severity of MAFLD, with higher prevalence of MAFLD in men, but higher incidence of liver fibrosis in women. Less is known about sex differences in MAFLD and liver fibrosis in the setting of HIV infection.
Purpose
The aim of this study is to investigate the sex differences in the prevalence of MAFLD in PWH and the severity of liver fibrosis among this population.
Method
This was a multicenter cohort study including consecutive PWH who underwent screening for MAFLD and liver fibrosis by liver stiffness measurement (LSM) with associated controlled attenuation parameter (CAP). MAFLD was defined as the presence of hepatic steatosis, diagnosed as CAP>270 dB/m, plus any among type 2 diabetes, overweight (BMI>25 Kg/m2) or two other metabolic abnormalities. Significant liver fibrosis was diagnosed as LSM>8 kPa. Incidence of MAFLD and significant liver fibrosis was assessed through survival analysis.
Result(s)
1359 PWH (25% females, 30% HCV coinfected) were included. Prevalence of MAFLD at baseline was lower in women than in men with HIV (17.7% vs. 24.3%, p=0.013). Conversely, there was no difference in prevalence of liver fibrosis (10.7% vs. 13.4%). Women with MAFLD were more frequently of black ethnicity (48% vs. 14%, p<0.001), had lower ALT (26.4+20.4 vs. 33.4+22.5; p=0.035), higher HDL cholesterol (1.46+0.57 vs. 1.11+0.33; p<0.001), lower triglycerides (1.69+0.96 vs. 2.47+2.63; p=0.035) compared to men with MAFLD. 485 of these PWH were followed for a median of 3.5 years. Incidence of MAFLD was similar between women and men with HIV. However, incidence of liver fibrosis was higher in women compared to men with HIV (7.0 per 100 person-years PY vs. 5.9 per 100 PY; p=0.035) (Figure 1). The higher incidence of significant liver fibrosis occurred particularly after the age of 50 years. On multivariable Cox regression analysis and after adjusting for age, presence of MAFLD (adjusted hazard ratio aHR 3.3, 95% CI 2.0-5.6) and female sex (aHR 2.2, 95% CI 1.3-3.5) were independent predictors of developing significant liver fibrosis while CD4 cell count was protective (aHR 0.99, 95% CI 0.99-0.99).
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Conclusion(s)
MAFLD seems a sexual dimorphic disease in PWH. Despite having lower rates of MAFLD, women with HIV have higher incidence of significant liver fibrosis compared to men, especially after 50 years of age. Future studies should target adequate consideration of sex differences in clinical investigation of MAFLD to fill current gaps and implement precision medicine for PWH.
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None
Disclosure of Interest
None Declared
Abstract
Background
Fecal calprotectin (FC) is a sensitive marker of intestinal inflammation, and is used to both discriminate inflammatory bowel disease (IBD) from non-IBD patients and to monitor ...patients with IBD. It is unclear whether normal values established in adult patients are applicable in pediatrics.
Purpose
To evaluate FC’s ability to differentiate IBD from non-IBD pediatric patients, and to understand factors influencing FC in pediatric IBD(pIBD).
Method
Stool FC samples collected on all patients<19 years of age in British Columbia(BC) from May 2020 to August 2022 were run using a Buhlmann ELISA at BC Children’s Hospital(BCCH). The BCCH GI database identified patients with IBD. FC’s ordered by adult IBD providers and patients awaiting endoscopy were excluded. The remaining samples were analysed as non-IBD. The sensitivity(Sn), negative predictive value(NPV) and false positive(FP) of FC were evaluated; comparisons were made using the Wilcoxon rank-sum test and chi-squared.
Result(s)
3506 FC samples met inclusion criteria: 1853 IBD and 1653 non-IBD. 221 IBD samples were from prior to diagnosis, with median (IQR) FC 2615ug/g(1090-4183); median FC for non-IBD patients was 54ug/g(24-122). Using the Buhlmann "normal" cutoff of 80ug/g, the Sn was 0.991 (NPV 0.998) with a FP rate of 37%. Young patients were more likely to have FP's: <6yo's (n=305) had a FP rate of 42% vs 36% in those >6yo (n=1348)(p=0.035). With a FC cutoff of 160ug/g, Sn was 0.973 (NPV 0.996), with a FP rate of 20% (24% <6yo vs 19% >6yo, p=0.025). At a threshold of 250ug/g, Sn was 0.959 (NPV 0.994) with a FP rate of 13% (18% <6yo vs 12% >6yo, p=0.01). For patients <2yo, all 4 new IBD diagnosis had FC>1900ug/g. In the non-IBD population (n=69 samples <2yo), the FP rate was 52%, 30%, and 22% using a threshold of 80, 160, and 250ug/g, respectively.
Evaluating FC as a disease-monitoring tool in IBD (667 patients, 1632 samples) found that at 6, 12, 18, and 24+ months post diagnosis, FC decreased from 750(159-1883), 505(110-1566), 351(87-1379), to 308ug/g(85-1129), respectively. Similarly, the proportion of FC’s <250ug/g increased from 4.1% at diagnosis to 30%, 39.7%, 41.5%, 47% during the follow up period.
Patients with UC/IBD-U had higher FC’s, and were less likely to achieve FC<250ug/g. By 12 months post diagnosis, median FC of CD patients was 347ug/g(96-1150) and UC/IBD-U was 745ug/g(191-2017)(p=0.036), and at 18 months, CD 273ug/g(61-902) vs UC/IBD-U 932ug/g(144-2229)(p<0.001). At 2+yrs, median FC for CD patients was 259ug/g(76-1038) vs 387ug/g(108-1577) for UC/IC (p=0.017).
Patients cared for by an IBD specialist had better FC outcomes vs those managed by non-IBD GIs: median FC at 12 months was 246ug/g(n=79) vs 677ug/g(n=153)(p=0.002), with 51% vs 34% achieving FC<250(p=0.014). Similarly, at 2+yrs post diagnosis, median FC was 243ug/g(n=437) vs 356ug/g(n=407)(p=0.02).
Conclusion(s)
Higher FC thresholds are likely required in younger populations compared to established adult cutoffs. In this pIBD cohort, <50% achieve FC levels <250ug/g.
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None
Disclosure of Interest
None Declared
Abstract
Background
Non-alcoholic fatty liver disease (NAFLD) is strongly associated with cardiovascular disease in the general population. Both NAFLD and cardiovascular diseases seem more frequent ...in patients with inflammatory bowel disease (IBD).
Purpose
We aimed to assess the effect of NAFLD and associated liver fibrosis on the cardiovascular risk in people with IBD.
Method
We prospectively included IBD patients undergoing a routine screening program for NAFLD by transient elastography (TE) with associated controlled attenuation parameter (CAP). NAFLD and significant liver fibrosis were defined as CAP >275 dB/m and liver stiffness measurement (LSM) by TE ≥8 kPa, respectively. Nonalcoholic steatohepatitis (NASH) with liver fibrosis was defined as Fibroscan-aspartate aminotransferase (AST) score (FAST) >0.35. Cardiovascular risk was assessed with the atherosclerotic cardiovascular disease (ASCVD) risk estimator proposed by the American Heart Association and computed from age, sex, race, lipid pattern, blood pressure, diabetes treatment and smoking. Based on the American Heart Association guidelines, the 10-year cardiovascular risk by ASCVD was categorized as low if <5%, borderline if 5%–7.4%, intermediate if 7.5%–19.9% and high if ≥20% or if previous cardiovascular event.Predictors of intermediate-high cardiovascular risk were investigated by multivariable logistic regression analysis.
Result(s)
We included 405 patients with IBD (54% female; mean age 45+15 years; mean BMI 26+5 Kg/m2; 31% with ulcerative colitis; 7% with diabetes; 14% with hypertension). Overall, 278 (68%), 23 (6%), 47 (12%) and 57 (14%) were categorized as at low, borderline, intermediate and high ASCVD risk, respectively. NAFLD and significant liver fibrosis were found in 129 (32%) and 35 (9%) patients, respectively. NASH with fibrosis was found in 11 (3%) patients. Patients with NAFLD and with significant liver fibrosis diagnosed by TE with CAP had higher proportion of intermediate-high ASCVD risk category (see Figure). These findings were confirmed also in young IBD patients <55 years old with NAFLD. No difference in ASCVD risk was detected for FAST score. After adjusting for IBD disease activity, significant liver fibrosis and BMI, predictors of intermediate-high ASCVD risk were NAFLD (adjusted odds ratio aOR 2.97, 95% confidence interval CI 1.56–5.68), IBD duration (aOR 1.55 per 10 years, 95% CI 1.22–1.97), and ulcerative colitis (aOR 2.32, 95% CI 1.35–3.98). Only 30% of IBD patients classified as intermediate-high ASCVD risk were on statin treatment, with no difference between patients with and without NAFLD.
Image
Conclusion(s)
NAFLD increases cardiovascular risk, independently of age, IBD-related factors and BMI.
A potential deliverable of our finding is the targeted cardiovascular assessment in IBD patients with NAFLD and appropriate initiation of statin, particularly if they have longer IBD duration and ulcerative colitis.
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None
Disclosure of Interest
None Declared
Abstract
Background
The colonic stem cell niche is established by a gradient of WNT, R-spondin, BMP factors and their antagonists along the colonic epithelial vertical axis. Telocytes (TCFoxL1+) are ...mesenchymal cell forming a 3D hub underneath the epithelium, identified as an important source of niche factors. Specifically, they express non-canonical (nc) WNT factors and are the richest source of BMPs. Disruption of the BMPs gradient has been shown to be related to the development of several gastrointestinal diseases like Inflammatory Bowel Diseases (IBD). Such chronic inflammation drives the onset of Colitis-Associated Cancer (CAC) in about 60% of IBD patients. We previously showed that following a chronic inflammatory stress, 50% of the KO mouse for the BMP receptor 1a in colon telocytes (Bmpr1a△FoxL1+) presented malignant epithelial transformations. These cancer-like regions showed an aberrant epithelial b-catenin localization and an enlargement of the double positive α-SMA+/Vimentin+ mesenchymal population.
Purpose:
Loss of BMP signaling in TCFoxL1+ affects the mesenchymal-epithelial crosstalk and makes the colonic epithelium vulnerable to injuries, promoting/perpetuating inflammation fostering CAC onset.
Method
Following a DSS-based chronic inflammatory challenge in mutant and control mice, TCFoxL1+ ultrastructure was analyzed using transmission electron microscopy. Expression levels of members of the WNT-BMP axis (BMPs, WNTs and associated antagonists) were evaluated by qPCR in tumor-like areas and adjacent tissue. YAP cellular localization was evaluated by immunofluorescence in colon after chronic DSS challenge in Bmpr1a△FoxL1+ mice and controls. To differentiate cancer-associated fibroblasts (CAFs) subtypes, myCAF (myofibroblastic) and iCAF (inflammatory), in tumor-like region and adjacent tissue, we used co-staining against gp38, ICAM, Tagln and αSMA.
Result(s)
Following a chronic DSS-challenge, electron microscopy analysis demonstrated that TCFoxL1+ in the control mice exhibited a shortening and erosion in their telopodes (Tp). TCFoxL1+ in Bmpr1a△FoxL1+ mice tumor-like regions presented an expanded endoplasmic reticulum with fragmented and dilated Tp. A significant increase in BMP 4, 5 and 7 and in Wnt5 (nc) was detected in Bmpr1a△FoxL1+ mice compared to controls. Confocal analysis revealed a strong nuclear accumulation of YAP in cancer-like regions in mutant mice compared to controls. Finally, tumour-like regions presented an heterogeneous distribution of iCAF and myCAF compared to controls.
Conclusion(s)
These results exposed that the disruption of TCFoxL1+ associated BMP signaling disturbs the WNT-BMP gradient essential for the optimal maintenance of the SC niche and thus impacting epithelial regeneration when under stress. Thus, defective TCFoxL1+ assume a key role in the poor regeneration process of the epithelium which in the end promotes the development and progression of CAC.
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CIHR
Disclosure of Interest
None Declared
Abstract
Background
The NAIP-NLRC4 inflammasome is an important innate defence mechanism in intestinal epithelial cells (IECs) that protects the gut from invasive pathogens. NAIP-NLRC4 activation ...triggers pyroptotic cell death, releases active interleukin (IL)-18, and promotes the expulsion of infected enterocytes. Dysregulated inflammasomes result in exaggerated inflammation of the intestinal mucosa which is characterisitic of inflammatory bowel disease (IBD). There is increasing evidence that inflammasomes in IEC behave differently than in immune cells. However, a majority of inflammasome pathway characterization is studied immune cells. In addition, NAIP-NLRC4 activity in macrophages has been shown to be important for regulating the expression of nitric oxide (Nos2). The potential transcriptomic impact of NAIP-NLRC4 activity in IECs has yet to be explored.
Purpose
We will use intestinal orgnaoids to systematically characterize the NAIP-NLRC4 inflammasome pathway and identify the transcriptomic impact of NAIP-NLRC4 activity in IECs. To better define the role of NAIP-NLRC4 activity during a physiological infection, we have developed a novel ex vivo model of Shigella infection in 3D organoids.
Method
Organoids were derived from the ileal crypts of wild type (WT) and Nlrc4-/-, Casp1-/-, Pycard-/-, or Tlr5-/- mice and stimulated with Pam3CSk4, LPS, or flagellin. Inflammasome activation was assessed by Western blot (WB) and propidium iodide uptake. WT and Nlrc4-/- organoids were infected with WT or a non-invasive Shigella mutant and the inflammasome response was evaluated by WB and a colony forming unit assay. WT and Nlrc4-/- organoids were stimulated with flagellin and gene expression was assessed by RT-qPCR.
Result(s)
Basolateral organoid stimulation with bacterial ligands revealed a novel response of IECs to bacterial flagellin that results in pyroptosis and IL-18 processing. Basolateral internalization of flagellin occurred in a TLR5-independent manner. Inflammasome activation by flagellin was fully abrogated in Nlrc4-/- and Casp1-/- organoids while only IL-18 processing was affected in Pycard-/- organoids. Infection with only WT Shigella induced inflammasome activation in an NLRC4-dependent manner. Interestingly, flagellin stimulation of WT but not Nlrc4-/- organoids led to increased expression of Nos2. Furthermore, Nlrc4-/- organoids had significantly lower expression levels of cytokine genes Ccl20, Cxcl1, and Tnf following inflammasome activation.
Conclusion(s)
Our study demonstrates an integral role for epithelial NAIP-NLRC4 inflammasomes in the response to bacterial flagellin and Shigella infection. We have uncovered a novel response of IECs to basolateral flagellin stimulation and revealed that NAIP-NLRC4 is important for the transcriptional regulation of inflammatory genes. Further work will examine how NAIP-NLRC4 activation controls inflammation and epithelial integrity by analyzing the NLRC4-dependent transcriptome and the effects on cell proliferation, differentiation, and barrier integrity.
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CCC, CIHR
Disclosure of Interest
None Declared
Abstract
Background
The circadian clock is a time keeping mechanism that drives daily rhythms in the body, including behavior and gastrointestinal digestion, inflammation, and metabolism. These daily ...rhythms are endogenous to cells, and they must adjust themselves to cyclic environmental cues like feeding schedules and light dark cycles. However, when the cyclical environment is chrono-disrupted (ex. shiftwork and jetlag), the temporal organization in the body is lost. Chrono-disruption is linked to an increase in the risk of Inflammatory Bowel Disease (IBD); studies in rodents shown that IBD is more severe in mice without a circadian clock and intestinal regeneration is compromised
Purpose
Recent studies have shown that time-restricted feeding (TRF, the provision of food at times of peak behavior activity – without caloric restriction), that aligns with circadian behavior improves daily organization in the body and health. Therefore, we hypothesized that time-restricted feeding would be an effective strategy to ameliorate IBD and improve regeneration in chrono-disrupted animals.
Method
To test this, we used the DSS induction of colitis in mutant mice without clock function (Bmal1-/-) and controls with an intact clock (Bmal1+/+) and compared ad libitum feeding vs. TRF conditions (food available from 7pm to 7am).
Result(s)
Our results show that TRF ameliorates IBD symptoms and protects against epithelium colon damage in animals without clock function. Our preliminary results suggest that TRF may function by increasing the expression of the anti-inflammatory signal PPARg in the colon. Currently, we are testing if the circadian clock controls the daily expression of PPARg in vivo and in vitro. To further test the effect of TRF during regeneration after colitis, we tested proliferation (Ki67) and regeneration in the colon. (Hopx). Our results show that TRF increases the number of Ki67 positive cells and the expression of fetal-like regenerative precursor Hopx in both animals with and without a functional clock.
Conclusion(s)
Together these data suggest that TRF enhances regeneration in the colon after colitis. Moving forward, we are testing transcriptional pathways that are activated during regeneration in animals under TRF. Our results show that time restricted feeding can ameliorate IBD and highlight the role of circadian rhythms in regeneration and intestinal stem cell biology.
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CIHR, Other
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Crohn's and Colitis Canada
Disclosure of Interest
None Declared
Abstract
Background
Individuals with IBD have a reduced quality of life and while treatment has improved, a cure remains elusive. Interleukin-4 differentiated macrophages (M(IL4)s) from healthy ...donors and IBD patients in remission promote epithelial wound repair and reduce the severity of colitis in a mouse model: findings that support the autologous transfer of M(IL4)s to treat IBD. However, issues remain if this novel therapy is to advance to clinical evaluation: a much greater understanding of the possible heterogeneity within the human M(IL4) population, the biology of any sub-groups therein and the biology of any M(IL4) subtypes is needed.
Purpose
To determine if cryopreservation, sex, and disease status in Crohn's disease (CD) impact the phenotype and function of M(IL4)
Method
Blood mononuclear cells were collected from healthy donors, and individuals with active CD or in remission (n=6/group), and differentiated to macrophages with GMP-grade recombinant human M-CSF (10 ng/ml, 7 days). Macrophages (2.5 x105) were activated with IL-4 (10ng/ml, 48h). Fresh and cryopreserved M(0) and M(IL4) were stained using a panel of 18 metal-conjugated antibodies. Data were acquired on a Helios CyTOFII mass cytometer. FCS files were imported into Cytobank for manual gating (viable CD45+CD3-CD19- singlet cell events) and exported for meta-clustering in R with the CATALYST package using the FlowSOM function with 10 metaclusters.
Result(s)
FlowSOM analysis on M(IL4) from healthy donors indicated clear differences between M(0) and M(IL4), with 8 metaclusters >1.5% of M(IL4)s: the most abundant metaclusters express high and intermediate CD206, PDL1, CD11b, CD33, CD64, HLA-DR and low CD14 levels. There were no significant differences in these M(IL4)s metaclusters between males and females, or fresh vs. cryopreserved cells using this characterization strategy. M(IL4) from patients with active CD, were markedly different with the absence of three high-intermediate CD206+PDL1+ metaclusters and a general lack of CD206+ metaclusters compared to healthy controls. Surprisingly, and despite M(IL4)s from CD patients in remission showing increased CD206 mRNA, CYTOF analysis revealed that the macrophages were more similar to those from individuals with active disease than those from healthy controls
Conclusion(s)
Here human macrophages activated with IL4 in vitro are revealed as a heterogeneous population, and subsets characterized by increased high CD206 expression are generally absent in active Crohn’s disease. Intriguing while prior experiments with M(IL4) from CD patients in remission reduced the severity of colitis in rag1-/- mice, the characterization of these cells via 18 protein markers indicates they are more like M(IL4)s from patients with active disease compared to healthy controls. While presenting autologous M(IL4) transfer as a novel approach for IBD, a much fuller understanding of the phenotype and function of M(IL4)s (and sub-populations therein) is required before this would become a therapeutic option
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CAG
Disclosure of Interest
None Declared
Abstract
Background
Live biotherapeutic products (LBPs) offer a more rationalized and multitargeted approach to treating gastrointestinal diseases. BioColoniz and BioPersist are two LBPs derived from ...the parental strains L. reuteri and E. coli Nissle 1917 (EcN), respectively. The parental strains are known to offer some benefit in preventing relapses in IBD patients however the results are heterogeneous. To overcome this, the parental strains were approached as LBPs by introducing traits to thrive under the inflammatory conditions of the colon. Therefore, our aim is to characterize the role of these LBPs in IBD.
Purpose
To evaluate the therapeutic effect of the LBPs BioColoniz and BioPersist in an acute model of colitis.
Method
Female C57Bl/6 mice were treated with BioColoniz or BioPersist via oral gavage for three consecutive days prior to DSS challenge. Then mice were exposed to 3.5% DSS via drinking water for seven days. As controls, we also included mice treated with vehicle or the parental strains L. reuteri or EcN. In order to compare the effect of LBPs in the onset of acute colitis to current maintenance therapies for UC, we also exposed another group of mice to DSS and simultaneously administer 5-ASA. Mice were monitored daily for signs of disease and at the end of the experiment, colon tissue was collected for histopathological and molecular analysis.
Result(s)
The administration of BioColoniz and BioPersist delayed and decreased the colitic phenotype of mice exposed to DSS. Differences in signs of disease, such as diarrhea and weight loss, were evident by day 4 for vehicle or 5-ASA groups, whereas mice in the LBPs groups were still gaining weight. When analyzing the histopathological changes, mice in the LBPs groups presented lower scores when compared to the vehicle and 5-ASA groups. Specifically, mice treated early with BioColoniz or BioPersist presented a more preserved mucosal architecture with visible crypts. Although 5-ASA-treated mice still had vestiges of crypts, the damage in the mucosal architecture was more severe, similar to the observed in mice treated with the parental strains EcN and L. reuteri. We also looked at the expression of proinflammatory cytokines, finding an increase in TNFα, IFNγ, and IL-17a in mice treated with 5-ASA but not in mice treated with LBPs. However, the expression of protective factors such as mucin Muc2 or the antimicrobial peptide Reg3γ was similarly high in 5-ASA and LBP-treated mice when compared to vehicle or parental strains groups, suggesting some therapeutic commonalities between 5-ASA and our LBPs.
Conclusion(s)
The early administration of the LBPs BioColoniz and BioPersist protect mice from severe acute colitis, being more protective than 5-ASA. Since some differences and similarities were observed between the LBPs- and 5-ASA-treated mice, such as crypt preservation versus increased expression of some protective factors, the next step will aim to identify which mechanisms are specifically triggered by the LBPs.
Disclosure of Interest
None Declared
