Urinary Fractional Excretion Research Articles

P1338 Defining the preclinical changes of microbial composition, gut barrier function, and subclinical inflammatory markers associated with future risk of ulcerative colitis

Abstract Background Ulcerative colitis (UC) is a chronic inflammatory condition of the rectum and colon with a complex aetiology involving genetic, immune, and microbial factors. Less is known about the pre-disease phase of UC compared to Crohn’s disease (CD). Investigating the preclinical biomarkers that precede the development of UC may provide insights into its pathogenesis. Methods Participants were recruited as part of the CCC-GEM project, a prospective cohort study following healthy first-degree relatives (FDR) of patients with CD. Baseline gut inflammation was assessed using faecal calprotectin (FCP), with a cut-off of 100ug/g, while baseline gut permeability was assessed by the fractional urinary excretion ratio of lactulose-to-mannitol (LMR). Baseline faecal microbiome composition was analyzed using 16s rRNA sequences, and functional pathways were inferred using PICRUSt2. Cox proportional hazards models were used to assess the association of baseline variables with UC onset, adjusting for age, sex, and family clustering. Significance was determined using p-values and false discovery rate-adjusted q-values. Results Among 3,596 FDRs, 16 developed UC during a median follow-up of 6.88 years. FDRs with elevated baseline FCP had a 3.1-fold higher risk (p=0.02) of developing UC, while no significant association was observed between baseline LMR and UC onset. The relative abundance of genus Bilophila (HR=0.33 per SD, q=0.010) and Bifidobacteria (HR=0.16, q=0.02) were associated with UC risk, but lost significance after adjusting for FCP. Interestingly, the relative abundances of genera Alistipes (HR=0.15, q=0.0007), Phascolarctobacterium (HR=0.08, q=0.027), Christensenellaceae R.7 group (HR=8.37, q=0.02) and Angelakisella (HR=5.28, q=0.004) remained significantly associated with risk of UC even after adjusting for FCP. Thirty-seven pathways, predominantly involved in specific metabolic processes such as carbohydrates, lipid, or folate metabolism, were also associated with UC risk. Fur example, pathways involving hydroxymethylglutaryl-CoA reductase (HR=0.44, q=0.03), choline monooxygenase (HR=0.22, q=0.005), and 3-phenylpropionate/trans-cinnamate dioxygenase (HR=4.70, q=0.001) remained significantly associated with UC risk even after adjusting for FCP. Conclusion We report that elevated FCP, but not LMR, was significantly associated with future risk of UC. Furthermore, specific microbial taxa and functional pathways exhibited strong associations with future risk of UC, independent of FCP. These microbial signatures were distinct from the previously reported pre-CD microbial changes, suggesting a unique gut microbial contribution to the development of UC. References Raygoza Garay JA, Turpin W, Lee S-H, et al. Gastroenterology 2023;165:670–681.

DOP041 The association between fermentable dietary fibre intake and risk of developing Crohn’s Disease: Results from the GEM Project

Abstract Background The cause of Crohn’s Disease (CD) remains unclear; however evidence suggests that dietary factors play a key role. With a rising incidence and no definitive cure, identifying modifiable risk factors is critical in preventing CD development. We aimed to study the association between fermentable dietary fibre intake and the future development of CD in an at-risk population. Methods Participants were recruited as part of the Crohn’s and Colitis Canada GEM study, a prospective cohort study of healthy first-degree relatives of patients with CD. At enrolment, a validated food frequency questionnaire was administered. Pectin, β-glucan, inulin, fructooligosaccharides (FOS), and arabinoxylan intake were quantified, and values energy-adjusted. Survival analysis was used to test the association between intake of fibre subtypes and risk of CD development. Generalized estimating equation tested the association between fibre intake and baseline impaired intestinal permeability (fractional urinary excretion of lactulose to mannitol ratio (LMR) >0.025); subclinical gut inflammation (faecal calprotectin (FCP) >250 μg/g); and faecal microbiome composition via 16s rRNA sequencing. Results 76 of the 2,659 participants developed CD with a median follow-up of 8.9 years (IQR=5.7-12.3). Median age at recruitment was 17 years (IQR=12-25). 47% were male. Higher intake of inulin (HR = 0.78; 95% CI=0.61–0.99; p=0.04) and β-glucan (HR = 0.78; 95% CI=0.61–0.99; p=0.04) were associated with a reduced risk of CD development. No fibre subtype was associated with a higher risk of CD development. Lower intake of pectin, inulin, FOS and β-glucan was associated with impaired intestinal permeability (0.005<p<0.04). Lower inulin intake was associated with high FCP levels (p = 0.046). Higher pectin intake was associated with increased alpha diversity of faecal microbiome (p = 0.028). Higher inulin, FOS and pectin intake was associated with a shift in microbial taxa previously shown to be protective against CD onset in the GEM cohort; a reduction in Ruminococcus torques and an increase in NK4A214 group (phylum Firmicutes) presence (3.25×10−10<q<0.04). Conclusion This study demonstrates that dietary intake of fermentable fibre subtypes is associated with a reduced risk of CD development as well as with pleiotropic changes in CD risk biomarkers including microbiome, intestinal permeability, and subclinical inflammation. This suggests that the potential benefit of fermentable fibres on CD risk may involve changes in barrier function, and microbiome composition and function. Targeted interventions based on these findings could prove effective in preventing the onset of CD.

Short Term Treatment Monitoring of Renal and Inflammatory Biomarkers with Naturally Occurring Leishmaniosis: A Cohort Study of 30 Dogs.

Various inflammatory and renal biomarkers have already been assessed for monitoring the response to anti-leishmanial therapy in canine leishmaniosis. This study assessed the parasite load, various inflammatory and renal biomarkers pre- and post-treatment, and any association between the studied variables and the degree of disease severity at diagnosis. This is a prospective cohort study of 30 client-owned dogs with leishmaniosis, classified according to LeishVet's guidelines as stage I (n = 2), stage IIa (n = 7), stage IIb (n = 6), stage III (n = 8), and stage IV (n = 7). In addition to Leishmania real-time PCR in the bone marrow, blood and urine, previously studied biomarkers, and several inflammatory and renal markers never investigated in canine leishmaniosis, such as fibrinogen, antithrombin, urinary fractional excretion of sodium, and urinary amylase-to-creatinine ratio were measured pre- and post-treatment (meglumine antimoniate or miltefosine + allopurinol). A positive Leishmania real-time PCR in the blood at diagnosis predicted a positive Leishmania real-time PCR in the bone marrow post-treatment (p = 0.003). Following treatment, antithrombin and urinary amylase-to-creatinine ratio were significantly changed (p < 0.001, respectively). Urinary amylase-to-creatinine ratio, total iron-binding capacity, and antithrombin were the variables most strongly associated with disease severity (p < 0.005, respectively). Urinary amylase-to-creatinine ratio can be a useful marker to monitor treatment response and to classify the degree of disease severity.

Effect of D,L methionine and ammonium chloride on urine acidification, urinary fractional excretion of calcium, and blood bicarbonate in clinically healthy goats.

Urinary acidification with ammonium chloride (AC) for urolith dissolution is a common treatment for goats with urolithiasis. Studies have reported increased fractional excretion of calcium (FECa) following AC administration, which could increase calcium-based urolithiasis. D,L methionine (MET) may result in similar acidification with less calcium excretion. To compare the effects of orally administered MET and AC on urine and blood pH, FECa, and blood HCO3- concentrations in male goats. Prospective, randomized, crossover study. 12 healthy, 5-to-6-month-old Boer-cross wethers were administered 200 mg/kg of AC or MET orally for 14 days with a 7-day washout period between trials. Venous blood and urine samples were collected every 2 days. The effects of treatment and treatment day on urine and blood pH, HCO3-, and FECa were assessed using linear mixed models. Ammonium chloride and MET lowered least squares means (LSM) urine pH on day 6 (LSM, 7.49; 95% CI, 6.44 to 8.54), 8 (LSM, 7.78; 95% CI, 6.73 to 8.83), and 10 (LSM, 7.53; 95% CI, 6.49 to 8.58) when compared to day 0 (LSM, 8.23; 95% CI, 7.18 to 9.28). Some goats' urine indicated acidification (pH < 7.0) in the first phase of the trial; however, for the entire trial, a significant treatment effect was not detected on urine pH, blood pH, blood HCO3- or log10 FECa. Ammonium chloride and MET acidified urine of some goats. Dietary cation-anion difference should be considered when treating healthy goats to acidify their urine.

Healthy First-Degree Relatives From Multiplex Families vs Simplex Families Have a Higher Subclinical Intestinal Inflammation, a Distinct Fecal Microbial Signature, and Harbor a Higher Risk of Developing Crohn’s Disease

Unaffected first-degree relatives (FDRs) from families with ≥2 affected FDRs with Crohn's disease (CD, multiplex families) have a high risk of developing CD, although the underlying mechanisms driving this risk are poorly understood. We aimed to identify differences in biomarkers between FDRs from multiplex vs simplex families and investigate the risk of future CD onset accounting for potential confounders. We assessed the Crohn's and Colitis Canada Genetic Environmental Microbial cohort of healthy FDRs of patients with CD. Genome-wide CD-polygenic risk scores, urinary fractional excretion of lactulose-to-mannitol ratio, fecal calprotectin (FCP), and fecal 16S ribosomal RNA microbiome were measured at recruitment. Associations between CD multiplex status and baseline biomarkers were determined using generalized estimating equations models. Cox models were used to assess the risk of future CD onset. There were 4051 participants from simplex families and 334 from CD multiplex families. CD multiplex status was significantly associated with higher baseline FCP (P= .026) but not with baseline CD-polygenic risk scores or the lactulose-to-mannitol ratio. Three bacterial genera were found to be differentially abundant between both groups. CD multiplex status at recruitment was independently associated with an increased risk of developing CD (adjusted hazard ratio, 3.65; 95% confidence interval, 2.18-6.11, P < .001). Within FDRs of patients with CD, participants from multiplex families had a 3-fold increased risk of CD onset, a higher FCP, and an altered bacterial composition, but not genetic burden or altered gut permeability. These results suggest that putative environmental factors might be enriched in FDRs from multiplex families.

Urinary Concentrations of BPA and Analogous Bisphenols (BPF and BPS) among School Children from Poland: Exposure and Risk Assessment in the REPRO_PL Cohort

Bisphenols are a group of chemical compounds widely used in the industry, e.g., in the production of plastics, and as precursor of epoxy resins. The three bisphenols investigated in this study, namely bisphenols A (BPA), F (BPF) and S (BPS), seem to have endocrine activity in the same order of magnitude. Thus, an exposure and risk assessment was performed based on urinary concentrations of these bisphenols in 150 urine samples of 7-year-old children from the Polish Mother and Child Cohort Study (REPRO_PL). Samples were analyzed using high performance liquid chromatography with online sample clean-up coupled to tandem mass spectrometry (online-SPE-LC-MS/MS). BPA, BPF, and BPS were above the LOQ (0.25 µg/L) in 100%, 95%, and 20% of the samples, respectively, with median concentrations of 2.5 µg/L, 1.0 µg/L and < 0.25 µg/L (below the LOQ), respectively. Median Daily Intakes (DIs) were calculated: 45.9 ng/kg bw/day for BPA, at maximum 33.1 ng/kg bw/day for BPF (depending on the urinary excretion fractions used for calculation) and below 4.4 ng/kg bw/day for BPS (calculated at LOQ/2). Hazard quotients (HQ) at the 95th percentile of 0.011 for BPF and 0.63 for BPS did not indicate health risks. However, based on the TDI of 0.2 ng/kg bw/day newly established by EFSA, the HQ for BPA was above 1 in 100% of the samples, thus indicating the exceedance of safe exposure levels. No statistically significant associations were found between bisphenol concentrations and socio-demographic and life-style characteristics, although environmental tobacco smoke at 7 years of age, maternal educational level and socio-economic status showed a positive trend. Further studies are required in order to recognize other BPA replacement exposures among children.

Association of urinary excretion rates of uric acid with biomarkers of kidney injury in patients with advanced chronic kidney disease.

The potential influence of hyperuricemia on the genesis and progression of chronic kidney disease (CKD) remains controversial. In general, the correlation between blood levels of uric acid (UA) and the rate of progression of CKD is considered to be modest, if any, and the results of relevant trials oriented to disclose the effect of urate-lowering therapies on this outcome have been disappointing. Urinary excretion rates of UA could reflect more accurately the potential consequences of urate-related kidney injury. Using a cross-sectional design, we investigated the correlation between different estimators of the rates of urinary excretion of UA (total 24-hour excretion, mean urinary concentration, renal clearance and fractional excretion)(main study variables), on one side, and urinary levels of selected biomarkers of kidney injury and CKD progression (DKK3, KIM1, NGAL, interleukin 1b and MCP)(main outcome variables), in 120 patients with advanced CKD (mean glomerular filtration rate 21.5 mL/minute). We took into consideration essential demographic, clinical and analytic variables with a potential confounding effect on the explored correlations (control variables). Spearman's rho correlation and nonlinear generalized additive regression models (GAM) with p-splines smoothers were used for statistical analysis. Multivariate analysis disclosed independent correlations between urinary UA concentrations, clearances and fractional excretion rates (but not plasma UA or total 24-hour excretion rates of UA), on one side, and the scrutinized markers. These correlations were more consistent for DKK3 and NGAL than for the other biomarkers. Glomerular filtration rate, proteinuria and treatment with statins or RAA axis antagonists were other independent correlates of the main outcome variables. Our results support the hypothesis that urinary excretion rates of UA may represent a more accurate marker of UA-related kidney injury than plasma levels of this metabolite, in patients with advanced stages of CKD. Further, longitudinal studies will be necessary, to disclose the clinical significance of these findings.

Unveiling the heightened susceptibility: Exploring early hypophosphatemia in critically ill trauma patients

Phosphorus is a vital mineral crucial for various physiological functions. Critically ill trauma patients frequently experience hypophosphatemia during the immediate post-traumatic phase, potentially impacting outcomes. This study aims to investigate the incidence of early hypophosphatemia in critically major trauma patients. In this prospective observational study, trauma patients admitted to the intensive care unit (ICU) within one day were enrolled. These patients were categorized into Hypo-P groups and Non-hypo groups based on the development of new-onset hypophosphatemia within 72h after feeding. The primary outcome assessed was the incidence of new-onset hypophosphatemia. The secondary outcomes included ICU and hospital stay, ventilation duration, and mortality. 76.1% of patients developed a new onset of hypophosphatemia within 72h after feeding. The Hypo-P group had significantly longer ICU stays (8.1 days±5.5 vs. 4.4 days±3.1; p=0.0251) and trends towards extended hospital stay, ventilation duration, and higher mortality. Additionally, they demonstrated significantly higher urine fractional excretion of phosphate (FEPO4) on the first ICU day (29.2%±14.23 vs. 19.5%±8.39; p=0.0242). Critically ill trauma patients exhibited a significantly higher incidence of early hypophosphatemia than typical ICU rates, indicating their heightened vulnerability. The significantly high urine FEPO4 underscores the crucial role of renal loss in disrupting phosphate metabolism in this early acute phase after trauma. A significant correlation was observed between hypophosphatemia and longer ICU stays. Monitoring and managing phosphate levels may influence outcomes, warranting further investigation.

#68 Novel clinical application of urinary angiotensin-converting enzyme assay in renal sarcoidosis: a retrospective observational study

Abstract Background and Aims Renal manifestations of sarcoidosis are observed in approximately 1–5% and characterized mainly by granulomatous interstitial nephritis, which could lead to kidney failure. Angiotensin-converting enzyme (ACE) reflects the presence of granuloma; hence, serum ACE and tubular injury markers are measured in renal sarcoidosis (RS); however, they either have low sensitivity or low specificity. Although its practicality has not been established, the use of ACE levels in cerebrospinal fluid has been reported for neuro-sarcoidosis. Therefore, we hypothesized that urinary ACE could reflect granuloma presence in the kidneys and could be a disease-specific marker for RS. Method This was a single-center, retrospective observational study. We evaluated clinical characteristics and kidney biopsy findings from electronic medical record surveys and kidney biopsy specimens. The control groups included sarcoidosis without renal involvement (SAR-control) and tubulointerstitial nephritis without a sarcoidosis etiology (TIN-control). Serum and urinary ACE levels were measured, and urinary fractional excretion of ACE (FEACE) was investigated. Serum ACE assay and urinary ACE levels were measured using the colorimetric method with p-hydroxybenzoyl-glycyl-L-histidyl-L-leucine. A total of six patients (three with RS; one with sarcoidosis as controls; and two with TIN controls) with undetectable urinary ACE levels were excluded from the urinary ACE and FEACE analyses. Tubulointerstitial lesions were visually assessed for essentially normal (%), inflammatory cell infiltrates (%), tubular atrophy (%), and interstitial fibrosis (%) by several board-certified nephrologists, and analyzed the correlation of these lesions with each marker. Results This study included 18 patients with RS, 18 as SAR-control, and 10 as TIN-control. Serum ACE, urinary ACE, and FEACE levels in the RS group were significantly higher than those in the control groups. Receiver operating characteristic analysis identified FEACE and serum ACE levels as useful biomarkers for diagnosing RS among patients with both controls, respectively. In the RS group, FEACE was positively correlated with the degree of tubulointerstitial injury (r = 0.69, P = 0.0045), and the cutoff value of FEACE for diffuse tubulointerstitial injury (50.0% or more) was 0.39%, with a sensitivity and specificity of 100.0%. Furthermore, obvious positive correlations were observed among FEACE, inflammatory cell infiltrates (r = 0.53, P = 0.044), and interstitial fibrosis (r = 0.56, P = 0.029) in the RS group but not in the TIN-control. Conclusion Combination with serum and urinary ACE could help diagnose and assess disease severity among patients with RS.

Phosphocalcic metabolism and its potential association with biomarkers of kidney disease in dogs with spontaneous hyperadrenocorticism

The pathogenesis of increased serum phosphate concentration and proteinuria in dogs with spontaneous hyperadrenocorticism (HAC) is unclear. A potential link between proteinuria and calcium/phosphate metabolism has never been studied in dogs with HAC. The aims of the study were: (1) To evaluate calcium/phosphate metabolism in dogs with spontaneous HAC and compare to healthy dogs as well as to dogs with non-HAC illness; (2) to look for associations between markers of calcium/phosphate metabolism and biomarkers of kidney disease in dogs with HAC. Fifty-four dogs were included in the study, classified as HAC (n=27), non-HAC disease (n=17), and healthy (n=10). Serum calcium, phosphate, 25(OH)Vitamin D, 1,25(OH)2Vitamin D, plasma intact parathyroid hormone concentration (iPTH), FGF23, and urinary fractional excretion of calcium and phosphate were evaluated in all dogs at diagnosis and compared between each group. The correlation between these variables and urine protein-to-creatinine ratio (UPC) and urinary N-acetylglucosaminidase-to-creatinine ratio (uNAG/C) was evaluated in the HAC group. Medians [range] of serum phosphate concentration, urinary fractional excretion of calcium (FE(Ca)), and iPTH were significantly higher in dogs with HAC than in dogs with non-HAC illness (P<0.01) and healthy dogs (P<0.01). Increased 1,25(OH)2Vitamin D/25(OH)Vitamin D was also observed (P<0.001). In HAC group, UPC was significantly negatively correlated with 25(OH)Vitamin D (r(s): -0.54; P<0.01). Urinary NAG/C was significantly positively correlated with serum phosphate (r(s): 0.46; P=0.019). Increased serum phosphate, urinary excretion of calcium, and hyperparathyroidism were observed in dogs with HAC. Vitamin D metabolism may be shifted towards increased 1-alpha hydroxylation.

Environmental Factors Associated With Risk of Crohn’s Disease Development in the Crohn’s and Colitis Canada - Genetic, Environmental, Microbial Project

To date, it is unclear how environmental factors influence Crohn's disease (CD) risk and how they interact with biological processes. This study investigates the association between environmental exposures and CD risk and evaluates their association with pre-disease biomarkers. We studied 4289 healthy first-degree relatives (FDRs) of patients with CD from the Crohn's and Colitis Canada - Genetic, Environmental, Microbial (CCC-GEM) project. Regression models identified environmental factors associated with future CD onset and their association with pre-disease biological factors, including altered intestinal permeability measured by urinary fractional excretion of lactulose to mannitol ratio (LMR); gut inflammation via fecal calprotectin (FCP) levels; and fecal microbiome composition through 16S rRNA sequencing. Over a 5.62-year median follow-up, 86 FDRs developed CD. Living with a dog between ages 5 and 15 (hazard ratio [HR], 0.62; 95% confidence interval [CI], 0.40-0.96; P= .034), and living with a large family size in the first year of life (HR, 0.43; 95% CI, 0.21-0.85; P= .016) were associated with decreased CD risk, whereas having a bird at the time of recruitment (HR, 2.78; 95% CI, 1.36-5.68; P= .005) was associated with an increased CD risk. Furthermore, living with a dog was associated with reduced LMR, altered relative abundance of multiple bacterial genera, and increased Chao1 diversity, whereas bird owners had higher FCP levels. Large family during participants' first year of life was associated with altered microbiota composition without affecting FCP or LMR. This study identifies environmental variables associated with CD risk. These variables were also associated with altered barrier function, subclinical inflammation, and gut microbiome composition shifts, suggesting potential roles in CD pathogenesis.

Toxicokinetics of homosalate in humans after dermal application: applicability of oral-route data for exposure assessment by human biomonitoring

Homosalate (HMS) is a UV filter used in sunscreens and personal care products as a mixture of cis- and trans-isomers. Systemic absorption after sunscreen use has been demonstrated in humans, and concerns have been raised about possible endocrine activity of HMS, making a general population exposure assessment desirable. In a previous study, it was shown that the oral bioavailability of cis-HMS (cHMS) is lower than that of trans-HMS (tHMS) by a factor of 10, calling for a separate evaluation of both isomers in exposure and risk assessment. The aim of the current study is the investigation of HMS toxicokinetics after dermal exposure. Four volunteers applied a commercial sunscreen containing 10% HMS to their whole body under regular-use conditions (18–40 mg HMS (kg bw)−1). Parent HMS isomers and hydroxylated and carboxylic acid metabolites were quantified using authentic standards and isotope dilution analysis. Further metabolites were investigated semi-quantitatively. Elimination was delayed and slower compared to the oral route, and terminal elimination half-times were around 24 h. After dermal exposure, the bioavailability of cHMS was a factor of 2 lower than that of tHMS. However, metabolite ratios in relation to the respective parent isomer were very similar to the oral route, supporting the applicability of the oral-route urinary excretion fractions for dermal-route exposure assessments. Exemplary calculations of intake doses showed margins of safety between 11 and 92 (depending on the approach) after single whole-body sunscreen application. Human biomonitoring can reliably quantify oral and dermal HMS exposures and support the monitoring of exposure reduction measures.

The Uricosuric Effect of SGLT2 Inhibitors Is Maintained in the Long Term in Patients with Chronic Kidney Disease and Type 2 Diabetes Mellitus.

(1) Background: Sodium-glucose co-transporter 2 inhibitors (SGLT2is) increase uric acid excretion. The intensity of uricosuria is linked to glycosuria. (2) Methods: We aim to analyze the effect of SGLT2 inhibitors on urinary fractional excretion (FE) of uric acid and glucose in patients with type 2 diabetes mellitus (T2DM) and chronic kidney disease (CKD) in a single-center retrospective study with patients with T2DM and CKD who started on treatment with SGLT2is. Patients on renal replacement therapy or with glucagon-like peptide-1 (GLP1) analogs were excluded. Subgroup analysis was performed according to the estimated glomerular filtration rate (eGFR), the SGLT2i molecule, the main comorbidities, and concomitant treatment. As a secondary objective, the study analyzed the effect of SGLT2 inhibitors on uricemia levels. (3) Results: Seventy-three patients were analyzed, with a mean follow-up of 1.2 years. Uric acid and glucose FE significantly increased after the initiation of SGLT2is. This increase remained stable during the follow-up without differences among eGFR groups. No significant reduction in uricemia was observed. However, a trend towards a decrease was observed. (4) Conclusion: The use of SGLT2is in patients with CKD and T2DM is associated with an increase in uric acid FE, which maintains stability irrespective of glomerular filtration loss at least during 24 months of follow-up.

A261 INVESTIGATING THE RELATIONSHIP BETWEEN AIR POLLUTION AND BIOMARKERS OF CROHN’S DISEASE IN CANADA

Abstract Background The cause of Crohn's disease (CD) remains unclear. Studies suggest a potential connection between CD and air pollution, with a higher occurrence of CD found in areas with higher pollution levels. Aims Our study sought to explore this potential connection by investigating the relationship between components of air pollution and biomarkers indicative of CD risk. Methods In total, 2,256 healthy first-degree relatives of patients with CD were recruited as part of the CCC-GEM Project. We assessed baseline samples of gut permeability using the urinary fractional excretion ratio of lactulose-to-mannitol (LMR), subclinical inflammation via fecal calprotectin (FCP), and stool microbiome using 16S-rRNA sequencing. Air pollutant (n=8) concentrations were obtained from the National Air Pollution Surveillance Database. We used postal code to estimate air pollutant exposure via inverse distance weighting, averaging data for one (pollutant1-month), three (pollutant3-months), and twelve months (pollutant12-months) prior to recruitment. We used generalized estimating equation models to find associations between pollutant exposure and LMR, FCP, and microbiome composition, adjusting for age, sex, air pollutant seasonality, familial income, and familial relationships. Significance was set at pampersand:003C0.05, and false discovery rate (q) correction was applied to microbiome analysis. Results We observed a negative association between particulate matter (PMµg/m3), PM101-month, and PM103-months and LMR (β=-0.06[-0.09, -0.01], p=6.0×10-3 and β=-0.05[-0.07, -0.01], p=7.6×10-4), and a positive association between PM103-months and FCP (β=0.05[0.001, 0.10], p=0.04). 12-month pollutant exposure was associated with seven microbial taxa in a logit model. Notably, Carbon monoxide (CO)12-months was associated with Oscillospiraceae UCG_003 (β=3.49[1.53, 5.46], q=0.03). PM2512-months was associated with Marvinbryantia (β=0.12[0.06, 0.18], q=0.02). PM1012-months was associated with Moryella (β=0.07[0.03, 0.10], q=0.02). Nitrogen dioxide (NO)12-months was associated with Coprobacter and Ruminococcaceae UBA1819 (β=0.05[0.02, 0.07], q=0.03 and β=0.05[0.02, 0.08], q=0.04). NOX12-months was also associated with Ruminococcaceae_UBA1819 as well as Intestinimonas (β=0.03[0.01, 0.04], q=0.04 and β=0.03[0.01, 0.04], q=0.02). Conclusions Air pollutant exposure was associated with biomarkers of gut inflammation, barrier function, and microbiome composition. A shorter exposure (1/3 months) primarily affected FCP and LMR, while longer-term exposure (12 months) exerted a discernible impact on the microbiome. These findings suggest that exposure to air pollution may modulate the gut barrier function, inflammation, and the gut microbiome, biomarkers that are associated with the development of CD. Funding Agencies Crohn’s and Colitis Canada Genetics Environment Microbial (CCC-GEM) III, Leona M. and Harry B. Helmsley Charitable Trust, Dr. Croitoru is the recipient of the Canada Research Chair in Inflammatory Bowel Diseases

A18 EARLY LIFE EXPOSURE TO PARENTAL CROHN'S DISEASE IS ASSOCIATED WITH GUT MICROBIAL COMPOSITION AND FUNCTION, GUT PERMEABILITY AND INCREASED RISK OF FUTURE CROHN'S DISEASE

Abstract Background Previous studies show that offspring of persons with Crohn’s disease (CD) have a 7-fold higher risk of incident CD than the general population. Newborns of women with CD (vs healthy women) have altered stool microbiome composition and elevated fecal calprotectin (FCP). The perinatal period (pregnancy and first year postpartum) is critical for the development of the gut microbiome and immune system. Aims We investigated whether perinatal exposure to parental CD (compared to exposure later in life) has an impact on offspring’s gut barrier function, microbiome composition, and risk of future CD. Methods We assessed 1252 healthy offspring of persons with CD who were recruited in the CCC-GEM Project. We classified offspring into "perinatally exposed (pregnancy and first year postpartum)" vs "exposed later in life" to parental CD diagnosis. We measured baseline FCP and urinary fractional excretion ratio of lactulose to mannitol (marker of intestinal permeability, LMR). Fecal microbiome composition was determined by 16S rRNA gene sequencing. Microbiome function was imputed by PICRUSt2 (q value defined as false discovery rate-adjusted p&amp;lt;0.05). Multivariable regression and Cox proportional-hazards analyses were used to assess if peri-natal exposure to parental CD is associated with LMR, FCP, altered microbial composition, and future CD onset. Results Offspring exposed to a CD parent perinatally had a 4.7-fold higher risk (aHR 95%CI 1.3-17.5) of developing CD compared to those exposed to a parent who developed CD later in life (11/586 vs 3/666). LMR was significantly higher in the offspring exposed to parental CD perinatally (p=0.003), while no significant difference was observed for FCP at recruitment (p=0.94). Five bacterial taxa were significantly increased in the perinatally exposed group (q&amp;lt;0.05). The microbial functions such as glycine amidinotransferase, serine protein kinase and cyanophycin synthase were significantly increased in the perinatally exposed group (q&amp;lt;0.05). Conclusions Offspring exposed to parental CD perinatally had a higher risk of developing CD than those exposed to parental CD later in life. Early life exposure to parental CD was associated with higher baseline LMR, altered bacterial taxa and functional capacities. Environmental exposure during the perinatal period may determine the offspring’s risk of developing CD. Funding Agencies CAG, CCC, CIHRCanada Research Chair

A275 THE POTENTIAL MECHANISTIC ROLES OF SPHINGOLIPIDS IN HEALTHY FIRST-DEGREE RELATIVES OF PATIENTS WITH CROHN'S DISEASE

Abstract Background Crohn's Disease(CD) features a complex etiology intertwining genetic, environmental, and immunological dimensions. Sphingolipids (SPs) are pivotal lipid molecules within cell membranes and various biological processes, including cell growth, apoptosis, and inflammatory responses. While our prior research identified SPs as significant risk factors for CD onset, their pre-clinical influence on CD risk biomarkers and potential role in pathogenesis remains unclear Aims To identify the relationship between SPs and established CD risk factors Methods We used samples from healthy first-degree relatives(FDRs), followed in a nested case-control cohort from the Crohn's Colitis Canada- Genes, Environment, Microbial(GEM) project, matching those who developed CD(n=77) with control FDRs(n=303) based on age, sex, follow-up time, and geographic location. 59 SPs were selected from serum metabolites measured via an untargeted metabolomics platform. We used partial Spearman correlation to identify relationships between SPs and CD risk factors: i)intestinal permeability via the urinary fractional excretion ratio of lactulose to mannitol(LMR); ii)gut subclinical inflammation using fecal calprotectin(FCP); iii)systemic inflammation with C-reactive protein(CRP); iv)microbiome composition through fecal 16S rRNA sequencing and v)serum protein profile using the Olink® Proximity Extension Assay platform. A two-sided qampersand:003C0.05 was considered significant Results We identified 38 positive correlations between CRP and SPs across various sub-pathways, including Ceramides, Sphingomyelins and Hexosylceramides(0.117≤rho≤0.342, 1.37×10−09≤q≤0.042). One SP Ceramide(d18:1/16:0) correlated positively with FCP(rho=0.201, q=0.012). Moreover, two SPs, N-palmitoyl-sphingosine (d18:1/16:0) and glycosyl-N-(2-hydroxynervonoyl)-sphingosine (d18:1/24:1(2OH)), correlationed with Peptoclostridium genus (rho=0.465 and -0.245, q=1.47×10−16 and 0.028 respectively). All SPs correlated with one or more proteins, most positively between Sphingosine-1-phosphate and non-receptor tyrosine kinase(rho=0.637, q=1.98 ×10−36) and most negatively between sphingadienine and Chymotrypsin-C protein(rho=-0.334, q=4.11×10−8). No significant correlations emerged between SPs and LMR Conclusions We identified correlations between SPs and CD risk factors. The correlation with CRP suggests SPs might contribute to systemic inflammatory pathways related to CD. Moreover, correlations with the bacterial taxa highlight SPs' potential role in regulating microbial composition. Extensive correlations with proteins emphasize the pivotal impact of SPs on their function. This study may offer new insights into CD prevention Funding Agencies CCC, CIHR

P321 Development and validation of an Integrated Risk Score for future risk of Crohn’s disease in healthy first-degree relatives: The CCC-GEM Project, a multicentre prospective cohort study

Abstract Background Although the cause of Crohn’s disease (CD) is unknown, recent studies have identified a number of biomarkers associated with the risk of developing CD in healthy at-risk individuals. Establishing a combined prediction model that stratifies the future risk of CD in healthy at-risk individuals is the first step towards disease prevention and a better understanding of the preclinical phase of CD. Methods We recruited healthy first-degree relatives (FDRs) of persons with CD from 2008-2017 as part of the global multicenter prospective Crohn’s and Colitis Canada Genetic Environmental Microbial (CCC-GEM) Project. After collecting demographic information, blood, urine, and stool samples at recruitment, participants were followed for the development of CD. A GEM-integrative risk score (GEM-IRS), using random survival forest modeling that combined the baseline variables (demographics, measures of gut inflammation – fecal calprotectin, intestinal barrier function – urinary fractional excretion ratio of lactulose to mannitol, and fecal microbiome composition and predicted functional capacities – based upon 16S rDNA sequencing and PICRUSt2) to estimate time-to-CD onset, was derived and subsequently validated in two independent testing cohorts. Results Among 2,619 FDRs followed for a median of 6.8 years, 61 (2.3%) developed CD. The GEM-IRS, developed on the training cohort (n= 1,170), upon validation, showed a c-statistic of 0.789 in the pooled-testing cohorts (0.786 in the North American Testing cohort (n=1,141) and 0.804 in the Israeli Cohort (n=308)). The 4th quartile group compared to the rest had significantly increased risk of CD (hazard ratio [HR] 6.42, 95% confidence interval [CI], 3.10-13.30) in the pooled-testing cohorts (HR 5.89, 95% CI, 2.70-12.85 in North American Testing cohort; HR 10.7, 95% CI 1.24-92.65 in Israeli cohort). Furthermore, the GEM-IRS predicted CD in pre-specified subgroups of FDRs with minimal subclinical inflammation (fecal calprotectin&amp;lt;50 ug/g) or normal barrier function measures (lactulose/mannitol ratio&amp;lt;0.025), and up to 7 years before diagnosis in the pooled-testing cohort. Conclusion The GEM-IRS, which includes biomarkers of gut inflammation, intestinal barrier function, and the gut microbiome, is a valid risk stratification tool for predicting future development of CD in healthy first-degree relatives of persons with CD. The score may be used to guide preventative care for healthy first-degree relatives.

Understanding Predictors of Crohn's Disease: Determinants of Altered Barrier Function in Pre-Disease Phase of Crohn's Disease.

The pathogenesis of Crohn's disease (CD) remains unknown. The current working theory is that genetic susceptibility influences host-microbe interactions, resulting in chronic inflammation. Case-control studies fail to explain the triggers or pathogenesis of the disease, notably due to confounding factors in patients with established disease. Investigating the pre-disease phase of CD improves the capacity to assess these confounding factors and enables us to identify biomarkers associated with increased risk of CD. The Crohn's Colitis Canada-Genes, Environment, Microbial (CCC-GEM) project is a prospective cohort of healthy first-degree relatives of patients with CD, allowing us to interrogate the pre-disease phase of CD. The CCC-GEM Project has led to the identification of several demographic, serological, and microbiome composition markers associated with an increased risk of disease in pre-clinical participants. Notably, altered intestinal barrier function, as measured by the fractional urinary excretion of lactulose mannitol ratio, is associated with a significantly increased risk of CD. We review the intrinsic and external factors that are associated with altered intestinal barrier integrity, including genetic risk, subclinical inflammation, serum proteomics, intestinal microbiome composition, and environmental components, such as diet and lifestyle. Providing insights into the factors and mechanisms of altered barrier function contributes to our understanding of the pathogenic mechanisms of CD. These advances may aid in developing strategies for preventing disease in high-risk individuals. Further research and personalized strategies are needed to optimize these mitigation strategies for individuals at-risk for CD.

Acute phase reactions in Daboia siamensis venom and fraction-induced acute kidney injury: the role of oxidative stress and inflammatory pathways in in vivo rabbit and ex vivo rabbit kidney models.

This study examines the direct nephrotoxic effects of Daboia siamensis venom (RVV) and venom fractions in in vivo and isolated perfused kidneys (IPK) to understand the role of inflammation pathways and susceptibility to oxidative stress in venom or fraction-induced acute renal failure. We administered RVV and its venom fractions (PLA2, MP, LAAO, and PDE) to rabbits in vivo and in the IPK model. We measured oxidative stress biomarkers (SOD, CAT, GSH, and MDA) in kidney tissue, as well as inflammatory cytokines (TNF-α, IL-1β, IFN-γ, IL-4, IL-5, and IL-10), MDA and GSH levels in plasma and urine. We also calculated fractional excretion (FE) for pro-/anti-inflammatory cytokines and oxidative stress biomarkers, including the ratios of pro-/anti-inflammatory cytokines in urine after envenomation. In both kidney models, significant increases in MDA, SOD, CAT, and GSH levels were observed in kidney tissues, along with elevated concentrations of MDA and GSH in plasma and urine after injecting RVV and venom fractions. Moreover, RVV injections led to progressive increases in FEMDA and decreases in FEGSH. The concentrations of IL-4, IL-5, IL-10, IFN-γ, and TNF-α in plasma increased in vivo, as well as in the urine of the IPK model, but not for IL-1β in both plasma and urine after RVV administrations. Urinary fractional excretion of TNF-α, IL-1β, IFN-γ, IL-4, IL-5, and IL-10 tended to decrease in vivo but showed elevated levels in the IPK model. A single RVV injection in vivo disrupted the balance of urinary cytokines, significantly reducing either the TNF-α/IL-10 ratio or the IFN-γ/IL-10 ratio. RVV induces renal tubular toxicity by increasing oxidative stress production and elevating inflammatory cytokines in urine. During the acute phase of acute kidney injury, the balance of urine cytokines shifts toward anti-inflammatory dominance within the first two hours post-RVV and venom fractions.

Role of Glycosuria in SGLT2 Inhibitor-Induced Cardiorenal Protection: A Mechanistic Analysis of the CREDENCE Trial.

SGLT2 inhibitors have been shown to provide pronounced reductions in cardio-renal outcomes, including cardiovascular death, heart failure, and renal failure. The mechanisms underlying these benefits remain uncertain. We hypothesized that the effects could be attributed to the elevated glycosuria induced by these drugs. Urine concentrations of glucose, creatinine, and ketones were measured at baseline and after 1-year treatment with either placebo or canagliflozin 100mg/day in ∼2600 individuals from the CREDENCE trial (enrolling patients with type 2 diabetes, chronic kidney disease, and albuminuria). Associations between glycosuria and the primary composite endpoint from CREDENCE and secondary outcomes were assessed using Cox proportional hazards models. Canagliflozin treatment increased fractional urinary glucose excretion from 3±9% at baseline to 30±26% at year 1 (vs 5±19% with placebo, p<0.001). Subjects on canagliflozin and in the top quartile of urine glucose/creatinine ratio at year 1 were significantly protected for the primary endpoint (HR=0.42 [95% C.I.:0.30-0.61]); similar results were seen for hospitalized heart failure (HR=0.45 [0.27-0.73]) and all-cause death (HR=0.56 [0.39-0.80]). These associations persisted on adjustments for multiple conventional risk factors. In patients with T2D and CKD treated with canagliflozin, individuals with the highest amount of glycosuria showed the strongest protection against multiple cardio-renal outcomes.