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Ustekinumab is increasingly used in pregnant women with inflammatory bowel disease (IBD). Existing safety data are reassuring, but the stability of ustekinumab levels in pregnancy, degree of transfer to the infant and time to infant clearance are unknown.In this prospective observational study, ustekinumab-exposed women with IBD had trough levels measured in each trimester of pregnancy and at delivery. Infant ustekinumab levels were measured at delivery and ongoing until clearance was achieved. Trough ustekinumab level stability in individuals across pregnancy was compared by Skillings-Mack test. Spearman coefficients were used to correlate maternal and infant delivery levels, and median time to infant ustekinumab clearance was defined.19 pregnant women receiving ustekinumab were included. There was no difference in ustekinumab levels across pregnancy in those with two or more representative trough levels (P = 0.83, n = 11). Infant delivery ustekinumab levels were higher than maternal levels, with a median infant:maternal ratio of 1.79 (IQR 1.26-3.1). There was a positive correlation between maternal and infant delivery ustekinumab levels (r = 0.75, P = 0.001) and an inverse correlation between the number of days from final antenatal dose and delivery infant ustekinumab level (r = -0.65, P = 0.006). Median time of infant ustekinumab clearance was 9 (range 6-19) weeks (n = 9).Ustekinumab drug levels appear stable in pregnancy, with a delivery infant:maternal ratio similar to that of anti-TNFs. Infant ustekinumab clearance was complete by 20 weeks post-partum, however, infants exposed in utero should avoid live vaccination before 12 months of age until further clearance data are obtained.

Background The optimal use of infliximab depends on the measurement of trough levels with subsequent appropriate dose adjustment. With the introduction of biosimilars, it is important to demonstrate that the biosimilar behaves similarly in the assay utilized as the originator-infliximab, for which the assays were developed. In this study, the authors aimed to compare the concentrations of SB2-infliximab (Renflexis®) with that of originator-infliximab (Remicade®) when added to serum from healthy subjects and those with inflammatory bowel disease (IBD) when measured by commonly employed commercial assays. Methods Sera from two healthy controls, two patients with ulcerative colitis (one with quiescent disease, one with active disease), and two patients with Crohn's disease (one with quiescent disease, one with active disease) were spiked with SB2-infliximab or originator-infliximab at 0-20 μg/mL. Concentrations were measured using three commonly used assay kits (Lisa-Tracker®, Shikari Q-Inflix®, Promonitor IFX®) and one rapid test (Quantum Blue®). The results were compared using Bland-Altman techniques. Results Close agreement was observed between measured concentrations for all assays, irrespective of the origin of the serum. Limits of agreement varied between at worst -0.302 and 0.465 μg/mL, with the mean difference between the molecules being at worst 0.04 μg/mL (95% confidence intervals -0.011, 0.093). Conclusions The originator and SB-2 biosimilar-infliximab behaved similarly in several currently utilized assays in terms of their concentrations in biological fluids. Clinicians can be confident that therapeutic drug monitoring using platforms designed and developed for the originator-infliximab can be applied to SB-2-infliximab.

Therapeutic monitoring of infliximab is limited by the time lag between drug-level measurement and dose adjustment, along with the cost of dose escalation. Strategies for dose reduction in stable patients on maintenance infliximab at supratherapeutic levels are uncertain. This study determined the feasibility of a pharmacist-driven strategy for immediate dose adjustment using a sliding scale at the point of care in stable patients with inflammatory bowel disease on maintenance therapy.Adult patients with stable disease undergoing maintenance therapy with infliximab infusions, 5 mg/kg every 8 weeks, were prospectively studied. Trough drug levels were assessed by a rapid assay (and later by ELISA) at all infusions for up to 12 months with immediate but quantitatively small dose adjustment according to a sliding scale targeting a therapeutic range of 3-7 µg/mL. Disease activity was assessed both clinically and biochemically.The rapid assay and ELISA detected similar infliximab levels, and the strategy added approximately 30 min to the duration of infusion events. Only 20% of 48 patients (77% with Crohn's disease) had baseline trough infliximab concentrations within the therapeutic range. This value increased threefold after 24 and 48 weeks of interventions. One in two patients had baseline supratherapeutic levels and most were brought into the therapeutic range without discernible impact on disease activity by one dose adjustment, but two or three adjustments were generally needed for 29% of patients with sub-therapeutic levels. Overall, drug costs were reduced by 4%.Immediate dose adjustment following infliximab rapid assay performed by a pharmacist using a sliding scale is a feasible strategy. Supratherapeutic infliximab levels can be safely and quickly brought into the therapeutic range using small dose adjustments without impacting disease activity, offsetting (at least partly) costs associated with dose escalation.

Background and Aims In symptomatic patients with ileoanal pouches, pouchoscopy is needed for accurate diagnosis but is invasive. We aimed to assess the utility of non-invasive gastrointestinal ultrasound and faecal calprotectin in ileoanal pouch patients. Methods Patients with an ileoanal pouch were consecutively enrolled in this cross-sectional study from clinics in Victoria, Australia. The pouchitis disease activity index was used as a reference standard. Video-recorded pouchoscopies were reviewed by three gastroenterologists. Pouch, pre-pouch, and cuff biopsies were reviewed by a single pathologist. Ultrasound was performed by a single gastroenterologist transabdominally and transperineally. Faecal calprotectin was measured from morning stool samples. All examiners were blinded to patients’ clinical history. Results A total of 44 participants had a pouchoscopy, of whom 43 had a faecal calprotectin test and 42 had an ultrasound; 17 had pouchitis, 15 had pre-pouch ileitis, and 16 had cuffitis. Pouch wall thickness of Conclusions Faecal calprotectin and ultrasound are accurate and complementary tests to diagnose and localise inflammation of the ileoanal pouch. Prospective studies are needed to validate proposed sonographic indices and calprotectin levels.

Background: The impact of pregnancy on levels of biologic agents in patients with IBD is undefined and time to elimination in vedolizumab-exposed infants is unknown. Aims: To determine the effect of pregnancy on infliximab, adalimumab and vedolizumab levels and to study infant vedolizumab clearance. Methods: In a prospective observational study, maternal drug levels were measured pre-conception, in each trimester, at delivery and postpartum. The association between drug levels and gestation in weeks was assessed using generalised estimating equation modelling. Infant vedolizumab levels were performed at birth (cord blood), 6 weeks and 3 months or until undetectable. Results: We included 50 IBD patients (23 on infliximab, 15 on adalimumab and 12 on vedolizumab) with at least two intrapartum observations, plus 5 patients on vedolizumab with only mother and baby samples at delivery. Modelling showed no change in adalimumab levels, an increase in infliximab levels of 0.16 (95% CI 0.08-0.24) µg/L/week (P < 0.001) and a decrease of 0.18 (95% CI: −0.33 to −0.02) µg/L/week (P = 0.03) for vedolizumab. In 17 mother-baby pairs, median infant vedolizumab levels at birth were lower than maternal levels (P < 0.05) with an infant:maternal ratio of 0.7 (IQR 0.5-0.9). Vedolizumab was undetectable between 15 and 16 weeks of age in all 12 infants completing follow-up testing. Conclusions: During pregnancy, adalimumab levels remain stable, while infliximab levels increase and vedolizumab levels decrease. However, the increments were small suggesting that intrapartum therapeutic drug monitoring and dose adjustment are not indicated. Unlike infliximab and adalimumab, infant vedolizumab levels are lower in cord blood than in mothers and appear to clear rapidly.

Background Optimal timing for the last intrapartum dose of biologic agents is controversial. We established that cord blood levels of anti-TNF agents in exposed infants are greater than maternal levels,1 and another study with limited observations suggested maternal infliximab (IFX) levels increased in pregnancy but adalimumab (ADA) levels did not.2 The effect of pregnancy on vedolizumab (VDZ) levels, and placental transfer and time to clearance of VDZ in exposed infants remain undefined. Methods We performed a prospective observational study of maternal anti-TNF and VDZ levels pre-conception, in each trimester of pregnancy, at delivery and post-partum where possible. Women with IBD on IFX, ADA or VDZ who were pregnant or planning pregnancy were recruited. Serum trough IFX and ADA levels were measured by ELISA (Q-INFLIXI and Q-ADA (Matriks Biotek, Turkey) or Promonitor (Grifols, Spain)) and VDZ levels by ELISA (Theradiag, France). Infant VDZ levels were measured from the umbilical cord at delivery and at around 6–8 weeks and 3 months to assess time to clearance. Results Forty-nine patients (24 IFX, 14 ADA, 11 VDZ) with at least two observations on stable dosing were included. The median number of levels per patient was 3 (range 2–5). Infliximab cohort (n = 24): Median IFX levels were stable (Figure 1). IFX was administered 6–8 weekly at doses of 5 mg/kg (n = 21) or 10 mg/kg (n = 2) except for one patient on 4-weekly dosing. The last dose was given at a median 32 weeks; 3 remain pregnant. Adalimumab cohort (n = 14): Median ADA levels were stable (Figure 1). Twelve were on ADA fortnightly and 2 on weekly. Eleven continued ADA throughout pregnancy and 3 stopped at 30–33 weeks; 2 remain pregnant. Three patients were induced in pregnancy at least 12 weeks before the earliest ADA level. Vedolizumab cohort (n = 11): Trough VDZ levels were stable (Figure 2). Eight patients were on 8-weekly dosing and three were on 4-weekly. The last dose was given at a median 30 weeks; 4 remain pregnant (Patients H-K). Vedolizumab in infants (n = 8): The median VDZ cord blood level was 4.95 (1–19) μg/ml with a median infant:maternal ratio of 0.6 (0.4–0.9). VDZ was cleared at 8 weeks of age in three infants and by 15 weeks in two infants. Conclusion The stability of maternal levels of IFX, ADA and VDZ during pregnancy indicate that intrapartum dosing adjustment and routine therapeutic drug monitoring for patients in remission are not needed. Unlike anti-TNF, infant VDZ levels were lower in cord blood than in mothers and were cleared by 15 weeks. References

Summary Background & aims Vitamin D at serum 25(OH)D concentrations above 100 nmol/L is associated with disease remission in patients with IBD, suggesting targeted dosing might be anti-inflammatory. This study aimed to assess the effectiveness, safety and predictors of a 12-week regimen of vitamin D supplementation to achieve such a target in patients with active disease. Methods In a pilot study, patients with active colitis and a serum 25(OH)D concentration Results Five patients each with Crohn's colitis or ulcerative colitis (UC) had mean 25(OH)D concentration 52 (range 27–73 nmol/L). Five reached the targeted level and four 89–95 nmol/L. One withdrew after 4 weeks (88 nmol/L). Target dose was met only in those with BMI 2 and total dose inversely correlated with initial serum 25(OH)D. One patient had developed a high level at 8 weeks (146 nmol/L) and another new hypercalciuria. There were no serious adverse events attributable to the therapy. Clinical disease activity consistently declined, but faecal calprotectin and circulating markers of inflammation did not. Conclusions A specified oral vitamin D regimen successfully and safely achieved target or near-target levels, improved symptom-based activity scores, but did not alter objective measures of intestinal or systemic inflammation. A modified version of this dose-escalating regimen would be suitable for a randomised placebo-controlled trial, but does require regular safety monitoring.

Vitamin D may mediate immunomodulatory effects in patients with inflammatory bowel disease (IBD). The relationships between disease activity and circulating levels of total, free, and bioavailable 25(OH) vitamin D (25(OH)D) are poorly defined. The aim of this study was to measure circulating components of the vitamin D axis in patients with IBD and healthy controls and to correlate these with markers of disease activity, adjusting for potential confounders.Clinical data were obtained and serum was analyzed for 25(OH)D and vitamin D-binding protein in patients with IBD and controls. Markers of systemic and intestinal (fecal calprotectin) inflammation were measured.Serum 25(OH)D concentration was similar across 23 controls, 40 patients with Crohn's disease, and 31 with ulcerative colitis. An inverse correlation between 25(OH)D and calprotectin was noted in Crohn's disease (Pearson's r = -0.35, P = 0.040), ulcerative colitis (r = -0.39, P = 0.039), and all IBD together (r = -0.37, P = 0.003), but not with systemic markers. A similar trend was noted for free and bioavailable 25(OH)D. This inverse correlation remained after partial correlation analysis correcting for sunlight exposure, total oral vitamin D intake, and obesity and was also noted among the subgroup without small intestinal involvement.Despite total, free, and bioavailable 25(OH)D concentrations being similar to those in a healthy control population, they inversely correlated strongly with intestinal inflammation. This was independent of potential malabsorption, sunlight exposure, and total vitamin D intake and obesity. Vitamin D may play an immunomodulatory role in IBD.

BACKGROUND & AIMS: Little is known about in utero exposure to and postnatal clearance of anti-tumor necrosis factor (anti-TNF) agents in neonates. We investigated the concentrations of adalimumab and infliximab in umbilical cord blood of newborns and rates of clearance after birth, and how these correlated with drug concentrations in mothers at birth and risk of infection during the first year of life.METHODS: We performed a prospective study of 80 pregnant women with inflammatory bowel diseases at tertiary hospitals in Denmark, Australia, and New Zealand from March 2012 through November 2014: 36 received adalimumab and 44 received infliximab; 39 received concomitant thiopurines during pregnancy. Data were collected from medical records on disease activity and treatment before, during, and after pregnancy. Concentrations of anti-TNF agents were measured in blood samples from women at delivery and in umbilical cords, and in infants for every 3 months until the drug was no longer detected.RESULTS: The time from last exposure to anti-TNF agent during pregnancy correlated inversely with the concentration of the drugs in the umbilical cord (adalimumab: r = -0.64, P = .0003; infliximab: r = -0.77, P < .0001) and in mothers at time of birth (adalimumab, r = -0.80; infliximab, r = -0.80; P < .0001 for both). The median ratio of infant:mother drug concentration at birth was 1.21 for adalimumab (95% confidence interval [CI], 0.94-1.49) and 1.97 for infliximab (95% CI, 1.50-2.43). The mean time to drug clearance in infants was 4.0 months for adalimumab (95% CI, 2.9-5.0) and 7.3 months for infliximab (95% CI, 6.2-8.3; P < .0001). Drugs were not detected in infants after 12 months of age. Bacterial infections developed in 4 infants (5%) and viral infections developed in 16 (20%), all with benign courses. The relative risk for infection was 2.7 in infants whose mothers received the combination of an anti-TNF agent and thiopurine, compared with anti-TNF monotherapy (95% CI, 1.09-6.78; P = .02).CONCLUSIONS: In a prospective study of infants born to mothers who received anti-TNF agents during pregnancy, we detected the drugs until 12 months of age. There was an inverse correlation between the time from last exposure during pregnancy and drug concentration in the umbilical cord. Infliximab was cleared more slowly than adalimumab from the infants. The combination of an anti-TNF agent and thiopurine therapy during pregnancy increased the relative risk for infant infections almost 3-fold compared with anti-TNF monotherapy. Live vaccines therefore should be avoided for up to 1 year unless drug clearance is documented, and pregnant women should be educated on the risks of anti-TNF use.

Background: Wholegrain grains and cereals contain a wide range of potentially protective factors that are relevant to gastrointestinal health. The prebiotics best studied are fructans [fructooligosaccharides (FOS), inulin] and galactooligosaccharides (GOS). These and other short-chain carbohydrates can also be poorly absorbed in the small intestine (named fermentable oligo-, di- and monosaccharides and polyols; FODMAPs) and may have important implications for the health of the gut. Methods: In the present study, FODMAPs, including fructose in excess of glucose, FOS (nystose, kestose), GOS (raffinose, stachyose) and sugar polyols (sorbitol, mannitol), were quantified using high-performance liquid chromatography with an evaporative light scattering detector. Total fructan was quantified using an enzymic hydrolysis method. Results: Fifty-five commonly consumed grains, breakfast cereals, breads, pulses and biscuits were analysed. Total fructan were the most common short-chain carbohydrate present in cereal grain products and ranged (g per portion as eaten) from 1.12 g in couscous to 0 g in rice; 0.6 g in dark rye bread to 0.07 g in spelt bread; 0.96 g in wheat-free muesli to 0.11 g in oats; and 0.81 g in muesli fruit bar to 0.05 g in potato chips. Raffinose and stachyose were most common in pulses. Conclusions: Composition tables including FODMAPs and prebiotics (FOS and GOS) that are naturally present in food will greatly assist research aimed at understanding their physiological role in the gut.

Objective Colonic fermentation in patients with UC in remission was compared with that in matched healthy subjects on habitual diets and when dietary fibre was increased. Design Fibre intake, faecal output of fibre (measured as non-starch polysaccharide (NSP)), starch, microbiota and fermentation products, and whole gut transit time (WGTT) were assessed in association with habitual diet and when dietary intake of wheat bran (WB)-associated fibre and high amylose-associated resistant starch (RS) was increased in an 8-week, randomised, single-blind, cross-over study. Results Despite a tendency to lower habitual fibre intake in UC patients, faecal NSP and starch concentrations were threefold higher than in controls, whereas concentrations of phenols and short-chain fatty acids, pH and WGTT were similar. Increasing RS/WB intake was well tolerated. In controls (n=10), it more than doubled faecal NSP and starch excretion (p=0.002 for both), had no effect on NSP usage and reduced WGTT (p=0.024). In UC patients (n=19), high intake of RS/WB tended to normalise gut transit, but did not increase the proportion of NSP fermented. Increasing intake of RS/WB had little effect on faecal fermentation patterns or the structure of the microbiota. However, faeces from the UC cohort had lower proportions of Akkermansia muciniphila and increased diversity within Clostridium cluster XIVa compared to controls. Conclusions Gut fermentation of NSP and starch is diminished in patients with UC. This cannot be explained by abnormal gut transit and was not corrected by increasing RS/WB intake, and may be due to abnormal functioning of the gut microbiota. Trial registration number Australian New Zealand Clinical Trials Registry: ACTRN12614000271606.

Background: Dietary polyunsaturated fatty acids influence the natural history of intestinal inflammatory diseases. Varying the types of long-chain fatty acids that are exposed to cells alters the physicochemical properties of cell membranes. This study aimed to determine whether such variations alter transcellular and paracellular permeability in intestinal epithelium. Methods: Monolayers of Caco-2 cells, allowed to differentiate by culturing for 7 days following confluence, were used as the model for intestinal epithelium. Paracellular permeability was assessed by measurement of transepithelial resistance, while transcellular permeability was assessed by the transepithelial flux of horseradish peroxidase. Results: Exposure of the cells to 100 μmol/L of palmitic acid, oleic acid, eicosapentaenoic acid, or linoleic acid, was not toxic to cells (measured by leakage of lactate dehydrogenase), and altered cell membrane fatty acid composition (as measured by gas chromatography). Flux of horseradish peroxidase was significantly affected by 24 h fatty acid exposure (P = 0.038, ANOVA), being decreased by 23 ± 6% (mean ± SEM) by eicosapentaenoic acid and 25 ± 3% by linoleic acid. Oleic acid, palmitic acid and butyrate, had no effect. Transepithelial resistance also varied significantly across the treatment groups (P < 0.001) due to a 28 ± 5% increase induced by butyrate. The long-chain fatty acids had no effect. Conclusions: Both ω-3 and ω-6 polyunsaturated fatty acids reduce transcellular, non-receptor-mediated permeation of proteins across differentiated Caco2 cell monolayers, without altering paracellular permeability. Alteration of intestinal barrier function should be considered as a possible mechanism of action of dietary polyunsaturated fatty acids.

The relationships between changes induced by diet in colonic epithelial kinetics and in the activities of brush border hydrolases are poorly defined. The aims of this study are to define these relationships, as changes in kinetics would be expected to influence differentiation, and to determine whether the type of ingested dietary indigestible carbohydrates influences hydrolase activities.Groups of eight rats were fed a low fibre diet +/- supplements of different types of indigestible carbohydrates for 4 weeks. Alkaline phosphatase (ALP) and dipeptidyl peptidase IV (DPPIV) activities and epithelial kinetics were measured in distal colonic mucosa.Median ALP activities correlated positively and DPPIV activity negatively with the median proportion of cells entering metaphase (r = 0.58 and -0.58, respectively; P0.05) and number of metaphase arrests per crypt column across the diets (r = 0.59 and 0.58, respectively; P0.05). Stepwise regression analysis showed that both hydrolases independently predicted these kinetic indices (R263% for each). Mucosal ALP activities were markedly elevated during consumption of raw potato starch, guar gum and methylcellulose, while only potato starch caused a significant elevation of DPPIV activities.The type of indigestible carbohydrate in the diet influences colonic mucosal hydrolase activities. The opposite relationship between kinetics and each of the two hydrolases indicates that these hydrolases do not reflect the same event; dipeptidyl peptidase IV might relate to differentiation status while ALP could also be influenced by epithelial irritation due to changes in luminal conditions.

1. This study aimed to determine the effect of luminal butyrate on proliferative kinetics, a differentiation marker (alkaline phosphatase), and a molecule that controls cell-substratum adhesion (urokinase) in histologically normal human rectal mucosa. 2. Ten subjects with a colonoscopically normal colon (seven had previous adenomas) were given either butyrate or saline enemas for 4 days in a double-blind cross-over manner. Rectal biopsies were taken before and after each course of enemas. Epithelial proliferative kinetics were measured immunohistochemically using antibodies to proliferating cell nuclear antigen. Urokinase and alkaline phosphatase activities were measured spectrophotometrically in biopsy homogenates. 3. Both saline and butyrate enemas were well tolerated and induced no histological change except for a significant increase in crypt length (P < 0.05). The number of proliferating cells per crypt also increased significantly after butyrate (P = 0.018). 4. Compared with saline enemas, butyrate did not affect kinetic indices nor alkaline phosphatase activities. However, mucosal urokinase activities were significantly lower in butyrate-treated patients (9.5 ± 2.0 i.u./g) than in saline-treated patients (12.8 ± 2.0 i.u./g; P = 0.045). 5. Delivering of extra butyrate to the distal colon in healthy subjects may stabilize cell-substratum adhesion in surface epithelium and therefore offer a potential mechanism by which elevating distal colonic luminal butyrate concentrations might be beneficial in patients with colitis or hyperproliferative large bowel epithelium.

The hypothesis that the colonic epithelium is diffusely abnormal in ulcerative colitis was examined by comparing disease related responses in expression of markers of differentiation by colonic crypt cells to culture with and without butyrate. Cells were isolated from patients with normal colon (15), cancer (24), ulcerative colitis (19), or Crohn's disease (16). Alkaline phosphatase activities were measured in cell homogenates and the rate of glycoprotein synthesis assessed at the end of 24 hours of culture and expressed relative to the rate of protein synthesis as the G:P ratio. Alkaline phosphatase activities, but not G:P ratios, differed across the groups before and after 24 hour culture (p < 0.05), activities being lowest in the cancer group and highest in inflammatory bowel disease groups. Butyrate (1 mM) suppressed alkaline phosphatase activities in the cancer group by mean (SEM) of 17 (4) (p = 0.006) compared with no change in the other groups. Butyrate suppressed G:P ratios only in the cancer (6 (3)%, p = 0.03) and ulcerative colitis groups (5 (3)%, p = 0.04) and the changes in both were different (p < 0.05) from those in normal cells (increase of 10 (7)%). Changes in ulcerative colitis were different from those in Crohn's disease (p = 0.029). Responses were independent of the presence or absence of mucosal inflammation. These data confirm the diffuse nature of epithelial abnormalities in colorectal cancer. In ulcerative colitis, a different pattern of abnormality occurs, supporting the notion that the epithelium is also diffusely abnormal independent of mucosal inflammation.

In colitis, colonic epithelial cells have a shortened life span but show normal or increased expression of phenotypic markers of differentiation. This study examined the effect of differing culture conditions on the expression of such markers in colonic crypt cells. Crypt cells were enzymatically isolated from macroscopically normal large bowel mucosa resected because of neoplasia, inflammatory bowel disease or non-neoplastic non-inflammatory conditions. Cells cultured in the presence of serum exhibited a doubling of the rate of protein synthesis (measured by 14C-leucine uptake; p < 0.001) compared with autologous cells cultured in the absence of serum without evidence of loss of cell viability (assessed by 51Cr release from prelabelled cells) or of change in the rate of cell proliferation (assessed by total DNA content and 3H-thymidine uptake). Irrespective of the underlying colonic disease, crypt cells cultured in the absence of serum exhibited increased expression of phenotypic markers of differentiation compared with those cultured with serum: the rate of glycoprotein synthesis relative to that of protein synthesis increased by a mean of 59% and the cellular expression of brush border glycoproteins, alkaline phosphatase, and carcinoembryonic antigen significantly increased. The effects seen could not be mimicked by addition of dexamethasone or insulin to serum free medium. Thus, under less optimal (serum free) culture conditions, colonic crypt cells express phenotypic markers of differentiation at an accelerated rate suggesting that unfavourable microenvironmental conditions themselves are probably in part responsible for the normal or increased expression of such markers in colitis.

Methods: A systematic literature search was conducted to identify studies that investigated the pregnancy outcomes among women with IBD on anti-TNF-a therapy. The primary outcome was the overall rate of unfavourable pregnancyrelated outcomes among women with IBD on anti-TNF-a therapy. Secondary outcomes included rates of abortions (spontaneous or elective), preterm delivery, low birth weight, and congenital malformations. Odds ratios (OR) with 95%CI are reported. Studies included met the following criteria: observational or treatment design; had subjects with IBD on anti-TNF-a therapy for at least one trimester; and comparison with appropriately matched controls. Results: Overall, five studies with a total of 1216 participants were eligible for inclusion in the meta-analysis. There was no significant difference in the rates of total unfavourable pregnancy outcomes between pregnant women with IBD who were on anti-TNF-a therapy compared to controls not on antiTNF-a therapy (OR 1.12, 95%CI 0.79 1.59) (Figure 1). Similarly, there were no statistically significant differences in the rates of abortion (OR 1.48, 95%CI 0.93 2.35), preterm birth (OR 1.00, 95%CI 0.62 1.62), low birth weight (OR 1.05, 95%CI 0.62 1.78), or congenital malformation (OR 1.10, 95%CI 0.58 2.09) (Figure 2). Conclusions: The use of anti-TNF-a therapy does not appear to increase the risk of unfavourable pregnancy outcomes among women with IBD, although the optimal timing of therapy through pregnancy and the post-partum period was not assessed in this analysis. This data can help counsel patients around family planning and perinatal management.

Summary Background Functional gut symptoms are induced by inclusion and reduced by dietary restriction of poorly absorbed short-chain carbohydrates (FODMAPs), but the mechanisms of action remain untested. Aims To determine the effect of dietary FODMAPs on the content of water and fermentable substrates of ileal effluent. Methods Twelve ileostomates without evidence of small intestinal disease undertook two 4-day dietary periods, comprising diets differing only in FODMAP content in a randomized, cross-over, single-blinded intervention study. Daytime (14 h) ileal effluent was collected on day four of each diet. Patients rated effluent volume and consistency on a 10-cm visual analogue scale. The FODMAP content of the diet and effluent was measured. Results Ingested FODMAPs of 32% (range 6–73%) was recovered in the high FODMAP diet effluent. Effluent collection weight increased by a mean of 22% (95% CI, 5–39), water content by 20% (2–38%) and dry weight by 24% (4–43%) with the high compared to low FODMAP diet arm. Output increased by 95 (28–161) mL. Volunteers perceived effluent consistency was thicker (95% CI, 0.6–1.9) with the low FODMAP diet than with the high FODMAP diet (3.5–6.1; P = 0.006). Conclusions These data support the hypothetical mechanism; FODMAPs increase delivery of water and fermentable substrates to the proximal colon.

Fermentable oligo-, di-, and monosaccharides and polyols (FODMAPs) are short-chain carbohydrates that can be poorly absorbed by the small intestine and may have a wide range of effects on gastrointestinal processes. FODMAPs include lactose, fructose in excess of glucose, fructans and fructooligosaccharides (FOS, nystose, kestose), galactooligosaccharides (GOS, raffinose, stachyose), and sugar polyols (sorbitol, mannitol). This paper describes an analytical approach based on HPLC with ELSD that quantifies the major FODMAPs in 45 vegetables and 41 fruits. Sorbitol and/or mannitol were measured in 18 vegetables (range = 0.09-2.96 g/100 g of fw), raffinose and/or stachyose in 7 vegetables (0.08-0.68 g/100 g of fw), and nystose and/or kestose in 19 vegetables (0.02-0.71 g/100 g of fw). Apple, pear, mango, clingstone peach, and watermelon all contained fructose in excess of glucose. Sorbitol was measured in 15 fruits (0.53-5.99 g/100 g of fw), mannitol was found in 2 fruits, and nystose or kestose was measured in 8 fruits. Understanding the importance of dietary FODMAPs will be greatly assisted by comprehensive food composition data.

Fructans are not digested in the small intestines of humans. While many health benefits have been attributed to these carbohydrates, they can cause gastrointestinal symptoms in some individuals. We measured the total fructans in 60 vegetables and 43 fruits using the Megazyme fructan assay. Vegetables with the highest quantity of fructans included garlic, artichoke, shallots, leek bulb, and onions (range, 1.2-17.4 g/100 g fw). Fruits with low, but detectable, fructans included longon, white peach, persimmon, and melon (range, 0.21-0.46 g/100 g fw). The fructan assay was modified to provide an estimate of the average chain length (degree of polymerization) for high fructan vegetables. d-Fructose can also be malabsorbed in the small intestine of humans, so the d-fructose content in some foods was measured to supplement the current food tables. Research in this area will be facilitated through the availability of more comprehensive food composition data.

Further, medical treatment, disease activity, and counselling were investigated in an international multicentre study. Methods: 115 women exposed to Infliximab (IFX) or Adalimumab (ADA) during pregnancy who had given birth during a 3 year period were identified at 14 hospitals in Denmark, Australia and New Zealand. Electronic questionnaires were used to investigate diagnosis, medical treatment, disease activity, counselling, and birth outcome. Recruitment is ongoing. Results: 86 (74.8%) of the women filled in the questionnaire of whom 3 had a miscarriage prior to gestational week (GW) 10, leaving a study population of 83 women. The pregnancy was planned in 60 (72.3%) of the cases. 54 (65.1%) and 29 (34.9%) received treatment with IFX and ADA, respectively. Co-medication with thiopurines were used in 25 (30.1%) of the pregnancies. 7 (8.4%) gave birth preterm (GW 32 36). None of the babies were small for gestational age or had congenital malformations. Median Apgar score at 5 minutes was 9.3 (range 7 10). Caesarean section was performed in 52 (60.5%) of the deliveries and in 36 (69.2%) of these cases the women stated IBD as the reason. 19 (22.3%) women experienced a relapse during pregnancy of whom 8 (42.1%) had disease activity at delivery. Anti-TNF-alpha therapy was discontinued during the 1st., 2nd. and 3rd. trimester in 3.9%, 38.2% and 57.9% of the cases, respectively. Counselling on medical treatment with anti-TNF-alpha therapy was given to 72 (83.7%) of the women most frequently by a gastroenterologist (88.9%). The vast majority (98.4%) of women were satisfied with the information provided. 75 (87.2%) of the women would accept treatment with anti-TNF-alpha during a future pregnancy. Conclusions: Maternal anti-TNF-alpha treatment for IBD did not seem to be a risk factor for adverse pregnancy outcomes. Anti-TNF-alpha was discontinued late in pregnancy. The vast majority of women had received counselling by a gastroenterologist on anti-TNF-alpha treatment during pregnancy and were satisfied with the information provided. A third of the pregnancies were unplanned suggesting risk benefit counselling should be offered to all fertile women at initiation of anti-TNF-alpha therapy.

Butyrate may have paradoxical effects on epithelial cells of similar origin. This study aimed to examine the hypothesis that one mechanism that dictates a cell's response to butyrate is its state of activation. First, the responses to 24 h exposure to butyrate (1-2 mM) of normal and neoplastic human colonic epithelial cells activated by their isolation and primary culture, and of colon cancer cell lines, LIM1215 and Caco-2, were examined. In primary cultures of normal and cancer cells, butyrate had no effect on alkaline phosphatase activities but significantly suppressed urokinase receptor expression by a mean +/- SEM of 30 +/- 12% and 36 +/- 9%, respectively. Interleukin-8 secretion was suppressed by 44 +/- 7% in normal cells (P0.05) but was unchanged in cancer cells. In contrast, the cell lines significantly increased alkaline phosphatase activities by50%, urokinase receptor expression2-fold and interleukin-8 secretion3-fold in response to butyrate. Secondly, the effect of butyrate on Caco-2 cells was examined with or without prior exposure to a specific activating stimulus [tumour necrosis factor alpha (TNF alpha)]. Interleukin-8 secretion increased by 145 +/- 23% and 132 +/- 17% on 24 h exposure to 2 mM butyrate or 0.1 microM TNF alpha alone, respectively. However, in cells pre-treated with TNF alpha, butyrate significantly inhibited secretion by 34 +/- 7% below unstimulated levels. The response to butyrate of urokinase receptor, whose expression was not stimulated by TNF alpha, was unchanged. These effects were mimicked by trichostatin A, an inhibitor of histone deacetylase, suggesting that butyrate's paradoxical effects may have been operating by the same mechanism. In conclusion, some of the paradoxical effects of butyrate do not appear to represent inherent differences between normal and transformed cells. Rather, the response may be determined by the state of activation of the cells.

Background—The functions of urokinase in intestinal epithelia are unknown.Aims—To determine the relation of urokinase expressed by intestinal epithelial cells to their position in the crypt-villus/surface axis and of mucosal urokinase activity to epithelial proliferative kinetics in the distal colon.Methods—Urokinase expression was examined immunohistochemically in human intestinal mucosa. Urokinase activity was measured colorimetrically in epithelial cells isolated sequentially from the crypt-villus axis of the rat small intestine. In separate experiments, urokinase activity and epithelial kinetics (measured stathmokinetically) were measured in homogenates of distal colonic mucosa of 14 groups of eight rats fed diets known to alter epithelial turnover.Results—From the crypt base, an ascending gradient of expression and activity of urokinase was associated with the epithelial cells. Median mucosal urokinase activities in each of the dietary groups of rats correlated positively with autologous median number of metaphase arrests per crypt (r=0.68; pr=0.75; pConclusions—Localisation of an enzyme capable of leading to digestion of cell substratum in the region where cells are loosely attached to their basement membrane, and the association of its activity with indexes of cell turnover, suggest a role for urokinase in facilitating epithelial cell loss in the intestine.

Background & Aims Patients with non-celiac gluten sensitivity (NCGS) do not have celiac disease but their symptoms improve when they are placed on gluten-free diets. We investigated the specific effects of gluten after dietary reduction of fermentable, poorly absorbed, short-chain carbohydrates (fermentable, oligo-, di-, monosaccharides, and polyols [FODMAPs]) in subjects believed to have NCGS. Methods We performed a double-blind cross-over trial of 37 subjects (aged 24−61 y, 6 men) with NCGS and irritable bowel syndrome (based on Rome III criteria), but not celiac disease. Participants were randomly assigned to groups given a 2-week diet of reduced FODMAPs, and were then placed on high-gluten (16 g gluten/d), low-gluten (2 g gluten/d and 14 g whey protein/d), or control (16 g whey protein/d) diets for 1 week, followed by a washout period of at least 2 weeks. We assessed serum and fecal markers of intestinal inflammation/injury and immune activation, and indices of fatigue. Twenty-two participants then crossed over to groups given gluten (16 g/d), whey (16 g/d), or control (no additional protein) diets for 3 days. Symptoms were evaluated by visual analogue scales. Results In all participants, gastrointestinal symptoms consistently and significantly improved during reduced FODMAP intake, but significantly worsened to a similar degree when their diets included gluten or whey protein. Gluten-specific effects were observed in only 8% of participants. There were no diet-specific changes in any biomarker. During the 3-day rechallenge, participants' symptoms increased by similar levels among groups. Gluten-specific gastrointestinal effects were not reproduced. An order effect was observed. Conclusions In a placebo-controlled, cross-over rechallenge study, we found no evidence of specific or dose-dependent effects of gluten in patients with NCGS placed diets low in FODMAPs. www.anzctr.org.au. ACTRN12610000524099

outcome was reported in patients refractory to multiple drugs or with extensive loss of the mucosal tissue (Sands, et al. Gastroenterology 2008). We thought that young UC patients should respond to GMA. Methods: We assessed the efficacy of GMA in growing patients as mono-therapy or in combination with low dose PSL if monotherapy was ineffective. Seventeen children and adolescents, age 11 19 years, each was to receive 5 GMA sessions, 2 sessions in the first week, and then weekly sessions. Patients who achieved a decrease of at least 5 point in the clinical activity index (CAI) were to receive additional weekly GMA, up to 6 sessions, while poor responders were to receive 20mg/day PSL plus additional GMA session. 5ASA was the only concomitant medication, which had started longer than 8 weeks prior to GMA. At entry and week 12, patients were clinically and endoscopically evaluated, allowing each patient to serve as her or his own control. Results: Five patients did not respond well to the initial 5 GMA sessions and were given PSL plus GMA, while 12 patients responded to the first 5 GMA sessions and received further sessions. At entry, the average CAI was 14.1±0.4, range 11 17, and the average endoscopic index was 9.2±0.3, range 7 11. The corresponding values at week 12 were 2.1±0.2, range 1 4 (P< 0.001) and 2.4±0.2, range 1 4 (P< 0.001), respectively. Therefore, 12 patients achieved remission with GMA as monotherapy and 5 patients achieved remission with GMA plus PSL. PSL was tapered to 0mg within 3 months. Conclusions: In this study, GMA in children and adolescents with UC, who were corticosteroid naive was associated with clinical remission and mucosal healing. Therefore, there are patients who may respond well to GMA and be spared from pharmacologicals. Additionally, GMA is favoured by patients for its good safety profile and being a non-drug intervention.

Epithelia from several sites exhibit inducible secretion of interleukin 8 (IL-8). This study aimed to characterise secretion of IL-8 by colonic epithelial cells in vitro. Colonic crypt cells were isolated enzymatically from resected colon and the IL-8 content of culture supernates was measured by ELISA. The rate of secretion of IL-8 accelerated and levels of IL-8 transcripts increased appreciably during culture. Exposure to tumour necrosis factor alpha (TNF alpha) failed to increase secretion further. Secretion was not induced by the enzymatic digestion or by serum used in the culture medium but was significantly inhibited by butyrate, by a mean of 23%. Control experiments indicated that colonic crypt cells were the likely source. The secretion of IL-8 over 24 hours by cells from uninflamed mucosa of patients with ulcerative colitis or Crohn's disease was more than twofold that from normal cells, while that from cancer bearing colons was normal. TNF alpha (10 mM) significantly suppressed IL-8 secretion only in the ulcerative colitis group and the change was different to those in the normal (p = 0.007) and Crohn's disease groups (p = 0.012). Cells from inflamed areas secreted more IL-8 than those from autologous uninflamed areas (p = 0.009) but responses to modulating factors were no different. The induction of IL-8 secretion by colonic crypt cells in vitro is probably a response to injury associated with isolation and culture. It is suppressed by butyrate and increased in inflammatory bowel disease independently of the presence of mucosal inflammation. Whether epithelial derived IL-8 plays a part in the pathogenesis of inflammatory bowel disease is not yet clear.

The amount and type of dietary fibre ingested influences colonic luminal characteristics, especially the concentration of carbohydrate fermentation products such as butyrate. This study aimed to assess whether diets supplemented with fibres of differing fermentability (delivering different amounts of butyrate to the colon) influence mucosal activities of urokinase and brush border hydrolases, and epithelial turnover. Groups of five rats were fed one of four diets containing low (2%), highly fermented (guar 10% or oat bran 10%) or slowly fermented fibre (wheat bran 10%) for 4 weeks. Activities of urokinase, alkaline phosphatase, dipeptidyl peptidase IV and maltase were measured in mucosal homogenates of proximal and distal colon and from rectum. Proliferative kinetics were assessed in distal and proximal colon by the metaphase arrest technique. Hydrolase activities were similar across all four dietary groups but a significant difference was found for urokinase (P = 0.014). This was due to a reduction in urokinase activities of > 30% at the three sites in the wheat bran group compared with the other groups. Of proliferative indices, only crypt column height differed across the groups (P = 0.038) and was highest in rats fed wheat bran and lowest in those fed the low fibre diet (P = 0.047). The proportion of mitoses in the top one-fifth of the crypt also differed across groups (P = 0.038) due to the high values in the distal colon of the low fibre group. Thus, addition of a slowly fermented (but not highly fermented) fibre to the diet of rats reduces net urokinase activity in large bowel mucosa and increases the life span of colonic epithelial cells without changing activities of brush border hydrolases.

Background/Aims: Because the neutral protease urokinase is important in control of cell adhesion and migration, the effects of the physiologically relevant fermentation product butyrate on urokinase secretion by colonic epithelium were examined. Methods: Secreted and cell-associated levels of urokinase and plasminogen activator inhibitor 1 were measured in colonic crypt cells within 24 hours of isolation from macroscopically normal mucosa of normal or cancer-bearing colons. Results: Butyrate caused a concentration-dependent inhibition of both secreted (56% ± 4% inhibition after 24-hour exposure to 1 mmol/L butyrate; n=20; mean ± SEM; P P = 0.003). Acetate and propionate had minimal effects. Butyrate also stimulated plasminogen activator inhibitor 1 secretion by 25% ± 7% ( P = 0.013). Net urokinase activities were suppressed in supernates and cell homogenates by butyrate. Levels of transcripts for urokinase and the inhibitor changed with butyrate exposure in parallel to the levels of secretion of the respective proteins. Cells from the cancer group showed significantly reduced inhibitor secretion and abnormal responses to butyrate (greater inhibition of urokinase secretion and no stimulation of inhibitor secretion), probably reflecting the diffuse disturbance of colonic epithelial biology associated with colorectal cancer. Conclusions: Butyrate has dual effects in markedly reducing colonic epithelial urokinase activity, and these may have important implications to understanding colonic epithelial physiology and the pathogenesis and treatment of colonic diseases.

Urokinase is a neutral protease whose major site of action is the external surface of the plasma membrane of cells and whose major function seems to be modulation of cell adhesion, such as that which occurs during cell migration. This study aimed to determine whether colonic epithelium is involved with the urokinase system. The contents of urokinase and one of its specific inhibitors, plasminogen activator inhibitor-1, were measured in culture supernatant and cell homogenates of isolated human colonic crypt cells. The amounts of both factors increased in supernatants over 24 hours, and approximately twice the amount was found in supernatants than in autologous cell homogenates. The secretion of both factors was similar in serum free and serum containing media. Northern blot analysis showed that messenger ribonucleic acid specific for urokinase and plasminogen activator inhibitor-1 was present in colonic crypt cells and that expression over 18 hours of culture was increased 12 fold for urokinase type plasminogen activator and two to fourfold for the inhibitor compared with values found in autologous freshly isolated cells. Urokinase activity was detected in crypt cell homogenates and supernatants indicating that it was present in excess of its inhibitors. Control experiments indicated that the epithelial cells themselves were responsible for the observations and excluded artefactual effects of the isolation procedure. In conclusion, isolated human colonic epithelial cells secrete urokinase and at least one of its specific inhibitors. Further investigation of the role of urokinase in the physiology and pathophysiology of colonic epithelium is indicated.

Expression of brush border hydrolases can reflect the state of differentiation of an epithelium. To determine if expression of these enzymes is disordered in patients with neoplastic or hyperplastic lesions, the activities of alkaline phosphatase, maltase, and dipeptidyl peptidase IV were measured spectrophotometrically in colonoscopic biopsies from the proximal and distal colon and rectum in 50 controls, 17 patients with large bowel adenomas, 29 with carcinoma, and 9 with hyperplastic polyps. In normal controls, a descending cecorectal gradient of alkaline phosphatase activities and an ascending gradient of maltase activities were seen (P < 0.001). Though regional patterns of expression were generally preserved in disease groups, there were significant differences of activities across patient groups for alkaline phosphatase (greater in cancer, adenoma, and hyperplastic groups than in normals; P < 0.05) and for dipeptidyl peptidase IV (greater in hyperplastic polyp group than normals, greater in adenoma than cancer group; P < 0.05). Compared with normal controls, abnormalities of site-specific activities were confined to the rectum in patients with adenoma (maltase decreased, P = 0.02; dipeptidyl peptidase IV increased, P < 0.01) or with carcinoma (alkaline phosphatase increased, P = 0.03) but dipeptidyl peptidase IV activities were increased in all regions in bowels bearing hyperplastic polyps (P < 0.01). These data suggest that neoplastic and hyperplastic lesions, while focal in nature, occur in large bowel epithelium, which is diffusely abnormal in terms of its expression of these enzymes.