Oral monosaccharides and disaccharides are used to measure in vivo human gut permeability through urinary excretion.
The aims were as follows: (1) to obtain normative data on small intestinal and ...colonic permeability; (2) to assess variance on standard 16 g fiber diet performed twice; (3) to determine whether dietary fiber influences gut permeability measurements; and (4) to present pilot data using 2 selected probes in patients with diarrhea-predominant irritable bowel syndrome (IBS-D).
Sixty healthy female and male adults, age 18–70 years, participated in 3 randomized studies (2 studies on 16.25 g and 1 study on 32.5 g fiber) in otherwise standardized diets. At each test, the following sugars were ingested: 12C-mannitol, 13C-mannitol, rhamnose (monosaccharides), sucralose, and lactulose (disaccharides). Standardized meals were administered from 24 hours before and during 24 hours post-sugars with 3 urine collections: 0–2, 2–8, and 8–24 hours. Sugars were measured using high-performance liquid chromatography–tandem mass spectrometry. Eighteen patients with IBS-D underwent 24-hour excretion studies after oral 13C-mannitol and lactulose.
Baseline sugars (>3-fold above lower limits of quantitation) were identified in the 3 studies: 12C-mannitol in all participants; sucralose in 4–8, and rhamnose in 1–3. Median excretions/24 h (percentage of administered dose) for 13C-mannitol, rhamnose, lactulose, and sucralose were ∼30%, ∼15%, 0.32%, and 2.3%, respectively. 13C-mannitol and rhamnose reflected mainly small intestinal permeability. Intraindividual saccharide excretions were consistent, with minor differences with 16.25 g vs 32.5 g fiber diets. Median interindividual coefficient of variation was 76.5% (10–90 percentile: 34.6–111.0). There were no significant effects of sex, age, or body mass index on permeability measurements in health. 13C-mannitol measurements are feasible in IBS-D.
Baseline 12C-mannitol excretion precludes its use; 13C-mannitol is the preferred probe for small intestinal permeability.
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Variations in genes that regulate bile acid (BA) synthesis are associated with colonic transit in patients with irritable bowel syndrome (IBS). We investigated features of BA synthesis and excretion ...and genetic features of patients with different types of IBS.
In 26 healthy volunteers, 26 patients with IBS and constipation (IBS-C), and 26 with IBS and diarrhea (IBS-D), we measured serum levels of 7α-hydroxy-4-cholesten-3-one (C4; a surrogate for BA synthesis) and fibroblast growth factor (FGF) 19 (an ileal hormone that downregulates BA synthesis). For stool samples, we measured concentration of BA, weight, and amount of fat when participants were given high-fat diets. Spearman correlations were used to explore relationships among factors. We analyzed 1 polymorphism in Klotho-β (KLB) and 3 in fibroblast growth factor receptor-4 (FGFR4) for all members of each group using analysis of covariance.
The concentration of BA in stool was associated with group (for a comparison of 3 groups; P = .057); it was higher in patients with IBS-D than IBS-C (P = .017). The serum level of C4 was higher in patients with IBS-D than IBS-C (P = .02) or healthy volunteers (P = .01); 38% of patients with IBS-D had increased serum levels of C4, compared with healthy volunteers. Serum level of C4 correlated with stool concentration of BA (rs = 0.606; P < .001), serum FGF19 (rs = -0.324; P = .007), and stool weight (rs = 0.366; P = .003). Stool concentration of BA correlated with weight (rs = 0.737; P < .001) and level of fat (rs = 0.528; P < .001). Body mass index correlated with serum level of C4 (rs = 0.423, P < .001) and stool concentration of BA (rs = 0.507, P < .001), and was higher in patients with IBS-D compared with other groups (overall P = .036). FGFR4 rs1966265 was associated with stool level of BA (P = .032).
Patients with IBS-D have greater body mass index and synthesize and excrete higher levels of BA than individuals with IBS-C or healthy volunteers. Serum levels of C4 might be used to identify patients with IBS-D who have BA malabsorption; studies are needed to determine if some patients have a genetic predisposition to this disorder.
The intestinal lumen contains several proteases. Our aim was to determine the role of faecal proteases in mediating barrier dysfunction and symptoms in IBS.
39 patients with IBS and 25 healthy ...volunteers completed questionnaires, assessments of in vivo permeability, ex vivo colonic barrier function in Ussing chambers, tight junction (TJ) proteins, ultrastructural morphology and 16 s sequencing of faecal microbiota rRNA. A casein-based assay was used to measure proteolytic activity (PA) in faecal supernatants (FSNs). Colonic barrier function was determined in mice (ex-germ free) humanised with microbial communities associated with different human PA states.
Patients with IBS had higher faecal PA than healthy volunteers. 8/20 postinfection IBS (PI-IBS) and 3/19 constipation- predominant IBS had high PA (>95th percentile). High-PA patients had more and looser bowel movements, greater symptom severity and higher in vivo and ex vivo colonic permeability. High-PA FSNs increased paracellular permeability, decreased occludin and increased phosphorylated myosin light chain (pMLC) expression. Serine but not cysteine protease inhibitor significantly blocked high-PA FSN effects on barrier. The effects on barrier were diminished by pharmacological or siRNA inhibition of protease activated receptor-2 (PAR-2). Patients with high-PA IBS had lower occludin expression, wider TJs on biopsies and reduced microbial diversity than patients with low PA. Mice humanised with high-PA IBS microbiota had greater in vivo permeability than those with low-PA microbiota.
A subset of patients with IBS, especially in PI-IBS, has substantially high faecal PA, greater symptoms, impaired barrier and reduced microbial diversity. Commensal microbiota affects luminal PA that can influence host barrier function.
•LC-MS/MS methods for vitamin D3-sulfate and 25OHD3-sulfate in breastmilk.•Vitamin D3-sulfate is more abundant in breastmilk than 25OHD3-sulfate.•Vitamin D sulfates are soluble in the aqueous portion ...of breastmilk.
Sulfated metabolites of vitamin D have been suggested to be in breastmilk, although current methods to measure sulfated vitamin D compounds in breastmilk by liquid chromatography-tandem mass spectrometry (LC-MS/MS) have not adequately accounted for increased aqueous solubility of these sulfated metabolites. The purpose of this study was to generate a method of LC-MS/MS for measuring vitamin D3-3-sulfate (VitD3-S) and 25-hydroxyvitamin D3-3-sulfate (25OHD3-S) specifically in human breastmilk. The resulting method uses methanol to precipitate protein and solid phase extraction to prepare the samples for LC-MS/MS. The limits of quantification for analytes in solvent were 0.23 ng/mL VitD3-S and 0.2 ng/mL 25OHD3-S. Various experiments observed concentrations ranging 0.53 to 1.7 ng/mL VitD3-S and ≤ 0.29 ng/mL 25OHD3-S. Both analytes were present in aqueous skim milk, demonstrating the enhanced aqueous solubility of these vitamin D sulfates. In conclusion, we describe an effective method for measuring VitD3-S and 25OHD3-S in breastmilk by LC-MS/MS.
ABSTRACT
Intestinal barrier function and microbiota are integrally related and play critical roles in maintenance of host physiology. Sex is a key biologic variable for several disorders. Our aim was ...to determine sex‐based differences in response to perturbation and subsequent recovery of intestinal barrier function and microbiota in healthy humans. Twenty‐three volunteers underwent duodenal biopsies, mucosal impedance, and in vivo permeability measurement. Permeability testing was repeated after administration of indomethacin, then 4 to 6 wk after its discontinuation. Duodenal and fecal microbiota composition was determined using 16S rRNA amplicon sequencing. Healthy women had lower intestinal permeability and higher duodenal and fecal microbial diversity than healthy men. Intestinal permeability increases after indomethacin administration in both sexes. However, only women demonstrated decreased fecal microbial diversity, including an increase in Prevotella abundance, after indomethacin administration. Duodenal microbiota composition did not show sex‐specific changes. The increase in permeability and microbiota changes normalized after discontinuation of indomethacin. In summary, women have lower intestinal permeability and higher microbial diversity. Intestinal permeability is sensitive to perturbation but recovers to baseline. Gut microbiota in women is sensitive to perturbation but appears to be more stable in men. Sex‐based differences in intestinal barrier function and microbiome should be considered in future studies.—Edogawa, S., Peters, S. A., Jenkins, G. D., Gurunathan, S. V., Sundt, W. J., Johnson, S., Lennon, R. J., Dyer, R. B., Camilleri, M., Kashyap, P. C., Farrugia, G., Chen, J., Singh, R. J., Grover, M. Sex differences in NSAID‐induced perturbation of human intestinal barrier function and microbiota. FASEB J. 32, 6615–6625 (2018). www.fasebj.org
Introduction
We aimed to provide cut points for the automated Elecsys Alzheimer's disease (AD) cerebrospinal fluid (CSF) biomarkers.
Methods
Cut points for Elecsys amyloid beta 42 (Aβ42), total tau ...(t‐tau), hyperphosphorylated tau (p‐tau), and t‐tau/Aβ42 and p‐tau/Aβ42 ratios were evaluated in Mayo Clinic Study of Aging (n = 804) and Mayo Clinic Alzheimer's Disease Research Center (n = 70) participants.
Results
The t‐tau/Aβ42 and p‐tau/Aβ42 ratios had a higher percent agreement with normal/abnormal amyloid positron emission tomography (PET) than the individual CSF markers. Reciever Operating Characteristic (ROC)‐based cut points were 0.26 (0.24–0.27) for t‐tau/Aβ42 and 0.023 (0.020–0.025) for p‐tau/Aβ42. Ratio cut points derived from other cohorts performed as well in our cohort as our own did. Individual biomarkers had worse diagnostic properties and more variable results in terms of positive and negative percent agreement (PPA and NPA).
Conclusion
CSF t‐tau/Aβ42 and p‐tau/Aβ42 ratios are very robust indicators of AD. For individual biomarkers, the intended use should determine which cut point is chosen.
Concentrations of vitamin D (VitD) and 25-hydroxyvitamin D (25OHD) in breastmilk are low despite the essential role of VitD for normal infant bone development, yet additional metabolic forms of ...vitamin D may be present. This study evaluates the contribution of sulfated vitamin D metabolites, vitamin D3-sulfate (VitD3-S) and 25-hydroxyvitamin D3-sulfate (25OHD3-S) for lactating women and assesses the response to high-dose VitD3 supplementation.
Serum and breastmilk were measured before and after 28 days with 5000 IU/day VitD3 intake in 20 lactating women. Concentrations of VitD3-S and 25OHD3-S in milk, and 25OHD2, 25OHD3, 25OHD3-S, VitD3 and VitD3-S in serum were determined by mass spectrometry.
Baseline vitamin D status was categorized as sufficient (mean ± SD serum 25OHD3 69 ± 19 nmol/L), and both serum VitD3 and 25OHD3 increased following supplementation (p < 0.001). 25OHD3-S was 91 ± 19 nmol/L in serum and 0.47 ± 0.09 nmol/L in breastmilk. VitD3-S concentrations were 2.92 ± 0.70 nmol/L in serum and 6.4 ± 3.9 nmol/L in breastmilk. Neither sulfated metabolite significantly changed with supplementation in either serum or breastmilk.
Sulfated vitamin D metabolites have prominent roles for women during lactation with 25OHD3-S highly abundant in serum and VitD3-S distinctly abundant in breastmilk. These data support the notion that 25OHD3-S and VitD3-S may have physiological relevance during lactation and nutritional usage for nursing infants.
We studied interrelationships between CSF biomarkers and associations with
ε4 genotype, demographic variables, vascular variables, and clinical diagnosis in Olmsted County, Minnesota.
We included 774 ...Mayo Clinic Study of Aging participants (693 cognitively unimpaired CU; 71 with mild cognitive impairment MCI). CSF β-amyloid 42 (Aβ42), total tau (t-tau), and hyperphosphorylated tau (p-tau) were analyzed using Aβ42 CSF, t-tau CSF, and p-tau (181P) CSF electrochemiluminescence immunoassays. Bivariate mixture models were used to evaluate latent classes. We used linear regression models to evaluate independent associations of
ε4, demographic factors, cardiovascular risk, and diagnosis with CSF biomarker levels. Results were weighted back to the Olmsted County population.
Interrelationships between CSF Aβ42 and p-tau/t-tau were consistent with 2 latent classes in the general population. In subgroup 1 (n = 547 71%), we found a strong positive correlation between Aβ42 and p-tau (ρ = 0.81), while the correlation was much smaller in group 2 (ρ = 0.26, n = 227 29%). Group 2 was associated with older age,
ε4 genotype, a diagnosis of MCI, and elevated amyloid PET. Overall,
ε4 genotype and MCI were associated with Aβ42, while age was associated with p-tau/t-tau. There were no associations with sex, education, or vascular risk.
We hypothesize the population without dementia can be subdivided into participants with and without biological Alzheimer disease (AD) based on the combination of CSF Aβ42 and p-tau/t-tau (represented also by the p-tau/t-tau/Aβ42 ratio). In those without biological AD, common factors such as CSF dynamics may cause a positive correlation between CSF Aβ42 and p-tau/t-tau, while AD leads to dissociation of these proteins.
The objective of this study was to determine whether constipation-predominant irritable bowel syndrome (IBS-C) is associated with changes in intestinal barrier and secretory function.
A total of 19 ...IBS-C patients and 18 healthy volunteers (all females) underwent saccharide excretion assay (0.1 g
C mannitol and 1 g lactulose), measurements of duodenal and colonic mucosal barrier (transmucosal resistance (TMR), macromolecular and Escherichia coli Bio-Particle translocation), mucosal secretion (basal and acetylcholine (Ach)-evoked short-circuit current (Isc)), in vivo duodenal mucosal impedance, circulating endotoxins, and colonic tight junction gene expression.
There were no differences in the in vivo measurements of barrier function between IBS-C patients and healthy controls: cumulative excretion of
C mannitol (0-2 h mean (s.e.m.); IBS-C: 12.1 (0.9) mg vs. healthy: 13.2 (0.8) mg) and lactulose (8-24 h; IBS-C: 0.9 (0.5) mg vs. healthy: 0.5 (0.2) mg); duodenal impedance IBS-C: 729 (65) Ω vs. healthy: 706 (43) Ω; plasma mean endotoxin activity level IBS-C: 0.36 (0.03) vs. healthy: 0.35 (0.02); and in colonic mRNA expression of occludin, zonula occludens (ZO) 1-3, and claudins 1-12 and 14-19. The ex vivo findings were consistent, with no group differences: duodenal TMR (IBS-C: 28.2 (1.9) Ω cm
vs. healthy: 29.8 (1.9) Ω cm
) and colonic TMR (IBS-C: 19.1 (1.1) Ω cm
vs. healthy: 17.6 (1.7) Ω cm
); fluorescein isothiocyanate (FITC)-dextran (4 kDa) and E. coli Bio-Particle flux. Colonic basal Isc was similar, but duodenal basal Isc was lower in IBS-C (43.5 (4.5) μA cm
) vs. healthy (56.9 (4.9) μA cm
), P=0.05. Ach-evoked ΔIsc was similar.
Females with IBS-C have normal colonic barrier and secretory function. Basal duodenal secretion is decreased in IBS-C.
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