An approach for evaluating the effects of dietary fiber polysaccharides on the human gut microbiome and plasma proteome

Increases in snack consumption associated with Westernized lifestyles provide an opportunity to introduce nutritious foods into poor diets. We describe two 10-wk-long open label, single group assignment human studies that measured the effects of two snack prototypes containing fiber preparations from two sustainable and scalable sources; the byproducts remaining after isolation of protein from the endosperm of peas and the vesicular pulp remaining after processing oranges for the manufacture of juices. The normal diets of study participants were supplemented with either a pea- or orange fiber-containing snack. We focused our analysis on quantifying the abundances of genes encoding carbohydrate-active enzymes (CAZymes) (glycoside hydrolases and polysaccharide lyases) in the fecal microbiome, mass spectrometric measurements of glycan structures (glycosidic linkages) in feces, plus aptamer-based assessment of levels of 1,300 plasma proteins reflecting a broad range of physiological functions. Computational methods for feature selection identified treatment-discriminatory changes in CAZyme genes that correlated with alterations in levels of fiber-associated glycosidic linkages; these changes in turn correlated with levels of plasma proteins representing diverse biological functions, including transforming growth factor type β/bone morphogenetic protein-mediated fibrosis, vascular endothelial growth factor-related angiogenesis, P38/MAPK-associated immune cell signaling, and obesity-associated hormonal regulators. The approach used represents a way to connect changes in consumer microbiomes produced by specific fiber types with host responses in the context of varying background diets.

Advances in our understanding of the role of the gut microbiome in regulating many aspects of human physiology hold the promise of evolving our view of human nutrition by establishing mechanistic connections between the foods we consume and how they affect health status. One manifestation of this effort is a series of studies, performed on well-phenotyped cohorts, that seek to relate features of gut microbial community composition (organisms, genes), dietary practices, and pre- and postprandial cardiometabolic responses to test meals (1–4). A key question raised by these initiatives relates to the nature of the “bioactive” components of foods. Specifically, what are the nutrients utilized by various gut community members or microbiome-encoded metabolic pathways? What products are produced by biotransformation of these nutrients? How are these products linked to specific host physiologic (or pathophysiologic) processes?Plant-derived dietary fibers represent a “poster child” for these efforts and illustrate the formidable challenges faced. The health benefits of dietary fibers are widely known, as is their inadequate representation in Western diets. However, natural fibers are structurally complex and highly diverse. They contain numerous, typically undefined polysaccharide structures and largely unspecified protein, lipid, and small molecule constituents. Their composition varies as a function of their origin (food staple and cultivar), the different methods employed to recover them from these sources, as well as the different techniques used to incorporate them into processed foods with acceptable organoleptic properties (5). Moreover, analyzing the host effects of metabolism of different fibers is confounded by the fact that there is substantial intra- and interpersonal variation in microbiome configuration (6, 7).Snacking is becoming an ever more dominant feature of daily life worldwide and thus provides an opportunity to introduce nutritious ingredients, such as fibers, into diets. However, obtaining structure-activity relationships for specific fiber types and their corresponding targets in the gut community is foundational for designing snack foods that evoke and/or reinforce microbiome responses that are beneficial to the host.Degradation of dietary polysaccharides is a function primarily performed by bacterial carbohydrate-active enzymes (CAZymes). The gut microbiome harbors tens of thousands of CAZyme genes belonging to at least 136 glycoside hydrolase (GH) and 29 polysaccharide lyase (PL) families [extrapolated and updated from El Kaoutari et al. (8)]. In contrast, the human genome only contains 98 GH and no PL genes (9), of which <20% contribute to the processing of dietary glycans.In the current study, we test the effects of dietary supplementation with two snack food prototypes, one containing pea fiber and the other orange fiber, in two pilot studies of overweight and obese individuals consuming their normal, unrestricted diets. Our strategy was to focus on fiber-associated changes in the abundances of microbial GH and PL genes to determine whether responses to the pea or orange fiber prototypes in the gut microbiome and host are decipherable against a background of varying dietary practices and starting microbiome configurations. Higher order singular value decomposition (10) was utilized as a feature selection tool to identify treatment-discriminating changes in GH and PL gene representation. Mass spectrometric assays of the levels of fecal glycan structures (glycosidic linkages) were subsequently performed and the results were correlated with changes in the abundances of treatment-discriminating GH and PL genes with known or predicted substrate specificities. Our analysis concluded by measuring changes in levels of 1,305 plasma proteins in each study participant as a function of fiber treatment and applying computational tools to identify links between these microbiome and plasma proteome changes in response to fiber consumption. Our results provide an approach, using pilot human studies, for selecting specific fiber preparations, plus informative microbiome and host biomarkers, that can be advanced to proof-of-concept clinical trials which assess their capacity for precise manipulation of microbiome and host features.     

Quantitative glycoproteomics of high-density lipoproteins

In this work, we developed a targeted glycoproteomic method to monitor the site-specific glycoprofiles and quantities of the most abundant HDL-associated proteins using Orbitrap LC-MS for (glyco)peptide target discovery and QqQ LC-MS for quantitative analysis. We conducted a pilot study using the workflow to determine whether HDL protein glycoprofiles are altered in healthy human participants in response to dietary glycan supplementation.

The optimized HDL glycoproteomics method was sensitive enough to detect the effects of dietary supplements on HDL protein glycoprofiles even in a small sample size.     

自身免疫甲状腺病循环和甲状腺内细胞毒性T细胞(T_8/S_6F_1)的实验研究

应用新型单克隆抗体T_8/S_6F_1和流体细胞测定仪双色荧光程序检测了15例Graves病和8例Hashimoto甲状腺炎患者循环和甲状腺内的细胞毒性T细胞(cytotoxic T ceils,Tc)的分布。传统的单克隆抗体CD_8(OKT_8或Leu_2)仅能检测含有两个次亚群——抑制性T细胞(Ts)和细胞毒性T细胞(Tc)的Ts/TcT细胞亚群,不能将它们区别开来。本文应用的单克隆抗体T_8/S-6F_1首次能够单独测定Tc亚群。Graves病和Hashimoto甲状腺炎患者的循环Tc亚群分别为27.6±11.2％和27.5％±12.2％,与正常对照组差异无显著性。两组患者甲状腺内的Tc亚群数目也未见增加。Graves病患者甲状腺内Te细胞为16.45±4.7％。本文结果提示:AITD患者循环内和甲状腺内的细胞毒性T细胞亚群的数目未见增加。     

Magnetic resonance imaging of prosthetic heart valves

To evaluate the safety of magnetic resonance (MR) imaging of prosthetic heart valves, nine different synthetic and tissue valves were studied ex vivo. Deflection was measured in 0.35-tesla (T) and 1.5-T superconducting magnets and at the edge of the bore of a 2.35-T electromagnet in field gradients of 5, 1.1, and 6.3 mT/cm, respectively. No valve deflected in the 0.35-T magnet; six synthetic valves deflected 0.25 degrees-3 degrees in the 1.5-T magnet; all valves deflected 1 degree-27 degrees at the edge of the 2.35-T magnet. Each valve was then submerged in a vial of water and the temperature was measured immediately before and after each of two spin-echo imaging sequences in the two superconducting magnets. No significant temperature rise followed exposure in either magnet. Image distortion varied from negligible to severe in both imagers; magnitude of distortion paralleled magnitude of deflection. These data suggest that patients with present-day prosthetic heart valves can be safely imaged in present-day MR imagers and that prosthesis-induced artifacts will not interfere with interpretation in most instances.     

A computerized method for rapid quantification of gallbladder volume from real-time sonograms

A computerized method that requires only 1-2 minutes to quantify gallbladder volume from real-time sonograms is described. This time is considerably shorter than that required using the hand-calculation method. There was a highly significant correlation between gallbladder volumes calculated by computer and hand (r = 0.97; P less than .001).     

ECG-synchronized cardiac MR imaging: method and evaluation

An electrocardiographic (ECG) sensing and gating device compatible with a 0.35-tesla (T) magnetic resonance (MR) imager has been developed and used to produce 802 MR images of the heart in 30 patients. The instrument consists of an isolated acquisition module, an electrically floating preamplifier, and a monitor gating module. Two spin-echo images were acquired for each of five, 0.7-cm thick, transaxial sections from the base to the apex of the heart during each ECG-synchronized imaging run. Image quality was assessed in a blind study by two investigators, on a scale from 0 to 3, as diagnostic [2-3] or nondiagnostic [0-1]. There was agreement in 91.4% of their assessments of diagnostic images (68.1% of the images studied). Resolution of heart anatomy on the MR images was adversely affected by prolonged spin-echo time delay, imaging in late diastole, image acquisition at the cardiac apex, irregular triggering, and artifacts. The synchronization of gradient pulses to the ECG at 0.35 T appears safe for patients, permits diagnostic resolution of images, allows image acquisition at distinct points during the cardiac cycle, and enables monitoring of patients during imaging.