bluuebunny/crossref_metadata_2025 huggingface dataset:
Exposure of intestinal epithelia to IFNg results in enhanced PMN transepithelial migration
Authors: Lee Winston Y , Nava Porfirio , Kolegraff Keli , Chin Alex , Nusrat Asma , Parkos Charles A
Published: June 2021

In inflammatory intestinal diseases, cytokines play a major role in regulating neutrophil (PMN) infiltration of intestinal mucosa and the degree of PMN migration correlates with the severity of mucosal injury. Previous studies using PMN isolated by a gelatin‐based method demonstrated decreased PMN transepithelial migration (TEM) across model intestinal epithelial cell (T84) monolayers treated with IFNg for 48h. This gelatin based method has recently been reported to enhance priming of PMN compared to dextran based methods. Priming of PMN has been shown to enhance oxidative burst and inhibit migration. Therefore, we evaluated dextran isolated (unprimed) PMN in TEM assays across IFNg activated T84 monolayers. In contrast to results obtained with gelatin isolated PMN, dextran isolated PMN exhibited enhanced TEM across T84 monolayers treated with IFNg. The effect on migration required at least 12h of cytokine treatment on T84 cells and was not due to enhanced epithelial permeability as transepithelial resistance of T84 monolayers remained above 1000Ω‐cm2 after 24h of IFNg exposure. Analyses of intercellular junction proteins in T84 cells after IFNg treatment revealed internalization and downregulation of desmosomal cadherins with no gross alteration in tight junction proteins such as occludin. We speculate that IFNg may regulate PMN TEM through alterations of desmosomal junctions.
Antibody‐Antigen Dissociation: A Novel Diagnostic Tool in Alzheimer Disease
Authors: Smith Mark A. , Gustaw Kasia , Garrett Matthew R. , Siedlak Sandra L. , Zhu Xiongwei , Friedland Robert P. , Lee Hyoung‐gon , Perry George
Published: June 2021

With the increasing population of aged individuals with Alzheimer disease (AD), the cost of assessment and ineffectiveness of treatment following diagnosis, there is an urgent need for a sensitive, non‐invasive, diagnostic standard. In the search for a biomarker, no consistent change in plasma amyloid‐beta levels has been found in AD patients. In biological fluids, antibodies and antigens are in a state of dynamic equilibrium that is concentration dependent. As such, when complexed, antigen effectually masks a proportion of its corresponding antibody, and limits both antibody and antigen detection. This interaction can be interrupted, thus freeing antibody and antigen and providing a more accurate analysis of antibody titers and antigen concentration. In this study, sera collected from 36 AD patients and 24 age‐matched controls were examined before and after dissociation. Statistical analysis shows no difference between AD and control patients before dissociation. After dissociation, however, the level of amyloid‐betafnantibody assessed was greater than before dissociation in all cases. Importantly, levels from AD cases are much greater than controls. The importance of measuring total amounts of antigen or antibody following unmasking, has been shown to be very important, and as shown here, could be an important diagnostic tool in AD.
Effect of Gelsolin on Cerebral Amyloid Angiopathy (CAA) in Transgenic Mice
Authors: Frosch Matthew P. , Prada Claudia M. , Garci‐Alloza Monica , Betensky Rebecca A. , Bacskai Brian J. , Greenberg Steven M.
Published: June 2021

Cerebral amyloid angiopathy (CAA), the deposition of cerebrovascular β‐amyloid (Aβ), is a cause of hemorrhagic stroke and is present in most cases of Alzheimer disease. We have previously developed a quantitative approach to measure CAA progression in mice and have shown that CAA can be reversed by passive immunotherapy. We now report the effect of peripheral administration of gelsolin, an agent with the ability to bind Aβ in the plasma and that may thus act as a peripheral sink for Aβ, on the rate of CAA progression. Cranial windows were placed in 11 month old mice, an age at which CAA deposition follows a mixed‐effects linear model. Seven days post‐craniotomy, mice were randomized to intraperitonal administration of either gelsolin (0.6 mg/kg) or vehicle every 2 days for 3 weeks. CAA in leptomeningeal arteries was imaged on day 0, 7, 14 and 21 post‐craniotomy using methoxy‐X04, with angiograms used to facilitate vessel alignment. Between day 0 and day 7 (prior to start of treatment) there was CAA progression as expected. Peripherally administrated gelsolin resulted in mild regression of CAA deposits. The effect of the peripherally administrated gelsolin was evident within a week after initial administration and this effect was sustained until day 21 (last imaging session). These data demonstrate that gelsolin, an agent with a peripherally acting effect on Aβ can cause regression of CAA in Tg2576. In addition, these data also suggest that plasma/brain Aβ dynamics may play a crucial role in the progression of CAA. Supported by NIH AG020570, EB00768 and AG21084, Caja Madrid fellowship (MG‐A), Lefler fellowship (CMP) and the Harvard NeuroDiscovery Center (RAB).
Creutzfeldt‐Jakob Disease in the Baltimore, Maryland Area: A Recent Survey
Authors: Gonzalez‐Cuyar Luis Francisco , Morrison Alan L , Castellani Rudy J
Published: June 2021

Objective: To provide an overview of cases of Creutzfeldt‐Jakob disease (CJD) referred to the University of Maryland for autopsy.

Methods: Autopsy tissue from subjects with CJD over two years was obtained and dissected per standard protocol. Routine histopathology, PrP immunohistochemistry, Western blot, and PrP gene analysis were performed. Results: Ten biochemically confirmed cases of CJD were identified. Age ranged from 51 to 77 years of age (5 male, 5 female). All were consistent with sporadic disease, with molecular phenotyping according to Parchi et al (Ann Neurol 1999 Aug;46(2):224‐33). 5 subjects had the classic phenotype (MM1); 3 had the kuru plaque variant (MV1), 1 had the ataxic phenotype (VV2), and 1 had the cortical phenotype (MM2). Of note, one subject had premorbid Parkinsonâ€™s disease, confirmed pathologically. A second subject (age 51) was an intravenous drug user who shared needles with an individual who expired from an uncharacterized neurological decline. No atypical clinical, pathological, or molecular features were noted. Conclusion: A recent survey of subjects with CJD in the Baltimore area demonstrated sporadic cases only, and a typical distribution of molecular phenotypes. One subject was had premorbid Parkinsonâ€™s disease, while a second subject was an intravenous drug user. Whether the latter comprises a significant risk factor for CJD awaits further study.
A Novel Human Disease with Abnormal Prion Protein Sensitive to Protease
Authors: Gambetti Pierluigi , Alshekhlee Amer , Castellani Rudy , Cohen Mark , Mastrianni James A , Kong Qingzhong , Zou Wen‐Quan
Published: June 2021

All human prion diseases are defined by the presence of protease‐resistant prion protein (PrPres); inherited forms are invariably associated with mutations in the PrP gene open reading frame (ORF). Twenty subjects that formed two groups were evaluated. The patients of the first group presented with behavioral and psychiatric manifestations on average at 62 years with a 21 months mean duration. Type of the spongiform degeneration and the PrP immunostaining pattern were distinct from those of the known prion diseases. PrPres was not detectable on standard tests. Following enrichment, PrPres was detected and formed a distinct electrophoretic profile. All the patients of this group were homozygous valine at codon 129 of the PrP gene. But although several subjects had family history of dementia, no mutations were found in the PrP gene orf suggesting that another gene or another region of the PrP gene may be involved. The patients of the second group were 129 methionine/valine heterozygous. They had a later disease onset and longer duration; otherwise clinical, histopathological and PrP features were similar except for the presence of PrPres. This novel prion disease raises issues concerning its nature and classification. Since many cases were initially diagnosed clinically as non Alzheimer's disease dementia, currently this novel condition may be hidden in this dementia group and be more prevalent than our data indicate.
Cystatin C: A potential surrogate biomarker in amyotrophic lateral sclerosis
Authors: Wilson Meghan E. , Darko Samuel , Lacomis David , Bowser Robert
Published: June 2021

Amyotrophic lateral sclerosis (ALS) is a fatal neurological disease characterized by progressive motor neuron degeneration. Currently, disease management is hindered by a lengthy diagnostic process which is based heavily on clinical criteria. A reliable panel of biomarkers is needed in order to accelerate the diagnosis of this disease and aid in monitoring its progression. We have previously reported proteomic evidence supporting a reduction of Cystatin C protien levels in the cerebrospinal fluid (CSF) of ALS patients relative to controls. In this study, we have evaluated the potential of Cystatin C as a diagnostic and/or prognostic biomarker by assessing its expression over the course of disease progression. We used a quantitative ELISA to evaluate Cystatin C concentration in both CSF and plasma at multiple time points following ALS symptom onset. Cystatin C levels in both fluids were found to correlate strongly with clinical measures of disease progression. This correlation was maintained during transient functional improvements observed in some patients during drug trial participation. Additionally, expression patterns were observed to vary between specific subpopulations of ALS patients. These data support the utility of Cystatin C as both a diagnostic biomarker and a surrogate biomarker for monitoring disease progression and treatment response.

Funded by: University of Pittsburgh Center for ALS Research
Large‐scale volume data analysis for meso‐ and macroscopic physiological modeling
Authors: Stiles Joel R , Nave Demian , Hood Greg , Dittrich Markus , Wetzel Arthur
Published: June 2021

Modeling is increasingly important to biological investigations, from basic cell biology to cognitive neuroscience. For many years, however, models and simulations have been severely limited by a lack of spatial, i.e. anatomical, realism. Simulation methods that incorporate highly detailed anatomy are expensive and complex, but developers are targeting ever increasing 3‐D spatial realism and have made major advances in recent years. Thus there is a renewed need for quantitative 3‐D anatomical data at subcellular to tissue scales, with many remaining challenges: (1) Automating detailed analyses from varied imaging modalities; (2) Handling potentially enormous datasets; (3) Correcting artifacts introduced by tissue preparation and imaging method; (4) Annotating datasets to include model parameters, e.g., molecular constituents and physical properties of intra‐ and extracellular spaces and membrane compartments; (5) Production of surface and volume meshes that include annotated properties; and (6) Formatting of meshes and properties for use with one or more simulation methods. We are developing PSC Volume Browser, DReAMM, and MCell software to address all of these challenges, with special emphasis on arbitrarily large volumetric datasets and Monte Carlo diffusion‐reaction simulation methods. We will present examples ranging from semi‐automated analysis of cardiac time series from micro‐CT imaging, to nonlinear registration and alignment of large‐scale serial EM sections, surface mesh generation using volume segmentation and computer‐aided design approaches, and quantitative simulation of microphysiological events such as calcium‐dependent neurotransmitter release from nerve terminal active zones. Supported by NIH P41 RR06009, R01 GM068630, NLM N01‐LM‐9‐3531, and NIH/NSF 0234002.
Animal models to investigate lung inflammation induced following exposure to swine barn air
Authors: Charavaryamath Chandrashekhar , Podborochynski Rachel , Janardhan Kyathanahalli S , Juneau Vanessa , Suri Sarabjeet Singh , Townsend Hugh , Singh Baljit
Published: June 2021

Modern swine barns house large number of pigs in confined buildings. The barn air contains microbes, dust, gases and endotoxin. Swine farmers experience loss of lung function and suffer from respiratory diseases. Exposure of naïve volunteers to swine barn air has shown to induce of lung inflammation and increased airway hyperresponsiveness (AHR) whose mechanisms remain unclear.

We exposed rats to either swine barn or ambient air for eight hours per day for 1, 5 or 20 days. Following 20 day exposure and either 2 or 5 week recovery, rats were challenged either with saline or lipopolysaccharide. We measured AHR and lung inflammation. Exposure to barn induced AHR, mucus‐positive airway epithelial cells and lung inflammation while prolonged exposure dampened the AHR. Following barn exposures and recovery, rats showed lung inflammation with an increased intensity following lipopolysaccharide challenge.

To clarify the role of Toll‐like receptor 4 (TLR4) in inducing lung inflammation and AHR, we exposed C3HeB/FeJ (intact TLR4, WT) and C3H/HeJ (defective TLR4, mutant) mice in a similar pattern. Both strains showed increased and dampened AHR following 5 and 20 exposures respectively while barn exposure induced lung inflammation in WT but not in mutant mice to show different roles of TLR4 in lung inflammation and AHR (CIHR's strategic training grant in PHARE and Lung Association of Saskatchewan).
Can we encourage our graduate students to develop a more scholarly approach to classroom teaching?
Authors: O'Loughlin Valerie Dean , Braun Mark , Kearns Katherine Dowell , Heacock Isaac
Published: June 2021

Previous research indicates that graduate courses in pedagogical methods produce specific teaching behaviors in the short term. However, little assessment has been done to examine how teaching behaviors and attitudes endure over time. Assessments of pedagogy courses should continue well past the completion of the course, in order to determine the persistence and ongoing development of the students' perceptions of teaching and learning. Indiana University faculty members Valerie O'Loughlin and Mark Braun developed and team‐taught a three credit hour pedagogy course (M620: Pedagogical Methods in the Health Sciences) to prepare graduate students to be reflective and scholarly teachers. They examined the impact of the course on how well graduate students:

* Develop more complex and refined attitudes regarding student learning;

* Evaluate their current teaching methods and explore alternative teaching strategies;

* Practice methods of assessment and evaluation; and

* Plan to be involved in a public teaching resource network, such as teaching workshops or a scholarship of teaching and learning program.

Data examined include students' teaching philosophy statements, teaching portfolios, pre‐ and post‐course surveys, and course evaluations. In addition, data from a six‐month follow‐up student interview was used to examine the lasting and ongoing influence the course has on graduate student attitudes about teaching and learning. The results of this study should contribute to the literature on short‐term and longer‐term effects of these courses on the development of future faculty.

This research was funded, in part, by an Indiana University Scholarship of Teaching and Learning Grant.
Creating a Teacher‐Scholar Training Program for Anatomy at the University of Utah School of Medicine
Authors: Albertine Kurt
Published: June 2021

Developing doctoral programs to train the next generation of anatomy teacher‐scholars will be essential to respond to the manpower shortage of medical gross anatomy, histology, embryology, and neuroanatomy teachers that is anticipated by 2010. At the program level, rigor is required to meet the expectations of traditional doctoral training programs, such as education in theoretical and practical disciplines. Our program provides training in educational research theory because advancement in academic medicine is dependent on research scholarship. That training goal is complimented by education in ethics of research, institutional review board (IRB) protocols, scientific writing, and biostatistics. Other courses, such as scientific lecturing, journal club, and research‐in‐progress, provide practical experience in presenting research. An IRB‐approved, educational research project is required, for which the results are to be submitted as a first‐authored, peer‐reviewed manuscript. Another requirement is innovation of teaching tools, such as paper or electronic textbooks, dissection guides, or educational software. Trainees also take and teach medical gross anatomy, histology, embryology, and neuroscience. At the institutional level, recognition of and reward for teacher‐scholar contributions and accomplishments is required. Our institution, like other institutions, has created as faculty track for teacher‐scholars, which reflects the growing change in attitude about the value of teaching in medical schools. Further programmatic growth requires curriculum integration with the College of Education, and establishment of endowed funding for trainee stipends. [Support from Dept of Neurobiol & Anat and Medical Scholars Program]
Membrane Lipid Biogenesis in B‐Lymphocytes
Authors: Jackowski Suzanne , Fagone Paolo , Brewer Joseph W.
Published: June 2021

Stimulated B‐lymphocytes differentiate into plasma cells committed to immunoglobulin production. The B‐cells proliferate and expand the endoplasmic reticulum to enhance immunoglobulin synthesis and secretion. Cellular membrane lipid components increase in response to stimulation using a genetic and biochemical program of events. Selected genes in fatty acid and phospholipid synthesis are up‐regulated, most notably the genes encoding Lipin1 and the choline phosphotransferase. The choline cytidylyltransferase (CCT) controls the rate of phosphatidylcholine production but its expression does not increase. Rather, the protein is stabilized and activated allosterically by association with intracellular membranes whose lipid composition changes. Two CCT isoforms are expressed in B‐cells. Knockout of the major CCTα isoform impairs proliferation, resulting in B cells that are unable to undergo immunoglobulin gene re‐arrangement and cannot switch from IgM to IgG production in response to immunization. IgM secretion is maintained in the knockouts, however, due to expression of the alternate isoform CCTβ2. These data suggest either different roles for the CCT isoforms or that a threshold of CCT protein is required to support both proliferation and secretion in B‐cells. Supported by NIH GM45737 (S.J.), GM61970 (J.W.B) and by ALSAC.
Using Conus venom peptides to understand nervous systems and discover drugs
Authors: Olivera Baldomero M.
Published: June 2021

The predatory cone snails have complex venoms; each of the 700 species has 100–200 venom components, mostly small (10–30AA), disulfide‐rich peptides. Conopeptides generally target receptors or ion channels in the nervous system, but there is virtually no molecular overlap between venom components from different Conus species, suggesting over 100,000 peptides in living Conus venoms. Because conopeptides are generally highly specific for a particular molecular target, they have been extensively used to investigate nervous system function, and have therapeutic and other biomedical applications as well. One conopeptide (Prialt) is an approved drug for intractable pain. Each individual peptide can be used as a molecular probe for a specific pharmacological site on a receptor/ion channel complex. The discovery work on Conus peptides has provided a general strategy for the systematic discovery of leads for drugs acting on the nervous system from animal biodiversity. This involves analyzing, in a phylogenetically informed manner, rapidly diversifying gene families (“exogenes”) encoding compounds that act on other animals. Supported by GM48677 from the National Institute of General Medical Science.
High‐Speed and Long‐Term Imaging of Cell Dynamics in the Developing Heart
Authors: Liebling Michael , Vermot Julien , Fraser Scott E.
Published: June 2021

We developed in vivo image acquisition strategies and digital post‐processing algorithms to follow tissue growth and track individual cells in the developing embryonic zebrafish heart. Cardiac imaging at high spatial resolution is notoriously difficult because the heart is beating while it develops and cell velocities can be as high as 0.5 mm/s. This dynamic environment is an important epigenetic factor for the proper formation of many of the heart's features (e.g. heart tube looping, valve formation). It is therefore essential to capture both the rapid dynamics of the heartbeat and the much slower dynamics of morphogenesis. Although state of the art microscopes provide high framerate capabilities, they still suffer from the inherent limitations of three‐dimensional imaging (images must be acquired successively at multiple depths) and the increased noise level as the number of photons per pixel decreases. We address the problems of imaging at multiple time scales, in three‐dimensions, and at low signal‐to‐noise levels via a general image acquisition and processing procedure that takes advantage of the pseudo‐cyclic motion of the heart and allows us to follow cells and tissue in their native environment as they grow while the heart beats.
Focus on Function: Single Molecule Enzymology of the VS ribozyme
Authors: Walter Nils G , Pereira Miguel J.B. , Collins Richard A.
Published: June 2021

Non‐coding RNAs of complex structure are involved in numerous aspects of the replication and processing of genetic information. Within the mitochondria of the bread mold Neurospora, the Varkud satellite (VS) RNA replicates parasitically by cycling between DNA and multimeric RNA forms. The RNA contains the VS ribozyme whose self‐cleavage is thought to be essential for replication. Models of VS ribozyme tertiary structure predict three parallel helical stacks built from two three‐way junctions and a kissing loop interaction. Whether or not such complex RNA architectures are dynamic and how catalytic activity emerges is ill‐understood. Here we exploit single molecule FRET to show that the VS ribozyme exhibits dynamic hierarchical folding. Reversible kissing loop formation involving the upstream substrate is required but only partially rate‐limiting for catalysis. Our results invoke a model whereby the VS ribozyme kinetically selects the distal downstream substrate for cleavage before the proximal upstream substrate, thus facilitating processing of the multimeric RNA replication intermediate. We are currently further testing this model.
Metabolic and genomic changes induced by macronutrient composition vs. caloric restriction
Authors: Kalupahana Nishan Sudheera , Giannone Richard J , Fletcher Sarah J , Kim Hyoung‐Yon , Stewart Allison , Hall Lorin , Saxton Arnold , Voy Brynn H , Moustaid‐Moussa Naima
Published: June 2021

The aim of this study was to investigate differential effects of dietary fat and energy restriction on metabolic and genomic differences in adipose tissue. Eight week‐old C57/BL6 mice were either given free access to a low fat (LF) or high fat (HF) diet, or were initially fed HF for 16 weeks followed by 8‐weeks of caloric restriction (HFR, 70% of HF ad lib caloric intake). As expected, the HF group gained more body and fat mass than the LF group, and energy restriction resulted in loss of body fat and weight. Although fasting glycemia was comparable between the 3 groups, HFR group had lower insulinemia, compared to HF and LF groups. Analysis of serum adipokines indicated that plasma leptin levels were lowest in HFR mice; surprisingly, plasma adiponectin levels were also lowest in this group, despite their apparent enhanced insulin sensitivity. Microarray analysis of gonadal fat indicated upregulation of genes involved in fatty acid synthesis, oxidation and glycolysis in the LF group. Differentially expressed genes in the HF group included several oncogenes, while those in HFR group included genes related to insulin sensitivity and apoptosis. These studies demonstrate differential effects of low fat diet and caloric restriction on fatness and insulin sensitivity, and on the molecular responses in adipose tissue.
Folic acid dose, MTHFR C677T genotype, and temporal effects on plasma folate species
Authors: Pfeiffer Christine M , Fazili Zia , Hao Ling , Berry R J , Bailey Lynn B , Zhu Jianghui , Crider Krista , Li Zhu
Published: June 2021

We evaluated the effect of chronic folic acid (FA) intake (100, 400, 4000 μg/d) and of the MTHFR 677C→T polymorphism on plasma folate species measured by LC‐MS/MS in 90 reproductive age Chinese women (30 women/treatment group, 50% CC, 50% TT genotype) not previously exposed to FA. 5‐Methyltetrahydrofolic acid (5CH3THF) was the major folate form in all treatment groups (~80% of total folate [TFOL]) at baseline and at 6 mo. 5‐Formyltetrahydrofolic acid (5CHOTHF) was a smaller contributor to TFOL, ~20% at baseline and ~13% at 6 mo. The increase in TFOL as a result of dosing was attributable to 5CH3THF. Plasma levels of FA were generally below the limit of detection at baseline and slightly above at 6 mo; mean levels were <1 nmol/L in all treatment groups. The distribution of folate forms did not change with dose or genotype. TFOL increased (p<0.001) from baseline (14, 16, 12 nmol/L, respectively) to 6 mo (19, 30, 31 nmol/L, respectively) for each treatment group. In this small subset, women with TT genotype had lower TFOL levels (~30%) than those with CC genotype, both at baseline (p<0.001) and at 6 mo (p=0.038). The increase in TFOL from baseline to 6 mo appeared lower in women with CC genotype (86%) than in those with TT genotype (113%). These data suggest that in fasting women who are exposed to only 1 daily dose of FA up to 4000 μg/d there are minimal levels of FA circulating in plasma. This study was supported by a collaborative CDC agreement.
The gastric intrinsic factor polymorphism, A68G, modifies the association between the transcobalamin polymorphism, C776G, and vitamin B12 status
Authors: Lee Kyung‐Seon , Garrod Marjorie G , Allen Lindsay H , Haan Mary N , Green Ralph , Miller Joshua W
Published: June 2021

Gastric intrinsic factor (GIF) and transcobalamin (TC) are required for absorption of vitamin B12 across the ileal enterocyte. Polymorphisms in GIF (A68G) and TC (C776G) have been identified. Previously, we observed differences in plasma holoTC and homocysteine levels among individuals with the TC C776G polymorphism in a cohort of elderly Latinos (Sacramento Area Latino Study on Aging ‐ SALSA). Little is known about the effect of GIF A68G on indicators of B12 status, although the polymorphism has been linked to risk of pernicious anemia. In this study we investigated the prevalence of GIF A68G and its association with B12 status and TC genotype in the SALSA cohort (N=813, age 60–93y). The distribution of GIF genotypes was 94% wild‐type (AA) and 6% heterozygotes (AG), with 1 subject homozygous for the variant isoform (GG). No differences among the GIF genotypes were observed in total B12, holoTC, or homocysteine. However, interactions between GIF genotype and TC genotype were observed on total B12 (p=0.007) and holoTC (p=0.05) such that subjects with both the GIF 68G allele and TC 776G allele had lower total B12 and holoTC than all other allelic combinations. There was no interaction between the GIF and TC genotypes on homocysteine. The GIF A68G polymorphism is prevalent in elderly Latinos and combined with the TC C776G polymorphism may influence B12 absorption and status. NIH AG12975; USDA 00–35200–9073
Soy isoflavone (IF) supplementation appears safe after a two‐year randomized controlled trial (RCT)
Authors: Steinberg Francene , Murray Michael , Lewis Richard , Cramer Margaret , Amato Paula , Young Ronald , Barnes Stephen , Fraley Kennard , Konzlemann Karen , Smith E.O. , Fischer Joan , Wong William
Published: June 2021

Although RCTs quantifying a benefit for menopausal soy IF supplementation abound, few studies have considered the safety of long‐term soy IF intake, particularly when exposure levels are in the form of a tablet, as opposed to food. To evaluate the effect of soy IF on bone loss, 403 early postmenopausal women (54.0±4.0 y) were enrolled in a multi‐site, 2‐year, double‐blind RCT with 1/3 on placebo, 1/3 on 80 mg/d IF and 1/3 on 120 mg/d IF. Baseline characteristics were similar in all groups. The study design focused on safety as a secondary outcome measure and included serum electrolytes, lipids, liver and renal functions, complete blood counts, reproductive and thyroid hormones and cancer (CA) screening. In the supplement groups versus placebo, there was a decrease in free thyroxine (T4) (p=0.024) and an increase in blood urea nitrogen (BUN) (p=0.048), but not thyroid stimulating hormone (p=0.264) or creatinine (p=0.145) after 2 years. Absolute changes in free T4 and BUN were neither outside the normal range nor clinically significant. Major adverse events included one subject with endometrial CA and one with breast CA, which were fewer than predicted in this study population. Soy IF supplementation up to 120 mg/d has no measurable deleterious effect over a 2‐year interval. Supported by Initiative for Future Agriculture and Food Systems Grant 2001‐52102‐11255 USDA CSREES.
Polypeptide from 7S soy globulin alpha' subunit up‐regulates LDL‐receptors in HepG2 cells
Authors: Lovati Maria Rosa , Manzoni Cristina , Pizzagalli Anna , Castiglioni Silvia , Duranti Marcello Maria
Published: June 2021

7S soy globulin and its alpha’ subunit have been shown to positively modulate LDL receptor activity and to decrease triglyceride synthesis both in vitro and in vivo. The aim of the study was to evaluate the effect of a short chain of alpha’ (S‐alpha’) from 7S globulin, on cell cholesterol homeostasis. HepG2 cells were incubated in the presence/absence of alpha’ subunit (3.5.10−6M) or the S‐alpha’ at concentration of 10−5 M and 10−6 M. A dose‐dependent increase in up‐regulation of LDL‐receptors has been detected in cells exposed to S‐alpha’ vs that found in controls. Moreover, the 125I‐LDL uptake (+ 192%) and degradation (+ 143%) of cells exposed to the highest dose of S‐alpha’ (8.10−6M) were similar to that found in cells incubated with 1 mM simvastatin; these results were confirmed by a dose‐dependent increase in the SREBP2 and LDL‐receptor mRNAs. Hep G2 cells exposed to S‐alpha’ for 24 h showed an increased uptake of fluorescent LDL vs both alpha’ and controls as detected by fluorescence microscopy: the fluorescence intensity was proportional to the amount of LDL uptaked by the cells as confirmed by FACS analysis.

These findings open new prospects in the studies aimed at identifying the regulatory peptide/s from soybean proteins and the mechanism/s involved in this biological response, in view of their potential utilization in the management of human lipid disorders.

Supported in part by a grant of MIUR of Italy.
Postprandial carbohydrate sensing in the liver involves components of the unfolded protein response
Authors: Pfaffenbach Kyle , Ellis Flannery , Wei Yuren , Wang Dong , Pagliassotti Michael
Published: June 2021

The Unfolded Protein Response (UPR) is a quality control pathway in the endoplasmic reticulum (ER) that senses and responds to the presence of unfolded proteins. Our understanding of the UPR is primarily based on studies that utilize severe pharmacologic insults in cultured cells. Less studied is the role of the UPR as a putative nutrient sensor in vivo. The aim of this study was to examine whether a high sucrose meal (HS), a carbohydrate source primarily metabolized by the liver, activates the UPR. Rats (n=12) were acclimated to a 3hr meal feeding paradigm using a high starch diet (ST). After 10–11 days, half of the rats were provided ST and half HS ad libitum for 3hr. Liver and blood samples were taken after an additional hour. Food intake (grams) was 14.6±2.0 in ST and 11.7±2.3 (mean±SD) in HS. Splicing of XBP‐1 mRNA, an early marker of UPR activation, was observed in livers from both groups. However, the amount of spliced XBP‐1 mRNA was greater in HS vs. ST. Several UPR gene markers that encode chaperone or pro‐apoptotic proteins were not increased in ST or HS. These data suggest that a feeding‐related stimulus can activate specific components of the UPR in the liver. Support by DK47416.
Activation of anabolic signal transduction pathways in L6 muscle cells in response to Amino Acids (AA) and Insulin
Authors: Patel Rekha , Smith Ken , Rennie Michael , Atherton Philip
Published: June 2021

Muscle protein synthesis (MPS) in vivo is stimulated by feeding‐associated increases in AA and insulin. However, there is limited knowledge of their separate and common effects on signal transduction pathways regulating MPS. We aimed to measure the activation and temporal responses of signalling proteins to insulin, AA or their combination in vitro.

After withdrawal of serum for 6h and of AA for 3h L6 cells were treated with either, AA (1× plasma concentration) or insulin (5 μIU/ml) alone, or a combination thereof. Proteins were extracted at intervals between 15–105 min. Activation of most proteins studied occurred by 15 min. Insulin stimulated PKB, mTOR, 4EBP1, eIF4G, eIF4B and eIF2Bε by >300% at all time‐points. AA alone only stimulated eIF2Bε phosphorylation by ~50%. Addition of AA with insulin potentiated peak insulin‐stimulated phosphorylation of PKB, p70 and 4EBP1 >200%. eEF2 was activated from 45 min onward.

Insulin with or without AA induced rapid activation of growth pathways and their co‐provision augmented insulin‐induced phosphorylation events. Conversely, AA provision alone was insufficient, to induce anabolic signalling, except for eIF2Bε. Delayed activation of eEF2 may be indicative of its role in elongation and not initiation. To conclude, insulin is required for AA‐induced stimulation of MPS, and their co‐provision potentiates activation of signalling proteins regulating MPS.
Effect of folate deprivation on WNT signaling pathway in human colon cell lines
Authors: Huang Min , Chen Hong
Published: June 2021

Human epidemiology studies and designed animal experiments have shown both timing and dose are critical for folate relation with colorectal cancer. The molecular mechanism of folate effect is not fully understood. In our study, we examined gene expression of several specific genes of wnt signaling pathway in human colonic cells lines at different stages with or without folate in culture media. Cell proliferation was inhibited at around day 8 after folate depletion. Quantitative real‐time RT‐PCR results showed that mRNA level of DKK1 was increased whereas AXIN2 and NKD1 decreased with time in SW480. No significant changes were observed in DLD1. Methylation sensitive PCR and bisulfite sequencing were used to examine whether DNA methylation level of these genes altered. The putative CpG island of DKK1 analyzed in this study is totally unmethylated in SW480 and hypermethylated in DLD1 and no significant changes were observed after folate depletion. In conclusion, folate deprivation altered expression of DKK1 in wnt pathway in SW480 human colonic cancer cell line and the effect is not due to DNA methylation levels.
Rapeseed and milk protein induce different patterns of dietary N tissue accretion despite a similar postprandial retention in rats
Authors: Boutry Claire , Chevalier Laure , Fouillet Hélène , Blachier François , Gaudichon Claire , Tomé Daniel , Bos Cécile
Published: June 2021

Rapeseed constitutes an emerging, promising source of dietary protein for human consumption. We previously found that these protein exhibit atypical nutritional properties in humans characterized by a low bio‐availability counterbalanced by a high postprandial biological value. The objective of this study was to further investigate the metabolic fate of rapeseed protein (RP) and their effect on protein synthesis rates (FSR) in various tissues as compared to milk protein (MP). Rats were killed after ingestion of a 15N‐labeled RP or MP meal and after injection of 13C‐Valine. Identical ileal and deamination losses of dietary N (4% and 12% of the meal, respectively) were measured between groups. In contrast, the incorporation of dietary N into intestinal mucosa was significantly higher after RP than MP (+19%) whereas it was lower in skin, plasma protein and muscle (−39%, −24% and −17%, respectively; P<0.05), but not different in liver and kidneys. FSR were not significantly influenced by the nature of protein in any tissue. These results confirm that RP and MP lead to the same level of postprandial retention in rats, consistently with our findings in humans. However, this global response conceals qualitative differences at the tissue level for dietary N accretion, but not for FSR, suggesting a differential modulation of proteolysis after rapeseed or milk protein ingestion or other mechanisms that warrant further study.
Serum retinol binding protein is more strongly associated with retinol than measures of insulin sensitivity and BMI in fasted obese adults
Authors: Mills Jordan P , Tanumihardjo Sherry A , Furr Harold C
Published: June 2021

Elevated serum retinol binding protein (RBP), the major carrier of vitamin A, has been linked to obesity and type 2 diabetes, suggesting therapeutic potential in lowering blood concentrations. Obese men and women (BMI > 30) (n = 76) were fasted for ≥ 8 hr prior to blood collection. Blood chemistries were obtained. Retinol and retinyl esters were quantified by HPLC and RBP by ELISA. The relations between serum RBP, retinol and retinyl esters as well as measures of insulin sensitivity and BMI were determined using the Pearson correlation for simple associations and multiple linear regression for multivariate models. Retinol and RBP were .0001) with a molar ratio (retinol:RBP) of 0.73 Â± 0.14. RBP was not associated with BMI, glucose, insulin or the homeostatic model assessment (HOMA) (P > 0.05), but was inversely associated with the glucose/insulin ratio (r = −0.31, P < 0.02). In multivariate models, only retinol and the glucose/insulin ratio were significant explanatory variables for RBP. Total serum retinyl ester concentration was positively associated with retinol (r = 0.30, P < 0.01) and the association was strengthened after the removal of three potentially hypervitaminotic subjects (retinyl ester ≥ 10% total retinol) from the sample (r = 0.48, P < 0.0001). When evaluating the therapeutic potential of lowering serum RBP, consequences on vitamin A metabolism should be considered.
Bioavailability of chlorogenic acids from green coffee extract in humans
Authors: Farah Adriana , Monteiro Mariana C. , Donangelo Carmen M. , Lafay Sophie
Published: June 2021

Chlorogenic acids (CGA) are phenolic compounds with biological properties mostly related to their antioxidant activity. Caffeoylquinic (CQA) and dicaffeoylquinic (diCQA) acids are the main CGA in nature. Because green coffee is a major source of CGA, it has been used for the production of nutraceuticals. Despite this, data on the bioavailability of CGA from green coffee in humans is inexistent. This preliminary study evaluated the pharmacokinetic profile and bioavailability of CGA isomers and metabolites in plasma and urine of 4 healthy adults for 8 h after the consumption of a green decaffeinated Robusta coffee extract (svetol®) containing 450 μmol of CGA. HPLC‐DAD‐MS results showed that in addition to three CQA, three diCQA and caffeic acid, previously identified after roasted coffee consumption, the metabolites ferulic, isoferulic and p‐coumaric acids were also identified in plasma. On average, 24% of ingested CGA were recovered in plasma, including metabolites, with peak levels observed 2.1 ± 2 h after treatment. CGA/metabolites identified in urine after green coffee consumption were 5‐CQA, gallic, vanillic, sinapinic, dihydrocaffeic, p‐hydroxybenzoic, isoferulic, ferulic, caffeic, and p‐coumaric acids, totaling 30% (average) of urinary recovery. This study shows that the major CGA compounds present in green coffee are highly absorbed and metabolized in humans. Financial support: Berkem, CNPq.
Dairy and related nutrient consumption among US adults and their association with obesity, central obesity and the metabolic syndrome
Authors: Beydoun May A. , Wang Youfa
Published: June 2021

We examined the association between dairy and its related nutrients with obesity, central obesity and metabolic syndrome (MS) using NHANES 1999–04 data collected from a nationally representative sample of US adults. Dietary intakes were assessed using 24‐hour recall and direct anthropometric assessment, blood pressure and lab tests were used. Sample sizes ranged between 14,616 for obesity and 4,519 for MS. Statistical analysis included multivariate linear and logistic models and structural equations modeling (SEM). We found a significant inverse association between intakes of some dairy products (high‐fat milk, yoghurt) and related nutrients (calcium, magnesium) and the main metabolic outcomes. Odds ratios for one yoghurt serving and 100 mg magnesium in relation to MS were 0.43 (0.20, 0.92) and 0.81 (0.71, 0.94), respectively. In contrast, intakes of cheese, low‐fat milk and phosphorus were associated with higher risks. Using SEMs, ethnic differences in some metabolic outcomes such as body mass index were partly explained by variations in intakes of dairy and related nutrients. Thus, future long‐term controlled intervention trials are needed to test the causality and should consider potential effects of various dairy products and the related nutrients in the diet.
Dietary patterns and body fat distribution in older adults
Authors: Anderson Amy L. , Sahyoun Nadine R. , Tylavsky Frances A. , Goodpaster Bret H. , Lee Jung Sun , Sellmeyer Deborah E. , Houston Denise K. , Harris Tamara B.
Published: June 2021

To examine whether dietary patterns are related to adipose tissue deposition in older adults, we used data from the Health, Aging, and Body Composition Study of well‐functioning adults aged 70 to 79 in 1998–99. Food intake was assessed with a modified Block food frequency questionnaire, and dietary patterns were derived by cluster analysis in 1,809 participants with complete data. Six dietary pattern clusters were identified: 1) Meat, snacks, fats and alcohol (n=480); 2) Sweets and desserts (n=257); 3) Refined grains (n=247); 4) Breakfast cereal (n=273); 5) Healthy foods (n=306); and 6) High‐fat dairy products (n=246). The ‘Healthy foods’ cluster was characterized by higher intake of lowfat dairy products, fruit, whole grains, poultry, fish and vegetables, and lower intake of red meat, sweets, added fats and high‐calorie drinks. Compared to the ‘Healthy foods’ cluster, the ‘Meat, snacks, fats and alcohol’ cluster had significantly greater abdominal visceral fat, and it and the ‘Breakfast cereal’ cluster had a significantly higher percent body fat, after controlling for age, gender, race, site, education, physical activity, smoking, calorie intake and PPAR‐γ2 Pro12Ala genotype. No differences were found in BMI or total lean body mass between the ‘Healthy foods’ cluster and other clusters. Results suggest a relationship between dietary patterns and body fat distribution in older adults. Supported by NIA/NIH.
Bacterial Regulation of Vitamin D Receptor in Intestinal Epithelial Inflammation
Authors: Sun Jun , Liao Anne P. , Xia Rick Y. , Kong Juan , Li Yan Chun , Sartor Balfour
Published: June 2021

Vitamin D deficiency has linked to the development of inflammatory bowel disease. Vitamin D receptor (VDR) is required for all known biologic effects of vitamin D. Previous works focused on the immune response. However, how bacteria regulate VDR expression in intestinal epithelial cells has not been investigated. We investigated the effect of VDR ablation on NF‐κB activation using VDR knockout mouse infected with pathogenic Salmonella or human commensal E. coli. We also investigated the intestinal VDR expression in the germ‐free mice. Mice lacking VDR were in the proinflammatory state and became more susceptible to bacterial infection. Intestinal epithelial cells lacking the VDR exhibit increased NF‐κB activity due to the reduction in IκB levels and the p65 nuclear translocation. VDR expression and distribution were specifically regulated by the bacteria‐induced inflammation. We found that pathogenic bacteria increased VDR expression, whereas commensal bacteria did not change VDR expression in colon epithelial cells. In germ‐free mice, VDR was localized at the top of the crypts of the colon. Bacterial colonization in conventionalized mice induced VDR relocation, and VDR staining was found in the middle and even at the bottom of the crypts. Our data indicate that VDR is involved in suppression of bacteria‐induced intestinal inflammation.
Protection of the intestinal epithelium from the direct effects of a shiga‐like enterotoxin is conferred by anti‐toxin serum IgG
Authors: Mantis Nicholas John , McGuinness Carolyn R.
Published: June 2021

Shiga‐like enterotoxins are extremely potent ribosome inactivating proteins (RIPs) associated with emerging foodborne E.coli infections and, in the case of ricin, are considered potential bioterrorism agents. A hallmark of this family of toxins is their ability to act directly on mature absorptive enterocytes, inducing epithelial cell sloughing and triggering pro‐inflammatory signaling cascades. We have developed a mouse model of intestinal ricin intoxication to better understand the effects of shiga‐like toxins on epithelial physiology and barrier function in vivo, and to determine the role of secretory IgA and serum IgG in protecting the epithelium from toxin‐induced damage. Preliminary studies using available chemokine and chemokine receptor knock‐out mice suggest ricin‐induced epithelial damage is the result of both direct cytotoxic effects and bystander inflammatory responses, possibly emanating from lamina propria leukocytes. We also report that anti‐toxin secretory IgA and serum IgG antibodies, elicited by vaccination or administered passively, are sufficient to protect the intestinal epithelium from ricin intoxication. We are currently investigating, both in vitro and in vivo, the mechanistic interplay between inflammation, epithelial barrier function and serum IgG‐mediated intestinal immunity. This work is supported by the NIH and the Northeast Biodefense Center.
Butyrate influences epithelial signaling via generation of reactive oxygen species
Authors: Kumar Amrita , Collier‐Hyams Lauren , Kwon Young Man , Hansen Jason M. , Neish Andrew S
Published: June 2021

Short chain fatty acids (e.g. butyrate) arise from bacterial fermentation of complex carbohydrates in the colon and are known to modulate innate immunity. We previously reported that commensal bacteria cocultured with intestinal epithelial cells stimulated generation of reactive oxygen species (ROS), with consequent suppression of ubiquitin‐mediated degradation of important signaling intermediates, including β‐catenin and the NF‐κB inhibitor IκB‐α. Ubiquitination of β‐catenin and IκB‐α is catalyzed by the SCFβ‐TrCP ubiquitin ligase, which itself requires regulated modification of the cullin‐1 subunit by the ubiquitin‐like protein NEDD8. Herein, we show that butyrate at physiological concentrations induced ROS in cultured intestinal epithelial cells within 30 minutes, transiently altering the intracellular redox balance. Butyrate induced ROS caused oxidative inactivation of Ubc12, the NEDD8‐conjugating enzyme, and resulted in loss of cullin‐1 neddylation and inactivation of the SCF‐βTrCPubiquitin ligase. Consistently, butyrate blocked TNF‐α induced nuclear translocation of the p65 subunit of NF‐κB and mediated the stabilization of β‐catenin. These data demonstrate how a natural product of the intestinal normal flora profoundly affects epithelial inflammatory and proliferative signaling pathways. This research was funded by NIH grant AI064462.