bluuebunny/crossref_metadata_2025 huggingface dataset:
Dietary patterns are associated with overweight and obesity in Mexican adults
Authors: Flores Mario , Macias Nayeli , Rivera Marta , Lozada Ana
Published: June 2021

The objective was to identify dietary patterns and to evaluate their association with overweight and obesity in Mexican adults.

Dietary information was obtained by a single, 101‐food item FFQ in 17,893 adults (20+ y) from the 2006 Mexican National Health & Nutrition Survey. Dietary patterns were defined by k‐means cluster analysis based on the % contribution to total energy intake of 30 food groups. Overweight and obesity were defined according to WHO criteria. Three dietary patterns were identified: Rural ("R", 40%), Urban‐poor ("U", 39%), and Western ("W" 21%). The "R" pattern was characterized by low dietary diversity, with maize and maize‐based foods accounting for ~50% of energy intake. This pattern had the lowest contributions of most food groups, with the exception of beans (~4.0 %). The "U" pattern had the highest contribution of alcohol (9.4%), sodas (9.3%), white bread (7.7%), fast food, sweets and snacks. The "W" pattern had the lowest contribution of maize (15.5%), and the highest contribution of dairy (12.6%), rice & pasta, meat, poultry, eggs, fats & oils, fruits, and vegetables. After adjusting for age, gender and SES, logistic regression analysis showed that "W" and "U" diets were associated with 30% and 25% increased risk of overweight/obesity, compared to the "R" pattern, respectively (p <0.001). Dietary patterns were associated with overweight and obesity in Mexican adults.

Supported by SSA, Mexico.
Fructose consumption during pregnancy and lactation induced elevated liver triglyceride content and glucose intolerance in rats
Authors: Zou Mi , Arentson Emily , Teegarden Dorothy , Donkin Shawn
Published: June 2021

Gestational diabetes and fatty liver are health risks for both mother and offspring. Dietary fructose is linked to hepatic steatosis and insulin resistance. We investigated the effects of fructose intake on hepatic steatosis and insulin resistance in pregnant and lactating rats and compared responses to a low protein diet, an established maternal dietary insult that predisposes offspring to risks for metabolic syndrome. Sprague‐Dawley rats (n=25) were fed either a control diet (C), a 63% fructose diet (F) or an 8% protein diet (LP) during pregnancy and lactation. Glucose tolerance test at day 17 of pregnancy revealed greater (P < 0.1) blood glucose at 10 (75.6 vs 64.0±2.5 mg/dl) and 20 (72.4 vs 58.6±1.7 mg/dl) min after glucose dose and greater area under the curve (4302.3 vs 3763.4±117.0 mg·dl−1·min) for F fed rats compared to C fed rats. The dams were sacrificed at 21 days postpartum. The F fed rats had enlarged (P < 0.05) livers (11.4, vs 4.8±0.6 % body weight). Both F and LP fed rats had elevated (P < 0.05) liver triglyceride (199.6, 130.0 vs 19.9±2.9 mg/g liver weight) compared to C fed rats. Fructose induced fatty liver and glucose intolerance in pregnant and lactating rats. Low protein induced fatty liver intermediate to F and C diets. The data demonstrate gestational diabetes coupled with hepatic steatosis in fructose fed dams, which is more severe than a fetal programming diet of low protein. Supported by NIH DK077581.
Proteomic, metabolomic and lipidomic approaches to unravel the roles of polyunsaturated fatty acid nutrition in the developing liver
Authors: Novak Elizabeth M , Keller Bernd O , Dyer Roger A , Innis Sheila M
Published: June 2021

Polyunsaturated fatty acids (PUFA) regulate metabolism in adult liver; PUFA accumulation in fetal and infant liver depends on diet. We used combined proteomics, metabolomics and lipidomics to address whether PUFA nutrition impacts metabolic development in neonatal rat liver. Rats were fed as % energy, 3.9% 18:2n‐6 and 1.5% 18:3n‐3 (high PUFA), or 1 % 18:2n‐6 and <0.1% 18:3n‐3 (low PUFA) in gestation and lactation. On day 3 postnatal, lipidomics using HPLC and GLC showed lower n‐3/n‐6 PUFA, but no difference in lipid classes in low compared to high PUFA offspring. Liver proteins were resolved on 2D gels, resolving over 800 proteins. PDQuest analysis showed 24 proteins up‐regulated and 1 down‐regulated over 3 fold in the high compared to low PUFA group, and these were identified by MALDI‐TOF MS. Up‐regulated proteins included F‐1,6‐biphosphatase 1, G‐3‐P dehydrogenase, galactokinase 1, catalase and 60 kDa heat shock protein, with argininosuccinate synthase down‐regulated. GC‐MS profiling of small molecules in liver extracts, with principal component analysis to address changes in flux through metabolic pathways showed higher gluconeogenic amino acids in the high PUFA group. Finally, integration of results from the proteomic and metabolomic analyses into metabolic pathways shows that maternal lipid nutrition impacts pathways of gluconeogenesis and oxidative stress in developing liver. Funded by CIHR

Grant Funding Source

Michael Smith Fdn Health Research Studentship
Early methoxychlor exposure reduces growth and bone mass in rats
Authors: Shupper Peter , Patel Sarav , Ambia‐Sobhan Hasina , Armenti AnnMarie , Uzumcu Mehmet , Shapses Sue A
Published: June 2021

Estrogens have a well known physiological effect on bone. Chemicals in the environment with estrogenic activity may thus influence bone during development. Methoxychlor (MXC) is a chlorinated hydrocarbon pesticide with estrogenic properties and is commonly used in the United States. This study examines early MXC exposure in 25 male Fischer 344 rats (6‐7/group) randomized to control [DMSO (CTL)], MXC [0.02 mg/kg/day, low environmentally relevant dose (M‐low)], MXC [100 mg/kg/day, high dose (M‐hi)], or E2B [positive control; 1 mg/kg/day of β‐estradiol‐3‐benzoate]. Treatments were administered in utero (days 19‐22) and postnatally (days 0‐7). Body composition, and the femur and tibia bone mineral density (BMD) and content were examined at postnatal day 84 using dual energy x‐ray absorptiometry and biomechanical properties are currently being tested. Body weight was 92‐94% (p ≤ 0.05) and 80% (p < 0.001) of CTL in the MXC and E2B groups, respectively. Only the M‐low group showed a reduced % body fat (p < 0.01) compared to CTL, whereas lean body mass was lower (p ≤ 0.01) only in the M‐hi and E2B groups compared to CTL. Whole body BMD was lower (p < 0.05) in M‐hi and E2B versus CTL, but not after correcting for body weight. Low and high dose MXC reduced femur BMD (p < 0.01). These data show that early exposure to MXC will reduce growth and bone mineral density, and may be associated with an increase in fracture risk.
Toll‐like receptor‐4 (TLR‐4) deficiency did not offer protection against trans fat or high fat diet‐induced hyperinsulinemia and hyperglycemia in mice
Authors: Ganji Vijay , Vijay‐Kumar Matam , Aitken Jesse , Gewirtz Andrew T
Published: June 2021

Low‐grade chronic inflammation is linked to insulin resistance, type‐2 diabetes, obesity, and atherosclerosis. TLR‐4 signaling plays an important role in inflammation. It has been reported that TLR‐4 deficiency offers protection against obesity and inflammatory response induced by diets high in fat. We investigated whether natural TLR‐4 mutation (BL‐10 background) protects against trans fat (TF) and high fat (HF) induced insulin resistance, hyperglycemia, and hyperleptinemia. TLR‐4 deficient and wild‐type (WT) mice were fed either low fat control (LF; 4.5%), TF (35%), or HF (35%) diets ad libitum for 12 weeks. Both groups of mice had significantly higher serum insulin, glucose, cholesterol, and leptin in response to TF and HF diets compared to LF control (P<0.05). Mice fed TF diet had hepatomegaly regardless of TLR‐4 genotype. There was no significant difference in serum insulin, glucose, and triacylglycerol between TLR‐4 deficient and WT mice fed LF, TF, or HF diets. There was a tendency toward higher serum cholesterol in TLR‐4 deficient mice compared to WT mice, in all three diet groups. Neither TLR‐4 genotype nor diet type had effect on serum adiponectin. These data suggest that lack of TLR‐4 signaling does not confer protection against TF or HF induced hyperinsulinemia and hyperglycemia in BL‐10 mice. Supported by Intramural Grant from the College of Health and Human Sciences, Georgia State University.
The Automated Self‐Administered 24‐hour Dietary Recall (ASA24): publicly available from the National Cancer Institute (NCI)
Authors: Subar Amy F , Potischman Nancy , Thompson Frances E , Tapia Ramsey , Zimmerman Thea Palmer , Hull Stephen , Mittl Beth , Buday Richard , Crafts Jennifer , McNutt Suzanne , Guenther Patricia , Willis Gordon , Islam Noemi , Bosire Claire , Morrissette Meredith , Schatzkin Arthur , Baranowski Tom
Published: June 2021

We will demonstrate the ASA24, an automated, web‐based, self‐administered 24 hour dietary recall instrument. The application uses state‐of‐the‐art computer technology, including a tutorial, graphic enhancements, and animated characters with audio‐language cues to guide participants. Respondents select foods consumed from a list based on the US Department of Agriculture's (USDA) survey database. Respondents find foods by browsing through food groups or typing and searching. Detailed questions about reported food, adapted from USDA's Automated Multiple Pass Method (AMPM) used in national dietary surveillance, collect information about preparation methods, additions to food, and portion size. Up to 8 sequentially‐sized digital pictures aid respondents in reporting portions. Respondents are given multiple opportunities to edit their food lists. A validation study to compare ASA24 to an interviewer‐administered recall using the AMPM is planned. ASA24, publicly available, can be accessed by researchers and sent to participants over the Internet or administered in a clinic/office. ASA24 makes feasible the inexpensive administration of multiple days of recalls in large‐scale studies or clinical research, enhancing researchers' ability to assess usual dietary intakes. Support: NCI.
Interaction design of a mobile phone food record for adolescents
Authors: Six Bethany L. , Mariappan Anand , Schap TusaRebecca E. , Kerr Deborah A. , Delp Edward J. , Ebert David S. , Boushey Carol J.
Published: June 2021

Mobile telephones with an integrated camera can provide a unique mechanism for collecting dietary information that reduces burden on record keepers. The purpose of this study was to receive feedback from users during the interaction design of a mobile phone food record (mpFR) for adolescents that will eventually translate to an accurate account of daily food and nutrient intakes. A total of 78 adolescents (26 males, 51 females) ages 11‐18 y were recruited to participate in breakfast and lunch meal sessions and provide feedback about the mpFR. Prior to their first meal, all participants received instruction for using the mpFR. Volunteers captured an image of their meals (or snacks) before and after eating using the mpFR. After using the mpFR, the majority of participants (79%) agreed that the software was easy to use. Also, the students participated in a training activity where they received additional instruction on capturing images in various snacking situations. As a result of the additional training, significantly more participants agreed that taking images before snacking would be easy (10% vs 28%, p=0.01), as well as, after snacking (19% vs 38%, p<0.001). Though adolescents readily adopt new technologies, formative evaluation is needed to tailor technology. Further, these results suggest that additional training in using a new technology may improve the accuracy among users.

Grant Funding Source

NIH‐NIDDK
Higher Leucine Content in an Essential Amino Acid Solution Enhances Human Skeletal Muscle mTOR Signaling
Authors: Glynn Erin Leigh , Fry Christopher S , Timmerman Kyle L , Dhanani Shaheen , Drummond Micah J , Volpi Elena , Rasmussen Blake B
Published: June 2021

Essential amino acid (EAA) ingestion stimulates mammalian target of rapamycin (mTOR) signaling in humans. This effect appears to be largely due to the branched‐chain amino acid leucine. We hypothesized that increasing the leucine content of an EAA solution would enhance mTOR signaling to a greater extent as compared to an EAA solution with leucine content typical of high‐quality proteins. Subjects were randomized into two groups, each ingesting 10 grams of EAA but with differing amounts of leucine (18% or 1.8g in EAA group, n=6; 35% or 3.5g in LEU group, n=6). Muscle biopsies were taken at baseline, 1‐, 2‐ and 3‐hr post‐ingestion, and blood was sampled throughout. mTOR signaling was assessed by immunoblotting of biopsies. Phosphorylation of mTOR was increased 1‐hr following nutrient ingestion in both groups, however, the increase tended to be higher in the LEU group. mTOR signaling to downstream effectors S6K1 and 4E‐BP1 was also increased in both groups, though the time course differed between groups. Phosphorylation of eEF2 tended to decrease more at 1‐hr post‐ingestion in the LEU group, and remained dephosporylated 2‐ and 3‐hr post, indicating enhancement of translation elongation. We conclude that increasing leucine content in an EAA solution appears to alter the time course and enhance skeletal muscle mTOR signaling in young human subjects.

Supported by NIH/NIAMS grant RO1 AR049877 & NIH/NIA P30 AG024832
Rapamycin mediated inhibition of mTOR prevents postprandial activation of the unfolded protein response and upregulation of fatty acid synthase
Authors: Pfaffenbach Kyle , Ellis Flannery , Wei Yuren , Wang Dong , Pagliassotti Michael
Published: June 2021

We have previously demonstrated that select components of the Unfolded Protein Response (UPR) are activated in the postprandial state and recent evidence suggests that the UPR is linked to the mTOR pathway and lipogenesis. The purpose of this study was to examine the effects of rapamycin on postprandial regulation of the UPR and lipogenic gene expression in rats. Rats (n=24) were acclimated to a 3hr meal feeding paradigm using a high starch diet over 17 days. On the study day, 1hr prior to the feeding period, 9 rats were injected with rapamycin (RAP) and 10 rats with vehicle (VEH). An additional 5 rats remained fasted (FTD). Liver and blood samples were taken either 1 or 7hrs after the meal feeding period. Several markers of UPR activation (GRP78, GRP94, ATF4, XBP1, and CHOP mRNA) were elevated 1.5‐6 fold in VEH vs. FTD. These same markers were not increased in RAP. Fatty acid synthase (FAS) mRNA was increased ~20‐fold in VEH vs. FTD, but only 10‐fold in RAP vs. FTD. Insulin but not glucose increased GRP78 mRNA and induced XBP1 splicing in H4IIE liver cells. This was prevented in the presence of rapamycin, but not inhibitors of PI3‐kinase or MAPK. These data suggest that the mTOR pathway is involved in the postprandial activation of the UPR and FAS, and that this may be mediated, in part, by insulin.
GCN2 is Essential for the Survival of Mice Born Deficient in Branched‐Chain alpha‐Ketoacid Dehydrogenase Kinase (BDK).
Authors: Bunpo Piyawan , Cundiff Judy K , Stoltz Aaron , Wek Ronald C , Harris Robert A , Anthony Tracy G
Published: June 2021

The branched‐chain alpha‐keto acid dehydrogenase enzyme complex catalyzes the irreversible catabolism of the branched‐chain amino acids (BCAA). Phosphorylation of the complex by its kinase, BDK, inhibits BCAA oxidation. Mice lacking BDK (BDK−/−) demonstrate decreased BCAA levels, impaired somatic and skeletal muscle growth, and neurological abnormalities. Dietary deficiency of BCAA increases phosphorylation of the translation factor, eIF2, by the protein kinase, GCN2, leading to activation of an amino acid stress response that serves to promote cell adaptation. To understand if GCN2 is involved in the development of the BDK−/− phenotype, GCN2−/− and BDK−/− mice were bred to produce GCN2‐BDK double null mice (GB) alongside wild‐type and single null littermates (all on C57BL/6J background). At d1 postpartum, GB pups are indistinguishable from littermates in size, weight, blood glucose and liver glycogen. However, by d7 GB pups stop gaining weight and blood glucose is reduced 44%. Between d7‐d14, GB pups fail to right themselves, demonstrate splaying of the limbs, uncontrollable shaking and weight loss. Despite cross‐fostering at birth and daily provision of glucose and BCAA, GB pups mice become progressively wasted and neurologically unresponsive, expiring by d15. All GB body tissues collected between d11‐d14 are significantly reduced in size, except brain, which is increased 33% as compared to non‐GB littermates. These data demonstrate that GCN2 is critical for surviving BCAA deficiency during early postnatal life.
Overweight with concurrent stunting in school children from Mexico.
Authors: Barquera Simón , Lozada Ana Lilia , Hernández Lucia
Published: June 2021

The objective was to document the prevalence of overweight or obesity concurrent with stunting in Mexican children and to identify demographic and socio‐economic characteristics that identify families in risk. A secondary analysis from the Mexican Health and Nutrition Survey 2006. All calculations were adjusted for the complex survey sampling design using the statistical software STATA (Stata, Corp), an a value p=0.05 was considered significant. School children (n=9537), aged 5‐11 years. Anthropometric measurements and socio‐demographic characteristics of the children were recorded. Height ‐for‐age Z scores were low, with a prevalence of stunting (10%). The children also exhibited a high prevalence of overweight/obesity (17%). Fifty‐one (51%) and thirty‐four (34%) children were both stunted and overweight in south and central area, respectively. After adjusting for age, gender and SES, logistic regression analysis showed that south area was associated with three times risk greater of overweight concurrent stunting, compared to the north area, respectively (p=0.036). Concurrent overweight or obesity concurrent stunting is an important public health issue in low‐income areas of Mexico beginning in childhood. Supported by SSA, Mexico
Grape‐Enriched Diets Alter Cardiac Pathology and Cardiac Glutathione Dynamics in Hypertensive Rats
Authors: Seymour Mitchell , Lewis Sarah K , Bolling Steven F , Bennink Maurice R
Published: June 2021

Increased fruit and vegetable intake can reduce hypertension, but its effect upon hypertensive cardiac pathology is unknown. This relationship was tested in the Dahl‐Salt Sensitive rat fed a high‐salt diet. Provision of 3% (w:w) whole table grape powder for 18 weeks reduced blood pressure, reduced cardiac hypertrophy, improved cardiac function, and increased cardiac glutathione, a key endogenous antioxidant. Provision of vasodilator hydralazine matched the blood pressure reduction of grape but failed to similarly reduce cardiac pathology. In vitro, phytochemicals found in grapes can alter genes related to Phase I/II metabolism, impacting glutathione dynamics. However, cardiac effects with a whole food are unknown. RT‐PCR analysis indicated that grape provision altered genes for oxidoreductases, transferases, sulfotransferases and peroxidases which affect glutathione dynamics, which on balance, would favor elevated glutathione. Key expression changes are confirmed by immunoblot. Importantly, these changes were conserved in grape‐fed, healthy (low‐salt fed) rats, further supporting the nutrigenomic effects of grape intake. In conclusion, physiologically relevant phytochemical intake reduced hypertensive cardiac pathology and altered cardiac transcripts related to glutathione dynamics, which may be vital to its cardioprotective effects. California Table Grape Commission, NIH‐NHLBI.

Grant Funding Source

NIH‐NHLBI, California Table Grape Commission
Inflammation, Collagen Deposition, and Healing in a Goldfish (Carassius auratus) Model of Heart Injury
Authors: Lafontant Pascal J , Golden Benjamin , Grivas Jamie , Cobb Adrienne , Frounfelter Tyler
Published: June 2021

Heart disease remains the number one cause of death in the United States. Mammalian and non‐mammalian models are used to study heart healing and regeneration following injury. In the teleost zebrafish, the removal of ventricular tissue by resection is fully replaced by thirty days with little to no scarring. We hypothesized that the goldfish (Carassius auratus) is able to regenerate its heart in a manner analogous to the zebrafish. We have performed cautery injury of 25% of the ventricle of goldfish that resulted in significant destruction of the ventricular wall and ventricular myocytes. We found an intense inflammatory response in the area of injury, followed the recruitment or proliferation of cells 3 days post injury. We also found a marked increased in collagen deposition at 3 days and 7 days that is resolved by 21 days. This observation is in contrast to the lack of collagen deposition reported in other models. Our data suggests that following cautery injury the goldfish ventricular wall is rebuilt by one week and a quasi complete healing is observed by 21 days. This is the first study showing healing following cautery injury in a fish heart.

Funding

DePauw University
Intravenous MSCs embolize to lung and improve myocardial infarcts in mice by activation to express the anti‐inflammatory protein TSG‐6
Authors: Prockop Darwin J. , Lee Ryanghwa
Published: June 2021

Improved assays for human DNA and mRNA were used to examine the paradox that intravenously infused human multipotent stromal cells (hMSCs) enhance tissue repair with limited engraftment. After 106 hMSCs were intravenously infused into mice, most of the cells were trapped as emboli in the lung. The cells in the lung gradually disappeared with a half‐life of about 24 hr but less than 2,000 cells appeared in other tissues. The hMSCs in lung up‐regulated expression of multiple genes with the largest increase in the anti‐inflammatory protein TSG‐6. After myocardial infarction, intravenous hMSCs or recombinant TSG‐6 decreased activities of serum plasmin and heart MMP‐9. Infusion of MSCs but not MSCs with a knock down of TSG‐6 reduced infarct size. The results suggest improvements seen in other animal models and patients after infusions of MSCs are at least in part explained by activation of MSCs to secrete TSG‐6. Supported in part by NIH grants HL 073755, P40 RR 17447, and P01 HL 075161
Taking a Bite out of the Lab Book: Stereoscopic Laboratory Models in Student's Hands
Authors: Hopkins Robin Marie , Wilson Timothy D
Published: June 2021

Decreasing time for anatomical teaching, coupled with a new generation of techno‐savvy students, has encouraged educators to explore new pedagogical approaches and instructional tools. An evolving tool is a stereoscopic 3D anatomical computer model of the head and neck. The objective of this study was to explore its effectiveness as an instructional and laboratory aid. Third year student volunteers (N=74) were randomly assigned to one of three lab groups. All groups studied the muscles of mastication and completed identical learning objectives during a 45‐minute lab. Each group utilized a different laboratory model: Group I‐gross prosections; Group II‐3D stereoscopic computer model; and Group III‐a hybrid model utilizing both resources. Model efficacy was measured with a pre‐post multiple‐choice quiz. We hypothesized that students using the 3D stereoscopic model or gross prosections would yield similar change scores (post‐test minus pre‐test), while students experiencing the hybrid model would have higher change scores. One week following the initial lab session, participants completed a second post‐test and a qualitative questionnaire to gain insights of knowledge retention and participant preferences respectively. Information gathered may help form new curricula direction.

Grant Funding Source

TSC‐Small Grant on Teaching
Interactive pathways of the central nervous system: a web based didactic multimedia
Authors: Brabec Jiri , Hromadka Rastislav , Nanka Ondrej , Kacvinsky Jan , Hoyt Brian , Rulseh Aaron , Petrovicky Pavel
Published: June 2021

Neuroanatomy is difficult, but successful and innovative use of interactive multimedia presentations can serve as another source for students of medicine to truly understand relationships of structures involved in the pathways. Using Adobe Flash CS3 to create images, videos, and simple animations allows interactive multimedia to be both integrated into lectures, and made available to students on the web. Our website, www.anatonomina.org, illustrates our work in such interactive media. In the photography section, students can look at familiar images, having been also published in a neuroanatomical atlas (Petrovicky P, Brabec J, 2007). By clicking on 'Video', students are presented with a list of main sensory and motor pathways. Choosing one of these, students find a brief overview of the pathway and the option to view sections of the appropriate levels of each pathway. Students can click on the diagram of gross sections and obtain histological as well as schematic views of the section with relevant structures highlighted, and even follow one pathway through MRI. Additionally, clicking on the 'Quiz' link, students are able to assess their knowledge through simple tests. To expect students to truly understand complex neuroanatomical relationships, we feel that the continued development of such interactive media are promising pedagogical techniques.

Supported by grants IGA MZ 2006 NR8931‐4 and FRVS 809 2007

Grant Funding Source

IGA MZ CR NR8931‐4
Neuronal Nogo‐A in the normal and post‐stroke brain
Authors: Cheatwood Joseph L
Published: June 2021

Originally identified as a component of central nervous system (CNS) myelin, we now know that Nogo‐A is expressed by a variety of cell types in the normal CNS, including neurons. The Nogo‐A protein is known to play a key role in limiting neuronal plasticity and functional recovery after stroke, but the precise mechanism of its inhibitory influence remains unknown. Post‐stroke administration of anti‐Nogo‐A antibodies results in functional recovery associated with lasting increases in dendritic complexity and spine density in key cortical areas. Due to the observable effects of anti‐Nogo‐A therapy on neurons, neuronal Nogo‐A represents a potential target for anti‐Nogo‐A therapeutics. Further, results of ongoing studies employing siRNA‐mediated knockdown of neuronal Nogo‐A and Nogo‐A knockout mice are beginning to yield important data on the functions of Nogo‐A in the intact and injured brain.
Calpain 2 functions upstream of MT1‐MMP activation to regulate endothelial cell invasion in 3D collagen matrices
Authors: Kwak Hyeongil , Bayless Kayla J
Published: June 2021

Pro‐angiogenic factors initiate endothelial sprout formation and coordinate intracellular signals to regulate these events. Here, we investigated whether calpain regulates sprout outgrowth in 3D collagen matrices. Global calpain inhibition significantly limited invasion responses. Pharmacological blockade of calpain 2 dose‐dependently reduced invasion, while blocking calpain 1 had no effect. Consistent with these data, calpain 2 knockdown using shRNA reduced invasion, while calpain 1 knockdown had no effect. Maximal invasion responses were stimulated by combining sphingosine‐1‐phosphate (S1P) and angiogenic growth factors (GFs). Quantification of calpain activation revealed control«S1P«GF «S1P+GF. Further, calpain 2 localized to areas of cell‐cell contact and lamellipodia following S1P, but not GF treatment, indicating that while GF treatment increased calpain activity, S1P induced intracellular calpain mobilization. To begin deciphering the mechanism through which calpain 2 mediates invasion, we investigated MT1‐MMP activation and localization. The activation and localization pattern of MT1‐MMP were same as calpain 2. Finally, calpain 2 blockade reduced MT1‐MMP activity, indicating that calpain acts as an intracellular upstream switch for MT1‐MMP.

Grant Funding Source

American Heart Association
Probing RNA sequence specificity and function of PUF proteins
Authors: Hall Traci , Miller Matthew , Wang Yeming , Zhu Deyu , Opperman Laura , Stumpf Craig , Wickens Marvin
Published: June 2021

PUF proteins, named for Drosophila PUMILIO and C. elegans FBF (fem‐3 binding factor), bind specifically to their mRNA targets and decrease expression by promoting mRNA decay or repressing translation. RNA recognition is critical to direct regulatory machinery to the appropriate target mRNAs. We have used a combination of structural and biochemical methods to examine the sequence specificity of PUF proteins from S. cerevisiae and C. elegans. Such studies have allowed us to appreciate how PUF proteins have been adapted to bind specifically to RNA using conserved and protein‐specific features.
RNase H activity and drug resistance to nucleoside and nonnucleoside reverse transcriptase inhibitors
Authors: Pathak Vinay K. , Delviks‐Frankenberry Krista A. , Nikolenko Galina N.
Published: June 2021

Antiretroviral nucleoside and nonnucleoside reverse transcriptase inhibitors (NRTIs and NNRTIs, respectively) initially control HIV‐1 infection, but rapid emergence of drug‐resistant variants limits their effectiveness. Elucidating the mechanisms that lead to drug resistance could facilitate better management of antiviral therapy and development of more effective treatments. Toward this end, we previously proposed that the rate of degradation of HIV‐1 RNA by RNase H (rh) defines the time available for reverse transcriptase (RT) to excise an incorporated NRTI from a terminated primer. This novel concept predicted that a reduced rate of template RNA degradation will increase NRTI resistance. Our recent studies have verified key predictions of this model. First, we identified 8 novel mutations in the connection subdomain (cn) of HIV‐1 RT that increased AZT resistance by as much as 536‐fold when the pol domain contains standard thymidine analog mutations (TAMs). Second, we showed that the cn mutations reduced RNase H cleavages in biochemical assays. Third, we demonstrated that the cn mutations increased AZT monophosphate (AZTMP) excision on an RNA template but had minimal effects on AZTMP excision on a DNA template. These studies indicate that including cn and rh portions of RT in clinical genotypic and phenotypic assays could lead to more accurate determinations of viral drug resistance.
Contrasting Effects of n‐3 EPA vs. Palmitic Acid on Endothelial Lipase and Inflammatory Cytokines in Murine Macrophages In Vitro
Authors: JUNG UN JU , Torrejon Claudia , Chang Chunchun , Hamai Hiroko , Deckelbaum Richard J.
Published: June 2021

Endothelial lipase (EL) links to increased inflammatory responses and atherogenesis. Because of its anti‐inflammatory properties we hypothesized that an n‐3 fatty acid (FA), EPA, in contrast to saturated palmitic acid (PA), would lower lipase and inflammatory markers in cells that produce EL, i.e., murine J774 and peritoneal macrophages (from C57BL/6 mice). Cells were cultured +/‐ lipopolysaccaride (LPS), 1 μg/ml, for 4‐21h after preincubation with increasing concentrations of PA or EPA for 3h. EL, IL‐6, IL‐12 and IL‐10 were assayed by RT‐PCR, and EL by Western analyses. LPS alone increased EL mRNA (>2.5 fold). PA alone as well as with LPS dose dependently increased EL mRNA by >2.5 fold. EPA totally abrogated effects of LPS on increasing EL mRNA. EL protein levels showed similar responses to LPS and to PA vs. EPA. EPA alone or with LPS also decreased pro‐inflammatory IL‐6 and IL‐12 mRNA by 70% and increased anti‐inflammatory IL‐10 mRNA (>2 fold), whereas PA decreased (by 46 %) IL‐10 mRNA (All p<0.05). Thus, n‐3 EPA has strong antiatherogenic compared to proatherogenic effects of PA in cells important in atherogenesis. We hypothesize that suppression of arterial EL and pro‐inflammatory cytokines with activation of anti‐inflammatory cytokines are important pathways underlying the effects of n‐3 FA in reducing cardiovascular events in humans.
Biological variation in the plasma concentrations of polyunsaturated long chain fatty acids (PUFA) in older adults
Authors: McCoy Leslie F , Jorgensen Wanda I , Chew Emily Y , Kim Jonghyeon , Wong Wai T , Schleicher Rosemary L
Published: June 2021

Within‐ and between‐subject biological variation is often used to set analytical quality specifications and to evaluate the significance of changes in serial results. A placebo‐controlled PUFA intervention trial allowed estimation of intra‐ (CVi) and inter‐ (CVg) subject coefficients of variation of plasma concentrations of four n‐3 and six n‐6 PUFA in older adults (> 60 years old) all of whom were being supplemented with lutein and zeaxanthin for macular degeneration. Fasting blood samples were collected from 40 subjects at month 0 and from 20 subjects at months 1, 2, 3, 6, and 9 (placebo group). PUFA concentrations (umol/L) were quantitated using negative ion chemical ionization GC‐MS (Mol Gen Metab 2001;73: 38) with some modification. Analytical imprecision (CVa), calculated using inter‐assay CV for three quality control samples measured in duplicate over 20 assays, ranged from 12‐20%. CVg were calculated from the baseline blood draw (n=40). CVi were calculated using data from baseline and 5 subsequent blood draws (n=20). CVg was largest for eicosapentaenoic acid (72%) and smallest for linoleic acid (22%). CVi was largest for eicosapentaenoic acid (33%) and smallest for arachidonic acid (11%). To our knowledge there is little information available about the biological variation of absolute PUFA concentrations, independent of total fatty acid composition, either in older persons or the general population.
Old obese GPX1 overexpressing mice do not develop overt type 2 diabetes
Authors: Roneker C A , Yan X , Huang L , Vatamaniuk M , Lei X G
Published: June 2021

Previously we reported that the GPX1 overexpressing mice (OE) developed a metabolic syndrome‐like phenotype at 6‐months of age. Subsequent characterization illustrated hyperinsulinemia as the primary effect of GPX1 overproduction. The objective of this study was to determine if the long‐term obesity and insulin resistance led to overt type 2 diabetes at old age. Five OE and wild‐type (WT) mice each were fed a Se‐adequate diet until 1 year of age and used for glucose and insulin analysis. Compared with the WT, the OE mice were 44% heavier (36.0 ± 4.2 vs. 51.9 ± 3.8 g, P < 0.05), had 5‐fold greater fasting plasma insulin concentration (0.21 ± 0.13 vs. 1.45 ± 0.33 ng/ml, P< 0.05), showed 50% less plasma glucose decrease (P < 0.05) at 30, 60, and 240 min after an injection of insulin (1 U/kg), and secreted 4 to 8 times more (P < 0.05) insulin at 15, 30, and 60 min after an injection of glucose (2 g/kg). However, the old OE mice exhibited only mild fasting hyperglycemia (100 ± 15 vs.144 ± 20 mg/dl) and improved glucose clearance, without glucosuria. In conclusion, the old OE mice seemed to be a novel obesity model that might be resistant to the development of overt type 2 diabetes (NIH DK53018).
Regularity of use of supplements over a one‐month period: results from the SURE Study
Authors: Wilkens Lynne R. , Murphy Suzanne P. , Monroe Kristine R. , Steffen Alana D. , Albright Cheryl L. , Yonemori Kim , Morimoto Yukiko
Published: June 2021

The Supplement Reporting (SURE) Study is intended to compare methods of supplement reporting. Participants were adults, aged 58 and over, who reported some supplement use. Three hundred and eighty‐three older adults completed a one‐month diary of supplement use, where they recorded the specific type, including brand and name, and number of supplement doses consumed on each day. A total of 1771 supplements were reported. In general, participants regularly took the supplements that they reported. On average, each supplement was taken on 74% of the diary days, and one‐third of supplements were taken each day. These statistics were similar for multivitamin/mineral combinations, single vitamins, single minerals and herbals. Regular use was more common in individuals consuming fewer supplements: on average, each supplement was taken on 80% of the days in individuals reporting 1‐4 supplements and 67% for those taking 10 or more. We define a run to be a series of consecutive days where a supplement was consumed. Supplements were consumed in an average of 2.3 runs across the one‐month period, with an average run length of 9.5 days There appear to be few intermittently consumed supplements, with only 11.5% being recorded 1‐3 times, thus indicating use of less than once a week. Intermittent use was more common for single minerals and herbals than for multivitamin and single vitamin formulations. Supported by NCI grant R01 CA106744.
Dietary folate intakes in the United States: Data from the National Health and Nutrition Examination Survey (NHANES) 2001‐2004
Authors: Bailey Regan Lucas , Wilger Jaime , Dodd Kevin , Dwyer Johanna , McDowell Margaret , Burt Vicki , Radimer Kathy , Sempos Chris , Picciano Mary Frances
Published: June 2021

Relative contribution of folate from foods and dietary supplements (DS) was tabulated and compared to Dietary Reference Intakes. Diet was assessed via 2, 24‐hour recalls and DS use during the past 30 days including product type, name, frequency of use, and dose was collected via home interview; all data were adjusted for measurement error. Folate from food and folic acid from DS were converted to Dietary Folate Equivalents (DFE) to account for differences in bioavailability. Mean folate intakes from DS were 210 and 232 DFE (2001‐2) and 239 and 237 DFE (2003‐4) for males and females, respectively. DS folate contributed 5% (19‐30 yrs), 10% (31‐50 yrs), and 16% (51+ yrs) to folate intakes for males and 11 % (19‐30 and 31‐50 yrs) and 21% (51+ yrs) to folate intakes of females. DS use lowered the prevalence of "at risk" intakes by up to 8%, and increased the prevalence of intakes above the Upper Limit (UL) by up to 6%. Among females, 11% (19‐30 yrs) and 10% (31‐50 yrs) had total folate intakes below the Estimated Average Requirement (EAR). A high percentage of reproductive age females do not meet the EAR for folate, while other age groups exceed the UL. Without consideration of the form of nutrients from DS, errors may occur when calculating total nutrient intakes. DS use is important to consider when assessing the prevalence of inadequate and excessive folate intakes.
Characterization of Transforming growth factor (TGF‐β) in human milk among US and Mexican mothers
Authors: Rosales Francisco Jose , McMahon Robert J , Ferguson Paul W , Morrow Ardythe L , Jouni Zeina E
Published: June 2021

Human milk TGF‐β plays an important role in maintaining appropriate immune function in infants. We assessed its morphology and ontogeny during lactation in milks from geographically different populations.

Milk samples from three cohorts, each of 20 clinically healthy mothers delivering term babies: donors from the Cincinnati, OH, donated during 2005 ‐07 while donors from Mexico City donated during 2006‐07. The entire content of one breast was collected between 9‐12 am over the first 26 weeks postpartum. TGF‐β1 and β2 were determined using a commercially available ELISA kit (Qunatikine®) validated for its use in human milk, and total protein concentration (PROT) by Commassie Plus.

Most TGF‐β (>90%) was in a latent form and ‐b2 was 5 to10‐fold higher than ‐b1, irrespective of lactation period and sampled region. TGF‐β (‐β1 + ‐β2) concentrations were positively skewed with high variability within and between mothers (figure), which was not explained by clinical status, PROT, lactation period or sampled region. A geometric‐mean and variance were calculated for each of the 13 times and locations, from which a grand geometric‐mean was calculated, 2,700 pg/ml. These results suggest that if an infant consumes 750 ml of milk daily his/her total TGF‐β intake would range from 0.4 to 9.8μg/day.

TGFb concentrations in human milk

* C= Cincinnati 2005‐06; M= Mexico City 2006‐07, & U= Cincinnati 2006‐07.
Effect of protein sources on glucose and insulin response in overweight and obese men
Authors: Courville Amber Beverlie , Stote Kim S , Rumpler William V , Paul David R , Leger Jayme , Novotny Janet A , Baer David J
Published: June 2021

Two randomized, double blinded, crossover designed studies were performed to examine the effect of whey protein on glucoregulation. In the first study, 15 healthy male subjects (age 43‐65, BMI 28‐38) consumed a shake containing whey protein concentrate (WPC‐80), soy protein isolate (SPI) or maltodextrin twice daily with their habitual diet. On the eighth day, fasted subjects underwent an oral glucose tolerance test. In the second study, 9 healthy male subjects (aged 47‐65, BMI 25‐31) were fasted for 12 hours before consuming a hot cereal containing sucrose + whey protein isolate (WPI), sucrose + SPI, lactose + WPI, or lactose + SPI. In both studies, glucose and insulin were measured every 30 minutes for 4 hours. Results of the first study showed no differences in glucose or insulin concentrations among treatments. However, in the second study when WPI was consumed with lactose, the area under the curve for glucose was lower than SPI consumed with lactose (24727 ± 475 vs. 25690 ± 475, p=0.02). Thus, the effect of dairy protein on glucoregulation may be due to the interaction of multiple nutrients.
Genetic Determinants of Responses to Selenium Supplementation
Authors: Combs Gerald F. , Zeng Huawei , Jackson Matthew I. , Johnson Luann K. , Hoeg Antonia , Schomburg Lutz , Davis Cindy D. , Milner John A.
Published: June 2021

In a cohort of healthy adults (106 M, 155 W) in eastern North Dakota, we determined the relationships of five biomarkers of selenium (Se) status (plasma Se, serum selenoprotein P [SePP], plasma glutathione peroxidase [GPX3] activity, buccal cell Se, urine Se) to genotype for four selenoproteins (cytosolic glutathione peroxidase [GPX1], phospholipid hydroperoxide glutathione peroxidase [GPX4], the 15 kD selenoprotein [SeP15], SePP) and two glutathione S‐transferases (GST‐M1, GST‐T1). The cohort had plasma Se of 141.5±23.7 (SD) ng/ml, SePP lof 3.55 (CI 2.61, 4.51) mg/L, GPX3 activities of 3.64±0.54 nmoles NADPH/min/mg protein, buccal cell Se of 10.2±6.5 ng Se/mg protein, and urinary Se of 58.2±21.5 ng Se/mg creatinine. Plasma Se was significantly related to GPX3 genotype (198Leu/Leu: 135.7a±19.0 ng/ml; 198Leu/Pro: 139.2a,b±23.8 ng/ml; 198 Pro/Pro: 145.9b±24.4 ng/ml; P<0.05). Buccal cell Se was significantly related to SeP15 genotype (C/C: 8.77a [5.85, 13.14]; T/C: 7.65b [5.06, 11.57]; T/T: 8.27a,b [6.16, 11.10] P<0.05). SePP was significantly related to SePP genotype at a polymorphism in the 3′‐UTR region (A/A: 3.49a,b [2.91, 4.18]; G/A: 3.24 a [2.44, 4.31]; G/G: 3.62 b [2.75, 4.76]), but not one in the coding region (A234T). No other effects were detected. These results show that genetic polymorphisms can significantly affect some biomarkers used to assess Se status.
Targeted Metabolomic Assessment of Postprandial Plasma Oxylipids
Authors: Zivkovic Angela M , Georgi Katrin , Hegedus Christine M , German J. Bruce , Hammock Bruce D
Published: June 2021

Postprandial lipemia, the prolonged circulation of lipid rich lipoproteins following a meal, is deleterious due to transient endothelial dysfunction and inflammation. Plasma was collected from 3 volunteers before and 1h, 3h, 6h and 8h after a liquid test meal, and 1h, 3h and 6h after a solid test meal. Plasma concentrations of 72 oxylipids were measured by LC/MS/MS. Several lipoxygenase metabolites including 9‐HODE, 9‐oxo‐ODE, 13‐oxo‐ODE, 9‐HOTrE, and 13‐HOTrE increased relative to baseline following the liquid meal, whereas 5‐HETE, 8‐HETE, and 15‐HETE decreased. The ratios of the cytochrome P450‐derived epoxides to their respective soluble epoxide hydrolase‐derived diols 9,10‐EpODE/DHODE, 12,13‐EpODE/DHODE, and 15,16‐EpODE/DHODE were significantly increased at 3h, 6h and 8h relative to baseline, whereas 9,10‐EpODE/ DHODE, 11,12‐EET/DHETre, and 9,10‐EpOME/DHOME were significantly decreased at 1h relative to baseline following the liquid meal. There were several key differences in response to the liquid vs. the solid meal and among subjects, indicating differential metabolism both at baseline and in response to meals that was measurable by targeted metabolomics. At least a part of the inflammation associated with postprandial lipemia is promoted by changes in the synthesis and metabolism of oxylipids in response to meals.
Protein biomarkers for amyotrophic lateral sclerosis (ALS) in the CSF
Authors: Bowser Robert , An Jiyan , Hood Brian , Conrads Thomas
Published: June 2021

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with no effective treatments and unclear pathogenic mechanisms. We have explored protein‐based biomarkers in the cerebrospinal fluid (CSF) of ALS patients, healthy controls, and neurologic disease mimics. CSF was collected using standardized protocols from a total of 250 subjects (80 sporadic ALS, 20 familial ALS, 90 healthy controls, 20 multiple sclerosis, 20 Alzheimer's disease, 10 upper motor neuron disease, and 10 lower motor neuron disease). For purposes of this study we generated 25 sample groups, each containing equal amounts of CSF pooled from 10 individuals within a specific subject type. Each sample group was controlled for age, gender, medication, and site of disease onset (for ALS patients). Liquid chromatography mass spectrometry (LC‐MS/MS) was performed on each sample group to identify proteins by mass peptide fingerprinting. We obtained semi‐quantitative data on 4,400 proteins, representing the most complete proteome analysis of the human CSF performed to date. Specific proteins were identified that exhibit alterations in ALS patients. Unsupervised cluster analysis distinguished and separated the sporadic ALS, familial ALS and various control subject groups. We noted proteins that were altered only in specific ALS age groups, suggesting different pathogenic mechanisms in different age populations. Sporadic and familial ALS sample groups were easily distinguished from pure upper or lower motor neuron disease subject groups. We also detected age‐associated changes in proteins within the healthy control population. These findings identified novel protein biomarkers for ALS and highlight biochemical pathways altered during ALS and potential therapeutic targets.
Linkage to the actin cytoskeleton is critical for α4β1‐mediated leukocyte rolling and chemokine‐induced arrest, but not prolonged adhesion
Authors: Rullo Jacob , Hyduk Sharon J , Digby Genevieve , Lee Leslie , Cybulsky Myron I
Published: June 2021

The avidity or strength of integrin‐mediated cell adhesion is attributed to affinity (strength of individual bonds) and valency (number of bonds formed). We investigated the role of the actin cytoskeleton in α4β1 integrin mediated adhesion to VCAM‐1 in assays that model distinct steps of leukocyte emigration.

In U937 cells stably transfected with the formyl peptide receptor, α4β1 integrin affinity up‐regulation induced by GPCR signaling remained intact following disruption of the actin cytoskeleton by cytochalasin D (cytoD) or latrunculin B (latB) (2 µM, 30 min incubation). Parallel plate flow chamber adhesion assays revealed that U937 cells pretreated with cytoD or latB failed to roll on surfaces coated with VCAM‐1 and arrest on surfaces co‐coated with VCAM‐1 and SDF‐1α. High affinity α4β1 induced by Mn2+ independent of GPCR signaling mediated spontaneous U937 arrest, which was blocked by cytoD or latB pretreatment. In contrast, U937 cell adhesion in 15 min assays was not inhibited by cytoD or latB. Fluorescence recovery after photobleaching demonstrated increased diffusion of α4β1 integrins in cells pretreated with cytoD, consistent with a disruption of integrin linkage to the actin cytoskeleton.

These studies suggest that constitutive linkage of α4β1 integrins to the actin cytoskeleton is critical for mediating α4β1‐dependent rolling and arrest, but not prolonged leukocyte adhesion.

Funded by CIHR.