bluuebunny/crossref_metadata_2025 huggingface dataset:
Porcine coronary collaterals after stimulated myocardial ischemia: count and location
Authors: Spaan Jos , Wijngaard Jeroen van den , Schulten Henny , van Horssen Pepijn , Siebes Maria , Post Mark
Published: June 2021

Introduction

Aim is to quantify number and location of intramural collateral connections following chronic coronary obstruction.

Method

In 7 anesthetized pigs an ameroid constrictor, AC, was placed around the left circumflex artery, LCX, stimulating coronary collateral formation. After six weeks the animals were anesthetized again and the heart harvested, the left main, LM, Right, RCA and LCX distal of the constrictor were cannulated. All arteries were flushed with saline and filled with fluorescent polymer. The hearts were processed in an Imaging Cryomicrotome and images (2000 x 2000 pixels) of the cutting plane taken before each cut of 40 µm thick. The stack of about 4000 images contains information on the coronary vessels down to 20 µm. An algorithm was developed to identify closed arterial pathways induced by collateral connections, CC.

Results

Hearts had 23 (8.3 sdev) subendocardial CC's , 12 midmyocardial and only 4 epicardial. Intramural connections always bridged between normal preexisting vessels but were not restricted to boundaries between boundaries of the LCX territories. 4 control hearts did not show more then 5 CC.

Conclusions

The diffuse presence of coronary collaterals in contrast to collaterals directly connecting the LCX with the LAD or the LCX with the RCA are suggestive of regional activation of coronary collateral growth stimulating pathways.
Negative Cooperativity in the EGF Receptor
Authors: Pike Linda Joy
Published: June 2021

The EGF receptor is thought to exist in the membrane as a monomer that dimerizes following ligand binding. Scatchard analysis of the binding of EGF to its receptor yields concave up plots that have been interpreted as indicating the presence of two classes of binding sites. By simultaneously fitting the binding isotherms from cells expressing increasing levels of EGF receptors, we have shown that 125I‐EGF binding data can be completely explained by a model involving negative cooperativity in an aggregating system. Analysis of the non‐dimerizing Y246D‐EGF receptor mutant showed that the negative cooperativity was dependent on the ability of the extracellular domain of the EGF receptor to dimerize. An EGF receptor lacking the entire intracellular domain did not exhibit negative cooperativity suggesting that cooperativity in ligand binding occurs as a result of interactions within the intracellular domain of the receptor. Mutations that impair EGF receptor kinase activity or that truncate the receptor to remove the sites of autophosphorylation produced a receptor that still exhibited negative cooperativity but also showed positive linkage. The data are consistent with a model in which ligand binding to the naive EGF receptor promotes dimerization of the receptor. Phosphorylation of the receptor as well as the negative cooperativity reduce ligand affinity and promote dissociation of receptor dimers.
Structural aspects of extracellular EGFR signaling
Authors: Ferguson Kathryn M.
Published: June 2021

In terms of dimerization mechanism, it is now clear that the EGFR family is an outlier. Whereas all other well‐studied RTKs are activated by ligand‐mediated dimerization, members of the EGFR family undergo a ligand‐stabilized conformational change that precedes entirely receptor‐mediated dimerization. A "dimerization arm" protrudes from domain II and "hooks" the other molecule in the dimer. In its inactive state the receptor adopts a very different conformation in which the dimerization arm is buried in an intramolecular interaction or tether. A major domain rearrangement is required to transition from this "tethered" state to the extended state that can dimerize. The implications of this mechanism for activation and inactivation of EGFR will be discussed in light of recent developments. It is well established that dysregulation of EGFR, and other members of this family, is linked to many cancers. Several EGFR targeted therapies are in use in the clinic and many more are in advanced stages of clinical trials. Our current understanding of the inhibition of EGFR dimerization and activation by clinically relevant antibodies will be discussed.

This research is supported by a Career Award in the Biomedical Sciences from the Burroughs Wellcome Fund and by NIH grant R01‐CA112552. Kathryn Ferguson is the Dennis and Marsha Dammerman Scholar of the Damon Runyon Cancer Research Foundation.
Are Weekends Necessary in Dietary Assessment?
Authors: Arab Lenore , Jardack Patricia , Liu Weiqing , Schoeller Dale
Published: June 2021

The existing dogma holds that, because of intra individual variability in intakes, dietary intakes on both weekdays and weekends are necessary for valid capture of usual intake. We explored this assumption in the Energetics Study, an NCI funded study designed to test the validity of a web‐based automated 24‐hour recall (DietDay). The study incorporated doubly labeled water as a marker of energy intake. A comparison of the correlations between the gold standard (doubly labeled water) and intakes averaged over 3 weekdays plus 1 weekend (r=0.37) vs. 4 weekdays (r= 0.43) vs. 2 weekends (r= 0.28) showed the strongest correlations to be with 4 weekdays. This general conclusion differed by age, race and gender. Preliminary analyses suggest that the addition of one weekend day to 3 weekdays weakens the correlation with doubly labeled water among women, subjects over 30 years of age and Caucasians and strengthens the correlation among those under 30 years of age and among African Americans when compared with 4 weekdays. This finding suggests that in general, and particularly among women and subjects older than 30, their energy expenditure is most strongly predicted by dietary intakes during the work week. Weekends may add superfluous variation. If confirmed, this finding will simplify efforts at capturing usual energy intakes through repeated 24 hour recalls. Supported by NIH R01CA105048.
Mice lacking Junctional Adhesion Molecule‐A (JAM‐A) develop severe acute lung injury in response to endotoxin
Authors: Koval Michael , Kwon Michael , Torres‐Gonzalez Edilson , Peatman Eric , Nusrat Asma , Parkos Charles A , Rojas Mauricio
Published: June 2021

Acute Lung Injury (ALI) is associated with inflammation and increased alveolar permeability. JAM‐A is localized to tight junctions and expressed by neutrophils, suggesting a role in barrier function and regulating neutrophil migration into the alveolus. To determine whether JAM‐A deletion increases susceptibility to ALI, JAM‐A‐deficient (JAM‐A−/−) and wild‐type (JAM‐A+/+) mice were intraperitoneally injected with 1 mg/kg endotoxin (LPS) to induce ALI. We found that JAM‐A−/− mice displayed elevated levels of alveolar permeability and inflammatory response to LPS‐induced ALI as compared to JAM‐A+/+mice. Both groups displayed the greatest level of inflammation at 24 h, however, lung tissues from JAM‐A−/− mice showed significantly higher vascular congestion and neutrophil infiltrate into the airspace at every time interval examined after LPS treatment when compared to JAM‐A+/+ mice. This suggests that JAM‐A plays an important regulatory role in modulating alveolar barrier function and neutrophil transmigration into the lung. It remains to be determined whether the effect of JAM‐A deficiency is primarily due to an effect on pulmonary endothelium, alveolar epithelium, neutrophils or whether all three cell types are impaired in JAM‐A−/− mice. Supported by: NIH‐HL83120 (MK), NIH‐HL72724 (CAP) and NIH‐ HL84683 (MR).
Inflammatory Cytokine Induced Immunosuppression in Mesenchymal Stem Cells
Authors: Shi Yufang , Zhang Liying , Zhao Xin , Xu Guangwu , Zhang Yingyu , Roberts Arthur I , Zhao Robert C , Ren Guangwen
Published: June 2021

Mesenchymal stem cells (MSCs) are strongly immunosuppressive in vivo and in vitro in both animals and humans. Using our recently generated clones of MSCs derived from mouse bone marrow, we have shown that MSCs potently inhibit the T cell receptor (TCR)‐triggered proliferation and cytokine production by freshly‐isolated T cells. However, the molecular mechanisms controlling the immunosuppressive capacity of MSCs are unknown. We found that the immunosuppressive function of MSCs is elicited by IFNγ and the concomitant presence of any of three other proinflammatory cytokines, TNFα, IL‐1α or IL‐1β. These cytokine combinations provoke the expression of high levels of several chemokines and inducible nitric oxide synthase (iNOS) by MSCs. Chemokines drive T cell migration into proximity with MSCs, where T cell responsiveness is suppressed by nitric oxide (NO). This cytokine‐induced immunosuppression was absent in MSCs derived from iNOS−/− or IFNγR1−/− mice. Blockade of chemokine receptors also abolished the immunosuppression. Wild‐type MSCs, but not IFNγR1−/− or iNOS−/− MSCs, prevented GVHD in mice, an effect reversed by anti‐IFNγ or iNOS inhibitors. Wild‐type MSCs also inhibited delayed‐type hypersensitivity, while iNOS−/− MSCs aggravated it. Therefore, proinflammatory cytokines are required to induce immunosuppression by MSCs through the concerted action of chemokines and NO. We believe that a better understanding of the mechanisms through inflammatory cytokine activated‐MSCs mediate immunosuppression will lead to better clinical protocols for immune modulation.
A role of Wnt/beta‐catenin signaling in the pathogenesis of renal interstitial fibrosis
Authors: He Weichun , Dai Chunsun , Li Yingjian , Zeng Gang , Monga Satdarshan P.S. , Liu Youhua
Published: June 2021

Wnt proteins are a family of signaling regulators that play an essential role in embryonic kidney development; however, their expression in adult kidney is thought to be silenced. In this study, we report a comprehensive analysis of the expression and regulation of Wnts, their frizzled (Fzd) receptors and antagonists in normal and fibrotic kidneys after obstructive injury. The vast majority of 19 different Wnts and 10 Fzd receptor genes were expressed at various levels in mouse adult kidney. Except Wnt5b, Wnt8b and Wnt9b, all members of the Wnt family were up‐regulated with distinct dynamics in fibrotic kidney after unilateral ureteral obstruction. The expression of most Fzd receptors and Wnt antagonists was induced as well. A dramatic accumulation of beta‐catenin in renal tubular epithelial cell cytoplasm and nuclei was observed after obstructive injury, indicating the activation of the canonical pathway of Wnt signaling. Numerous Wnt/beta‐catenin target genes, such as c‐Myc, Twist, LEF1 and fibronectin, were induced, and their expression was closely correlated with renal beta‐catenin abundance. Delivery of Wnt antagonist Dickkopf‐1 gene significantly reduced renal beta‐catenin accumulation and inhibited the expression of Wnt/beta‐catenin target genes. Furthermore, gene therapy with Dickkopf‐1 inhibited myofibroblast activation, suppressed type I collagen and fibronectin expression, and reduced total collagen content in mouse model of obstructive nephropathy. These results suggest a pivotal role of Wnt/beta‐catenin signaling in the pathogenesis of renal fibrosis; therefore, targeting this signal pathway may be a novel strategy for therapeutic intervention of fibrotic kidney disorders.
Intestinal Regeneration after Irradiation: Stem cells and IGF‐I
Authors: Scull Brooks Patrick , Magness Scott , Lee Ginny , Lund P. Kay
Published: June 2021

Abdominal irradiation causes damage and loss of intestinal epithelium that requires proliferation of surviving stem cells for regeneration. Evidence for this role of stem cells is indirect due to lack of stem cell biomarkers. This study tested the hypothesis that SOX9 marks proliferating stem cells after irradiation and that insulin‐like growth factor I (IGF‐I) promotes survival or expansion of SOX9 positive cells.

Wild type or SOX9‐GFP mice were killed 1 through 9 days after 14Gy of abdominal irradiation and intestines collected. Morphology was assessed by H&E. Immunofluorescence for SOX9 and Ki67 and visualization of SOX9‐GFP tested if SOX9 marked proliferating microcolonies or regenerating crypts. Effects of IGF‐I on SOX9 were evaluated by immunoblot.

There was almost complete loss of small intestinal crypts and damage to colon epithelium by days 3‐5 after irradiation. Hyperplastic regenerating crypts were observed on days 5‐9. SOX9‐GFP marked and SOX9 co‐localized with Ki67 in microcolonies and hyperplastic, regenerating intestinal crypts. IGF‐1 treatment increased SOX9 protein levels.

SOX9 provides a new biomarker of putative stem cells in microcolonies and regenerating crypts in the intestine after irradiation. The SOX9‐GFP reporter mouse provides a useful model to evaluate stem cell survival and expansion, and effects of trophic therapies during regeneration after irradiation.

NIH DK40247‐16
Regulation of MicroRNA Biogenesis
Authors: Kim Narry
Published: June 2021

MicroRNAs (miRNAs) play key roles in diverse pathways in eukaryotes, and the dysregulation of miRNA biogenesis is often associated with human diseases. Biogenesis of canonical miRNAs is initiated by RNA polymerase II‐mediated transcription. The primary transcript (pri‐miRNA) gets processed into mature miRNA by two RNase III type enzymes; Drosha and Dicer. Drosha cleaves a pri‐miRNA and releases a small hairpin (called pre‐miRNA). Pre‐miRNA is subsequently cleaved by Dicer into ~22 nt mature miRNA. Mature miRNA is then accepted by the Argonaute protein to constitute the RNA silencing complex.

Regulation of miRNA biogenesis can be achieved at either transcriptional or post‐transcriptional level. Post‐transcriptional control is widespread and can take place at multiple steps including Drosha processing, nuclear export, Dicer processing, RNA editing, uridylation, and decay. We have been investigating the regulatory mechanisms of cancer‐associated miRNAs. We identified several specific RNA binding proteins that post‐transcriptionally regulate miRNA biogenesis. This talk will focus on the roles of the RNA binding proteins in miRNA biogenesis in cancer and stem cells.
Using Anatomy to teach medical students teamwork skills
Authors: Lerner Susan M , Laitman Jeffrey T. , Reidenberg Joy S , Friedman Erica S
Published: June 2021

An aim of our medical curriculum is to teach teamwork skills. We implemented a team building curriculum for first year students in 2007 using anatomy as a vehicle for learning these skills. The course was restructured to involve both team teaching and its assessment. Dissection teams were split into two groups that alternated dissection responsibilities and teaching the cohort not present. To reinforce the team concept, standard practical exams were replaced by "Table Conferences" (faculty/teaching assistant run in‐lab oral quizzes) with all members receiving the same grade. This year they will also do a team graded project based on CT images of their cadavers. Students completed peer and self evaluations of team dynamics and a survey of the team building curriculum. 79% felt that they developed teaching skills that supported team learning. 79% agreed that teamwork enhanced their educational experience in anatomy and that the curriculum provided additional benefits outside of anatomy. We conclude that educating and assessing students in a group environment improves teamwork skills. The anatomy experience encourages students to use a team approach to problem solving that will last throughout medical school and beyond.

Grant Funding Source

N/a
Human IL1B gene induction requires transcription factor‐mediated DNA looping
Authors: Wang Kent Z.Q. , Unlu Sebnem , Adamik Juraj , Su An‐Jey , Auron Philip E.
Published: June 2021

Human IL1B gene transcription is regulated by a core promoter along with a 2.8 kbp upstream induction sequence (UIS) enhancer which generates potent monocyte‐specific LPS‐dependent activity. Several approaches were used to investigate how these two distant sequences cooperate in IL1B gene induction. These included the use of non‐monocytic cells with ectopically expressed putatively relevant transcription factors, as well as monocytes, in order to assay the levels of mRNA expression from the endogenous IL1B gene and the activities of human IL1B promoter‐driven luciferase reporters. Consistent with earlier studies, our data reveal that Spi‐1 is the most important factor, demonstrating a dose‐dependant induction of IL1B. Ectopic expression of other transcription factors such as C/EBPβ, IRF8 and NF‐κB cooperatively enhance the effect of suboptimal Spi‐1, especially in the presence of dominant‐positive TRAF6 acting as a TLR4/IL1R surrogate. The results argue for the importance of constitutive Spi‐1 and activated NF‐κB and C/EBPβ in supporting IL1B induction. Investigation of LPS‐dependent genomic DNA conformation in monocytes reveals an inducible loop between the enhancer and promoter. Furthermore, the DNA loop may be a critical element in IL1B induction and is hypothesized to involve a Spi‐1 interaction at the promoter with C/EBPβ at the UIS.
AKAP220 Links the cAMP Signaling Pathway to Cell Adhesion
Authors: Logue Jeremy S , Scott John D
Published: June 2021

A‐kinase anchoring proteins (AKAP) target the cAMP dependent protein kinase PKA to precise locations inside cells. AKAP220 has been shown previously to interact with the protein kinase GSK3β [1]. Co‐anchoring of PKA and GSK3β permits robust phosphorylation of GSK3β by PKA, inhibiting GSK3β activity. Little is known, however, regarding the biological significance of this complex and its role in inhibiting GSK3β activity. Our recent evidence suggests that AKAP220 links the cAMP signaling pathway to cell adhesion. Mass spectrometry experiments identified a potential interaction between AKAP220 and IQGAP1, an effecter of rac/cdc42 that controls cell adhesion by modulating cadherin attachment to the actin cytoskeleton [2]. We have shown by co‐immunoprecipitation that endogenous AKAP220 is in a complex with both IQGAP1 and E‐cadherin in MCF‐7 cells, and that AKAP220 colocalizes with IQGAP1 and E‐cadherin at cell‐cell junctions. Binding between IQGAP1 and its partners is often regulated by binding to calmodulin through its IQ domain [2]. We find that AKAP220 preferentially binds IQGAP1 lacking the IQ domain. IQGAP1 does not contain a site for PKA phosphorylation; however a site for GSK3 was identified and can be phosphorylated in vitro. Interestingly, the GSK3 phosphorylation site in IQGAP1 is in proximity to its cadherin binding domain. AKAP220 may therefore control cell adhesion by coordinating a novel kinase cascade that links PKA regulation of GSK3β to GSK3β regulation of IQGAP1 and cadherin. Evidence that the IQ domain of IQGAP1 regulates binding to AKAP220 also suggests that this is a calcium regulated complex.
National Report on Biochemical Indicators of Diet and Nutrition in the U.S. Population 1999‐2002
Authors: Pfeiffer Christine M , Schleicher Rosemary L , Osterloh John D , Jain Ram B , Wong Lee‐Yang , Sampson Eric J
Published: June 2021

Using advanced laboratory science and innovative techniques, the Division of Laboratory Sciences (DLS) at CDC has been in the forefront of efforts to assess the nutritional status of the U.S. population through monitoring nutritional indicators. In August 2008, DLS released the National Report on Biochemical Indicators of Diet and Nutrition in the U.S. Population 1999‐2002, the first CDC publication that offers reference information on a wide range of biochemical indicators (27) in the U.S. population in a single document (www.cdc.gov/nutritionreport). These indicators are representatives of water‐soluble and fat‐soluble vitamins, iron‐status indicators, trace elements, and isoflavones and lignans. CDC scientists measured these levels in the blood and urine of people who took part in CDC's National Health and Nutrition Examination Survey (NHANES) during all or part of the four‐year period from 1999 through 2002. The information is presented by age, sex, and race/ethnicity. Overall, the purpose of this report is to improve understanding of the levels of biochemical indicators of diet and nutrition in the general U.S. population and in selected subpopulations. These data will help assess inadequate or excess intake and will inform analyses on the relation between biochemical indicators and health outcomes. They are expected to be a valuable resource for the nutrition community.
Maternal characteristics differentiating first grade children above 75th versus below 50th percentile on BMI
Authors: Hubbs‐Tait Laura , Babb D'Lee , Page Melanie C. , Topham Glade L. , Kennedy Tay S. , Rutledge Julie , Barrett Aimee J. , Shriver Lenka Humenikova , Harrist Amanda W.
Published: June 2021

Study objective was to identify maternal characteristics discriminating children above 75th versus below 50th percentile on BMI ‐for‐age. Participants were 444 rural first‐grade children and mothers. Maternal measures were Child Feeding Questionnaire (CFQ), Parenting Styles and Dimensions Questionnaire (PSDQ), Center for Epidemiologic Studies Depression Scale (CESD). Child BMI was measured in fall and spring: 42.6% (189) children had fall BMI > 75th percentile and 28.4% (126) had BMI < 50th percentile. ANOVAs revealed the following higher means for the fall BMI > 75th percentile group: all CFQ items about mother weight status, all p < .001; PSDQ permissive parenting, F(1, 312) = 4.793, p = .029; CESD F(1, 310) = 3.208, p = .074. BMI > 75th percentile group had lower mean CFQ responsible for feeding/eating scores, F(1, 312) = 8.577, p = .004. Regressions showed CFQ responsible explained significant variance in child BMI percentile (R2 = .02, p = .005) after maternal current and childhood weight status were controlled (R2 = .09, p < .000001). Children in fall BMI > 75th group with more permissive mothers had a closer link between mother childhood weight status and child BMI spring percentile (r = .303, p = .007) than children with less permissive mothers (r = .188, p = .070). Results extend previous findings on early childhood BMI > 75th percentile status as linked to greater risk for later childhood obesity.
Effects of Fluoxetine on the Acquisition of Methamphetamine Self‐Administration in Rats
Authors: Lambert Chase Steven , Goeders Nicholas , Guerin Glenn
Published: June 2021

Fluoxetine is a SSRI commonly used to treat depression. Like other SSRIs, it blocks the reuptake of serotonin, a neurotransmitter involved in mood regulation, at the neuronal synapse. Methamphetamine users experience a "high" due in part to the release of excess serotonin in the brain. Therefore, it can be assumed that some of the same neurological pathways that cause the excess release of serotonin due to fluoxetine are also involved in dopamine release due to methamphetamine. So, what would happen if someone that had taken or was currently taking fluoxetine tried methamphetamine? Would they be more or less sensitive to the drug and its effects? More importantly, would they need the same does of methamphetamine to achieve the same "high?" Would it take a weaker dose, meaning that they were more sensitive to the drug's effects? Or would they require a stronger dose, meaning either that they were developing a tolerance to the drug, or that its effects were less reinforcing? Are people suffering from depression and other psychological disorders at a greater risk for methamphetamine abuse? In order to further examine this possible risk, groups of Wistar rats were used and subjected to varying pre‐treatments and continued treatments of vehicle, fluoxetine, and methamphetamine throughout the course of the experiment.
Mechanical and Mesenchymal Mechanisms of Secondary Chondrogenesis
Authors: Solem Richard Christian , Eames Brian Frank , Tokita Masayoshi , Schneider Richard A
Published: June 2021

Strategies for treatment of skeletal malformation and injury benefit from an understanding of the molecular and biomechanical mechanisms of secondary cartilage development. We conduct a series of experiments using two species of quail and duck, which exhibit considerably different musculoskeletal morphologies. In duck, mandibular elevator muscles attach to a pronounced secondary cartilage on the jaw. An equivalent cartilage is absent in quail. We hypothesize that species‐specific differences in jaw morphology related to the mode of feeding, and a concomitant dissimilarity in the local mechanical environment promote formation of secondary cartilage in duck versus quail. We test our hypothesis by employing two experimental approaches. First, we alter the mechanical environment in embryonic duck by paralyzing skeletal muscles, or by inhibiting stretch‐activated Ca+2 channels. Second, we re‐pattern the duck jaw complex to resemble that found in quail by transplanting neural crest mesenchyme destined to form skeletal and connective tissues of the jaw. This mesenchyme is known to generate species‐specific pattern. Both experimental approaches inhibit secondary cartilage development, as evidenced by genetic and immunohistochemical data. We conclude that mesenchyme‐dependent changes in gene expression, musculoskeletal pattern, and mechanical forces control secondary cartilage induction.

Grant Funding Source

None
Targeted DNA damage in specification and maintenance of CENP‐A at centromeres: is it required?
Authors: Zeitlin Sam G. , Baker Norman M , Chapados Brian R , Soutoglou Evi , Wang Jean Y , Berns Michael W , Cleveland Don W
Published: June 2021

The essential, histone H3 variant CENP‐A is required for epigenetic specification of centromere identity through a loading mechanism independent of DNA sequence. We used short‐pulse green laser light and the enzyme I‐SceI in live human cells to demonstrate that CENP‐A is recruited to DNA 1 minute after damage, including double‐strand breaks.The centromere‐targeting domain of CENP‐A is both necessary for CENP‐A recruitment to sites of DNA damage, and sufficient to drive H3 recruitment to sites of DNA damage. Furthermore, three proteins associated with CENP‐A at centromeres (CENP‐N, CENP‐T, and CENP‐U) are also recruited to sites of DNA damage. CENP‐A accumulation at sites of DNA damage is enhanced by non‐homologous end‐joining (NHEJ), but NHEJ is not required for CENP‐A targeting. In human and Xenopus cells, nuclear uracil DNA N‐glycosylase (UNG2) localizes to centromeres, and UNG2 activity is required for CENP‐A targeting. Taken together, these observations suggest that DNA damage is sufficient to recruit site‐specific CENP‐A targeting in each cell division, independent of DNA sequence.
Dietary resistant starch increases Bifidobacterium spp./ Lactobacillus spp. and Clostridia spp. in the gut of mice fed low and moderate fat diets
Authors: Senevirathne Reshani N , Janes Marlene , Keenan Michael J , Martin Roy J , Raggio Anne M , McCutcheon Kathleen L , Tulley Richard T , Zhou Jun
Published: June 2021

Our previous work showed that resistant starch (RS) failed to reduce obesity in rodents fed a high fat diet. The high fat diet (41% dietary energy) prevented fermentation of RS and reduced bacterial populations in the gut compared to a low fat diet (18% dietary energy). The current 10‐wk study used thirty‐six C57Bl/6J mice to determine the effects of RS with low vs. moderate fat diets (18% vs. 28% dietary energy) on growth of bacterial populations. We measured three types of bacteria involved in RS fermentation and butyrate production. The study was analyzed as a 2x2 factorial with control versus RS diets and moderate versus low fat diets (significance at p<0.05). Ceca with contents were aseptically collected with an anaerobic gas pack. After serial dilutions, populations of Bifidobacterium spp./Lactobacillus spp. and Clostridia spp. in cecal contents were determined using appropriate media to enumerate Colony Forming Units (CFU). Moderate dietary fat did not diminish the growth of bacterial populations. The presence of RS in the diet promotes increased growth of bacterial populations in low and moderate fat diets, but not in high fat diets. This is important in formulating diets that support healthy gut microflora and reduced obesity. Supported by the NIH and the LSU AgCenter.
Isotopic measurement of muscle triglyceride synthesis and breakdown rates
Authors: Zhang Xiao‐jun , Chinkes David L , Aarsland Asle , Herndon David N , Wolfe Robert R
Published: June 2021

Whereas intracellular lipid disregulation is linked to insulin resistance, quantitative measurement of lipid metabolism in the muscle has not been adequately described. We developed a stable isotopic method to quantify muscle triglyceride (TG) kinetics. L‐[U‐13C16]palmitate was injected as a bolus in 4 anesthetized rabbits; arterial blood and muscle samples were taken before the injection and at 2, 5 and 10 min after the injection. Muscle TG and phospholipid (PL) fractional synthesis rate (FSR) was measured using the tracer incorporation method and its fractional breakdown rate (FBR) was measured using the tracee release method. The absolute synthesis or breakdown rate was calculated by FSR (or FBR) x pool size. Intramyocellular palmitoyl‐coenzyme A was used as the precursor for lipid synthesis. In the postabsorptive state muscle TG FSR was 0.10 ± 0.02%/h (6.81 ± 1.50 nmol/g.h), FBR was 0.30 ± 0.07 %/h (20.07 ± 4.68 nmol/g.h). Thus, there was a net TG loss of 0.20 ± 0.05 %/h (13.20 ± 3.32 nmol/g.h). Muscle PL FSR was 0.91 ± 0.06 %/h (20.42 ± 1.35 nmol/g.h), greater (p<0.01) than the corresponding values of muscle TG synthesis. This method measures both the synthesis and breakdown rates of muscle TG as well as muscle lipids pool sizes, and could be a useful tool in studying the metabolic mechanism of peripheral insulin resistance. Supported by Shriners grants 8560, 8600 and 8490.
Improvement of skin health status when collagen‐rich cattle knee bone soup was consumed by young adults
Authors: Lee Bog‐Hieu , Cho Kyung‐Dong , Kim Mi‐Young , Yang Seung‐Hye , Jeon Ki‐Hong , Han Chan‐Kyu
Published: June 2021

The study was conducted to evaluate the effects of collagen‐rich cattle knee bone soup on the skin health status of 14 young adults (mean age: 23.7 yrs. old). Cattle knee bone soup was fed twice weekly (300 g/ meal) as part of lunch for 6 weeks. The study included anthropometry, body composition, skin status, dietary intake and blood analyses for before‐and‐after the cattle knee bone soup consumption trial. After the trial, body weight, BMI, WHR, body fat mass and percent body fat tended to decrease, while total body water to increase. Spot, evenness and wrinkles of the face tended to attenuate, but moisture, elasticity and sebum T‐zone and U‐zone levels to increase (p<0.05). The intakes of vitamin C, vitamin B6, niacin, phosphorus, iron and energy were increased and especially protein and zn intakes were well above the KDRIs after the trial (p<0.05). The serum triglyceride and MDA levels tended to decrease. From the findings, intake of cattle knee bone soup may be beneficial for maintaining good skin health status. However, further studies would be required to identify the effects of collagen rich food intake on skin health status.
Community‐based multidisciplinary weight management program in Mexico: Determinants of success
Authors: Kaufer‐Horwitz Martha , Vázquez Verónica , Reynoso Ricardo , Zamora Margarita , Soriano Ruth , Manjarrez Iliana , González Lizbeth , Rocha Ingrid , Soto Valeria , Eduardo García‐García
Published: June 2021

Background

Strategies to improve success rates in weight managements programs are needed.

Objective

To identify the determinants of success (weight loss =5%) of patients attending a 6 month (7 appointments) multidisciplinary weight management program at the National Institute of Medical Sciences and Nutrition in Mexico City.

Subjects and methods

1216 obese patients who voluntarily entered the program were evaluated. Assessment at baseline included general and socioeconomic (age, gender, schooling, civil status, appointments, previous treatment), anthropometrical (BMI, waist circumference, previous weight loss), clinical (blood pressure), metabolic (glucose, lipid profile) psychological (emotional eating, conscience of disease, anxiety and depression scores, family support, motivation stage, and diagnosis of food disorders) measurements. A stepwise logistic regression model for success was developed, with socioeconomic, anthropometrical, metabolic, clinical and psychological characteristics as independent variables.

Results

Patients were 38.8±11.7 years old with a BMI of 43.8±8.4; 74% were female; 62.9% of participants had BMI=40. Success rate was 43%. Determinants of success were socioeconomic class (OR=1.51, 95%CI 1.11‐2.07), number of visits (OR=2.09, 95%CI 1.44‐3.04), previous treatment (OR=0.49, 95%CI 0.25‐0.97) and anxiety score (OR 0.94, 95%CI 0.88‐0.99).

Conclusions

Strategies to help patients deal with stress should be implemented and efforts to ensure compliance should be made to ensure the expected weight loss.
Protein kinase A and Epac (Exchange Protein Activated by cAMP) are Pro‐and Anti‐Apoptotic Mediators, respectively, in Chronic Lymphocytic Leukemia
Authors: Murray Fiona , Zhang Lingzhi , Zahno Anja , Anastasopulos Alexandra , Rassenti Laura , Kipps Thomas J , Insel Paul A
Published: June 2021

Chronic lymphocytic leukemia (CLL), which can be classified as "indolent" or "aggressive", is generally characterized by the accumulation of B‐cells. Increasing the level of cAMP can promote apoptosis of CLL cells but mechanisms for this pro‐apoptotic response are not known. The actions of cAMP are largely mediated by protein kinase A (PKA) and exchange protein activated by cAMP (Epac). We sought to investigate the role of these two downstream mediators in cAMP‐induced apoptosis using CLL cells and EHEB cells, a CLL‐cell line. Activation of PKA by N6‐ Phenyladenosine‐cAMP (50 μM, 48 hr) and 8‐Chloro‐cAMP (50 μM, 48 hr) both induced apoptosis of CLL cells and decreased the proliferation of EHEB cells. In contrast, Epac activation by 8‐pCPT‐2Me‐cAMP (50 μM, 48 hr), which stimulated Rap‐1 activation, protected the CLL from spontaneous apoptosis and increased EHEB proliferation. Further analysis revealed that CLL cells (isolated from both indolent and aggressive patients) have increased Epac‐1 mRNA and protein expression compared to normal PBMC and normal B‐cells. These data suggest that increased expression of Epac‐1 contributes to anti‐apoptosis in CLL. Efforts to decrease the expression and function of Epac‐1 may thus be beneficial for the treatment of CLL by increasing the pro‐apoptotic effects of drugs that act through raising cAMP and activating PKA. Supported by the Lymphoma and Leukemia Society and NIH.
Hydrogen Sulfide Preconditioning Attenuates Ischemia/Reperfusion‐Induced Mitochondrial Dysfunction in Rat Intestine by a Bkca Channel‐Dependent Mechanism
Authors: Liu Yajun , Kalogeris Theodore , Yusof Mozow , Davis Michael , Korthuis Ronald
Published: June 2021

The objectives of this study were to determine whether treatment with the hydrogen sulfide (H2S) donor NaHS (NaHS‐PC) 24 hrs prior to ischemia/reperfusion (I/R) would prevent postischemic mitochondrial dysfunction in rat intestinal mucosa and, if so, whether calcium‐activated, large conductance potassium (BKca) channels were involved in this protective effect. BKCa expression in intestinal mucosa was detected by immunohistochemistry and western blotting. I/R was induced by 45‐min occlusion of the superior mesenteric artery (SMA) followed by 60‐min reperfusion in rats preconditioned with NaHS (0.8 mg/Kg, ip) 24 hrs earlier. Mitochondrial function was assessed by measuring the mitochondrial membrane potential (JC‐1), mitochondrial dehydrogenase function (MTT), and cytochrome c release. The protective effect of NaHS‐PC on postischemic mitochondrial function was abolished by coincident treatment with a BKca channel inhibitor, paxilline (2.5 mg/Kg, ip). In addition, preconditioning with a BKca channel activator (NS‐1619, 1 mg/Kg, ip) in lieu of NaHS‐PC was also effective in preventing I/R‐induced mitochondria dysfunction. Moreover, the protective effects of NS‐1619 preconditioning were abolished by coincident treatment with paxilline. Our data demonstrate that NaHS‐PC prevents postischemic mitochondrial dysfunction by a BKca channel‐dependent mechanism.
AGING AND LYMPHATIC CONTRACTILITY
Authors: Gashev Anatoliy A. , Gasheva Olga Yu. , Nepiushchikh Zhanna V. , Moore James E. , Muthuchamy Mariappan , Rahbar Elaheh
Published: June 2021

All functions of lymphatic system require the lymph flow, which can not exist without the driving force generated by contractions of lymphatic vessels (LV). However the mechanisms regulating the lymphatic contractility and particularly the mechanisms of age‐related alterations in lymphatic pumping remain greatly under discovered. Using confocal imaging we observed the profound reduction of muscle cells in aged rat LV. In vivo flow measurements using fast video microscopy demonstrated greatly decreased basal lymph flow; maximal lymphocyte velocities in aged mesenteric LV are 4‐6 times lower than in the adult LV, contraction amplitude diminished by 50‐60%; while the contractions are irregular with long periods of inactivity. The profound inhibition of the contractile activity has been observed also in isolated aged rat thoracic duct (TD) and mesenteric LV. The basic flow/eNOS‐dependent regulation is abolished in aged TD, protein message is greatly depleted. At the same time in aged TD the substantial iNOS activation, confirmed by Western blotting and immunohistochemistry, occurs; its functional importance confirmed by pressure/flow tests. We concluded that depletion of the contractile reserves in LV in elderly diminishes their ability to provide the adequate transport of lymph during the periods of the increased volumetric loads. Support: NIH RO1 AG030578, HL070308, HL080526, TAMU CERH 5 P30 ES09106‐08.
The temporal evaluation of proteomic research on angioplasty‐induced neointimal formation in rat carotid arteries
Authors: Wu Chieh‐Hsi , Huang Hsianh‐Tien
Published: June 2021

Angioplasty is the major clinical application for restoring blood circulation of coronary stenosis. However, there have been 40% patients suffered from restenosis within 6 months. We aimed in this study to explore the related proteins involved in restenosis. The neointimal formation of the injured vessels at various time points was evidenced by the histological assay. The protein profiles from the injured vessels at various time points were analyzed by two‐dimensional gel electrophoresis (2DE) and mass spectrometry. Western blot analysis was subjected to confirm the presence of these proteins in the vessels affected by angioplasty. The neointimal formation was evidenced starting from 7 days after angioplasty. Four proteins have been identified by mass spectrometry as PDIA3, GSTp, GLUD1, and PGK1 with protein expression level in a time‐dependent manner after angioplasty. Of which, PDIA3 was most expressed at 7 days after angioplasty while GSTp, GLUD1 and PGK1 were all induced the most at 2 hours after angioplasty. The activity of MMP3 was most potent at 2 hours after angioplasty whereas that of MMP2 was at 7 days after angioplasty. FAD and PI3K were most expressed at 14days after angioplasty.

Our results indicate that these proteins are importantly involved in the process of neointimal formation, suggesting their potential roles as biomarkers and therapeutic targets for restenosis.
Cardiac Diabetic Autonomic Neuropathy is Associated with Insulin Regulated GIRK4 Expression via Activation of Akt and SREBP‐1
Authors: zhang yali , Park Ho‐Jin , Picard Kristen , Galper Jonas B
Published: June 2021

Diabetic Autonomic Neuropathy is characterized by parasympathetic dysfunction of the heart. Although GIRK4 is required for parasympathetic response, no data on the regulation are available. Here we describe insulin up‐regulation of GIRK4, via an Akt/GSK3β/SREBP‐1 dependant pathway. Type 1 diabetic Akita mice demonstrate parasympathetic dysfunction. Expression of GIRK4, SREBP1, and phosphorylation of Akt and GSK3β were decreased in the Akita mouse atrium, and was reversed by insulin. Insulin stimulated Akt phophorylation, increased GIRK4 promoter activity and protein in atrial myocytes, which was inhibited by the PI3 kinase inhibitor, LY294002. Active myristoylated Akt (MyrAkt) induced the expression of SREBP1 and GIRK4. Insulin increased nuclear SREBP level and DN SREBP1 reversed the insulin effect on GIRK4 protein. Reporter assays demonstrated that SREBPs stimulated GIRK4 promoter activity in a dose dependent manner, which was inhibited by LY. Inhibition of GSK3β also stimulated SREBP and GIRK4 protein. Our data suggest that insulin increases GIRK4 expression by activation of Akt, which phosphorylates and inhibits GSK3β, which involved in SREBP degradation. And SREBP up‐regulates GIRK4. So hypoinsulinemia in type 1 diabetes may result in parasympathetic dysfunction due to decreased SREBP and GIRK4 levels. The role of Akt and GSK3β may offer new therapeutic targets for this complication of diabetes.
The Outs and the Ins of the Pleiotropic Lipid Mediator Sphingosine‐1‐Phosphate
Authors: Spiegel Sarah
Published: June 2021

Sphingosine‐1‐phosphate (S1P) is now widely recognized as a bioactive sphingolipid metabolite that plays a key role in regulation of many fundamental cellular processes important for human health and diseases. Sphingosine kinases, which catalyze the phosphorylation of sphingosine to S1P, are central regulators of the cell fate determining sphingolipid rheostat. Activation of sphingosine kinases by a variety of agonists increases intracellular S1P, which in turn can function intracellularly, and importantly, be secreted out of the cell and act extracellularly by binding to and signaling through a family of specific S1P receptors in autocrine and/or paracrine manners. Recent studies suggest that this "inside‐out" signaling by S1P may play a role in cancer, atherosclerosis, inflammation, and autoimmune disorders such as multiple sclerosis. In this lecture, I will summarize progress made in understanding metabolism of S1P, mechanisms of sphingosine kinase activation, potential new intracellular targets of S1P, how S1P is exported out of cells to activate its receptors and their downstream signaling pathways, and examine relationships to multiple disease processes. I will also describe some recent preclinical and clinical trials of therapies targeting S1P signaling. Supported by NIH grants, R37GM043880, R01CA61774, R01AI50094, and 1U19AI077435.
Anti‐inflammatory effects of camelina sativa oil in postmenopausal women
Authors: Campbell Christina Gayer , Picotte Mindi S. , Syndergaard Sarah , Filipowicz Rebecca , Thorland William G.
Published: June 2021

Camelina sativa is a novel plant source of alpha‐linolenic acid (ALA) with a similar fatty acid profile to flaxseed. A double‐blind parallel‐arm pilot study investigated the anti‐inflammatory effects of daily consumption of camelina sativa oil for 6 weeks. Twenty overweight women (mean ± SD; 28.0 ± 2.6 kg/m2) were randomized to one of two groups: 2T/d camelina oil (CO)(8g ALA/d) (n=10) or 2T/d sunflower oil (SO)(0g ALA/d)(n=10). Subjects consumed an ad libitum diet; test foods provided as baked goods. Weight significantly increased over time for both groups (CO: 70.8 ± 4.9 kg vs. 75.3 ± 7.8 kg; SO: 77.2 ± 10.2 kg vs. 79.4 ± 10.7 kg)(P<0.05). C‐reactive protein decreased 19.4% and increased 57% for the CO and SO groups, respectively (P>0.05). Plasma interleukin‐6 concentrations increased 2.8% for the CO group and 17.5% for the SO group (P>0.05). Results from this pilot study suggest that CO, an omega‐3 rich oil may provide anti‐inflammatory responses as compared to SO despite weight gain. Future research is warranted with more controlled feeding interventions that minimize weight gain. Funding was provided by Biobased Institute of Montana (USDA‐CREES 2005‐34510‐15726).
How not to lose your students with concept maps
Authors: Benay Francoise , Semsar Katharine , Kennedy Sarah
Published: June 2021

In the reformation of teaching practices in physiology and other science courses, concept maps have been increasingly introduced as a tool that promotes meaningful learning and integration of ideas. Following a faculty training session on the use of concept maps in the classroom, four faculty members independently began using concept maps in four different courses. Students in one course in particular had substantially higher perceived value in the concept map technique on the end‐of‐term surveys. Following this result, we used additional end‐of‐term survey questions, faculty and teaching assistant interviews, and a review of course materials to assess what factors lead to greater student value in the technique. Based on a review of these materials, we identified three key factors that students indicate are important for their acceptance of the technique. One, the mapping activity needs to be appropriately designed to meet the educational goals, i.e. not excessively complex. Two, there needs to be an adequate amount of teacher or teaching assistant feedback. Three, the students have to see that what they are learning from the process of concept mapping is also being assessed on exams. When implemented properly, students view concept mapping as a valuable tool in their learning of physiology, but this requires careful attention to the above issues.
NMR based structure and enzymatic insight into diacylglycerol kinase, an alpha‐helical membrane protein
Authors: Van Horn Wade D. , Kim Hak‐Jun , Sonnichsen Frank , Ellis Chuck , Karra Murthy , Sulistijo Endah S. , Sanders Charles R.
Published: June 2021

E. coli diacylglycerol kinase (DAGK) is a trimeric 40 kDa a‐helical integral membrane protein comprised of 9 transmembrane (TM) helices, which is important in phosphatidic acid metabolism. In this work, we present the 3D structure and corresponding structure‐function data which detail the active site of the protein. We determined the structure of DAGK, which was folded in detergent micelles using NMR. We were able to generate an ensemble of DAGK structures with an RMSD of less than 1Å. The resulting structure represents a unique fold that has a threefold symmetry axis made of the 2nd TM of each monomer, forming a three helix bundle core. DAGK has an intricate fold with helices from different monomer subunits contributing to each of the three active sites. To map the active site, DAGK was titrated with substrates and followed by NMR. These data are in agreement with biochemical data which were generated by examining the effects of cysteine mutations on protein activity. Our results provide a platform for future studies of the folding and enzymatic mechanism of DAGK and illustrate the utility of NMR to investigate challenging membrane protein targets. Supported by NIH RO1GM47485.
Characterization of inter‐domain electron transfer in Ncb5or, a redox enzyme involved in fatty acid desaturation
Authors: Deng Bin , Parthasarathy Sudharsan , Xu Ming , Wang WenFang , Benson David R , Zhu Hao
Published: June 2021

Microsomal cytochrome b5 (b5) and cytochrome b5 reductase (b5R) are redox proteins that donate electrons to stearoyl‐CoA desaturase (SCD) in fatty acid desaturation. NADH cytochrome b5 oxidoreductase (Ncb5or) contains b5 and b5R domains, and a linkage domain similar to the CS (CHORD‐SGT1) family members. Our recent studies show that Ncb5or functions as an alternative electron donor for SCD. Ncb5or knockout mice display lipoatrophy and diabetes as a result of impaired fatty acid desaturation. In this study, we aim to characterize the properties of individual human Ncb5or domains, the inter‐domain electron transfer and the role of CS domain. Recombinant b5, b5R, and CS+b5R domains were generated from a bacterial system. Native gel electrophoresis revealed that b5 is stable under heat treatment, but b5R and CS+b5R are not. Inter‐domain electron transfer can be reconstituted by mixing b5 with b5R or CS+b5R under excess NADH, indicating a non‐essential role of CS in electron transfer. The percentage of heme reduction increases with decreasing salt concentrations, suggesting an electrostatic interaction. In the same buffer, b5R reduces more heme than CS+b5R. Both b5R and CS+b5R alone can reduce cytochrome c with the same rate. The addition of b5 increases the rate, more with b5R than CS+b5R. We conclude that electrons can flow from both b5 and b5R to cytochrome c but only the b5‐b5R interaction is specific. Supported by NIH.