bluuebunny/crossref_metadata_2025 huggingface dataset:
Oxygen Consumption and Substrate Oxidation Rates Following Passive Acclimation to a Hot Environment
Authors: Etter Clara Vada , Beaudin Andrew E. , Walsh Michael L. , White Matthew D.
Published: April 2020

Introduction

Acclimation to a hot environment via prolonged or repeated, daily exposure to a heat stress results in adaptations of the human thermoregulatory responses. In this study, it was investigated whether metabolic responses to passive heat acclimation (HA) adapted similarly to thermoregulatory heat loss responses including following a 10‐day, 2 h·day−1 passive heat acclimation (HA) to 50°C and 20 % RH. This study was approved by the Simon Fraser University Office of Research Ethics.

Purpose

It was hypothesized that, similar to that seen during active heat acclimation with exercise, there would be an elevation of oxygen consumption following a passive heat acclimation. It was also hypothesized that following heat acclimation there would be increased lipid oxidation relative to the pre‐acclimation state.

Methods

Nine male participants performed two immersions in a 40°C bath separated by a 10‐day passive heat acclimation. Heat acclimation was performed in a climatic chamber controlled at 50°C and 20 % RH where participants remained seated at rest for 2 hr·day−1 with their rectal temperature maintained between 38.5 – 39.0°C. A two‐way repeated measures ANOVA with factors of Acclimation State (Pre and Post) and delta TES (0 to 1.4°C above rest) was employed with post hoc analysis completed with two‐tailed, paired t‐tests. The significance level was p<0.05.

Results

HA was confirmed by a significant decrease in resting esophageal temperature (TES) from 37.60 ± 0.19 (mean ± SD) to 37.31 ± 0.11°C (p=0.001). During the 40°C immersions significant decreases in the TES thresholds for temple cutaneous blood velocity from 37.34 ± 0.12 to 37.14 ± 0.16°C (p=0.008) and for forehead eccrine sweating from 37.61 ± 0.21 to 37.24 ± 0.24°C (p=0.007) also confirmed heat acclimation. HA did not influence oxygen consumption from 0.21 ± 0.07 to 0.22 ± 0.07 L/min (p=0.15). Carbohydrate oxidation rate increased from 0.21 ± 0.21 to 0.28 ± 0.26 g·min−1 (p=0.002). Lipid oxidation rate decreased from 1.16 ± 0.22 to 1.09 ± 0.24 g·min−1 (p<0.001).

Conclusion

Following a passive heat acclimation the evidence reports no change in oxygen consumption, and contrary to that hypothesized, there was a decrease in lipid oxidation rate and an increase in carbohydrate oxidation rate.

Support or Funding Information Funding Source

This project was supported by Natural Sciences and Engineering Research Council of Canada and the Canada Foundation for Innovation.
IL‐4 Stimulated Macrophages Induce a Pro‐Inflammatory Secretome in Fibroblasts
Authors: Julian Dana Rose , Buechel Heather , Kulkarni Mangesh , Ismail Nahed , Brown Bryan , Yates Cecelia C.
Published: April 2020

The precis continuum manifold of cell state of macrophages and fibroblast cells during skin fibrosis, described as a chronic inflammatory and tissue remodeling microenvironment, is not well understood. Both cell populations respond directly to damage and present a coordinated chain of activation states that dictates the reparative outcome. The spectrum change in phenotype during late remodeling is important, because it is by both cells that autocrine and paracrine communication contribute to the deposition of extracellular matrix (ECM) and chronic inflammation. Here, we provide evidence that the anti‐inflammatory M2‐like (IL‐4 induced) macrophage cell state drives activated fibroblasts to secrete important pro‐inflammatory mediators and thus changing the fibroblast cell state to one that promotes a continuous fibrotic milieu. Indirect and direct co‐culture systems were used to elevate the effects paracrine and autocrine feedback of secreted factors and cell‐contact dependent factors. These studies showed the exposure of TGF‐β‐activated fibroblasts to M2‐like macrophages altered the cytokine/chemokine secretion profile of fibroblasts to express IL‐1β, TNFα, and CXCL10 and enhanced its expression of profibrotic matrix production by using transcriptomic analysis. To understand the necessity of fibroblast‐macrophage crosstalk in the fibrotic process a gain of function approach was used by reconstituting the mice with IL‐4 polarized macrophage in an in vivo sterile wound/fibrosis bleomycin model. Tissue fibroblasts increased expression of pro‐inflammatory cytokines included IL‐10, INF‐γ, and IL‐13, as confirmed by transcriptomic analysis. These analyses additionally identified an imbalance in proteolytic remodeling actions of TIMPs and MMPs. These findings identify an important role of macrophage‐specific cell state and phenotype‐directed crosstalk with tissue remodeling activated fibroblast in signaling to orchestrate a chronic inflammatory and fibrotic microenvironment.

Support or Funding Information

This work was supported by NIH grants R01AR68317, R01AG055564, and R01GM121558.
On the Orbit Orientation and Visual Resolution of Mammalian Gliders
Authors: Heesy Christopher Paul
Published: April 2020

In this study, I analyze the comparative visual system of gliders through orbit and skull dimensions and geometry between gliding rodents, marsupials, and colugos with their non‐gliding relatives. Gliders need: (1) the depth, distance, size, and orientation of target landing substrates prior to takeoff; (2) heading and acceleration data on movement in order to adjust while in air; and (3) to re‐position for landing based on visual approach to the landing substrate. Previous work has suggested that gliding mammals have higher orbit convergence to expand their visual fields or that these animals have larger interorbital distances to increase binocular parallax; both suggestions rely on greater depth perception as underlying functional causes. Additionally, I model the limitations imposed by visual resolution, sensitivity, and visual field sizes on distance and heading perceptions in gliders. Morphometric results do not support the suggestion that orbit dimensions and geometry vary with locomotor type; gliders do not have more convergent or widely spaced orbits. Perhaps more importantly, modeling of visual resolution of substrates at the distances commonly traversed by gliders suggests that these animals cannot rely solely on visual data for locomotor judgments.
Behavioral and histopathological examination of antidepressant‐like effect of Vitamin D in rats
Authors: Filali Houda , Kadil Youness , Agoub Mohamed
Published: April 2020

Exposure to repetitive stressful life situations is clearly associated with the onset of depression. Convergent evidence demonstrates that vitamin D has surprisingly diverse functions in the Nervous central system. The aim of the present study is to investigate the behavioral and the histopathological effect of Vitamin D in rats been exposed to the Unpredictable Chronic Mild Stress.

Twenty‐four rats were randomly divided into three groups of eight received respectively Nacl, Vit D (200 UI/kg/day) and Fluoxetine (10mg/kg/day) intraperitoneally during 6 weeks. All animals were exposed to various stressful situations, with a frequency of two stressors per day throughout the 5‐weeks. Forced swim test and histopathological evaluation were used to evaluate the antidepressant potential effect of vitamin D versus fluoxetine.

The administration of Vitamin D daily reduces significantly the immobilization time during the forced swim test versus the control groupe (p=0.003). The histopathological analysis showed that supplementation by vitamin D prevent against apoptosis by a significant reduction of apoptotic neurons. Vitamin D provides a significant protective effect against neuronal loss and cellular disorganization at the hippocampus.

The interpretation of our results supports our theory which suggest the antidepressant‐like effect of Vitamin D. Other investigations in the same field are being conducted to demonstrate the mechanisms of action involved in this approach
Unraveling the Pathology of the Blood‐Tumor Barrier in an Experimental Model of Brain Metastases of Lung Cancer
Authors: Auld Claudine , Reddy Alexandra , Uzunalli Gozde , Dieterly Alexandra , Lyle L. Tiffany
Published: April 2020

Lung cancer is the leading cause of cancer‐related deaths worldwide (1.76 million deaths in 2018) and the leading cause of brain metastatic disease among all primary cancers. As tumor cells colonize the neuroparenchyma, they breach the tightest and most effective vascular barrier in the body, the blood‐brain barrier (BBB). The functional components of the BBB include endothelial cells, basement membranes, pericytes and astrocytes. Herein, we hypothesized that dynamic transformation of the BBB to the blood‐tumor barrier (BTB) would correlate with changes in paracellular permeability. An experimental model of lung cancer brain metastases was developed using twelve 6‐week‐old athymic nude mice. Mice were injected with 250,000 brain‐seeking cells, which colonized the brain for 4–6 weeks. Animals were injected with 3 kd Texas Red dextran at the time of euthanasia, and brains were harvested, cryosectioned and prepared for immunofluorescence analysis. Metastatic tumors were characterized as highly or poorly permeable based on the diffusion of Texas Red dextran within and around the tumor parenchyma. Brain metastases were roughly spherical and measured between 50–500 μm in diameter, irrespective of their permeability status. The most striking BTB pathology was identified in tight junctions and associated adapter proteins. Highly permeable tumors exhibited haphazard expression of claudin‐5 and a loss of zona occludens‐1 adapter protein compared to metastases with low paracellular permeability. Understanding the cellular and molecular alterations occurring within the BBB during brain metastatic disease from non‐small cell lung cancer is imperative for the identification of novel therapeutic targets and improved methods of drug delivery.

Support or Funding Information

Claudine Auld was supported by the Summer Research Opportunities Program, sponsored by the American Society of Investigative Pathology. This research was supported in part by the Showalter Research Trust.
Angiotensin‐Converting Enzyme I/D Polymorphism on Hematological Changes after Marathon Race
Authors: Cury-Boaventura Maria Fernanda , Rogante Julliane Izidoro , Sierra Ana Paula Renno , Oliveira Rodrigo Assunção , Lima Giscard Humberto Oliveira , Benetti Marino Pereira , Kiss Maria Augusta , Pesquero João Bosco
Published: April 2020

The aim of this study was to determine the influence of ACE I/D polymorphisms on hematological changes induced by marathon race. One hundred and eleven Brazilian amateurs’ male endurance runners participated in the study. Blood samples and urine were collected before, immediately after, one, three and fifteen days after the marathon race. Urine (hematuria), hematological parameters (erythrocytes, hemoglobin, Hb, hematocrit, Ht, red blood cell distribution width, RDW, mean corpuscular volume, MCV, mean corpuscular hemoglobin, MCH, and mean corpuscular hemoglobin concentration, MCHC) and ACE I/D genotyping analyses were performed. Immediately after the race we observed hematuria in ID and DD genotypes but not in II genotypes. The marathon induced a decrease in erythrocyte counts and hematocrit; and an increase in MCV one and three days after marathon in ID and DD genotypes, but there was no change in athletes with genotype II. There was also a reduction of hemoglobin 3 days after the marathon in genotypes ID and DD, but there was no significant change in genotype II runners. We observed an increase in RDW and CHCM three and fifteen days after the marathon, respectively, in runners with genotype ID, suggesting compensation for a reduction in the number of erythrocytes. ACE II genotype has a protective role on hematological changes induced by marathon avoiding the risk of athletic anemia and acute renal failure.

Support or Funding Information

This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) [Grant Number 2014/21501‐0].
BRAIN DERIVED NEUROTROPHIC FACTOR CHANGES IN RESPONSE TO SUCROSE INTAKE IN ADOLESCENT RATS
Authors: Simsek Zinar Darius , Sanroman Dolores Vazquez , Curtis Kathleen
Published: April 2020

Sugar intake is a natural stimulus, like sodium or stress, and activates similar neural circuits involved in mediating reward. Sugar intake may lead to a sensitization of the reward system, changing the response to subsequent rewarding experiences. BDNF plays an important role in the survival, maintenance and growth of many types of neurons and is expressed abundantly in the nucleus accumbens (NAcc) and prefrontal cortex (PFC). Rodents maintained on energy‐rich diets containing high levels of sugar show impaired learning and memory performance, and recent evidence indicates that such deficits may be related to the effects of these diets on the regulation of brain‐derived neurotrophic factor (BDNF) in the brain. Nevertheless, it is not clear how sucrose intake translates into decreased BDNF levels in brain reward areas. The specific aim of this study, therefore, addresses the lack of information on the role of sugar intake, as a natural reward, in BDNF expression in the PFC and NAcc.

Hypothesis

Animals pre‐exposed to sucrose intake in early adolescence are more likely to maintain elevated levels of sucrose intake which is related to changes in BDNF levels in PFC and NAcc.

Methods

1) We have determined preferred sucrose concentration in adolescent rats using a chronic intermittent sucrose intake model. 2)We have determined if early sucrose intake affects adult sucrose intake and examined changes in BDNF levels of expression in the PFC and NAcc. On PND28, (pre‐adolescents period) rats were assigned to sucrose (5% or 15%) or tap water groups. From PND28 (adolescent period) to PND32 rats had 2hr access to sucrose or water each day. After 8 weeks, rats were re‐exposed to sucrose or water and after the last sucrose intake session, animals were sacrificed. Brains were extracted and prepared for immunoassay analysis (ELISA) for identifying Pro and Mature BDNF levels.

Results

Adolescent rats showed significantly higher levels of sucrose intake at the 5% concentration than the 15% concentration. Both ProBDNF levels and Mature BDNF levels in the PFC showed statistically significant differences among the groups, whereas there is not any statistically significant change among groups in NAcc.

Conclusion

Our study demonstrated for the first time that sucrose ingestion affects BDNF levels in PFC but not in NAcc in the brain when adolescent rats are pre‐exposed to sucrose intake.

Support or Funding Information

Oklahoma State University Vice President office of research 2019
Effects of food intake on relaxin 3 (Rxn3) mRNA expression in the muscle of Atlantic salmon
Authors: White Taylor , Moyers Macy , Kostner Danica LeAnn , Wertz Blaine , Peterson Brian C. , Kobayashi Yass
Published: April 2020

The Rxn3 is a protein that belongs to the insulin‐like/relaxin superfamily and affect various physiological functions, including growth, food intake, and reproduction. Evidence in rodents suggests that Rxn3 is involved in the regulation of food intake and critical for proper development of muscle and bones. The roles that the insulin‐like/relaxin superfamily plays in regards to growth and metabolism is largely unknown in fish, including Atlantic salmon. Recently, we identified the mRNA sequence for the Atlantic salmon Rxn3 by screening the database using channel catfish insulin‐like 5 polypeptide mRNA sequence. Given the possible role of Rxn3 in muscle function and nutrient metabolism, changes in the expression of Rxn3 mRNA in response to the changes in food intake were examined. Juvenile Atlantic salmon (n=72, 6 fish per tank) were assigned to either receive food every day (control, n=4 tanks) or fasted for 14 days (fasted, n=4 tanks). Additionally, fish were fasted for 7 days and fed daily for the subsequent 7 days (refed, n=4 tanks). Muscle samples were collected from fish at the end of 14 days, and expression of Rx3 mRNA was measured using real‐time PCR. Expression of Rxn3 mRNA was similar among the three feeding treatments, suggesting that muscle expression of Rxn3 is not affected by changes in food intake. Given the preliminary nature of the study, further investigations are warranted to define the role of Rxn3 on the regulation of growth, food intake, and nutrient metabolism in Atlantic salmon.

Support or Funding Information

This project was partially funded by Kansas IDeA Network of Biomedical research Excellence (K‐INBRE) under the NIGMS/NIH grand P20GM103418.
Impact of Natural and Anthropogenic Perturbations on Animal Behavior: A Cellular Approach
Authors: Mattei Gabriela Angelica Rosado , Miller Mark W.
Published: April 2020

The nervous systems of gastropod molluscs contain large neurons that can be identified in all specimens. They are therefore useful for elucidating the cellular basis of behavior and its modification due to extrinsic factors. Our studies examine two tropical marine gastropod species, Bursatella leachii and Dolabrifera dolabrifera, to determine the impact of natural perturbations and anthropogenic contaminants on identified neurons and motor circuits. Here, we report a census for Bursatella leachii during 2019 and compare it to previous years. We find a reduced population that may be recovering during the two years since the major hurricanes of 2017. Specimens were collected to perform immunohistochemistry with the goal of characterizing the localization of specific neurotransmitters in the nervous system. We found that the catecholamine dopamine is likely to be involved in the regulation of feeding and reproductive behaviors. RNA was also extracted from the CNS of Bursatella in order to generate a nervous system transcriptome. These observations provide a platform for future investigation of the impact of natural and man‐made disturbances on the cellular control of behavior in these model organisms.

Support or Funding Information

Funding: NIH (USA): RCMI MD007600, NIGMS‐RISE R25 GM061838 NSF (USA): CREST HRD‐1137725
Higher Weekly Walking Volume Improves Cardiac Autonomic Profile and Arterial Stiffness Index in Hypertensive Elderly
Authors: Neto Octávio Barbosa , Bilharinho Frederico , Resende Elisabete Aparecida Mantovani , Resende Luiz Antônio Pertili , da Silva Marco Antônio Vieira , Sordi Carla Cristina
Published: April 2020

Background

The practice of daily aerobic exercise such as healthy walking could have an important effect on cardiovascular parameters. However, there is still little evidence of how much weekly walking volume is enough to influence cardiovascular autonomic modulation and arterial stiffness on hypertension.

Purpose

To assess effects of weekly walking volume on cardiac autonomic function and arterial stiffness index in elderly hypertensive.

Methods

Sixty hypertensive elderly with 71.7 ± 3.4 years old were allocated in physically inactive (PI, n=30) and active (PA, n=30) groups. Hemodynamic parameters of heart rate (HR), peripheral blood pressure (SYS, DIA, MBP), cardiac autonomic modulation by the method of HR variability in both the time (root mean square of successive differences (RMSSD) and percentage of RR intervals differing by <50ms (pNN50)) and frequency domains (low and high frequency (LF, HF), and LF/HF ratio), stiffness (augmentation index; AIX, pulse wave velocity; PWV) and total vascular conductance (TVC), and central blood pressure (cSYS, cDIA) were measured in baseline conditions.

Results

The average of weekly walk in PA group was 312.4 ± 36.5 min. vs. 57.7 ± 47.9 min. in PI group (p <0.001). Pronounced resting bradycardia was observed in PA group (62.1 ± 8.7 bpm) in comparison with PI group (75.7 ± 9.1; p=0.0). Brachial SYS (124.3 ± 7.6 mmHg) DIA (76.5 ± 7.4 mmHg) and MBP (92.4 ± 7.4 mmHg) were lowers in PA than PI group (133.6 ± 6.0 mmHg; p=0.003), (82.7 ± 7.6 mmHg; p=0.032) and (99.4 ± 7.0 mmHg; p<0.05), respectively. The PA group showed higher cardiac parasympathetic modulation than PI group, evidenced by an increase in RMSSD (PI: 43.8 ± 23.9 vs. PA: 69.4 ± 29.4 ms; p=0.047), pNN50 (22.7 ± 3.9 vs. 40.7 ± 5.1 %; p=0.038) and in cardiac HF band (PI: 59.9 ± 5.8 vs. PA: 67.0 ± 3.7 nu; p=0.005). There were no differences in sympathetic modulation between PA and PI groups. The PI group had higher PWV (10.3 ± 1.2 m/s), AIX (36.4 ± 8.0 %) and TVC (1.4 ± 0.1 s*mmHg/ml) when compared to PA group (PWV = 9.1 ± 0.7 m/s; p = 0.005), (AIX = 23.5 ± 13.6 %; p = 0.003) and (TVC = 1.2 ± 0.1 s*mmHg/ml; p <0.001). Additionally, the cSYS (119.5 ± 14.0 mmHg) and cDIA (76.2 ± 18.3 mmHg) were lower in PA than PI (135.8 ± 20.6 mmHg; p=0.045 and 91.8 ± 20.5 mmHg; p=0.034), respectively.

Conclusion

These findings suggest that a higher weekly volume of walking improves baseline hemodynamic parameters and provides greater vagal modulation at rest, concomitant with attenuation of arterial stiffness evidencing that this type of intervention has important cardiovascular safety.

Support or Funding Information

CAPES, CNPq, FAPEMIG, UFTM.
The synthesis and investigation of the catalytic properties of symmetric Schiff‐base ligand metal complexes
Authors: Wolfe Charles A. , Mueller Paul H. , Deifel Nicholas P.
Published: April 2020

An ongoing project investigating the synthesis and catalytic properties of symmetric heterometallic Schiff‐base ligand complexes has been instigated. A novel symmetric Schiff‐Base ligand metal complex has been synthesized, characterized through IR spectroscopy, NMR spectroscopy, and LC mass spectrometry. The catalytic activity of this Schiff‐Base ligand has been investigated using cyclic voltammetry, mass spectrometric study, and UV‐vis spectroscopy. With the long‐term goal being a publication of results, this study aims to contribute to the fields of inorganic and biochemistry.

Support or Funding Information

Barry Goldwater Scholarship Foundation
Identifying Best Target Pocket for Small Molecule Inhibition of Beta Catenin/E‐Cadherin Binding
Authors: Khan Faiha Rafia , Zhang Di , Koes David
Published: April 2020

The Wnt/beta catenin signaling pathway has been linked to the growth of certain types of cancer cells secondary to an uncontrolled increased expression of beta catenin. Finding a molecule that can inhibit the activity of beta catenin and reduce its concentration in cells could treat cancers involving this pathway. E‐cadherin is a protein involved in maintaining the integrity and adhesiveness of a cell. Disrupting the interaction between E‐cadherin and beta catenin may inhibit beta catenin expression, which may reduce the survivability of the cancer cell. In this study, we aimed to find the best target pocket for beta catenin/E‐cadherin binding based on large volume and high “druggability” score. The “druggability” score measures the binding potential of small molecules to the pocket of interest. We analyze the pocket of interest in both the monomer (beta catenin) and the complex (beta catenin with E‐cadherin bound) in order to identify the best target for small molecules. We hypothesize that the pocket volume and “druggability” score will increase when comparing the pocket of interest in the monomer versus the complex. First, we used the software PyMol and PocketQuery to search for key residues in the E‐cadherin and beta catenin interaction. The residues chosen were the amino acids Leucine 661 and Aspartate 665. Then, we used the website Pharmit to add these residues as features to the beta catenin receptor to screen for potential molecules that may bind using these residues. The pocket that these molecules were bound to was the pocket we chose to focus on. In order to eventually obtain more potential molecules, we ran a pocket volume analysis using the software MDPocket to see if the pocket volume fluctuates over time in both the monomer and the complex. We also used the software Fpocket to calculate a “druggability score” for both the monomer and the complex as the more “druggable” a pocket is, the better target it is. The results showed that the volume fluctuates over time in the monomer and the complex, with the complex having overall larger volumes. Furthermore, the highest “druggability” scores from a scale of −1.0 (“undruggable”) to +1.0 (most “druggable”) in the monomer and complex were 0.601 and 0.984, respectively. With these results, we conclude that the complex may be a better target for future analysis as the properties of having both a larger pocket volume and a higher “druggability” score correlates with an increased chance for a small molecule to bind.
Mechanism of Reproductive Effect of Acetyl acetate‐methanol Fraction of Adansonia digitata (Linn) Root Bark Extract in Female Wistar Rats
Authors: Chibuogwu Ijeoma Chika , Nwannenna Agnes Ifeyinwa , Ogwu David , Ubah Simon Azubuike , Agbonu Adikpe Oluwa , Columbus Philemon Kwinjoh , Abah Kenneth Owoicho , Agbalaka Samson Eneojo , Abayomi Samuel Bankole , Enem Simon Ikechukwu , Itopa Ajayi Etudaye , Ejiofor Charles Ejike
Published: April 2020

The aim of this work was to identify the exclusive components of Acetyl acetate‐methanol fraction (AAMF) of Adansonia digitata (Linn) root bark extract and to elucidate its mechanism of action in modulating hormone signaling. Gas chromatography and Mass spectrometry was done to identify the compounds present in fractions of A. digitata (Linn) root bark extract. Thirty female Wistar rats had their estrous cycles synchronized and were randomly distributed into three groups of ten rats each. Each group received 0, 150, and 300mg/kg body weight of AAMF per os from the day observed as proestrus by vaginal cytology to day‐seven. On day‐one, day‐four, day‐five, day‐six and day‐seven, blood was collected from rats in each group. Blood collected was analyzed for estrogen, progesterone and follicle stimulating hormone levels. Uterus and ovary were processed for histopathology and immunohistochemistry to study the regulation of estrogen and progesterone receptor proteins. Where a receptor protein was regulated, further investigation was done to determine its effect on the gene expression of that receptor in endometrial stromal cells through RNA extraction, OneStep reverse transcription‐pCR amplification, Agarose gel electrophoresis and quantification of pCR product. Results showed that AAMF uniquely contained 15.25% of oleic acid and caused mass atresia of antral follicles at proestrus. AAMF treated rats had significantly (p<0.05) lowered serum estradiol levels at proestrus when compared to control rats and had up‐regulated estrogen receptor beta proteins in endometrium and downregulated estrogen receptor alpha (ERα) gene expression in endometrial cells. It was concluded that the oleic acid component of AAMF may have lowered estrogen level by inhibiting aromatase and hydroxysteroid 17 beta dehydrogenase enzyme activities in estrogen steroidogenesis. Also, oleic acid may have induced overexpression of Tumour Suppressor Factor (P53) which promotes follicular apoptosis and indirectly downregulates ERα when p53 activities become exacerbated. It is recommended that AAMF be investigated as a possible therapy for some ERα sensitive clinical conditions like cancer.
Types of Neurons in the Spinal Trigeminal Nucleus of the Camel Brain: Golgi Study
Authors: Bataineh Ziad M.
Published: April 2020

The camel lives in hot desert and eats thorny plants. It is exposed to severe environmental conditions such as dust, high temperature and thorny plants. These sensations from the face area may affect the development of neurons in the camel’s spinal trigeminal nucleus which contain the second‐order neurons of the sensory pathways for pain and temperature from the face area. Neurons in the spinal trigeminal nucleus of the camel were studied by Golgi impregnation.

Aims

The aims of this study were to describe the different types of neurons in the spinal trigeminal nucleus and to compare these neurons with their counterparts in other species.

Method

Blocks (2–4mm thick) of camel brainstem preserved in 10% formalin for months were cut and processed by two variations of Golgi Kopschi method (Fox and Braintenberg staining methods). Transverse sections (100 μm thick) were cut and examined under the light microscope. Well‐impregnated neurons were studied, measured and photographed.

Results

Neurons were classified based on differences in size and shape of their cell bodies, density of their dendritic trees, morphology and distribution of their appendages.

In the spinal trigeminal nucleus of the camel, at least twelve types of neurons were identified. These neurons include, stalk, islets, octubus‐like, lobulated, boat‐like, pyramidal, multipolar, round, oval and elongated neurons. They have large number of different forms of appendages not only for their dendrites but also for their cell bodies. Neurons with unique large dilatations especially at their dendritic branching points were found. These neuronal types were found in the three divisions of the spinal trigeminal nucleus namely the spinal trigeminal caudalis, spinal trigeminal interpolaris and spinal trigeminal oralis. The morphological features of these neurons were described and compared with their counterparts in other species.

Conclusion

Finding of neurons with variation in number and shape, in addition, to variation in number and shape of the appendages on the cell bodies and dendrites may indicate the presence of a very complex information processing for pain and temperature at the level of the spinal trigeminal nucleus in the camel. Together with the presence of cells with unique features such as large dilated parts for dendrites, may suggest that, the spinal trigeminal nucleus in the camel serves as center for sensations processing and not only as relay nucleus for the pain and temperature pathways from the face. This study will improve our understanding of the function of the spinal trigeminal nucleus and contribute to our understanding of comparative neuroanatomy and neurophysiology.

Support or Funding Information

Research Grant 100/2011 from the Deanship of Scientific Research at Jordan University of Science and Technology
Targeting the Upstream Open Reading Frame of Gadd34 to Treat Sepsis‐induced Acute Kidney Injury
Authors: Kidwell Ashley , Maier Bernhard , Dagher Pierre , Hato Takashi
Published: April 2020

Sepsis‐induced acute kidney injury (AKI) remains a major clinical problem with no effective therapies established to date. We have previously shown that bacterial sepsis causes global translation shutdown via phosphorylation of the eukaryotic translation initiation factor 2α (eIF2α). Under physiological conditions, eIF2α phosphorylation is tightly counter‐regulated by two eIF2α holophosphatases since excessive phosphorylation of eIF2α is deleterious. Of the two holophosphatases, growth arrest DNA‐inducible gene 34 (Gadd34) is the only stress‐inducible regulatory subunit. Using ribosome profiling, or Ribo‐Seq, we found that Gadd34 is translationally repressed during late phase sepsis even though eIF2α is already heavily phosphorylated. This failure of Gadd34 induction could explain the sustained phosphorylation of eIF2α and translation shutdown that contributes to delayed renal recovery in sepsis. The 5′‐untranslated region (UTR) of Gadd34 has multiple upstream open reading frames (uORFs) that are conserved across mammalian species. Our Ribo‐Seq analysis of kidneys from septic animals revealed high ribosomal occupancy of a specific Gadd34 uORF, but not the main protein coding sequence (CDS), consistent with a model in which the uORF serves as a translational inhibitor of the downstream CDS. With these findings, the objective of the investigation was to analyze the effect of disrupting translation in the uORF of Gadd34 in septic states.

To investigate the inhibitory role of the Gadd34 uORF, we designed plasmid constructs, which consisted of a full length Gadd34 5′‐UTR and luciferase reporter CDS in‐frame with a single nucleotide mutation introduced to abolish the uORF start codon. The uORF point mutation led to a two‐fold increase in luciferase signal compared with the wild‐type control, confirming the inhibitory property of the Gadd34 uORF. Next, we designed antisense oligonucleotides (ASOs) complementary to the upstream portion of the uORF. Interestingly, masking the uORF with ASOs resulted in sequence‐specific increases in translation of the downstream CDS, possibly due to enhanced leaky ribosomal scanning. Finally, we tested the applicability of the ASO approach in vivo using a mouse model of endotoxin‐induced kidney injury. Despite late intervention at eight hours post‐endotoxin challenge, intravenous administration of Gadd34 uORF ASOs significantly reduced renal tissue damage as determined by the levels of hepatitis A virus cellular receptor 1/kidney injury molecule 1 (Havcr1/KIM1). Collectively, these findings indicate that translational suppression of Gadd34 in late phase sepsis is a maladaptive response that could be therapeutically modulated by targeting its uORF.

Support or Funding Information

This work was supported by NIH grants K08‐DK113223, R01‐DK080063 and a Veterans’ Affairs Merit grant 1I01BX002901.
Up‐Regulation of Interleukin‐10 Expression in Dendritic Cells as an Important Role in The Immunomodulation By Strongyloides stercoralis
Authors: Sutaveesup Vittawin , Sodsai Pimpayao , Nuchprayoon Surang , Sanprasert Vivornpun
Published: April 2020

Strongyloidiasis is a parasitic disease caused by Strongyloides stercoralis. Infection in the immunocompromised hosts such as organ transplants, SLE patients, or HIV and HIPV infected patients can cause severe strongyloidiasis. About 90% of patients with severe strongyloidiasis are dead. Infected patients have high levels of Th2, and Treg related cytokines, including IL‐4, IL‐5, IL‐9, IL‐10, IL‐13, and TGF‐β. These cytokines are then decreased to normal levels after the treatments. Parasites have several mechanisms to suppress or evade the host’s immune response, such as suppressing the expression of co‐stimulatory molecules on antigen presenting cells (APCs), stimulating apoptosis of APCs, or stimulating Th2 and Treg responses. Dendritic cells (DCs) play a crucial role in linking the innate and adaptive immune systems against pathogens by engulfing antigens and presenting to naïve T cells with appropriate cytokines. To understand the mechanisms of S. stercoralis to evade the host’s immune responses, we studied the alteration of antigen presentation process and cytokines productions by DCs stimulated with S. stercoralis. We treated murine dendritic cell line (DC2.4) with S. stercoralis L3s crude antigen and measured the expressions of genes related with antigen presentation and cytokine productions by quantitative real‐time PCR; including MHC‐II, CD40, CD80, TLR‐2, TLR‐4, IL‐6, IL‐10, TNF‐α, and TGF‐β. The results showed that the expression of TLR2 and CD40 were significantly increased after the stimulation by crude antigens for 1 hr and 3 hrs, respectively. Likewise, the significant up‐regulation of IL‐10 expression of dendritic cells was also detected. While the expression levels of IL‐6 were not changed, TNF‐α expression levels were significantly increased. Our result suggested that S. stercoralis modulate the host’s immune response through the induction of Treg response by IL‐10 production by dendritic cells, not by the suppression the expression of co‐stimulatory molecules on antigen presenting cells.

Support or Funding Information

1. The Ratchadaphiseksompotch Fund, Faculty of Medicine, Chulalongkorn University, grant number RA62/026 2. Overseas Academic Presentation Scholarship for Graduate Students from the Graduate School, Chulalongkorn University
A New Mouse Strain to Study Motoneurons in Pompe Disease
Authors: Singer Michele L. , Lavassani Emmanuelle , Pope Megan K. , Harfe Brian D. , Haney Meghan Green , Corti Manuela , Byrne Barry J. , Fuller David D.
Published: April 2020

Pompe disease is a lysosomal storage disorder resulting from a deficiency in the lysosomal enzyme acid α‐glucosidase (GAA). The most severe form of the disease is associated with complete or near complete absence of functional GAA which results in cardiorespiratory failure and death within the first year of life. The only approved treatment for Pompe disease is human recombinant GAA (enzyme replacement therapy (ERT)) which improves overall survival rates but the recombinant protein does not reach the central nervous system. Respiratory impairment in Pompe disease has historically been attributed to muscle weakness but an accumulation of studies of Pompe patients and animal models indicate that motoneuron pathology is functionally relevant. In an effort to better understand the role of motoneurons in Pompe disease, we used breeding strategies to create a new mouse model with motoneuron histopathology features seen in Pompe disease while also allowing easy visualization of motoneurons via a fluorescent tag. Chat‐cre mice (Jackson) were crossed with Gaa−/− mice (Taconic). The resulting heterozygous strain were crossed with eYFP+/+ mice (courtesy Dr. Brian Harfe). Further breeding resulted in homozygous mice which were Chat‐cre+/+/Gaa−/−/eYFP+/+. These mice were then crossed to a Flox‐Gaa+/+ (Taconic) to create a heterozygous strain (Chat‐cre+/−/Gaa+/−/eYFP+/−/Flox‐Gaa+/−). Further inbreeding produced mice that were Chat‐cre+/+/Gaa+/−/eYFP+/+/Flox‐Gaa+/+. The intent of this final cross was to produce mice in which alpha motoneurons express eYFP but do not express GAA. Histological evaluation of the brainstem and spinal cord of these mice showed robust eYFP expression in the brainstem including the hypoglossal motor nucleus, nucleus ambiguous, and facial motor nucleus. In the cervical spinal cord, alpha motoneurons throughout the ventral horn also had robust fluorescence. This included putative phrenic motoneurons as determined based on location and size. The robust eYFP expression was restricted to motoneurons which indicates that motoneurons are Gaa−/− (based on the breeding scheme). Ongoing studies are evaluating GAA protein expression in peripheral tissues, glycogen accumulation in motoneurons, and also brainstem and spinal cord motoneuron loss over the lifespan. In addition, the eYFP signal in motoneurons will permit rigorous evaluation of motoneuron transduction efficacy in gene therapy experiments.

Support or Funding Information

2R01HD052682‐11A1 (DDF/BJB)
Involvement of transcriptional/epigenetics mechanisms in mediating the inhibitory effect of chronic alcohol exposure, and effect of alcohol metabolites on intestinal uptake of biotin.
Authors: Srinivasan Padmanabhan M. , Ramamoorthy Kalidas , Chu Brain D. , Juburi Saleh A.L. , Said Hamid M.
Published: April 2020

Biotin is indispensable for normal cellular metabolism, proliferation and survival. The vitamin acts as a co‐factor for multiple carboxylases that play critical roles in fatty acid, glucose and amino acid metabolism. Emerging evidence has also shown important roles for biotin in immune function and in gene expression, as well as in the maintenance of intestinal integrity and homeostasis. Humans and other mammals cannot synthesize biotin, and thus, must obtain the vitamin from exogenous sources (diet and the gut microbiota) via absorption in the gut. We had previously observed that chronic alcohol feeding/exposure leads to a significant inhibition in small intestinal and colonic biotin uptake (Am J Physiol Gastrointest Liver Physiol 300: G494–G501, 2011). The aim of this study was to elucidate the role of transcriptional/epigenetic factors in mediating the inhibitory effect of chronic alcohol on intestinal uptake of biotin, and to study the effect of alcohol metabolites on biotin uptake in the large intestine. Our results showed that the inhibition in biotin uptake by intestinal epithelial Caco‐2 cells upon chronic exposure to alcohol is associated with a decrease in the expression of KLF‐4, a key transcription factor involved in the regulation of SLC5A6. We also observed (using Chip‐qPCR) that exposure of Caco‐2 cells to alcohol to lead to decrease in the euchromatin histone markers (activators‐H3K4me3 and H3K9Ac) and an increase in the heterochromatin histone marker (repressor – H3K27me3) at the SLC5A6 P1 promoter. However, DNA methylation status appeared to be unaltered in these cells upon alcohol exposure. We confirmed the above effects of alcohol on histone modifications in vivo using small intestinal mucosa from mice chronically fed alcohol. We also examined the effect of the alcohol metabolites ethyl oleate (3 mM), ethyl palmitate (3 mM) and acetaldehyde (500 μM) on biotin uptake by human colonic epithelial NCM460 cells and observed a significant inhibition in carrier‐mediated biotin uptake in these cells. Studies are in progress to delineate the exact molecular mechanism(s) involved in this inhibitory effect. This study show, for the first time, that exposure of small intestinal epithelial cells to alcohol alters the epigenetic profile at the SLC5A6 promoters and the expression of an important nuclear factor that drive promoter activity, and that these changes contribute to the inhibition in biotin uptake. Further, the results show that relevant alcohol metabolites inhibit biotin uptake by colonic epithelial cells.

Support or Funding Information

DVA and the NIH (DK‐56061, DK58057, and AA 018071
Inhibition of bee pollen on acetylcholine esterases and cytotoxicity of human neuroblastoma, cerebral glioma, and macrophages
Authors: Lee Je-Hyuk
Published: April 2020

Inhibition of Actinidia argute, Quercus dentate, the mixed plant‐bee pollen extracts on acetylcholineesterase (AChE) and butyrylcholineesterase (BChE), which are indicators of anti‐Alzheimer’s activity, were investigated. Actinidia argute, Quercus dentate, the mixed plant‐bee pollen extracts (0–3mg/ml) did not show inhibitory activity of AChE and BChE. Additionally, the inhibitory activities of cathechin, rutin, hesperidin, and quercetin, which are flavonoids in bee pollen, on AChE and BChE were examined. Catehchin and rutin inhibited about 54 and 86% of AChE activity at a concentration of 1 mg/ml, respectively. Also, cathechin and rutin(1 mg/ml) inhibited about 42 and 86% of BChE, respectively. Quercetin was effective in inhibiting AChE and BChE, but the inhibitory pattern was not observed in proportion to the treatment concentration. However, hesperidin did not show a decrease for AChE and BChE activity. In addition, we investigated the inhibitory activity of Actinidia argute, Quercus dentate, the mixed plant‐bee pollen extracts against H2O2‐induced cytotoxicity for human neuroblastoma(SK‐N‐SH), cerebral glioma(C6), and macrophages(RAW 264.7). Cytotoxicity of SK‐N‐SH, C6, and RAW 264.7 by Actinidia argute, Quercus dentate, the mixed plant‐bee pollen extracts were examined by MTT in the range of 0–3 mg/ml. For SK‐N‐SH, C6, and RAW 264.7, there was a difference in the toxic concentrations by Actinidia argute, Quercus dentate, the mixed plant‐bee pollen extracts. Cytotoxicity by H2O2(0–10 mM) was partially improved by Actinidia argute, Quercus dentate, the mixed plant‐bee pollen extracts.
Cell Cycle‐Dependent Cleaving of Holliday Junctions via Yen1 in S. cerevisiae
Authors: Hadi Hafridha , Fischhaber Paula
Published: April 2020

Double‐strand breaks (DSBs) in DNA can be harmful if left unrepaired. Eukaryotes use repair mechanisms, some of which involve intact chromosomes that template repair via an intertwined intermediate known as a Holliday Junction (HJ). In S. cerevisiae, recruitment of the endonuclease enzymes that cleave HJs back into individual chromosomes is not well understood. Yen1 resolves HJs but it’s unclear whether it localizes to anaphase bridges, the final intertwined regions during chromosomal separation. Yen1 appears in anaphase which occurs during a small time frame within the cell cycle, making it difficult to observe in an asynchronous cell population. Further, Yen1 is present in lower abundance relative to other proteins, therefore, microscopy experiments with fluorescently labeled Yen1 will suffer from low signal‐to‐noise. Our aim is to clone a yeast strain with a bright version of red fluorescent protein under the strong galactose (GAL1) promoter so that Yen1‐yEMRFP will be in higher abundance, thereby allowing us to track the position and timing Yen1 during HJ resolution. To construct this strain, we used adaptamer‐mediated Polymerase Chain Reaction (PCR), DNA digestions and ligations to prepare two DNA plasmids. Plasmid One contains a gene for yeast Enhanced Monomeric Red Fluorescent Protein (yEMRFP), a partial URA3 auxotrophic selection marker gene, and an upstream flanker sequence of the target yeast chromosome to direct integration. Plasmid Two contains the other half of the partial URA3 selection marker, the galactose promoter, and the downstream flanker sequence to target integration. Candidate clones were screened by PCR and DNA sequencing which showed that the candidate plasmids had the desired sequences and are ready to be integrated into the yeast chromosome. These DNA plasmids will allow cloning of a specialized yeast strain that will produce an improved signal‐to‐noise resolution to elucidate Yen1’s role at anaphase bridges.

Support or Funding Information

This research was funded by NIH grant, SC1GM127204.
Investigating SLX4‐Dependent Recruitment of a DNA Repair Protein to DNA Double Strand Breaks in Dividing Cells
Authors: Hakimian Jonathan , Cureg Gyana , Fischhaber Paula
Published: April 2020

Cancer is a disease that functions as a consequence of abnormal cell growth, generally caused by DNA damage that triggers abnormal cell function. One form of DNA damage is the double‐strand break (DSB) in which both strands of a chromosome are severed in near proximity. In normal cells, several forms of DSB repair, including single strand annealing (SSA) and synthesis dependent strand annealing (SDSA), fix the double strand break (DSB), restoring an intact chromosome, albeit sometimes with an altered DNA sequence. These DNA repair processes require the sequential action of numerous proteins that enable their many steps. In S. cerevisiae these include the Rad1‐Rad10 endonuclease complex, which cleaves nonhomologous DNA flaps in SSA and SDSA as one of the final steps.

We have evidence that the Slx4 protein is partially required for the recruitment of Rad1‐Rad10 to DSB sites when cells are dividing (in S, G2 or M phases). Those experiments did not show the basis for the “partial” nature of the recruitment, but our hypothesis is that the Slx4‐dependency is tied to a specific phase of the cell cycle, possibly S phase. A plausible explanation is that when Slx4 exercises its checkpoint signal dampening role in S phase, which promotes homologous recombination, it indirectly recruits Rad1‐Rad10 to double‐strand break repair sites. Since, following DNA damage, checkpoint signal dampening is triggered (in part) by phosphorylation of Slx4 in S phase and the specific phosphorylation sites for this are known, we will test our hypothesis by creating mutant strains which lack these sites and repeating the earlier experiments using synchronized cell cultures. We began creating the mutant strains by cloning DNA plasmids containing the slx4‐7MUT gene, our focus, in which wild‐type SLX4 has been mutated to alanine at the seven phosphorylation sites activating checkpoint signal dampening. Candidate clones were screened by PCR and DNA sequencing. The successful candidate plasmids were transformed into a panel of W303 background yeast strains and integration of the mutant cassettes was confirmed by PCR and DNA sequencing screens. The strains were then crossed to strains bearing the fluorescence microscopy assay markers we will use in the fluorescence microscopy assay being employed. We have carried out preliminary fluorescence microscopy assays in which cultures of the progeny of these crosses were synchronized at the G1/S boundary and released back into the cell cycle that show acceptable arrest but the limited release back into the cell cycle that was observed requires further optimization. Using this separation‐of‐function mutant strain will allow us to compare recruitment of Rad1‐Rad10 in SLX4 wild‐type strains with Slx4 checkpoint signal dampening mutants. Understanding the process of Rad1‐Rad10 recruitment may aid in the design and development of a more effective means of cancer treatment.