bluuebunny/crossref_metadata_2025 huggingface dataset:
Investigating Oncogenic Roles of Long Intergenic Non‐Protein Coding RNA 461 (LINC00461) in HDAC6‐Mediated Glioblastoma Malignancy
Authors: Wu An-Chih , Yang Wen-Bin , Chang Wen-Chang , Chuang Jian-Ying
Published: April 2020

Glioblastoma (GBM) is the most complex, difficult‐to‐treat, aggressive, and lethal type of brain tumor. Thus, studies are urgently needed to determine the molecular mechanisms of GBM malignancy and develop novel concepts for treating the disease. According to our previous study, the selective histone deacetylase 1, 2, 6 (HDAC1, 2, 6) inhibitor MPT0B291 prevents the development of temozolomide (TMZ) resistance in GBM cells and prolongs the survival of mice bearing orthotopic GBM xenografts. Although HDACs are known to regulate the balance of lysine acetylation on histones and nonhistone substrates, the role of HDACs in regulating long non‐coding RNAs (lncRNAs) affecting GBM malignancy remains unclear. By performing microarray analysis, we identified an evolutionary conserved lncRNA LINC00461 that was clearly reduced by MPT0B291 in both TMZ‐resistant GBM cells and stem‐like tumorspheres. Additionally, we found that LINC00461 depletion suppressed GBM cell proliferation via G1 phase arrest. Because the miRNA‐411‐5p/topoisomerase II alpha (TOP2A) axis is known as a target of LINC00461 for controlling chromosome segregation during mitosis, we examined the TOP2A levels and verified that both MPT0B291 treatment and LINC00461 silencing caused dramatic decreases in TOP2A expression. Furthermore, we identified the interaction between HDAC6 and subunit 6 of CCR4‐NOT exoribonuclease complex (CNOT6), an acetylation‐activated deadenylase that promotes RNA degradation. In conclusion, this study will increase the understanding of the role of LINC00461 in GBM malignant properties and reveal whether targeting the HDACs/CCR4‐NOT/LINC00461 axis and its downstream effectors with a selective inhibitor is an effective therapeutic strategy against GBMs.

Support or Funding Information

This work was supported by grants from the Ministry of Science and Technology (MOST 106‐6601‐003‐112_15060508367900001 & 107‐6601‐004‐112_15553117427110001).
Altered oligomeric states in pathogenic ALS2 variants associated with juvenile motor neuron diseases cause loss of ALS2‐mediated endosomal function
Authors: Sato Kai , Otomo Asako , Ueda Mahoko Takahashi , Hiratsuka Yui , Suzuki-Utsunomiya Kyoko , Sugiyama Junya , Murakoshi Shuji , Mitsui Shun , Ono Suzuka , Nakagawa So , Shang Hui-Fang , Hadano Shinji
Published: April 2020

ALS2, the causative gene product for a number of recessive motor neuron diseases, forms a homophilic oligomer and acts as a guanine nucleotide exchange factor (GEF) for the small GTPase Rab5. This oligomerization is crucial for both Rab5 activation and ALS2‐mediated endosome fusion and maturation in cells. Recently, we have shown that pathogenic missense ALS2 variants retaining the Rab5 GEF activity fail to properly localize to endosomes via Rac1‐stimulated macropinocytosis. However, the molecular mechanisms underlying dysregulated distribution of ALS2 variants still unknown. In this study, we clarify the relationship between intracellular localization and oligomeric states of pathogenic ALS2 variants. Upon Rac1 activation, all variants moved from the cytosol to membrane ruffles, but not to macropinosomes and/or endosomes. Further, most wild‐type ALS2 in cultured cells forms tetramers. Importantly, the sizes of an ALS2 complex carrying missense mutations in the N‐terminus of RLD or in‐frame amino acid deletion in the PH domain were shifted toward high molecular weight, whereas the C‐terminal VPS9 domain missense mutant existed as a smaller dimeric or trimeric smaller form. Furthermore, in silico mutagenesis analyses using the RLD protein structure in conjunction with a cycloheximide chase assay in vitro disclosed that these missense mutations led to decrease in the protein stability. These results suggested that disorganized higher structures of ALS2 variants might explain their impaired endosomal localization and the stability, leading to loss of the ALS2 function. Currently, to further clarify the in vivo structure, gel‐filtration analyses of the ALS2 complex in murine tissues, including the brain, spinal cord and liver, are underway.

Support or Funding Information

This work was supported by Japanese Society for Promotion of Science Grants 26290018 and 24650189 and in part by the National Natural Science Foundation of China and JSPS Bilateral Joint Research Project.
Anatomy Faculty Views and Opinions about Curricular Reform at Allopathic Medical Schools in the United States
Authors: Taylor Melissa
Published: April 2020

Introduction

Medical education in the United States has changed dramatically in the last few decades. Medical schools have moved away from a traditional, didactic, and discipline‐based curriculum, to a more integrative, active, and student‐centered curriculum. While much research has documented the specific changes in medical curriculum, few have researched the perceptions towards these changes.

Aim

This study examines specific changes that have occurred in anatomical science courses at allopathic medical schools in the United States due to curricular reform and how faculty have reacted to those changes.

Methodology

A survey was distributed to anatomical science faculty who teach gross anatomy, microscopic anatomy, and/or neuroanatomy at allopathic medical schools in the United States. The survey included both quantitative and qualitative questions related to the change in content coverage, the change in lecture and lab experience, and faculty perceptions of those changes.

Results

Findings from the survey showed that most anatomical science courses are currently taught in an integrated manner with various active learning pedagogical techniques used in the classrooms. The survey also found that most of the anatomical science courses had their contact hours reduced, leaving much less time for the faculty to teach their students the relevant information. Open‐ended survey responses showed that faculty believed the integration of basic and clinical science information was helpful for their students. However, using Mann‐Whitney U and Kruskal Wallis tests relating faculty perceptions on a Likert Scale to changes in the anatomical science medical curriculum, it was found that faculty whose medical school had undergone recent curricular reform had more negative perceptions about their students receiving adequate instruction in anatomical science material.

Discussion and Conclusion

These data provided valuable, though contrasting, information about medical curricular changes and faculty perspectives about those changes. While some faculty had positive things to say about the curricular changes, others believed it was not beneficial to their students. These more negative views should not go unnoticed by administration designing the medical curriculum, and more heed should be taken to listening to all the stakeholders in a medical curriculum.
Chronic Low‐Dose ICV Infusion of Agrp in Mice May Suppress Indices of Resting Metabolism in Brown Adipose Tissue Independent of Effects on Feeding Behavior
Authors: Oliveira Vanessa , Balapattabi Kirthikaa , Patil Chetan N. , Hei Megan Vande , Silva Sebastião D. , Reho John J. , Sigmund Curt D. , Grobe Justin L.
Published: April 2020

Agouti‐related peptide (Agrp) acts within in the central nervous system to stimulate feeding behavior and suppress energy expenditure, which results in body weight gain. Acute and chronic intracerebroventricular (ICV) administration of recombinant Agrp (0.2 to 1.0 nmol/d) induces hyperphagia in rats (Wissig et al. 2003; Tallam et al. 2004), but studies in mice are lacking. Therefore, we tested the effects of this route and range of infusion doses upon food intake, body mass, and adiposity in wildtype male C57BL/6J mice (Jackson Labs, 10–12 wk). Mice were implanted with chronic indwelling ICV cannulae connected to osmotic minipumps (Alzet) to deliver artificial cerebrospinal fluid vehicle (n=6), or 0.1 nmol/d (n=8) or 1.0 nmol/d (n=6) recombinant Agrp(82–132) peptide for 21 days. Unexpectedly, despite the efficacy of these dosing schedules published in rats, these infusion rates had no effect on food intake, body mass, or overall body composition (by NMR) in the present cohort of mice. Interestingly, interscapular “brown” adipose tissue (BAT) was hypertrophied in mice infused with the lower dose of Agrp (aCSF: 0.07±0.01 mg, Agrp 0.1 nmol/d: 0.10±0.01, Agrp 1.0 nmol/d: 0.09±0.01 (mean±sem); p<0.05 by ANOVA), and this difference remained after correction by body mass or tibia length. Although underpowered to detect statistically significant changes with the limited sample size, physiologically relevant trends toward reduction in the mRNA expression of Ucp1 (0.75 (0.92–0.62) fold of aCSF (±1 sem), n=5), Adrb3 (0.65 (0.74–0.57), n=7), and Dio2 (0.49 (0.8–0.42), n=6) were observed in the BAT from mice receiving 1.0 nmol/d Agrp. We conclude that the effects of ICV Agrp on BAT mass (and possible effects on thermogenic gene expression profiles in BAT) are independent of changes in feeding behavior in male C57BL/6J mice. These findings support species‐specific differences in sensitivity to ICV Agrp, and suggest that Agrp influences resting energy expenditure independently from its effects on feeding behavior.

Support or Funding Information

HL134850, HL084207
Assessing the Effects of Leptin and Angiotensin II Upon Expression of Agouti‐Related Peptide in the GT1‐7 Cell Model
Authors: Hei Megan Vande , Patil Chetan N. , Sapouckey Sarah A. , Balapattabi Kirthikaa , Oliveira Vanessa , Deng Guorui , Grobe Justin L.
Published: April 2020

Leptin acts at its receptor (Lepr) within the arcuate nucleus of the hypothalamus (ARC) to reduce feeding behavior and increase resting metabolic rate (RMR) in part through the inhibition of Agouti‐related peptide (Agrp) signaling. Previously we demonstrated that a unique subset of Agrp neurons express the angiotensin II (ANG) type 1a receptor (Agtr1a). Genetic deletion of Agtr1a from Agrp neurons of mice was sufficient to disinhibit expression of Agrp within the ARC, and to attenuate adipose sympathetic nerve activity responses to leptin. In contrast, RMR responses to brain renin‐angiotensin system (RAS) stimulation are intact in the dysfunctional Lepr db/db mouse model, and acute leptin administration does not additively exaggerate RMR activation in a transgenic mouse model of increased brain RAS activity. New unpublished data also indicate that ANG signaling within the ARC stimulates RMR through suppression of Agrp expression and Agrp neuron electrical activity, and thereby disinhibition of melanocortin Mc4r receptor signaling. To further dissect the interaction of Lepr and Agtr1a within Agrp neurons, here we examined the effects of leptin and ANG stimulation of the “GT1‐7” immortalized cell culture model of mouse Agrp‐expressing cells. We confirmed that cultured GT1‐7 cells express critical markers of the Agtr1a‐positive Agrp neuron subtype, including Agrp, Agtr1a, and Neuropeptide‐Y (Npy), but not pro‐opiomelanocortin (Pomc). Dose‐ and time‐course experiments were performed to identify mid‐range doses and time courses for Agrp gene expression responses to acute application of leptin and ANG. Subsequently, Agrp expression responses to low doses of leptin (100 nM) and ANG (100 nM), which would allow detection of any additive or synergistic effect, were assessed after 1 and 2–2.5 hours to evaluate the hypothesis that leptin and ANG signaling are non‐additive for the suppression of Agrp expression. After 1 hour, ANG (0.66 (0.49–0.91), fold of vehicle (± 1 sem), n=0.5) and leptin (0.61 (0.44–0.84), n=6) had similar effects on Agrp mRNA expression compared to vehicle (1.00 (0.86–1.17), n=7), but co‐administration of both peptides had no synergistic effect (0.96 (0.78–1.18), n=5). Likewise after 2–2.5 hours, compared to vehicle (1.00 (0.78–1.28), n=6), ANG (0.56 (0.34–0.94), n=6) and leptin (0.69 (0.57–0.85), n=6) had similar effects that were not additive (0.81 (0.67–0.98), n=6). These findings support the preliminary conclusion that the effects of leptin and ANG upon Agrp expression are not additive in the GT1‐7 cell culture model of Agrp neurons. These findings support our working model in which leptin action at the ARC results in increased Agtr1a signaling to suppress Agrp, ultimately to disinhibit Mc4r signaling and thus increase resting energy expenditure.

Support or Funding Information

HL134850, HL084207
The Glucosome: A Metabolic Complex for Glucose Metabolism and Its Functional Contribution to Cellular Metabolism in Living Cells
Authors: An Songon
Published: April 2020

There have been growing evidence that metabolic enzymes not only compartmentalize in the membrane‐bound organelles, like mitochondria, but also form spatially‐resolved, membraneless granules in the cytoplasm of the cells. Recently, we have demonstrated that all the cytoplasmic, irreversible enzymes in glycolysis and gluconeogenesis (i.e. glucose metabolism) dynamically organizes into a macromolecular metabolic complex, namely the “glucosome,” whose granules form in various sizes in human cells. Our biophysical characterization reveals that the enzymes inside the glucosome granules directly interact with each other while the granules themselves also dynamically undergo fusion and fission in the cytoplasm. Importantly, quantitative high‐content imaging assays support that glucosome granules are metabolically functional to regulate glucose flux between energy metabolism and building block biosynthesis in a size‐dependent manner. Furthermore, our single‐cell analysis reveals that glucosome granules functionally oscillate during the cell cycle in a size‐dependent manner. Collectively, we propose that the enzymes in glucose metabolism reversibly form phase‐separated glucosome granules, which are functionally relevant metabolic complexes for glucose metabolism in human cells.

Support or Funding Information

• GM125981 (An) and CA219609 (An)
Site‐Specific Opening of the Blood‐Brain Barrier by Extracellular Histones
Authors: Villalba Nuria , Baby Sheon , Yuan Sarah Y.
Published: April 2020

Increased extracellular histones in the blood are known as a biomarker for vascular dysfunction observed in patients with severe trauma or sepsis. While many previous studies have been focused on histone‐associated coagulopathy and organ (lung, kidney, liver) failure, limited information is available regarding the effects of histones on the brain or cerebral vasculature. Brain endothelial cells and cell‐cell junctions constitute the blood‐brain barrier (BBB) providing a critical paracellular seal that restricts the transfer of harmful substances from the circulation into the brain. It remains unclear whether circulating histones affect BBB structure or permeability function. To address these gaps, we applied a combination of in vitro functional analyses to evaluate the direct effects of histones (10–100 micrograms/mL) on brain microvascular endothelial cells by measuring transendothelial electrical resistance and solute flux. We also used in vivo approaches to obtain evidence of their influence on BBB integrity by quantifying brain tissue accumulation of intravenously injected tracers of different sizes: sodium fluorescein (376 Da), Alexa fluor 555‐cadaverine (1 kDa) and 3 kDa Texas‐Red dextran, in mice receiving a sublethal dose of histones (45 mg/Kg, iv). In mouse brain endothelial cells, treatment with 100 micrograms/mL histones resulted in increased paracellular permeability to sodium fluorescein and reduced barrier resistance, accompanied by discontinuous staining of the tight junction proteins zona ocludens‐1 (ZO‐1) and claudin‐5. The histone effect did not seem to result from cell death or toxicity, as indicated by negative propidium iodide staining. In mice infused with histones, the BBB permeability to small solutes of <1 kDa was increased, whereas tracers larger than 3 kDa remained impermeable across brain microvessels. Further analysis of coronal sections in different brain regions showed that histone‐induced tracer leakage mainly occurred in the hippocampus, but not in the cerebral cortex. Moreover, BBB dysfunction was associated with a reduced expression of several junction proteins in the hippocampal region. Together, these data suggest for the first time that circulating histones cause a region‐specific disruption of the BBB which may be responsible for impaired cognitive function observed in sepsis survivors.

Support or Funding Information

This work is supported by NIH grants: GM097270, HL070752, HL126646 and HL070752 to SYY.
Annilin regulates breast cancer cell migration, growth and metastasis by non‐canonical mechanisms involving control of cell stemness and differentiation.
Authors: Naydenov Nayden , Wang Dongdong , Dozmorov Mikhail , Koblinski Jennifer , Ivanov Andrei
Published: April 2020

Breast cancer metastasis is driven by profound remodeling of the intracellular cytoskeleton that enables efficient cell migration and invasion. Anillin is a unique scaffolding protein regulating major cytoskeletal structures, such as actin filaments, microtubules and septin polymers. It is markedly overexpressed in breast cancer and high anillin expression is associated with poor prognosis. The aim of this study was to investigate the role of anillin in breast cancer metastasis. CRISPR/Cas9 technology was used to deplete anillin in highly metastatic MDA‐MB‐231 and BT549 cells and to overexpress it in poorly invasive MCF10AneoT cells. The effects of anillin depletion and overexpression on breast cancer cell motility in vitro were examined by a combination of wound healing and Matrigel invasion assays. Assembly of the actin cytoskeleton and matrix adhesions was evaluated by immunofluorescence labeling and confocal microscopy. In vitro tumor development was monitored by soft agar grow, whereas cancer stem cells were examined using a mammoshpere formation assay and flow cytometry. The effects of anillin knockout on tumor growth and metastasis in vivo was determined by injecting control and anillin‐depleted breast cancer cells into NGS mice. The loss‐of‐function and gain‐of‐function studies demonstrated that anillin is necessary and sufficient to accelerate migration, invasion and anchorage‐independent growth of breast cancer cells in vitro. Furthermore, loss of anillin markedly attenuated primary tumor growth and metastasis of breast cancer in vivo. In breast cancer cells, anillin was localized in the nucleus, however knockout of this protein affected the cytoplasmic/cortical events, such as the organization of actin cytoskeleton and cell‐matrix adhesions. Furthermore, we observed a global transcriptional reprogramming of anillin‐depleted breast cancer cells that resulted in suppression of their stemness and induction of the mesenchymal to epithelial trans‐differentiation. Such trans‐differentiation was manifested by upregulation of basal keratins along with increased expression of E‐cadherin and P‐cadherin. Knockdown of E‐cadherin reversed attenuated migration and invasion of anillin‐deficient breast cancer cells. Our study demonstrates that anillin plays essential roles in promoting breast cancer growth and metastatic dissemination in vitro and in vivo and unravels novel functions of anillin in regulating breast cancer stemness and differentiation.
TRAF3IP2 ablation protects against obesity‐associated glycemic dysregulation, elevated blood pressure, and endothelial dysfunction
Authors: Grunewald Zachary Ian , Woodford Makenzie L. , Ramirez-Perez Francisco I. , Morales-Quinones Mariana , Manrique-Acevedo Camila , Siebenlist Ulrich , Martinez-Lemus Luis A. , Bysani Chandrasekar , Padilla Jaume
Published: April 2020

Obesity and type 2 diabetes (T2D) are leading independent risk factors for cardiovascular disease morbidity and mortality. However, the precise molecular mechanisms underlying the development of cardiometabolic dysfunction in obesity and T2D remain largely elusive. The recently identified cytoplasmic adapter molecule TRAF3 Interacting Protein 2 (TRAF3IP2) is known to regulate various pathological stress pathways, including inflammation and oxidative stress, associated with obesity and T2D. Accordingly, we tested the hypothesis that TRAF3IP2 mediates metabolic and vascular dysfunction associated with chronic high fat/high sucrose (HFHS) feeding. The results show that HFHS feeding for 16 weeks (starting at 5 weeks of age) increased body weight and impaired glucose tolerance in wild‐type (WT) male and female mice relative to control chow‐fed mice (P<0.05). Notably, glucose intolerance was attenuated in HFHS‐fed male and female homozygous TRAF3IP2 knockout (KO) mice (P<0.05) compared to WT counterparts despite greater weight gain. Interestingly, male TRAF3IP2‐KO mice exhibited protection against HFHS feeding‐induced elevations in blood pressure (tail cuff plethysmography) and ex vivo aortic endothelial dysfunction (P<0.05). However, female mice fed HFHS were protected against increases in blood pressure and impairments in aortic endothelial function and, thus, deletion of TRAF3IP2 was inconsequential (P>0.05). In summary, our data show that TRAF3IP2 plays a causal role in obesity‐associated glucose intolerance, elevated blood pressure and endothelial dysfunction. These findings implicate TRAF3IP2 as a promising therapeutic target in cardiometabolic dysfunction associated with obesity.

Support or Funding Information

Dept. of Nutrition and Exercise Physiology, University of Missouri, Columbia, MO, USADalton Cardiovascular Research Center, University of Missouri, Columbia, MO, USA Dept. of Biological Engineering, University of Missouri, Columbia, MO, USA Division of Endocrinology and Metabolism, Department of Medicine, University of Missouri, Columbia, MO, USA Harry S. Truman Memorial Veterans’ Hospital, Columbia, MO, USA Laboratory of Immunoregulation, NIAID/NIH, Bethesda, MD, USA Dept. of Medical Pharmacology and Physiology, University of Missouri, Columbia, MO, USA Division of Cardiovascular Medicine, Department of Medicine, University of Missouri, Columbia, MO, USA
The Role of Lifelong Physical Activity on Cardiovascular Function
Authors: Mortensen Stefan , Munch Gregers , Boushel Robert , Calbet José
Published: April 2020

Introduction

Ageing is associated with vascular dysfunction and a reduced capacity of the central circulation to deliver O2 to the contracting muscles. The purpose of present study was to investigate the role of lifelong physical activity level on vascular function and hemodynamics during whole‐body exercise.

Methods

Vascular function was determined by femoral arterial Ach infusion (10, 25 and 100 μg/min/kg leg mass) and central (ICG) and limb (thermodilution) hemodynamics were measured during incremental cycling to exhaustion with and without simultaneous arm cycling (50w) were determined in 9 young (Y) (23±1 years; mean±SEM), 9 lifelong sedentary elderly (SE) (66±2 years), 8 lifelong moderately‐active elderly (MAE) (64±2 years) and 8 lifelong very active elderly (VAE) (65±2 years) men. Data was analyzed with one‐way repeated measure ANOVA (within groups) and two‐way ANOVA (between groups).

Results

The vasodilatory response to ACh was lowest in the SE, higher in MAE and VAE (P <0.05) and highest in the Y (P <0.05). During incremental cycling, peak workload was higher in the VAE (313±10 W) and MAE (274±14 W) compared to Y and SE (247±13; 191±3 W; P<0.05). Leg blood flow (LBF) increased similarly in VAE, MAE and SE, but peak values were higher in VAE, MAE and Y compared to SE (7.8±0.5; 7.9±0.9; 8.6±0.4 vs. 6.4±0.4 L/min; P<0.05). Leg a‐vO2 difference was higher in VAE and MAE compared to the Y and SE at all workloads (P<0.05), whereas leg VO2 was higher in VAE, MAE and Y compared to SE at exhaustion (P<0.05). During combined arm and leg cycling LBF, leg a‐vO2 difference and VO2 was lower in Y and SE compared to leg cycling alone (P<0.05). Mean arterial pressure increased in all groups, but was lower in Y (P<0.05).

Conclusions

These results demonstrate that lifelong moderate physical activity can maintain aerobic power despite a lower vascular function and exercise hyperemia by an increase in O2 extraction.
In vitro antioxidant, anti‐inflammatory and antihypertensive activities of methanol leaf extract of Peasea americana
Authors: Adejumobi Olumuyiwa Abiola , Omobowale Temidayo Olutayo , Oyagbemi Ademola Adetokunbo , Adedapo Adeolu Alex , Yakubu Momoh Audu , Ashafa Anofi
Published: April 2020

Hypertension is a cardiovascular disease of remarkable health and economic importance. Management of hypertension with orthodox medicine is associated with side effects and is unaffordable to a larger percentage of people in Africa. P. americana leaf extract (PALE) is commonly used in the management of hypertension in Nigeria trado‐medicine. In this study, we investigated the antioxidant, anti‐inflammatory and anti‐hypertensive activities of the crude methanol extract, fractions and phytochemicals of P. americana leaf. The in vitro‐antioxidant assays were carried out to evaluate the free radicals scavenging activities of PALE on 1, 1‐diphenyl‐2‐picrylhydroxyl (DPPH), 2, 2‐azenobis‐3‐ethyl benzothiozoline‐6‐sulfonic acid (ABTS), hydroxyl and nitric oxide radicals. In addition, metal chelating and ferric reducing power assay were done. Anti‐inflammatory activities were evaluated using 15‐1ypoxygenase enzyme inhibition assay. Absorption were read spectophotometrically and 1C50 was calculated using a standard calibration curve. Phytocompound isolation was by extract fractionation, column, thin layer and preparatory thin layer chromatography. Crude methanol extracts, N‐hexane, ethyl acetate, chloroform, N‐butanol and aqueous methanol fractions had significantly higher anti‐oxidant activities when compared with gallic acid. There anti‐inflammatory activity is comparable to that of Indomethacin and antihypertensive activity for PALE (with IC50 of 135.99±10.06) is significantly lower when compared with captopril (with IC50 41.86±2.10). Four pure phytochemicals were isolated and named LEACA, LEACB, LEACC, and LEACD. The isolated compounds had significantly higher antioxidant activities on DPPH, ABTS, and hydroxyl and nitric oxide radicals when compared with gallic acid. The anti‐hypertensive activities of the isolated phytocompounds from the ethyl acetate fraction of the leaf were significantly higher (with IC50 of 24.83±0.63, 9.48±1.45, 2.79±1.35 and 1.59±0.48 respectively) than that of captopril (with 1C50 of 41.89±2.10). The result obtained from this study suggest that P. americana leaf has free radical scavenging and antioxidant activities, anti‐inflammatory and anti‐hypertensive activities and can ameliorate oxidative stress, and inflammation associated with hypertension.
Impact of Reduced ASIC2a on Cortical Thickness, Microglia Density, and Brain Water Content in E18.5 Fetuses
Authors: Sherrod Akia M. , Shao Qingmei , Thompson Rachael , Griffin Ashley D. , Jones-Muhammad Maria , Warrington Junie Paula
Published: April 2020

Acid‐sensing Ion Channels (ASICs) are ligand‐gated cation channels that respond to acidic stimuli. ASIC2 is widely expressed in neurons of the central and peripheral nervous system as well as on microglia. Although ASIC2 plays a role in cell migration, their role in brain development is not fully known. We tested the hypothesis that reduction or absence of ASIC2a contributes to changes in microglia density, cortical thickness, and brain water content during development. Brains were collected from ASIC2a wild type (WT, n=6), heterozygous (HET, n=4), and homozygous (HOMO, n=3) fetuses at E18.5. We used hematoxylin & eosin (H&E) staining for cortical thickness measurements, immunofluorescence staining using ionized calcium‐binding adapter molecule (Iba‐1) to measure microglia density, and wet weight to dry weight ratio for brain water content. Reduction of ASIC2a had no impact on cortical thickness (WT=499±21, HET=509±7, and HOMO=478±21 μm; p=0.60) or microglia density in the subventricular zone near the lateral ventricle (WT=22±2, HET=18±2, and HOMO=24±3; p=0.34) at E18.5. Interestingly, microglia morphology in HET fetuses were different compared to WT and HOMO mice with thinner and longer processes. Brain water content increased in HOMO mice (88.2±0.7%) compared to WT (86.5±0.3%, p<0.01) and HET (86.8±0.2%, p=0.02) fetuses. Our results indicate that while reduced ASIC2a does not contribute to changes in microglia density or cortical thickness in E18.5 mouse fetuses, reduced ASIC2a induces morphological changes in microglia and ASIC2 knockout increases brain water content at E18.5. Increased brain water content could be an indicator of hydrocephalus. In ongoing studies, we are determining whether reduced ASIC2a mediates structural and functional impairments in the neurovasculature of the maternal and developing brain.

Support or Funding Information

NIH Grants: R00HL129192, R00HL129192‐S1
Vasoreactivity of the Murine External Jugular Vein and Carotid Artery
Authors: Cho Jae Min , Shiu Yan-Ting , Symons J David , Lee Timmy
Published: April 2020

Few studies have characterized vasoreactivity of the murine external jugular vein. This segment often is used in preclinical models to evaluate vascular remodeling in response to arteriovenous fistula (AVF) maturation. When an AVF is established, the venous segment is exposed to arterial pressures and disturbed flow patterns that have strong potential to impact vasoreactivity. Before testing this hypothesis in the context of AVF, we sought to establish procedures to comprehensively characterize endothelium‐dependent and vascular smooth muscle function of the carotid artery and external jugular vein from healthy mice. External jugular veins and ipsilateral common carotid arteries were isolated from five‐month‐old male C57BL/6 mice and vasomotor responses were evaluated using isometric tension procedures. External jugular veins developed tension (p<0.05) to the voltage‐gated Ca2+ channel opener potassium chloride (KCl) and the thromboxane A2 receptor agonist U‐46619, but not to the α‐1 receptor agonist phenylephrine (PE), whereas common carotid arteries responded to all three vasocontractile agents in a more robust (p<0.05) manner. While maximal responses (~70%) to the endothelium‐dependent vasodilator acetylcholine (ACh) were similar between the venous and arterial segments, the dose required to achieve this value was lower (p<0.05) in the artery versus the vein. Nitric oxide synthase (NOS) inhibition using L‐NMMA attenuated (p<0.05) but did not abolish ACh‐evoked vasorelaxation in both vascular segments. Endothelium‐independent vasorelaxation to sodium nitroprusside (SNP) was similar in the artery and the vein. Endothelium‐dependent and vascular smooth muscle function can be reliably assessed in the murine external jugular vein of healthy male C57Bl/6 mice. Compared to common carotid arteries, external jugular veins are resistant to PE‐evoked vasocontraction, less responsive to KCl and U‐46619‐evoked vasocontraction, less sensitive to ACh‐evoked vasorelaxation, and equally responsive to NOS inhibition and guanylate cyclase activation via SNP. Ongoing studies will determine if and how external jugular vein function is altered by arterial pressures and disturbed flow patterns that are encountered subsequent to establishing an AVF.

Support or Funding Information

JMC (University of Utah Graduate Research Fellowship); YTS (R01DK100505; I01BX004133); JDS (AHA16GRNT31050004, NIH RO3AGO52848, RO1HL141540); TL (R44DK109789, R01HL139692, I01BX003387).
An Alternative Method of Expressing Appreciation to Families of Body Donors
Authors: Corson Andrew , Ohara Peter
Published: April 2020

INTRODUCTION

For over fifteen years, the UCSF Willed Body Program (WBP) has sponsored an annual student‐led body donor memorial service. The service provides first year UCSF medical, pharmacy, physical therapy and dental students with an opportunity to appreciate and process their experiences in the anatomy lab. Due to a number of limiting factors, including a lack of facilities to host the expected number of attendees, donor families are not currently invited to the service. Donor families have expressed disappointment to WBP staff over not being included. Recognizing the importance of expressing appreciation to donor families, the WBP has sought alternate methods to honor donors and include families in an expression of gratitude. To this end, the WBP prepared a book featuring personal notes and artwork from medical students that is sent to donor families within a few weeks after the death of a donor.

METHODS

At the memorial service students are asked to write notes to donors or donor families on 3″ × 5″ cards, a practice now in its seventh year. WBP staff reviews the notes to ensure they are legible, appropriate and contain nothing that will identify the donors. A number of notes are selected, scanned, and formatted using Adobe Photoshop, imported into Adobe Acrobat and then exported as PDF files for printing. The book also includes student artwork and an introduction written by WBP staff explaining the purpose, value and appreciation for the generosity of body donation. The 2019 book contains 24 pages plus the cover. The cost per book is approximately $3.50 and is printed by UCSF Documents and Media. The WBP mails a copy of the book to each donor family or representative upon a donor’s death. Since January 2019, when the WBP began sending the books, the WBP has received dozens of phone calls expressing appreciation for the book, as well as requests for additional copies. Families regularly comment that they understood that body donation was important but that personal reflections from the students emphasize the profound impact of body donation in a much deeper way.

CONCLUSION

The gift of a whole body to medical education and scientific research is a significant act of generosity that deserves recognition. While memorial events that include donor families are ideal, not all universities have the facilities to hold such events. Alternative opportunities to express appreciation and connect students and donor families are available and can be created with minimal resources while remaining deeply meaningful.
Knowing Real Cardiac Anatomy Enhances Understanding of Cardiac Development
Authors: Bolender David
Published: April 2020

New ideas about cardiac chamber formation, interatrial septum formation and development of the outflow region can be better understood by knowing real cardiac anatomy. The definitive chambers of the heart can be regionalized into components that provide a basis for understanding their formation during prenatal embryonic development. Each atrium has a venous inlet, a body, an appendage and a vestibule, while each ventricle consists of an inlet, an apical trabeculated portion and an outlet. During development several precursors combine to form the components of definitive chambers. Recent studies have challenged the classical version of interatrial septum formation, particularly the presence of a secondary atrial septum (septum secundum), and the closure of the primary atrial foramen (ostium primum). These studies suggest that the primary atrial septum (septum primum) merges with a fold of myocardium from the roof of the right atrium rather than with a second septum. This fold can be demonstrated in the dissection laboratory. Closure of the primary atrial foramen is accomplished by fusion of the ingrowing vestibular spine fusing with the mesenchymal cap of the primary septum and the merged AV endocardial cushions. This forms the anterior inferior atrial buttress that might be considered a secondary atrial septum. Finally, formation of the outflow region can be better understood using a tripartite explanation rather that the traditional bipartite version. During cardiac looping, the outflow region elongates and develops a bend in the middle. This flexure can define the outflow region into proximal, intermediate and distal segments. From these segments the outflow regions of the ventricles, the aortic valves and the intrapericardial roots of the great vessels are formed. The septum dividing the proximal segment is formed exclusively from endocardial cushion tissue, while the septum that partitions the intermediate and distal outflow regions is derived from cushion tissue and neural crest ectomesenchyme. Traditionally the aorticopulmonary septum was described as partitioning the entire outflow region. More recent examination of outflow tract development suggests that the aorticopulmonary septum only partitions the distal portion of the outflow region and the aortic sac. The intrapericardial and extrapericardial roots of the great vessels form from the division of the distal outflow region and the aortic sac.
The Influence of Physical Therapy on Microbiotic Dysbiosis in Patients with Rheumatoid Arthritis
Authors: Fincher Colton Ray , Muraca Katherine , Torres Denise , Raynes Edilberto A.
Published: April 2020

Rheumatoid arthritis (RA) is an autoimmune disorder caused by genetic and environmental factors. It causes joint inflammation due to an increase in pro‐inflammatory T‐ lymphocytes. Natural bacteria residing in the gut microbiota have a regulatory role on the immune system. Alterations in the gut microbiome occur due to dietary changes, illness and other disturbances leading to dysbiosis of the gut microbiome. In patients with RA, studies have shown increases in Prevotella copri, which induces differentiation into pro‐inflammatory T‐lymphocytes causing a symbiotic imbalance. The use of exercise has shown to modulate the composition and functions of the human gut microbiota; however, few studies have explored the effects of exercise on the human gut microbiota. The purpose of the study was to examine the effects of alterations in the gut microbiota on the development of RA and the influence of physical therapy (PT) intervention in relieving inflammation. Using the Oxford Center for Evidence‐Based Medicine, levels of evidence of literature were assessed. Results of the study revealed patients with RA were found to have different microbiota composition and diversity. The effect of physical therapy interventions in the form exercise resulted in increased concentrations of short chain fatty acids and butyrate‐producing bacteria which assist in regulating anti‐inflammatory phenotypes and provide protection against microbial dysbiosis. Also, improvements in cardiorespiratory fitness were positively correlated with lower pro‐inflammatory biomarkers. The researchers conclude microbial dysbiosis contributes to the progression of RA, and PT interventions may promote microbiome symbiosis and may improve inflammation through aerobic exercise‐based interventions.
BIOMEDICAL ENGINEERING – AN INTEGRATED APPROACH TO PHYSIOLOGY IN HIGH SCHOOL
Authors: Anjur Sowmya
Published: April 2020

The Illinois Math and Science Academy (IMSA) is a residential school for students of Illinois gifted in Math and Science. Several programs are offered during summers through the Center for Teaching and Learning (CTL), the outreach arm of IMSA, to ignite student curiosity and passion for science and math and foster interest in STEM. An additional benefit of these programs is that they provide participants with hands‐on inquiry driven laboratory experiences. One of the most popular programs offered is the week‐long Physiology and Biomedical Engineering program, which I have taught for the past several years. Stemming from a successful student activity in my Physiology course, the curriculum is designed to enable students to apply physiological concepts to real world applications using engineering techniques. Students are tasked with visualizing the relationships between the structure and function of the cardiovascular system through biomedical engineering ‐ the creation of heart models, which are designed, engineered, and built during the course of the week. This is followed by a discussion of the biophysics of the fluid dynamics of a closed circuit ‐ how pressure, thickness and resistance factor into blood flow. Although only the mechanical system of the heart is modeled due to time constraints, the electrical system is discussed in the context of the heart models built. Students also have the opportunity to model different pathological conditions in their heart models such as atherosclerosis and Crohn’s disease, and explain the biophysical, molecular and biochemical basis of their creations. Through model‐building and testing, students learn how to apply and integrate engineering principles and biophysics to understand the complex workings of the cardiovascular system.

Support or Funding Information

Center for Teaching and Learning, Illinois Math and Science Academy
Pharmacomicrobiomics of Hepatic Drug Processing Genes During Pregnancy
Authors: Han Lyrialle W. , Wang Lu , Shi Yuanyuan , Dempsey Joseph L. , Pershutkina Olesya V. , Dutta Moumita , Bammler Theo K. , Cui Julia Y. , Mao Qingcheng
Published: April 2020

At present, little is known about how gut microbiome and pregnancy interplay to regulate expression or the activities of drug processing genes (DPGs). The objective of this study was to investigate the effects of gut microbiome on host hepatic DPGs during pregnancy using conventional (CV) and germ‐free (GF) mice. Four groups of C57BL/6 female mice were used: CV non‐pregnant (CVNP, n=6), GF non‐pregnant (GFNP, n=6), CV pregnant (CVP, n=5), and GF pregnant (GFP, n=5) mice. Hepatic transcriptome and proteome for DPGs were profiled using multi‐omics approach. Plasma bile acids and steroid hormone levels were determined using LC‐MS/MS. Cyp activities were measured using hepatic microsome incubations. Overall, RNA‐seq analysis revealed a similar trend of gene regulation by pregnancy, but different magnitudes of change in hepatic DPGs, in CV and GF mice. Of note, the Cyp3a11 mRNA levels were decreased 1.7‐fold and 3.2‐fold by pregnancy in CV and GF mice, respectively. On the contrary, the Cyp3a11 protein abundance was increased 1.2‐fold and 1.6‐fold by pregnancy in CV and GF mice, respectively. The Cyp3a activity was also significantly induced by pregnancy in CV and GF mice; however, the magnitude of induction in CV mice was >5‐fold greater than that in GF mice. Plasma bile acid and steroid hormone levels were significantly impacted by microbiome and pregnancy‐status, respectively, which may contribute to the differential effects of pregnancy in CV and GF mice. This is the first study to demonstrate that the gut microbiome may play a role in altering hepatic drug disposition in pregnancy.

Support or Funding Information

This project was supported by the University of Washington Drug Metabolism, Transport and Pharmacogenomics Research Program (DMTPR)
Lysophosphatidic acid promotes lung carcinoma cell migration via ARHGEF17, a RhoGEF directly controlled by Gβγ
Authors: García Irving
Published: April 2020

Cell migration plays an essential role in multiple physiological processes, such as the immune response. However, this process contributes to pathological situations such as inflammation and metastatic cancer. During cell migration, cells reshape their actin cytoskeleton, a process finely regulated by Rho family GTPases (RhoGTPases). RhoGEFs activate RhoGTPases by promoting the exchange of GDP for GTP. Thus, RhoGEFs represent a crucial point between activation of chemotactic receptors and remodeling of the actin cytoskeleton. In this sense, several RhoGEFs have been characterized as important for cancer cell migration. Here, we demonstrate that RhoGEF‐ARHGEF17 is important for lung carcinoma cell migration stimulated with lysophosphatidic acid (LPA). In addition, pull down experiments, to directly assess GEF activation, demonstrated that ARHGEF17 is stimulated by LPA via Gi (sensitive to pertussis toxin) and by Gβγ. Mechanistically, Gβγ recruits ARHGEF17 to the plasma membrane and disrupts intramolecular inhibitory interactions. Our results indicate a possible participation of ARHGEF17 in the metastatic dissemination of lung carcinoma cells, a possibility under investigation.
Preconditioning Effects of Forced and Voluntary Running on Stress– and Indomethacin–induced Gastric Injury in Rats
Authors: Filaretova Ludmila , Komkova Olga , Punin Yuriy , Yarushkina Natalia
Published: April 2020

Physical activity and, especially, running, is natural stressor which is increasingly recommended as preconditioning for prevention of the number of diseases. Although regular physical activity can be beneficial in the prevention and the management of several diseases, the intensive exercise may promote or exacerbate the gastrointestinal injury. The aim of the study was to investigate preconditioning effects of forced and voluntary running on a vulnerability of the gastric mucosa to ulcerogenic action of cold‐restraint stress (3 h, 10°C) or indomethacin (IM, 35 mg/kg, sc). For forced treadmill (0 incline) running two protocols were used: high‐intensity (25 sm/s, 30 min) and moderate‐intensity (15 sm/s, 15 min) running. For voluntary running the rats were placed in the cages with wheels (2 h/day) during 4 days before experiment. On a Day of experiment preliminary fasted (24 h) rats were subjected forced running in treadmill or voluntary running in the wheel (2 h), then, they were given back in their cages and resting at room temperature for 1 h before the onset of ulcerogenic stimulus. Control animals were in their rcages, but they were also given ulcerogenic stimulus. Stress‐induced and IM‐induced gastric erosions were examined 3 h after the onset of cold‐restraint or 4 h after IM administration, respectively. A single high‐intensity treadmill session attenuated IM‐induced gastric erosions but aggravated stress‐induced gastric erosions. A single moderate‐intensity treadmill session didn’t influence IM or stress‐induced gastric injury, but moderate‐intensity treadmill in training regime (during 4 days before experiment) attenuated stress‐induced gastric erosions. Voluntary wheel running resulted in a decrease of IM‐induced erosions but an increase of stress‐induced gastric injury. The results obtained suggest that moderate‐intensity treadmill running in training regime exerts gastroprotective effect on stress‐induced gastric injury whereas by high‐intensity treadmill running as well as voluntary wheel running has proulcerogenic action in this model. At the same time high‐intensity treadmill as well as voluntary wheel running produced gastroprotective effect on IM‐induced gastric erosions. Thus, preconditioning effect of forced or voluntary running on the gastric injury is dependent on an intensity of treadmill running as well as a nature of ulcerogenic stimulus. We suppose that a significance of the compensatory gastroprotective action of glucocorticoids during IM‐induced prostaglandin deficiency which was demonstrated in our previous studies may explain the gastroprotective effects of the preconditioning in the IM ulcerogenic model.

Support or Funding Information

The study was supported by grant of Russian Science Foundation (RSF) N 19‐15‐00430.
Co‐expression and purification of a toxin‐antitoxin VapC–VapB complex from the E. coli O145:H28 strain RM12581 associated with a Romain lettuce outbreak.
Authors: Jones Seth Patrick , Goossen Christian , Lewis Sean , Delaney Annie , Gleghorn Michael
Published: April 2020

VapC and VapB are the toxin and anti‐toxin proteins, respectively, of the VapCB toxin–antitoxin system found in bacteria and associated with stunting cell growth during stressful conditions. VapC homologs typically are tRNA endonucleases, and VapB homologs are known to bind to double‐stranded DNA, as well as VapC to inihibit its endonuclease acitivty. No cognate binding and/or substrate partners have been identified for E. coli strain here VapC or VapB homologs. Our goal is to determine the X‐ray crystal structure of an E. coli O145:H28 strain RM12581 VapC complexed with antitoxin VapB or to any tRNA substrate that we are able to identify. I will present our efforts to produce soluble relevant proteins and/or complex thereof. A DNA co‐expression vector encoding both VapB and VapC mimicking its natural expression context was transformed into E. coli BL21 (DE3) competent cells and protein expression was induced. Proteins were purified using nickel affinity chromatography and size exclusion chromatography. Currently crystal screening is underway to crystallize relevant proteins for subsequent X‐ray diffraction analysis.