bluuebunny/crossref_metadata_2025 huggingface dataset:
The Deletion of AMPK β1 Worsens Glucose Tolerance and Liver Lipid Accumulation, but Doesn’t Impact the Metabolic Effects of Olanzapine in Male Mice
Authors: Shamshoum Hesham , Ashworth Kristen E. , Medak Kyle D. , Kemp Bruce E. , Wright David C.
Published: April 2020

Second generation antipsychotic (SGA) medications, like olanzapine (OLZ), are an important component in the pharmacological management of many conditions including schizophrenia. These drugs, have been linked with a neuroendocrine signaling axis that, leads to weight gain, however, peripheral mechanisms and predictors are currently poorly understood. 5’AMP activated protein kinase (AMPK) is a ubiquitously expressed, energy‐sensing enzyme and significant associations between polymorphisms in AMPK subunit genes and OLZ‐induced weight gain have been demonstrated. Conversely, others have suggested that the central activation of AMPK mediates the deleterious metabolic effects of SGAs. The purpose of this study was to determine if reductions in AMPK activity, primarily in the liver, would impact the chronic effects of OLZ treatment. We hypothesized that the absence of AMPK β1, a subunit which is primarily found in the liver, at least in mice, would worsen the metabolic side effects of OLZ. To address this question, we housed male AMPK β1 knockout (KO) and wildtype (WT) mice at thermoneutrality (29°C) and fed them either a control diet (HFD, 45% kcal from fat) or a diet supplemented with OLZ (HFD+OLZ, 50mg/kg diet) for 10 weeks. OLZ treatment did not increase food intake or weight gain to a greater extent than the control diet in either genotype. There was a main effect of the OLZ containing diet to increase heat production, TEE and fat oxidation during the light and dark cycles. The OLZ‐fed mice also demonstrated suppressed RER during the dark cycle. Interestingly, the OLZ‐fed KO mice were more physically active during the light cycle than the OLZ‐fed WT mice. The AMPK β1 KO mice were more glucose intolerant than the WT control mice. On the other hand, the AMPK β1 KO male mice on both diets demonstrated lower circulating serum glycerol and NEFA levels but increased liver TAG accumulation. In conclusion, our findings provide evidence that the absence of AMPK β1 does not potentiate the effects of OLZ on weight gain, but worsens liver lipid accumulation and glucose tolerance.

Support or Funding Information

This study was supported by Canadian Institutes of Health Research
Identification of the Gut Microbiome and its Role in Paradoxical Anaerobism in Desert Pupfish
Authors: Hillyard Stanley , Bhute Shrikant , van Breukelen Frank , Hedlund Brian
Published: April 2020

The Devils Hole pupfish (Cyprinodon diabolis) frequently enters paradoxical anaerobism (PA) in response to endogenously produced or exogenously supplied ethanol in a dose‐dependent manner. PA is characterized by extended periods (e.g. up to 149 min continuous) of negligible oxygen consumption despite its availability. To decipher the role of the gut microbiota in ethanol‐associated paradoxical anaerobism, gut microbial communities of pupfish were depleted using a cocktail of antibiotics. Compared to the fish in the control group, microbiota‐depleted fish spent more time in paradoxical anaerobism. Although the gut microbial communities with and without antibiotic treatment were similarly diverse, they were distinct and the greatest contribution to the dissimilarity (27.38%) was the common fish commensal Cetobacterium. Although Cetobacterium is known as a primary fermenter, we speculate that it may consume ethanol through secondary fermentation and therefore regulate transition from paradoxical anaerobism to aerobic respiration, thereby playing a critical role in regulating the metabolic status of these fish. This study reports the first characterization of gut microbial communities of critically endangered pupfish and suggests the microbiome may play a critical role in regulating metabolic strategies that are critical for survival in extremes of temperature and oxygen concentration. Given the wide distribution and abundance of Cetobacterium in warm‐water fishes, this process may be of broad importance, and suggests that the microbiome be carefully considered for conservation and aquaculture alike.
Promoting Cultural Capitals and Self‐Determination Through Reflective Journaling in First‐Year Biology Labs
Authors: Tran Khanh , Coats Victoria Buenconsejo , Pelaez Nancy
Published: April 2020

First‐year biology students have preconceived notions of what it’s like studying biology. However, these dominant narratives can hinder students’ success and their pathway into the STEM field. Here we had students use reflective journaling as a form of counter‐narrative, informed by the Alma Project that aims to increase students’ persistence and sense of belonging in STEM. In the intervention, students enrolled in a first‐year biology laboratory were asked to do journalling for five minutes four times during the semester, to reflect on their personal experiences. Then, students spent another five minutes “sharing their stories” with their lab group and peers. Analysis of reflective essays, informed by the Community Cultural Wealth framework, identified 11 cultural capitals utilized by first‐year biology students: aspirational, attainment, community consciousness, familial, filial, first‐generation, navigational, perseverance, resistant, social, and spiritual. Moreover, preliminary data suggest that reflective journaling may also allow expression of students’ self‐determination. With these implementations, biology students engaged in meaningful reflections that identified their cultural wealth which may help them stay focused on their path to a STEM degree.
Concentration‐Time profiles of Glyburide in Rats: Predictions using a Continuous Absorption Model
Authors: Radice Casey , Korzekwa Ken , Nagar Swati
Published: April 2020

Most drugs are administered orally due to cost‐efficacy and increased patient compliance. However, the absorption of these drugs is subjected to complex biological processes and depends on the drug’s physicochemical properties. Prior to human trials, preclinical species such as rodents are utilized to evaluate efficacy and safety in vivo. Because animal studies are expensive and time‐consuming, it is beneficial to predict a drug’s pharmacokinetic profiles prior to in vivo studies. Pharmacokinetic modeling and simulation is useful for such predictions.

The small intestine and colon of male Sprague Dawley rats were divided into 10 unequal segments to measure the change in luminal pH. Luminal contents were collected and directly measured under fed (n=4) and 12‐hour fasted (n=4) conditions. Glyburide is an oral anti‐diabetic medication that stimulates insulin secretion by blocking ATP‐dependent potassium channels in pancreatic beta cells. In vivo pharmacokinetic studies were performed to determine the effect of food. Fasted (n=4) and fed (n=4) male Sprague Dawley rats were dosed 5mg/kg of glyburide by oral gavage. Rats (n=2) were dosed intravenously to collect pharmacokinetic parameters. Blood samples were collected at 0.1, 0.25, 0.5, 1, 2, 4, 6, 10, and 24 hours post dose. Data were analyzed with non‐compartmental analysis. A continuous rodent absorption model was developed to predict the concentration‐time profiles of compounds based on rodent anatomical data collected in‐house and from the literature. Several physiological parameters were incorporated in the model, including small intestinal transit time, luminal pH, and transporter expression along the intestines.

Rats fasted for 12 hours had a higher pH along the intestine that increased slowly, compared to fed rats. While the pH of the colon was higher in fasted rats, the cecum was not altered by fasted conditions. Fasted rats had an average glyburide AUC(0–24) of 2.7 μg*h/ml, while fed rats had an AUC(0–24) of 0.97 μg*h/ml. The mean apparent clearance (CL/F) was 1.8 L/min/kg for fasted rats and 5.1 L/min/kg for fed rats. The mean half‐life increased slightly from 10.6 hours in fasted rats to 11.6 hours in fed rats. Current predictions using the rodent continuous absorption model do not accurately describe the exposure profile of glyburide. To refine the model, further anatomical data collection and incorporation of fasted versus fed conditions are underway.

Support or Funding Information

This work was supported by NIH grants R01GM114369 and R01GM104178 to Swati Nagar and Ken Korzekwa.
TRPML channel activation partially rescues Ca2+ spark activity in sheep fetal pulmonary arterial myocytes following intrauterine long‐term hypoxia
Authors: Ang Dylan , Romero Monica M. , Puglisi Jose L. , Zhang Lubo , Wilson Sean M.
Published: April 2020

Ca2+ sparks result from the activation of ryanodine receptors located on the sarcoplasmic reticulum of pulmonary arterial smooth muscle cells. These Ca2+ sparks activate potassium channels on the plasma membrane and are important to arterial vasodilation and increases in vascular blood flow. Previous studies showed that long‐term high altitude hypoxia restricted membrane depolarization elicited Ca2+ spark activity in fetal lamb pulmonary arterial myocytes, but had little effect on pulmonary myocytes in adult sheep. Moreover, the studies also indicated that there was preservation of the sarcoplasmic reticulum Ca2+ stores in pulmonary arterial myocytes of both fetuses and adults as determined through direct activation of ryanodine receptors with caffeine. Given that the sarcoplasmic reticulum Ca2+ stores are intact and have functional ryanodine receptors we hypothesize that aberrations to the canonical membrane depolarization Ca2+ spark triggering pathway in fetal pulmonary arterial myocytes due to intrauterine hypoxia can be resolved through activation of alternate pathways. Recent studies illustrated that Ca2+ sparks were triggered in various vascular myocytes by releasing Ca2+ from lysosomes through activation of TRPML (transient receptor potential cation channel, mucolipin subfamily) channels. To evaluate the potential that TRPML activation may elicit Ca2+ sparks in fetal and adult pulmonary arterial myocytes we studied isolated pulmonary arteries from near term fetal and adult sheep from low altitude (700 m) or those that had been exposed to high altitude (3801 m) for 110+ days. Ca2+ sparks were recorded using laser scanning confocal microscopy while 10 μM MLSA1 was used to activate TRPML channels. MLSA1 enhanced the activity of Ca2+ sparks in myocytes of adult pulmonary arteries to a greater extent than those from fetuses, suggesting developmental changes in the communication of lysosomes with the sarcoplasmic reticulum. Of importance, TRPML activation by MLSA1 partially recovered the aberrant Ca2+ spark activity in fetal pulmonary arterial myocytes caused by hypoxia. The findings suggest that ryanodine receptor activation through pathways alternate to membrane depolarization may hold promise for improving pulmonary vascular function following intrauterine hypoxic exposure.

Support or Funding Information

Imaging was performed in the LLUSM Advanced Imaging and Microscopy Core with support of NSF Grant MRI‐DBI 0923559 (SMW) and the Loma Linda University School of Medicine. Additional support provided through NIH grant HD083132 (LZ).
Illuminating Spatiotemporal Regulation of AMPK with a Genetically Encoded Excitation‐Ratiometric Biosensor for AMPK
Authors: Schmitt Danielle L. , Zhang Jin-fan , Chen Mingyuan , Leung Allen , He Catherine Y. , Curtis Stephanie D. , Mehta Sohum , Shaw Reuben J. , Rangamani Padmini , Zhang Jin
Published: April 2020

AMP activated protein kinase (AMPK) is a heterotrimeric protein responsible for moderating cellular energy homeostasis. In cancer, AMPK has been implicated as a context‐dependent tumor promoter or suppressor. To illuminate spatiotemporal regulation of AMPK, which is critical for context dependent roles of AMPK, we developed a single‐fluorophore excitation‐ratiometric AMPK activity reporter (ExRai AMPKAR), which exhibits greater sensitivity and dynamic range compared to our previous AMPK biosensor. We used pharmacological perturbations to activate AMPK through both nucleotide‐dependent (2‐deoxyglucose) and ‐independent (MK‐8722) mechanisms, finding context‐dependent AMPK activity throughout the cell and fast AMPK activity dynamics around membranous organelles. A 3D computational model suggests high AMPK activity around organelles such as the lysosome are due to active transport across the lysosomal membrane. Furthermore, we observed robust AMPK activation in the nucleus following various pharmacological stimulation. To further investigate the mechanism regulating nuclear AMPK activity, we used fluorescence recovery after photobleaching experiments which showed that a catalytic subunit of AMPK, AMPK‐2, had no obvious change in subcellular location following pharmacological stimulation. Nuclear AMPK activity is enhanced upon LKB1 expression in LKB1‐null cells, indicating this important upstream regulator of AMPK is involved in nuclear AMPK activity. Altogether, using ExRai AMPKAR with complimentary computational and biophysical methods, we have characterized subcellular AMPK activity around membranous organelles, and unveiled a resident population of AMPK in the nucleus, which could be acutely controlled to provide fast regulation of transcription. Beyond the current study, our new AMPK activity reporter has the potential to provide further insight into localized AMPK activity in many cellular contexts.
Molecular Determinants of Signaling and Posttranslational Modifications of GPR64
Authors: Gad Abanoub A. , Azimzadeh Pedram , Balenga Nariman
Published: April 2020

Some of the adhesion G protein‐coupled receptors (aGPCRs) show constitutive activity in the absence of their large N‐terminal fragment (NTF). We previously demonstrated that GPR64/ADGRG2, an orphan aGPCR, regulates the secretion of parathyroid hormone by interacting with the calcium‐sensing receptor and elevating the cAMP levels in parathyroid adenoma cells. In HEK293 cells, we described the role of beta‐arrestins, dynamin, and GPCR kinases (GRKs) in the signaling of NTF‐truncated GPR64. Whether structural domains and motifs other than the NTF also regulate signaling, trafficking, and posttranslational modifications of GPR64 remains unknown. In the current study, we mutated a number of extracellular motifs that are conserved among zebrafish, mouse, and human GPR64. We measured cAMP production (time‐resolved FRET), surface expression (ELISA and biotinylation assays), and ubiquitination (co‐immunoprecipitation) of WT and mutant GPR64 in HEK293 cells. We discovered that several residues in the extracellular domains alter the surface expression, ubiquitination, and degradation of GPR64 at basal and tethered agonist‐stimulated conditions. The molecular mechanisms by which these residues regulate trafficking, signaling, and ubiquitination of GPR64 are currently under investigation.

Support or Funding Information

This work was supported by NIH grant 1R01GM13061701A1 (to N.B.). Student support was provided by an NIH‐NIGMS Initiative for Maximizing Student Development Grant (R25‐GM55036) (to A.A.G.).
Structure‐Based Insights into Control of Heme Biosynthesis
Authors: Brown Breann L. , Kardon Julia R. , Baker Tania A.
Published: April 2020

Protein assembly drives many aspects of cellular and mitochondrial physiology. We are interested in uncovering and characterizing the protein interactions that support proper heme biosynthesis. Heme is a critical biomolecule that carries out several functions in nearly all life forms, including humans where its most widely‐known role is mediating oxygen transport in blood. It is imperative that heme production is tightly controlled as alterations in cellular heme levels can have drastic consequences for human health. The first and rate‐limiting enzyme controlling heme biosynthesis is aminolevulinic acid synthase (ALAS). ALAS is conserved in α‐proteobacteria and non‐plant eukaryotes, there are two isoforms in vertebrates, with the mitochondrial protein ALAS2 being responsible for heme synthesis during erythropoiesis. There are two separate diseases that may result from one of over 50 mutations in the ALAS2 gene. Mutations decreasing optimal activity lead to the disease X‐linked sideroblastic anemia (XLSA), whereas other mutations causing hyperactivity underlie X‐linked protoporphyria (XLPP). Importantly, several disease‐causing mutations are located in a eukaryote‐specific C‐terminal extension, a region absent from bacterial ALAS enzymes. Certain mutations also result in an inability of ALAS2 to assemble with other proteins, including proteins involved in separate metabolic pathways. However, we lack an understanding of how these mutations lead to a change in ALAS2 structure, and therefore, function. We seek to understand how ALAS interacts with accessory proteins as well as organism‐specific differences in assembly that may alter regulation of heme production. Therefore, we use X‐ray crystallography combined with in vitro activity assays of various eukaryotic ALAS enzymes to parse apart the role of this key regulatory region in eukaryote ALAS function. Our work is beginning to reveal key structure‐function relationships between the orientation and molecular contacts mediated by the C‐terminus and overall ALAS enzyme function, thus controlling heme biosynthesis.

Support or Funding Information

Jane Coffin Childs Memorial Fund for Medical Research Award and a Burroughs Wellcome Postdoctoral Enrichment Program Fellowship Award 1015092 (B.L.B); NIH Ruth L. Kirschstein National Research Service Award (F32DK095726) (J.R.K.); National Institutes of Health Grant (R01 DK115558) (T.A.B.), and the Howard Hughes Medical Institute. X‐ray data collected at NE‐CAT (P41 GM103403).
CaMKII is Necessary for Proliferation and Migration of Pulmonary Arterial Smooth Muscle Cells
Authors: Huetsch John , Jiang Haiyang , Shimoda Larissa
Published: April 2020

Objective

Ca2+/calmodulin‐dependent protein kinase II (CaMKII) is a calcium‐ and oxidation‐activated serine/threonine kinase, with four known isoforms. CaMKII has been shown to be necessary for remodeling involving smooth muscle cell proliferation and migration in response to injury in the systemic vasculature. However, the role of CaMKII in pulmonary arterial smooth muscle cell (PASMC) function remains unknown.

Methods

PASMCs were isolated from male Wistar rats and expanded in culture. In cell lysates, mRNA expression of CaMKII isoforms was measured by real‐time quantitative reverse transcription PCR; protein expression was assessed via immunoblot. PASMC proliferation was measured via BrdU incorporation. PASMC migration was assessed by the Transwell assay. PASMCs were treated with 10 μM KN‐93 in vitro to inhibit CaMKII or with either vehicle or 10 μM KN‐92 (an inactive analog of KN‐93) for control experiments. To deplete CaMKIIδ protein, PASMCs were treated with 100nM siRNA targeted to CaMKIIδ while PASMCs treated with scrambled siRNA were used for control experiments.

Results

The CaMKIIδ and CaMKIIγ isoforms showed mRNA and protein expression in rat PASMCs. Both proliferation and migration of rat PASMCs were inhibited by pharmacologic inhibition of CaMKII using KN‐93, but were not inhibited by the inactive analog KN‐92. siRNA‐mediated knockdown of CaMKIIδ resulted in reduction of CaMKIIδ protein expression but did not have an effect on either PASMC proliferation or migration.

Conclusions

CaMKII activity is necessary for PASMC proliferation and migration. As the CaMKIIδ isoform is not required for these functions, future work will examine the role of the CaMKIIγ isoform.

Support or Funding Information

Funding Sources

NIH K08HL133475 & Pulmonary Hypertension Association K08 Supplement
The Effects of In Vitro Seizure‐like Activity on Structural Plasticity of Dendritic Spines
Authors: Nieder Cassidy , Ramon Alfonsina , Getzfread Mollie , Lipmann-Bell Jocelyn
Published: April 2020

Most excitatory synapses in the mammalian brain connect to dendritic spines. These membranous protrusions function as chemical and electrical micro‐compartments, transducing individual signals of excitatory neurotransmitters from axonal boutons. Structural plasticity, the ability of spines to change their shape and thus their function in response to synaptic activity, underlies learning and memory. This shape change requires regulation of actin mesh‐works, which are regulated by actin binding proteins (ABPs) within spines. Decreased concentrations of functional ABPs induce abnormalities in the density, morphology, and dynamic nature of dendritic spines. Early‐life seizure models demonstrate altered spine morphology, and a decrease in ABPs hinders synaptic signal amplitudes. Further, neonatal seizures are linked to cognitive deficits later in life, including autistic‐like behaviors, and patients with autism spectrum disorder express depleted concentrations of endogenous ABPs and exhibit dendritic spine dysgenesis. Based upon these findings, we hypothesized that seizure‐like activity causes deficits in the structural plasticity of dendritic spines via depletion of functional ABPs. To investigate the physiological consequences of uncontrolled, synaptic excitability on intracellular actin regulation at the molecular level, we induced in vitro seizure‐like activity in primary, rat hippocampal neurons using a zero magnesium model. At multiple time points after this event, we assessed localization of ABPs using immunocytochemistry, actin turnover employing fluorescent recovery after photo bleaching, and spine and actin dynamics in response to stimulation utilizing live‐cell imaging in GFP‐actin transfected neurons. Preliminary results show a decrease in localization of ABPs to dendritic spines and an increase in spine motility after in vitro seizure‐like activity in cultured hippocampal neurons. Taken together, these data indicate that seizures may alter ABPs in a model that also shows changes in spine dynamics and structural plasticity. Ongoing work will examine this link further to determine if ABPs could be a possible therapeutic target to decrease cognitive deficits that can result from early‐life seizures and lead to intellectual disabilities.

Support or Funding Information

CCDA: Center for Chronic Disorders of Aging
Training Students on Effective Small Group Dynamics Impacts Student Behavior and Learning Outcomes
Authors: Singh Atul K.
Published: April 2020

PURPOSE

Schools have invested significantly in training faculty to become proficient in developing and facilitating student small group learning. However, small group approach continues to receive mixed reviews, and assessing effectiveness of this type of learning experimence is challenging. An important factor that may be overlooked is lack of student training in this methodology. The aim of this study was to determine if training students on effective small group dynamics would result in changes in student behavior and attitude as well as improved outcomes.

METHODS

IRB approval was obtained to engage students in three activities: 1) training session focused on effective small group dynamics; 2) audiotape recording of small group interactions during two learning sessions prior to training (Pre), and two sessions following training (Post); and 3) survey of student attitudes and perceptions. Training involved a one‐hour session focused on the students’ responsibility for success in small group, nature of high performing individuals in group work, self‐identification of group participant type (initiator, over participant, blocker, deserter, etc.), patterns of effective communication in assent and dissent, sharing roles within the group, setting guidelines and best practices for the group, and reflection and assessment of group effectiveness. For the recorded small group sessions, three trained evaluators independently used an original rubric to assess each recorded session. Rubrics scored various items including participation, role playing, time on task, identification of learning objectives, and patterns of effective communication. Rubric scores were averaged for each session both pre and post training. Additionally, a 15‐question, Likert scale survey was developed to assess student attitudes. This survey was administered three times to each student: 1) prior to participation in any small group activity; 2) following the two Pre small group sessions, and immediately prior to training; and 3) immediately after recording the Post small group sessions. Survey items gathered student perceptions related to preparation, peer interactions, and general effectiveness of the activity.

RESULTS

Preliminary analysis of audio recordings suggest that following training, students participated more often and more equally. Students also encouraged and requested information from peers more frequently. However, although students were instructed to vary roles and focus on stated objectives, little change was noted in these areas when comparing Pre to Post sessions. Student survey responses also revealed that general enjoyment increased, students felt more empowered to contribute, and were able to avoid tangential discussions more effectively following training.

CONCLUSION

This study indicates that training students on effective small group dynamics influences behaviors and perceptions that impact student enjoyment and learning success.

Support or Funding Information

No funding to report
Comparing Muscular System Coverage in Undergraduate Human Anatomy and A&P Courses at Community Colleges and Four‐year Universities
Authors: Reynolds Amberly , O'Loughlin Valerie , Goodwin Michael
Published: April 2020

Anatomical content coverage across undergraduate human anatomy and A&P courses is widely variable. A previous study by Saladin (2008) assessed muscle coverage (n~20) via self‐report in response to a listserv question. In response to the need for a more global understanding of the content taught in undergraduate anatomy courses, we prepared a muscular system survey where human Anatomy (or Anatomy & Physiology) faculty could report the extent to which they discuss skeletal muscles in these classes. The survey also collected demographic institutional data such as the type of institution (community college versus 4‐year), course format, and geographic location of the undergraduate institution, so comparisons could be made. The muscular system coverage survey was created and administered to participants online via Qualtrics. Muscles that were surveyed were those listed and described in a typical undergraduate human anatomy text (Human Anatomy, 5th edition textbook by McKinley, O’Loughlin, and Pennefather‐O’Brien (2017)). Participants were recruited via a poster presentation at HAPS 2018 and recruitment emails sent out through AAA, HAPS and AACA listservs. The survey has been examined and the study deemed exempt by Indiana University IRB ‐ Protocol # 1805378498. All data was de‐identified prior to analysis. Presented at EB 2019 were three posters detailing the general trends in muscle system coverage across all participating courses, comparisons between anatomy only and anatomy & physiology courses, and comparisons between international and U.S. courses. In response to questions posed at the posters last year, this analysis dives into comparisons of muscular system coverage trends between community college (n~90) and four‐year university (~130) Anatomy (and A&P) courses. In general, there are more similarities in content coverage than differences. Yet one difference found shows that across most regions (excluding muscles of respiration and abdominal wall), community college Anatomy and/or A&P courses use identification and action content coverage independently, whereas university courses assess all specifics (identification, action, attachment, and innervation. These similarities and differences in muscular system content coverage can expand the conversations on and increase collaborations between course instructors of Anatomy and A&P courses.
The Role of Pyruvate Kinase Isoform M1/2 in Colorectal Cancer
Authors: Park Bohye , Kim Ji Yeon , Crumbley Christopher , Wirthwein Kaci , Riffey Olivia , Beaird Jacob , Smith Mary , Donohoe Dallas
Published: April 2020

Colorectal cancer cell (CRC) increases the utilization of glucose and decreases the oxidation of butyrate, which is a primary energy source for non‐cancerous colonocytes. Butyrate is a short‐chain fatty acid that is a bacterial‐derived fiber fermentation product. It takes years for the colorectal adenoma to manifest as it progresses toward invasive carcinoma. Within this timeframe, the adenoma must acquire key genomic and metabolic modifications that promote tumor growth and progression. Understanding the metabolic shift mechanisms away from butyrate oxidation to glycolysis in the CRC is significant. The overexpression of the pyruvate kinase isoform M2 (PKM2) is observed in biopsies of colorectal cancer patients, which is a hallmark of cancer. Pyruvate kinase is a metabolic enzyme that catalyzes the last step in glycolysis, which converts phosphoenolpyruvate to pyruvate generating ATP. PKM1 is generally expressed in adult differentiated tissues, such as normal non‐cancerous colonic tissue, where it promotes oxidative metabolism. In contrast, PKM2 is expressed in proliferating cells, including colorectal cancer cells, where it enhances aerobic glycolysis. However, the function of PKM2 in colorectal cancer tumorigenesis remains controversial. Our preliminary studies suggest that PKM2 drives colorectal metabolism away from butyrate and toward glucose utilization. A stable knockdown of PKM2 in colorectal cancer cells increased the oxidation of butyrate. The major hypothesis for this project is that through enhancing glucose utilization and suppressing butyrate, utilization PKM2 promotes colorectal tumor progression. Here, we demonstrate that, when expressed rather than PKM2, the PKM1 increases butyrate oxidation in cancerous colonocytes.

Support or Funding Information

USDA R011770176
Purification of a Plant Lectin from Kohlrabi/Brassica oleracea var. gongylodes and Identification of Binding Specificity
Authors: Vue Mary , Rydberg Melinda , Talaga Melanie L.
Published: April 2020

Plant lectins are proteins that agglutinate cells. They are involved in cancer detection, cell toxicity, and cell growth inhibition. Kohlrabi or Brassica oleracea var. gongylodes is a vegetable that has been associated with a trypsin inhibitor, antifungal peptides, and proteins that have potential anti‐diabetic, anti‐inflammatory, antioxidant properties. A lectin from the same plant species (Brassica oleracea Italica x Alboglabra) was found to have anticancer effects. To our knowledge, we are the first to observe lectin activity from this subspecies known as kohlrabi.. The protein was extracted from kohlrabi cabbage and leaves with Phosphate Buffered Saline (PBS) pH 7.4. Hemagglutination assay was used to detect lectin activity in the crude extraction using 2% (v/v) sheep erythrocytes in PBS. The lectin was purified using size exclusion chromatography Sephadex G‐75. Column fractions were tested for presence of protein by using a UV spectrophotometer at 280 nm and testing for presence of lectin activity using 2% (v/v) sheep erythrocytes. The molecular weight of the lectin was determined using SDS‐PAGE Electrophoresis. Plant lectins bind to specific carbohydrates; the binding specificity of this new plant lectin was identified and in future research the medical application of this lectin can be investigated further.
Amplification of Love bugs (Plecia nearctica) Fungal Community DNA using the ITS86F and ITS4 primers
Authors: Grant Caylin Marie , Stamper Fre’Keliah Breana , Haggins Danielle Keyandra , Sakura-Lemessy Adia Issa , Okafor Uzoma
Published: April 2020

Plecia nearctica better known as love bugs or honeymoon flies are species of march fly that originated from parts of Central America and the Southern eastern United States. This research is aimed at investigating the selectivity of using the ITS86F and ITS4R, in amplifying fungi community on lovebugs and sex differentiation (male or female). Libraries will be constructed from PCR‐amplified fragments, sequenced and compared against sequences deposited in GenBank. The results will show the specific selectivity of the following fungal: Ascomycetes, Basidiomyces and Zygomycetes. Amplified products generated by the ITS86F and ITS4, which will be aligned with a range of fungi species. This will be the first reported use and assessment of the ITS86F and ITS4, primers in amplifying fungal community from love bugs.

Support or Funding Information

Albany State University STEM Center for Undergraduate Research
Structure of the E6/E6AP/p53 Complex in Human Papillomavirus
Authors: Fassler Amy , Bhat Tara , Rens Davis , Chojnacki Kennan , Neil Uma
Published: April 2020

Human Papillomavirus (HPV) is the most common sexually transmitted pathogen in humans and the primary cause of cervical cancer. The tumor suppressor p53, in its normal state, regulates cell growth and survival through mechanisms including DNA repair and apoptosis. The E6 oncoprotein directly inhibits p53s function by attaching itself to a ubiquitin ligase, E6AP, which in turn forms a complex with p53. E6 and E6AP form a heterodimer by binding at a leucine‐rich docking site, known as the LxxLL motif. The heterodimer then forms a binding pocket with p53, which is the cause of the inactivation of p53 function. Researchers are attempting to identify potential binding pocket function in the complex in order to form a protein inhibition mechanism that can block further proliferation of cervical cancer. Understanding the interactions of the E6/E6AP/p53 complex is leading to the development of unique antiviral and chemotherapeutic agents for cervical cancer. The Marshfield High School MSOE Center for BioMolecular Modellng for SMART Team used 3D modelling and printing technology to examine the structure‐function relationships of the E6/E6AP/p53 complex.

Support or Funding Information

Marshfield Clinic LaboratoriesMSOE Center for BioMolecular ModelingMarshfield Area Community Foundation
Blood Pressure, Vascular Reactivity, and SOD Expression/Activity in Mas1 Receptor Knockout Rats in the Dahl Salt‐Sensitive Genetic Background
Authors: Zheleznova Nadya N. , Terashvili Maia , Bullis Jack , Stumpf Megan , Nguyen Linh , Geurts Aron , Lombard Julian H.
Published: April 2020

Dahl salt‐sensitive (SS) rats are the most widely used rodent genetic model of human salt‐sensitive hypertension. Earlier studies have established the importance of impaired regulation of the renin gene in contributing to endothelial dysfunction in SS rats and the protective effect of low dose angiotensin II (ANG II) infusion in restoring endothelial function via the ANG II type1 receptor (AT1R) in the SS genetic background. Activation of the Mas1 receptor (Mas1R) for angiotensin (1‐7) [ANG (1‐7)] generally opposes ANG II‐mediated effects of AT1R activation. In Sprague‐Dawley rats fed high salt diet, binding of ANG (1‐7) to the Mas1R can activate NRF2 defenses and restore impaired endothelial function. The goal of the present study was to provide initial phenotyping of a Mas1R knockout (Mas1R KO) rat in the Dahl SS genetic background. Mas1R KO rats and wild type (WT) controls (12–15 weeks old) were maintained on a low salt (LS; 0.4% NaCl) or switched to a high salt (HS; 4% NaCl) diet for 3 days or 10 days. Arterial pressure measured via telemetry was similar in Mas1R KO rats (121±7 mmHg, n=3) and WT rats fed LS diet (124 mmHg, n=2) and increased in WT (138 mmHg, n=2) and Mas1R KO rats (132±5 mmHg, n=3) fed HS diet for 10 days. In rats fed LS diet, Mas1R KO eliminated vasodilator responses to ANG (1‐7) and AVE0991 (Mas receptor agonist) in middle cerebral arteries (MCA) and mesenteric resistance arteries (MRA) and reduced acetylcholine (ACh) reactivity in MCA. Because superoxide dismutase (SOD) enzymes contribute to antioxidant defense by catalyzing the dismutation of superoxide radicals and help to prevent diseases related to oxidative stress and endothelial dysfunction, we measured superoxide dismutase 1 (SOD1) and superoxide dismutase 2 (SOD2) activity in the liver of WT and Mas1R KO rats after 3 days of HS diet. High salt diet increased SOD1 activity (WT‐LS 1.04±0.16 U/ml vs WT‐HS 1.65±0.12 U/ml) and SOD2 activity (WT‐LS 0.76±0.03 U/ml vs WT‐HS 1.06±0.11 U/ml) in WT rats, but not in Mas1R KO rats. SOD1 and SOD2 mRNA expression (evaluated via RT‐PCR) tended to increase in WT rats fed HS diet vs. LS diet (1.3 – 1.8 fold change, p > 0.05). SOD1 mRNA expression was significantly higher in HS‐fed Mas1R KO rats vs. Mas1R KO rats fed LS diet (2.8 fold change, p = 0.01) and SOD2 mRNA tended to increase as well (1.7 fold change; p = 0.1). SOD1 and SOD2 protein expression measured by Western blot analysis was similar in WT and Mas1R KO rats fed either LS or HS diet. The present study using the Mas1 receptor knockout in the Dahl SS rat genetic background suggests that Mas1R activation by its agonists (AVE 0991) and ANG (1‐7) improves endothelial function and may provide novel therapeutic strategies for the treatment of low‐renin salt‐sensitive hypertension.

Support or Funding Information

NIH #HL‐128242 and NIH #R21‐OD024781
Visualization of Stereoscopic Model of the Liver and Gallbladder from Clinical Perspective
Authors: Yang Gongchao , Garza Andrew W. , Sinning Allan R. , Cui Dongmei
Published: April 2020

The hepatobiliary system is the most common site for many diseases, and anatomy of this region could be complicated. Laparoscopy surgery has now become the preferred option for cholecystectomy and many other gastrointestinal conditions. These procedures require advanced laparoscopic techniques and anatomical knowledge of the region. Understanding and familiarizing with the anatomical relationship of the liver and gallbladder will help clinicians in the evaluation and management of such diseases as gallbladder stones, bile duct stones, pancreatic duct stones, and liver cancers, etc. The development of 3D stereoscopic models usually requires the acquisition and conversion of voxel‐based CT or MRI data into 3D models. It has been reported recently that the computed tomography angiography (CTA) was used to create vascular models using volume rendering, and surface rendering for educational purpose. CTA images have been extensively used for vasculature anatomy, detection and evaluation of intracranial aneurysms, and diagnosis of ischemic stroke, however, the CTA data has not been well explored for creating a hepatobiliary and pancreatic stereoscopic virtual model in contemporary anatomy education. In this study, we utilize CTA data to develop a stereoscopic liver, gallbladder and pancreas model using the semi‐auto combined rendering in addition to the volume and surface rendering methods. The visualization of the liver, associated with other organs and their relationship can be viewed via the stereoscopic presentation. The two‐dimensional (2D) and 3D measurements can be applied to measure the length, angle, and volume of the liver. The stereoscopic visualization and comparison of the measurement might have potential benefit for helping students and clinicians to better understand anatomy knowledge in medical education and clinical training. The stereoscopic view of surgical field might as well enhance the surgeon’s dexterity.
Rapid Accumulation of Triacylglycerol in the Liver is a Component of the Acute Exercise Response and May Serve a Protective Role Against Lipotoxicity
Authors: Henderson Gregory C. , Tuazon Marc A.
Published: April 2020

We have previously shown that the content of triacylglycerol (TAG) in the liver transiently increases after a single bout of exercise in untrained mice, and this phenomenon has been observed in human subjects as well. This discovery depicted a new phenomenon in the metabolic exercise response. Here we propose the term “lipogenic flexibility” to describe this ability to rapidly alter hepatic TAG content in response to FFA supply. To explore lipogenic flexibility after exercise, we performed lipidomics analysis on liver samples obtained 3 hours after exercise and the following day. Female mice performed a single bout of continuous exercise (CE), high intensity interval exercise (HIIE), or no exercise (CON). The total concentration of hepatic TAG rose 3 hours after exercise (P < 0.05 for both CE and HIIE vs CON), but the total concentration of all other measured lipid classes remained unchanged after exercise (NSD). Hepatic TAG declined back to control levels by the day after exercise. We performed correlation analysis to test the association of TAG (an inert lipid depot) with lipotoxic lipid classes in the liver. Hepatic TAG was positively correlated with hepatic diacylglycerol (DAG) in CON (P < 0.05); however, during the hepatic TAG pool expansion after exercise, liver TAG and DAG were no longer significantly correlated with one another (NSD). Exhibiting lipogenic flexibility after exercise appears to break the equilibration between DAG and TAG, allowing fatty acids to be drawn toward the inert lipid pool (TAG) and away from the lipotoxic lipid pool (DAG). The results suggest that the TAG pool actively expands to buffer elevated FFA supply after exercise and may act to prevent lipotoxicity.

Support or Funding Information

This work was supported by the Faculty Research Grant Program of Rutgers University, the American Diabetes Association grant # 7‐13‐JF‐27‐BR to G Henderson, and the Purdue University College of Health and Human Sciences.
The Effect of Menopause Status on the Skeletal Muscle Metaboreflex.
Authors: villarraga nicolas , Smith Joshua R. , Randall Nicholas R. , Bruhn Eric , Olson Thomas P.
Published: April 2020

Introduction

It is well established that the exercise pressor reflex contributes to cardiovascular control (e.g. mean arterial pressure (MAP)) during exercise. One arm of the exercise pressor reflex includes the metabolically sensitive group IV afferents (i.e. metaboreflex). The metaboreflex‐induced cardiovascular responses to exercise are incompletely understood, especially in women across the aging spectrum. For example, post‐menopausal women have greater increases in MAP and systemic vascular resistance (SVR) during locomotor muscle exercise compared to pre‐menopausal women. However, it is unclear if an exaggerated metaboreflex arising from the locomotor muscles is contributing to these cardiovascular adjustments with exercise in post‐menopausal women. We hypothesized that 1) post‐menopausal women will have an augmented MAP response with isolated metaboreflex activation compared to pre‐menopausal women and 2) the exaggerated increase in MAP in the post‐menopausal women will be primarily due to greater increases in SVR.

Methods

Pre‐ (n= 10, 25±3yrs, 24±3 kg/m2) and post‐menopausal women (n=12, 58±4yrs, 24±3 kg/m2) performed 2 sessions of cycling exercise at 20 W for 5 min with bilateral upper thigh pressure tourniquets inflated to 90 mmHg. At end exercise, the recovery was randomized to post‐exercise circulatory occlusion (PECO) to isolate the metaboreflex (via bilateral upper thigh pressure tourniquets rapidly inflated to suprasystolic pressure for 2 min) or normal recovery (NR) without intervention. Systolic and diastolic blood pressure were measured via manual sphygmomanometry and MAP was calculated. Cardiac output was measured via open circuit acetylene wash‐in and systemic vascular resistance (SVR) was calculated. Data is reported as the absolute change from rest to recovery.

Results

MAP was elevated during PECO compared to NR in both groups (p<0.05), while MAP during PECO was greater in post‐ compared to pre‐menopausal women (20±9 vs. 13±5 mmHg, respectively) (p<0.05). Similarly, SVR was elevated during PECO compared to NR in both groups (p<0.05), while SVR was greater during PECO in post‐ compared to pre‐menopausal women (5±3 vs. 3±1 mmHg/L/min, respectively) (p<0.05). In contrast, cardiac output was not different between NR and PECO in either pre‐ (NR: 0.3±1 vs. PECO: −0.2±1 L/min) and post‐menopausal (NR: −0.1±1 vs. PECO: −0.7±0.9 L/min) (all, p>0.05).

Conclusion

These data demonstrate that isolated metaboreflex stimulation following whole‐body exercise results in exaggerated increases in MAP and SVR in post‐ compared to pre‐menopausal women. These data suggest that exercise pressor reflex‐mediated control of MAP during exercise is accentuated in post‐menopausal women with menopause status.
Association between dietary parameters and cognitive functions in first year undergraduate Health Sciences students
Authors: Ortíz-Cabrera Guadalupe , Estrada José Antonio , López-Meza Ma. Sagrario , Sánchez-Delgado Marcela , Contreras Irazú
Published: April 2020

Adolescence is a stage characterized by emotional, psychological, physical and social changes. When entering the university, it is common for adolescents to change their lifestyles along with their eating habits due to increased coursework, homework and socializing factors. These issues lead them to have less control over quotidian things such as diet quality and sleep. Unhealthy diets are a well‐known risk factor for developing metabolic, psychological and cognitive dysfunction, affecting their behavior, attention, learning, memory and decreasing school performance. The present study will describe the dietary and cognitive parameters in first‐year undergraduate students of health sciences at the Autonomous University of the State of Mexico. 125 18 to 21 year‐old freshmen, with variable body‐mass index participated in the study. Participants’ eating habits were evaluated with 24‐hr food diaries, thrice a week. Food consumption frequency questionnaires were used to identify dietary parameters according to diet quality indexes. The Wechsler intelligence scale for adults‐IV (WAIS‐IV) was used to analyze four cognitive parameters: verbal comprehension, perceptual reasoning, working memory and processing speed. Preliminary results from 50 individuals have been completed. Healthier dietary parameters have been identified in men in comparison with women, having more balanced diets overall, according to national guidelines. Similarly, men showed higher average scores in the cognitive parameters assessed. Positive correlations were found between intelligence quotient and total calorie and saturated fatty acid intake in men (p=0.0001, Rho 1.000), as well as between processing speed and saturated fatty acids, lipids, cholesterol and carbohydrate intake in women (p=0.049, Rho 0.350). A negative correlation was found between working memory and vegetable intake (p=0.036, Rho −0.426) in men. Our preliminary data suggest that male freshmen eat healthier diets than women at the authors’ academic institution and that their diets affect their cognitive performance. Additional assessments for sleep quality, exercise habits and self‐image are currently under study in this population.