bluuebunny/crossref_metadata_2025 huggingface dataset:
xCT and CD44v9 Co‐Expression Mediates Pancreatic Ductal Adenocarcinoma Survival in the Presence of ROS Inducing Chemotherapeutics
Authors: Chang Julie , Holokai Loryn , Chakrabarti Jayati , Lanfranca Mirna Perusina , Frankel Timothy , Zavros Yana
Published: June 2021

Background

Pancreatic ductal adenocarcinoma (PDAC) is the third leading cause of cancer‐related death in the United States. It has been shown that KRAS mutations, often found in PDAC, can lead to the up‐regulation of the expression of the cysteine/glutamate transporter xCT. Transporter xCT has been shown to allow cancer cells to evade ROS mediated cell death. CD44 is a transmembrane protein with many different splice varients. CD44 variant isoform 9 (CD44v9) is expressed during cellular damage and has been shown to mediate cell survival. CD44v9 stabilizes xCT expression on the surface of cancer cells. Therefore, CD44v9 stabilized the xCT transporter, which allows cancer cells to survive following treatment with chemotherapeutics that induce ROS.

Hypothesis

Transporter xCT and CD44v9 co‐expression mediates PDAC survival in the presence of ROS inducing chemotherapeutics.

Methods

Human‐derived pancreatic cancer organoids (HuTPOs) were generated from the resected tissue of patients with PDAC. A dose response curve was performed by treating the huTPOs with eight different micromolar concentrations of oxaliplatin, gemcitabine, 5‐fluorouracil (5‐FU). These chemotherapeutics have been shown to induce ROS within PDAC cells. The susceptibility or resistance of the huTPOs to the chemotherapeutics was determined by measuring cell death by an MTS proliferation assay. HuTPOs were analyzed by immunofluorescence for the expression of the cysteine‐glutathione transporter, xCT, and cancer stem cell marker, CD44v9.

Results

There was divergent expression of xCT and CD44v9 in huTPO lines generated from individual patients. Dose response curves performed on huTPO lines revealed that organoids not expressing xCT or only xCT were sensitive to ROS inducing chemotherapeutics (5‐FU, oxaliplatin and gemcitabine). Interestingly, huTPOs that expressed both xCT and CD44v9 were resistant to 5‐FU, oxaliplatin and gemcitabine.

Conclusion

CD44v9 may stabilize xCT in PDAC cancer organoids allowing for resistance to ROS inducing chemotherapeutics. Pancreatic organoids may be a good screening based model to determine efficacy of ROS inducing chemotherapeutics in PDAC.

Support or Funding Information

Steven Goldman Memorial Pancreatic Cancer Research Grant (Zavros) 1RO1DK083402

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Evaluation of Antimicrobial Properties of African Basil (Ocimum gratissimum) and Tassel Flower (Emilia coccinea) Leaf Extracts
Authors: Besong Mary M. , Chintapenta Lathadevi K. , Ozbay Gulnihal , Aikins Anthea , Besong Samuel A. , Aryee Alberta N.
Published: June 2021

The use of plant extracts and phytoconstituents with antimicrobial properties may provide therapeutic benefits and application in consumer products such lotions and soaps, and active food packaging. The objective of this study was to assess the efficacy of water, methanol, butanol and ethyl acetate extracts of Ocimum gratissimum leaf (OGL) in inhibiting the growth of selected bacteria (E. coli O157:H7 (ATCC 43894), Salmonella enterica serovar Typhimurium DT‐104 (CDC H3380), Aeromonas hydrophila K144, and Pseudomonas aeruginosa (ATCC 7700)). In a parallel study, water, methanol, and butanol extracts of Emilia coccinea leaf (ECL) and OGL were also tested against E. coli, Pseudomonas aeruginosa, and Staphylococcus aureus. To screen for antimicrobial activity, the susceptibility of these selected pathogens and non‐pathogenic bacteria was determined using the disk diffusion assay. Selected antibiotics such as tetracycline, streptomycin, ampicillin and kanamycin were used as positive control. Following incubation, zone of inhibition differed between control and extracts, with larger zone of inhibition in the OGL butanol extract than the water, methanol, and ethyl acetate extracts in all the four pathogens tested, and a smaller zone of inhibition for E. coli, Salmonella and Pseudomonas cultures. Aeromonas hydrophila K144 cultures were found to be susceptible to butanol extract showing 10 mm of clearance zone. Incubation of ampicillin with Aeromonas hydrophila K144, Salmonella enterica serovar Typhimurium DT‐104 (CDC H3380) and Pseudomonas aeruginosa (ATCC 7700) led to minor changes in the zone of inhibition of these antimicrobials. P. Aeruginosa and S. aureus were resistant and susceptible to all antibiotic controls tested, respectively. E. coli, P. aeruginosa, S. aureus were all susceptible to the methanol and butanol extracts of OGL and ECL. Butanol and methanol extracts of Ocimum gratissimum and Emilia coccinea leaf could be suitable alternative to some chemical antimicrobials, satisfying consumer demand for natural products.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Self‐serving Contagion and Mimicry in Buffet Lines: Whom You Follow Influences What and How Much You Take and Eat
Authors: Wansink Brian
Published: June 2021

Eating companions can influence eating behavior, but can they influence self‐serving? Less is known how strangers who self‐serve food in a public place may encourage self‐serving mimicry, whether this varies with food‐type and gender, and how self‐serving behavior influences eating behavior. This study investigates self‐serving mimicry in a buffet line in an attempt to determine what people and foods are most prone to serving mimicry and what is the impact of these on subsequent items. 163 strangers were ordered female‐female‐male male and then consecutively served themselves six food items from a lunch buffet. Amounts served and eaten were measured along with demographic measures. The amount a person serves and eats is influenced by the amount served and eaten by the person immediately in front of them. This result appears to be driven mostly by females following females, and can result in an increase in caloric intake. Among these women, mimicry appeared to be stronger with hedonic foods (French fries, applesauce, and macaroni salad) than with less indulgent foods (chicken and celery). Serving contagion and mimicry exist in public places like buffet lines. There are specific implications for dieters suggesting the most indulgent foods are the ones most prone to mimicry.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Neuropeptide Y and Dipeptidyl Peptidase IV in Pre‐ and Post‐Menopausal Women
Authors: Mann Elise , Kluess Heidi A. , Neidert Leslie E. , La Mantia Anna M. , Sandage Mary , Plexico Laura
Published: June 2021

The purpose of this study was to investigate changes in neuropeptide Y (NPY) protein and dipeptidyl peptidase IV (DPP‐IV) activity in the plasma and saliva in normally cycling women and women after menopause. We recruited seven premenopausal women and seven post‐menopausal women for the study. Blood via venipuncture and saliva samples were taken at each point in the menstrual cycle (pre‐menopausal) or once per week (post‐menopausal) for 2 months. Blood and saliva were analyzed for estrogen, neuropeptide Y using ELISA's and DPP‐IV activity using a fluorometric assay. Plasma β‐estradiol was an average of 96.45±57.04 pg/ml over 2 cycles in the pre‐menopausal group and 1.72±0.35 pg/ml over 2 months in the post‐menopausal group (p<0.05). In the pre‐menopausal group, there were no significant differences in saliva or plasma NPY or DPP‐IV over the cycle. For the post‐menopausal group, salivary NPY and DPP‐IV did not change over 2 months. Plasma NPY was lowest in the middle 2 weeks (average: 0.52±0.10 ng/ml) compared to the first and 4th weeks (average of week 1 and 4: 0.60±0.14 ng/ml; p<0.05). Plasma NPY in postmenopausal women was higher overall (0.56±0.13 ng/ml) compared to pre‐menopausal women (0.30±0.11 ng/ml; p<0.05). Plasma DPP‐IV activity was unchanged by time in the post‐menopausal group. Saliva DPP‐IV and saliva NPY in the pre‐menopausal group had a significant negative correlation (R=−0.95; p<0.05). We found that saliva measures of NPY and DPP‐IV activity are poor estimates of plasma concentrations and activities. Differences in plasma NPY concentrations between the groups and the relationship between salivary NPY and DPP‐IV suggests that there are some unique differences between these groups.

Support or Funding Information

The work presented in this poster session was supported by Award Number 1R03DC013664‐01A1 from the National Institute on Deafness and Other Communication Disorders (NIDCD). The content of this presentation is solely the responsibility of the authors and does not necessarily represent the official views of the NIDCD or the National Institutes of Health.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chronic Intermittent Hypobaric Hypoxia Ameliorates Pulmonary Hypertension Induced by Monocrotaline in Rats
Authors: Zhang Yi , Gao Lei , ZHOU JINGJING , Li De‐Pei
Published: June 2021

The anti‐hypertensive effect of chronic intermittent hypobaric hypoxia (CIHH) has been demonstrated, but the effect of CIHH on pulmonary hypertension (PAH) is not elucidated. This study investigated the effect of CIHH on monocrotaline (MCT)‐induced PAH in rats. Adult male rats were randomly divided into control group, PAH group, CIHH group, and CIHH+MCT group. The CIHH rats were treated with a hypobaric hypoxia simulating 5000 m altitude for 28 days, 6 hours per day in a hypobaric chamber. The PAH rats were subjected to a single‐dose injection of MCT (60 mg/kg, intraperitoneally). The CIHH+MCT rats were subjected to 28‐days CIHH treatment before MCT injection. The pulmonary arterial pressure and right ventricular pressure were measure through catheterization, and the wall thickness of pulmonary artery and right ventricle were measured. Micro‐vascular tension recording technique was used to determine the relaxation in pulmonary arterioles. Western blot and quantitative PCR were used to determine the expression of eNOS protein and mRNA in pulmonary arterioles. The mean pulmonary artery pressure and right ventricular systolic pressure were increased in MCT‐treated rats but did not changed significantly in CIHH+MCT rats compared with control rats. The wall thickness of pulmonary arterial and right ventricle were significantly increased in MCT‐treated rats and these increases were smaller in rats treated with CIHH and MCT. The endothelial‐dependent and endothelial‐independent relaxation in pulmonary arterioles rings were significantly decreased in MCT‐treated rats compared with untreated rats. The decreased relaxation was significantly ameliorated by CIHH treatment. The expression levels of eNOS mRNA and protein in pulmonary arterioles were decreased in MCT‐treated rats compared with untreated rats and this downregulation of eNOS was prevented by CIHH treatment. In conclusion, CIHH ameliorates pulmonary hypertension in MCT‐treated rats. This effect may be is related to up‐regulation of eNOS by CIHH treatment.

Support or Funding Information

This work was supported by the National Natural Sciences Foundation of China (31271223 and 31071002), National Basic Research Development Program of China (2012CB518200).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Microscopic Surface Geometries and Blood Clotting: A Computational Analysis
Authors: Riggenbach Zachary William
Published: June 2021

Biofouling and thrombogenesis present a challenge tobiomedical products that contact blood plasma. These devices include heartvalves and artificial blood vessels. Excessive blood‐borne protein depositionon device surfaces likely lead to clot formation and eventual device failure. Toprevent clot formations, an effective anti‐thrombotic surface is needed toallow for more viable regenerative medical devices and procedures. Previousresearch suggests superhydrophobic surfaces serve as one method to reduceprotein deposition and clot formation. The hierarchical surface features of superhydrophobicsurfaces allow fluids to rest in the fakir state which produces substantiallyreduced fluid‐to‐surface contact area and increased droplet mobility. Simple engineering techniques have successfully demonstrate that superhydrophobicsurfaces reduce clot formation compared to control surfaces. This study uses a predictive computational program to model blood droplet behavior on microscopic surface features. The computational software uses the gradient descent method to predict equilibrium droplet positions and geometries based on the surface energies of the fluid and surface features. The software predicts and visualizes the impact various surface feature dimensions have on the ability of human blood to maintain the fakir state. The new optimization builds off of prior work from previous literature by considering the non‐homogenous nature of whole human blood and the varying surface energies of coagulation cascade proteins. The optimized surface feature geometries potentially provides a more efficient anti‐clotting surface for biomedical devices.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hypergravity selectively augments neuronal in vitro differentiation
Authors: Brungs Sonja , Liemersdorf Christian , Lichterfeld Yannick , Frett Timo , Anken Ralf , Jordan Jens , Hemmersbach Ruth
Published: June 2021

Disturbed neuronal connectivity is the ultimate cause of disability in individuals with neurological disease including spinal cord injury, head trauma, and stroke. Functional neurological recovery is limited through an unfavorable balance between neuronal regrowth and glia scar formation. Neuronal growth requires dynamic cytoskeletal protein rearrangements. Because hypergravity stabilizes microtubules while de‐stabilizing actin filaments, we hypothesized that experimental hypergravity would shift the balance between neuronal and astroglial growth in vitro.

We exposed murine primary hippocampal neurons during different developmental stages to 2g using the DLR hypergravity platform. The platform unlike commercial laboratory centrifuges models physiological hypergravity and allows for cell cultivation and live‐cell imaging. We assessed neuritogenesis, neuronal polarization and maturation processes including synaptogenesis and synaptic integration in mature neural networks. Moreover, we studied primary astrocytes to shed light on their role during glial scar formation at neural lesion sites.

Exposure of hippocampal neurons to 24h of 2g hypergravity increased neurite number by 30% and neurite projection length by 20% compared to 1g. At later developmental stages, mature synaptic contacts were formed under hypergravity conditions. In contrast, astrocytes showed decreases in cell spreading and lamellipodial protrusions with hypergravity.

We conclude that experimental hypergravity ameliorates neuronal cell growth and synaptic contacts while halting astrocyte spreading and migration in vivo. Given the importance of this balance for neuronal regeneration in human neurological disease, we will now study the underlying mechanisms in more detail using, both, DLR hypergravity and clinostat‐induced hypogravity platforms.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Endothelial TRPA1 Channels Are Activated by Hypoxia in Cerebral Arteries and Protect Against Ischemic Damage
Authors: Pires Paulo Wagner , Earley Scott
Published: June 2021

Acute drops in cerebral perfusion can lead to rapid neuronal death, as seen during transient ischemic attacks and ischemic strokes. The extent of brain damage following hypoxia is linked to the ability of collateral blood vessels to adequately supply blood to the peri‐infarct region and thus prevent neuronal loss and infarct expansion. During hypoxia, reactive oxygen species (ROS) are increased and may activate the Ca2+‐permeable transient receptor potential ankyrin 1 (TRPA1) in endothelial cells, promoting vasodilation. Here, we tested the hypothesis that TRPA1 channels in endothelial cells are activated by hypoxia‐derived ROS, leading to cerebral artery dilation and reduced ischemic damage. Using a novel mouse model expressing the Ca2+ indicator GCaMP6f in endothelial cells (Tek:GCaMP6f), we show that 4‐hydroxynonenal (4‐HNE), a lipid peroxide, increased TRPA1 channel activity, detected as elementary Ca2+ influx events (TRPA1 sparklets). The TRPA1 inhibitor A967079 prevented 4‐HNE induction of TRPA1 sparklets in endothelial cells. Acute exposure of cerebral arteries to hypoxia (5% O2, 6% CO2, 89% N2) significantly increased 4‐HNE accumulation in cerebral arteries, which was linked to an increase in TRPA1 sparklets. Hypoxia induction of TRPA1 sparklets was blocked by the TRPA1 inhibitor A967079, and by PEGylated superoxide dismutase (PEG‐SOD), suggesting a role for intracellular generation of ROS. Incubation of arteries with the mitochondria‐targeted antioxidant mitoTEMPO prevented the hypoxia‐induced increase in TRPA1 sparklets, whereas extracellular SOD or the NADPH oxidase inhibitor apocynin had no effect. In pressure myography studies, hypoxia caused endothelium‐dependent dilation of isolated cerebral arteries, an effect that was prevented by the membrane‐permeable SOD mimetic tempol and by mitoTEMPO. Hypoxia‐induced dilation of cerebral arteries was partially inhibited by the TRPA1 antagonist A967079 and was reduced in arteries isolated from endothelial cell‐specific TRPA1 knockout mice (eTRPA1−/− mice). In vivo, loss of TRPA1 channels in endothelial cells increased infarct size following middle cerebral occlusion, a model of ischemic strokes. Reversely, TRPA1 activation with cinnamaldehyde (50 mg/kg) reduced infarct size in wildtype, but not eTRPA1−/−, mice. Together, these data suggest that TRPA1 channels in cerebral artery endothelial cells are early sensors of hypoxia and orchestrate a response that culminates with vasodilation, thereby reducing ischemic damage. Further, pharmacological activation of TRPA1 channels may be a putative therapeutic target to reduce cerebral ischemic damage.

Support or Funding Information

Supported by the National Institutes of Health (R01HL091095 to SE) and the American Heart Association (15POST2472002 to PWP)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Acid Sensing Ion Channel 1 Contributes to Endothelium‐Derived Hyperpolarizing Factor Induced Vasodilation in Small Mesenteric Arteries.
Authors: Garcia Selina , Naik Jay S. , Resta Thomas C. , Jernigan Nikki L.
Published: June 2021

Acid sensing ion channel 1 (ASIC1) belongs to the amiloride‐sensitive degenerin/epithelial sodium channel superfamily, and is permeable to both Na+ and Ca2+. ASIC channels have been found to regulate blood pressure through mechanotransduction in arterial baroreceptors and are also pH sensors in carotid body chemoreceptors. ASIC1 is additionally expressed in vascular smooth muscle and endothelial cells in a variety of vascular beds; yet little is known regarding the role of ASIC1 in the regulation of local vascular resistance. Although extracellular H+ is the recognized activator of ASIC1, we have previously shown this channel can be activated following G protein‐coupled receptor stimulation. In small mesenteric arteries, we found that ASIC1 contributes to acetylcholine (ACh)‐induced endothelial cell Ca2+ entry and vasodilation. However, the mechanisms involved in activating ASIC1 and its downstream effects on vasodilation are unknown. We hypothesized that ACh activates ASIC1‐mediated endothelial cell Ca2+ entry to mediate endothelium‐derived hyperpolarizing factor (EDHF)‐type vasodilation through activation of small (SK) and intermittent (IK) conductance Ca2+‐activated K+ channels. To test this hypothesis, we used pressure myography to study the effects of ASIC1 on ACh‐induced vasodilation (10−9 – 10−5 M) of freshly isolated mesenteric arteries (100–200 μm inner diameter), in the presence of Nω‐nitro‐L‐arginine (nitric oxide synthase inhibitor) and indomethacin (cyclooxygenase inhibitor). ACh‐induced vasodilation was abolished by specific inhibitors of SK and IK channels (apamin and TRAM‐34, respectively), confirming that the ACh‐induced dilation is EDHF‐dependent. The specific ASIC1 antagonist, psalmotoxin 1 (PcTX1; 20 nM) similarly blocks the EDHF‐mediated vasodilatory response, suggesting ASIC1 contributes to ACh‐induced vasodilation through an EDHF‐mediated pathway. Future studies will examine the upstream mechanisms involved in activation of endothelial ASIC1 and subsequent downstream effects to mediate EDHF‐dependent vasodilation. These studies suggest a role for ASIC1 as a novel contributor to agonist‐induced mesenteric vasodilation and cardiovascular homeostasis.

Support or Funding Information

Supported by National Heart, Lung, and Blood Institute Grants R01 HL‐111084 (to N.L. Jernigan), T32 HL007736 (to T.C. Resta), and R01 HL132883 (to T.C. Resta).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Maternal Protein Restriction Mediates the Development of the Musculoskeletal System in the Offspring
Authors: Vasilaki Aphrodite , Kanakis Ioannis , Giakoumaki Ifigeneia , Pollock Natalie , McArdle Anne , Sayer Avan Aihie , Whysall Katarzyna
Published: June 2021

The age‐related progressive decline of muscle mass and strength plays a major role in the pathogenesis of frailty which results in a substantial reduction of the quality of life of older people. Age‐related changes in motor neurons, muscle fibres and the interactions between nerves and muscles are responsible for the loss of strength. There is considerable evidence that poor maternal nutrition leads to a number of changes in muscle of the offspring that result in reduced function. Muscle strength is also compromised in older individuals who did not grow well in early life, and studies suggest that maternal, developmental and nutritional factors are important. The aim of this study was to examine the effect of reduced maternal protein intake on the development of the neuromuscular system of the offspring as well as on bone mass.

Studies were undertaken in Thy1‐YFP16 mice that express yellow fluorescent protein only in neuronal cells. Two weeks before mating, female mice were fed either a low protein diet (5% Crude Protein) or a control diet (20% Crude Protein). After acclimation to the chow, breeders were placed together. Newborn pups were cross‐fostered to different lactating dams (maintained on either normal or low protein diet) within 24 h after birth. Pups were culled either at weaning (21 days) by cervical dislocation or were weaned onto either the deficient or the control diet until 12 weeks of age. The extensor digitorum longus (EDL) muscle and tibial trabecular bone were analysed using histological and μCT techniques respectively. The maximum force generated by the EDL was also determined.

Our data indicate that mice born from dams maintained on a normal protein diet but fed post‐natally by a foster dam maintained on a low protein diet (N‐L) show significant reductions in body size, body weight, muscle fibre number and muscle size at 21 days when compared with pups from dams maintained on a normal protein diet throughout the in utero and post‐natal periods (N‐N). Maintenance of mice on a low protein diet until 12 weeks of age resulted in reduced muscle strength. Additionally, μCT analyses revealed changes in trabecular bone mass and the growth plate between the two groups.

Thus, these data suggest that, in mice, protein deficient diet in the post‐natal period can negatively affect musculoskeletal development.

Support or Funding Information

The authors would like to thank BBSRC, the MRC‐Arthritis Research UK Centre for Integrated research into Musculoskeletal Ageing (CIMA) and The Physiological Society for financial support.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Co‐expression of Orexin‐B and Corticotropin‐Releasing Factor in Hypothalamic Neurons of the Ovariectomized Female on a High Fat Diet
Authors: Ciriello John , Iqbal Waseem , Durand Nathan Jeffrey
Published: June 2021

The hypothalamic paraventricuar nucleus (PVH) is known to play a pivotal role in energy balance and in autonomic and neuroendocrine control. The neuropeptides orexin‐A (OX‐A) and orexin‐B (OX‐B) have been implicated in these homeostatic mechanisms. This study was done to investigate whether 17β‐estradiol (E) treatment coupled with ingestion of a high fat diet (HFD; 45 kcal fat) altered OX immunoreactivity in PVH of ovariectomized (OVX) female Wistar rats treated (OVX+E) or not‐treated with E (OVX only). Animals were placed on a normal diet (ND) or HFD for a period of 7 or 20 weeks. Twenty‐four hours before sacrifice all animals were given an intracerebroventricular injection of colchicine and the forebrains were processed for OX‐A‐and OX‐B‐ immunoreactivity. In all cases, relative OX‐B immunoreactive fiber density was greater throughout PVH compared to OX‐A in both female groups regardless of the diet. OX‐A‐immunoreactivity was also found to be greater in females fed a HFD compared to a ND. Furthermore, a distinct cluster of OX‐B immunoreactive neurons was found within the medial parvocellular PVH in OVX only animals, and the number of these neurons was found to higher in those fed a HFD for the longer period. Pre‐pro‐orexin protein expression was up‐regulated in PVH in OVX only females on the HFD compared to ND and compare to OVX+E on a HFD or ND. As the location of these OX‐B neurons in the medial parvocellular PVH overlapped the area known to contain corticotropin‐releasing factor (CRF) neurons, a double immunofluorescence studied was done to determine whether the OX‐B neurons co‐expressed CRF. All medial parvocellular PVH neurons that expressed OX‐B co‐expressed CRF, while not all neurons expressing CRF co‐expressed OX‐B. As OX has been shown to function as a neuromodulatory hormone that stimulates the adrenal corticotrophin hormone‐mediated response to stress, these findings suggest that in females without E, co‐release of OX‐B and CRF in response to a HFD stressor may affect anterior pituitary function.

Support or Funding Information

Supported in part by the Heart and Stroke Foundation.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
EFFECT OF DELTAMETHRIN ON THE FRAGILITY OF RED BLOOD CELLS IN ALBINO RATS
Authors: Umunnakwe Shalom E. , Chidi Nosiri , Friday Uhegbu
Published: June 2021

This study investigated the toxic effect of Deltamethrin dust Insecticide on the fragility of red blood cells using albino rats. Deltamethrin is an insecticide belonging to the Pyrethroid family used to kill bed bugs, ticks, fleas and spiders. In this study animals were divided into four groups: the test group1 as the control, group II recieved 0.03ml of the insecticide, groups III and IV recieved 0.05ml and 0.08ml of the insecticide respectively. The results showed a slight decrease in packed cell volume (PCV), Heamoglobin (Hb), and Red Blood Cell (RBC) concentrations with the values thus: PCV (40.80 +2.94% to 36.94±0.20%), HB (13.60±0.98g/dl to 12.31±0.06g/dl) and RBC concentration from 5.80±0.20 to 5.33±0.12(x1012 mm3). For Mean corpuscular volume (MCV) and Mean Corpuscular Hemoglobin (MCH) they experienced slight increase from 68.79±4.72 to 69.38±1.32(fl) and 22.87±1.32 to 23.13 ±0.44(pg) respectively. The Mean Corpuscular Heamoglobin Concentration (MCHC) had a constant value at 33.33g/dl. Oxidative stress from environmental toxicants like Deltamethrin dust insecticide may have affected the membrane intergrity of the RBC by the membrane shrinking or hemolysing showing that osmotic fragility has taken place in the animals used.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Calcium Regulation of Mitochondrial Respiration is Substrate Dependent and Tissue Specific
Authors: Kandel Sunil M , Tomar Namrata , Audi Said H , Camara Amadou K S , Cowley Allen W , Dash Ranjan K
Published: June 2021

Mitochondrial Krebs cycle dehydrogenase enzymes, such as pyruvate dehydrogenase, isocitrate dehydrogenase, and alpha‐ketoglutarate dehydrogenase, have been shown to be stimulated by calcium ion concentration, [Ca2+], in isolated enzyme experimental systems. However, the extents of the [Ca2+] stimulation of these enzymes and the resulting integrated stimulatory effects on mitochondrial energy metabolism (ATP synthesis) in situ or in vivo in the heart and other tissue/organ systems, such as kidney, is controversial and/or not yet well‐understood. This may be due to several compounding factors, such as the use of specific respiratory substrates in isolated mitochondrial experimental system to energize the mitochondria, and/or the way the mitochondrial [Ca2+] is perturbed. In this study, we hypothesize that the extents of the [Ca2+] stimulation is critically dependent on the specific choice of the respiratory substrates and the modes of mitochondrial [Ca2+] perturbations, and that this substrate‐dependent [Ca2+] stimulation is also tissue specific. Consequently, alterations in the respiratory substrates can lead to dramatically different respiratory rates in isolated mitochondria from the heart and kidney. To test this hypothesis, we used isolated mitochondria from two different organs (i.e. heart and kidney) from Sprague‐Dawley rats and measured oxygen consumption rates in different respiratory states: leak state (energized mitochondria in the absence of ADP) and ADPstimulated state (energized mitochondria in the presence of saturating concentrations of ADP) with/without bolus injections of different concentrations of CaCl2 using an Oroboros Oxygraph‐ 2k Instrument in the presence of a variety of respiratory substrates, including pyruvate + malate (PM), glutamate + malate (GM), alpha‐ketoglutarate + malate (AM), palmitoyl‐L‐carnitine + malate (PCM), and succinate (SUC). We found that CaCl2 at a certain range of concentrations differentially enhance the mitochondrial respiration (ATP synthesis) when the isolated mitochondria was fed with different respiratory substrates and their response was tissue dependent. Calcium significantly increased the mitochondrial state III respiration with GM and AM substrates compared to PM substrate in both organs, but the calcium effect on the mitochondrial respiration was more prominent in the kidney compare to the heart. These results suggest that the energized isolated mitochondria in the presence of calcium do not respond the same when the different substrates are used and the effective energy metabolism and the production of ATP may require a suitable substrate combination to be comparable to in vivo condition.

Support or Funding Information

NIH grants P01‐GM066730, U01‐HL122199

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Simultaneous Stabilization of Protein and RNA from a Single Tissue Homogenate
Authors: Pirman Natasha L
Published: June 2021

Maintaining activity and stability of both proteins and RNA from biological samples remains a major concern and challenge during sample preparation and storage. Many researchers employ a myriad of difficult and time‐consuming strategies to preserve important and costly samples. Moreover, with more studies requiring investigation of both RNA and protein, additional sample preparation steps are required; therefore, researchers are often relegated to performing specific and separate sample preparations to study both RNA and protein, or unfortunately at times required to choose one type of analysis. To overcome these issues, we developed the unique Stabilyser reagent that yields enhanced stability and activity of both protein and RNA from homogenized tissue. This reagent enables the ability to obtain both genetic and protein information from a single, uniform homogenized sample, significantly alleviating laborious sample preparation.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Acetylation of Lysine Residues within the MT‐binding Repeats Specifically Modulates the Structure Ensemble of Tau
Authors: Yao Zhiyang , Gao Meng , Huang Yongqi
Published: June 2021

The microtubule associated protein tau is expressed primarily in the central nervous system and promotes microtubule (MT) assembly. The aberrant accumulation of tau protein and formation of insoluble aggregates are pathological hallmarks of neurodegenerative tauopathies including Alzheimer's disease. Six tau isoforms are generated by mRNA alternative splicing containing either three (3R tau) or four (4R tau) MT‐binding repeats and various number of N‐terminal exons (0N, 1N and 2N). K18 and K19 are the truncated constructs of 4R tau and 3R tau, consisting of only the four and three repeats, respectively. K18 and K19 form amyloid fibrils with cross‐β structure, constituting the core of tau filaments. Recent studies have shown that acetylation of specific lysine residues within the MT‐binding repeats could either promote or inhibit tau aggregation and tau mediated microtubule assembly. Here, we investigated the effect of lysine acetylation on the structure of tau K18 by a combination of biophysical structure characterization and molecular dynamics simulation. Radius of gyration derived from small angle X‐ray scattering (SAXS) showed that the structures of acetylation mimic tau species (K274Q/K321Q and K280Q/K281Q/K369Q) were more extended than the WT protein. Although the influence of lysine acetylation on the intramolecular interactions was mainly local, long range perturbations were also observed from NMR chemical shift analysis. Molecular dynamic simulations in explicit solvent revealed site specific modulation in the structure ensemble of tau repeats, where K280Q/K281Q mutation led to compaction of repeat R2 and K321Q mutation led to extension of repeat R3. Statistics on the lifetime and distance of residue contacts and secondary structure components were performed to investigate the changes in structure and dynamics upon acetylation. These results show that the effect of acetylation on tau structure was site specific, which may promote or reduce aggregation by exposing or shielding the aggregation prone segments.

Support or Funding Information

This work was supported by National Natural Science Foundation of China (Grant number: 21603121) and Hubei University of Technology.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Pinealectomy Reduces Gene Expression of Caspase 9 in Subcutaneous Adipose Tissue of Rats
Authors: da Costa Tatienne Neder Figueira , Andreotti Sandra , Lima Fábio Bessa
Published: June 2021

Previous studies have shown that pinealectomy induces insulin resistance in rats. The present study aimed to verify whether pinealectomy modulates the expression of apoptosis‐related genes in rat subcutaneous adipose tissue (TA‐Sc). For this purpose, samples from the bank of frozen tissue of animals (Ethics Committee/Institute of Biomedical Sciences (ICB) Protocol No. 896/2017), belonging to the Laboratory of Adipose Tissue Physiology, University of Sao Paulo (USP), were used. The material came from Wistar rats (male and adults), whose were kept at the Experimentation Hall of the Department of Physiology and Biophysics of the ICB/USP and handled according to a certificate approved by the Ethics Committee on the Use of Animals (CEUA)/(03/10/2011, protocol no. 129) and standards established by the National Council for Control of Animal (CONCEA). The rats were housed in polypropylene boxes under controlled environmental conditions (25 ± 2 ºC) and light (12 h light/12 h dark), with water and standard ration for Nuvilab® rodents (Nuvital S/A, Colombo, PR) ad libitum. The animals, when completed 10 weeks old, were anesthetized and submitted to pinealectomy (P) by the method of Hoffman and Reiter (1965) or to the fictitious operation (C) and, following their recovery, they were kept in individual boxes. After four weeks of the surgery (14 weeks old) the animals P and C were killed by decapitation and inguinal TA‐Sc dissected. For the evaluation of gene expression related to apoptosis, samples of the Sc fat pad were submitted to the Total RNA Extraction stages; Synthesis of cDNA by Reverse Transcription and Polymerase Chain Reaction from cDNA (Real Time PCR). PCR reactions were performed by the StepOnePlus® Real Time PCR System (Applied Biosystems). The evaluated genes were: Caspases (3, 8, 9), Bax, Fas and BCL‐2. The data were analyzed according to the requirements of homoscedasticity and normality by means of the Shapiro‐Wilk test and later the Student's t‐test, unpaired and parametric. The 95% confidence interval and p value <0.05 were considered significant. The statistical program used was Graph Pad Prism version 6.0. The results showed that pinealectomy did not alter the expression of the anti and pro‐apoptotic genes evaluated (p> 0.05), with the exception of Caspase 9, whose gene expression was significantly reduced in group P in relation to C (Figure 1). In conclusion, pinealectomy is capable of selectively interfering with the gene expression of the apoptotic pathway, specifically in the caspase activator 9.

Support or Funding Information

Conselho Nacional de Desenvolvimento Científico e Tecnológico‐CNPq

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
[PIN+]ning down [PSI+] inducibility
Authors: Brechtel Teal M , Villali Janice , Davis Jason , Pei Fen , Sindi Suzanne S , Serio Tricia R
Published: June 2021

Prion proteins confer new phenotypes on cells by adopting an alternative conformation that changes their function and by templating the conversion of normally folded protein to the same state. While we understand how prion proteins in yeast maintain themselves in cells and colonies once they arise, the mechanism by which the alternative conformations of prion proteins first appear is poorly understood. To address this open question, our studies focus on the [PSI+] prion, an alternative conformation of the Sup35 protein. [PSI+] can only appear in yeast cells that also contain the [PIN+] prion, an alternative conformation of the Rnq1 protein. Our genetic studies suggest that [PSI+] appearance is sensitive to the level of the molecular chaperone Hsp104. Moreover, existing [PSI+] is sensitive to changes in Hsp104 levels in a [PIN+]‐ dependent manner. Together, our observations are consistent with a model in which [PIN+] acts as a decoy for chaperones and thereby promotes the persistence of nascent [PSI+].

Support or Funding Information

This work was supported by NIH Training Grant T32 GM008659 to TMB and NIH Research Project Grants R01 GM100740 and R35 GM118042 to TRS.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Antioxidant activities of anthocyanin and polyphenol fractions of black raspberry (Rubus occidentalis) in inflamed human umbilical vein endothelial cells
Authors: Park Sun Young , Jung Hana , Hwang Keum Taek , Kwak Ho‐Kyung
Published: June 2021

Black raspberry (Rubus occidentalis) contains a large amount of polyphenols. Among them, anthocyanin and phenolic acids are known to inhibit oxidative stress and inflammatory reaction, which are risk factors of chronic inflammation diseases. The aim of this study was to investigate antioxidant activities of anthocyanin (AF), phenolic acid (PF) and total polyphenol fractions (APF: combining AF and PF) from black raspberry in inflamed human umbilical vein endothelial cells (HUVEC) which were exposed to conditioned medium (CM) obtained from lipopolysaccharide‐stimulated RAW 264.7 cells. Intracellular reactive oxygen species (ROS) and DNA fragmentation were determined using 2′, 7′‐dichlorofluorescin diacetate and Hoechst staining methods, respectively. When HUVEC was treated with CM for 20 h, intracellular ROS and DNA fragmentation significantly increased (P<0.05). The HUVEC pretreated with AF, PF, and APF produced significantly less intracellular ROS and DNA fragmentation than CM‐treated control cells (P<0.05). In addition, comet assay was used to determine DNA damage of the cells. The cells pretreated with the three tested fractions (20 μg/mL) were less damaged. There were no significant differences in superoxide dismutase and catalase activities among the groups of the two control cells and three black raspberry fractions. The results suggest that AF, PF, and APF of black raspberry may have antioxidant effects in inflamed HUVEC.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Fibroblast Growth Factor 9 (FGF9) is Expressed in An Osteocyte‐like “Mini‐bone” Cell Line and Inhibits C2C12 Myogenesis via Overexpression of Myostatin
Authors: Huang Jian , Wang Kun , McCormick Lora , Brotto Leticia , Dallas Sarah , Brotto Marco
Published: June 2021

Musculoskeletal diseases affect about 130 million Americans and 1.7 billion humans worldwide. Sarcopenia and osteoporosis occur concurrently. Muscle and bone are connected anatomically and functionally and interact as a functional unit. Previously we reported that PGE2 and WNT3a are secreted factors from bone that accelerates myogenesis and β‐aminoisobutyric acid, l‐BAIBA, is a muscle‐derived osteocyte survival factor. Recently, we found that FGF9 was enriched in the OmGFP66 “mini bone” cell line.

To study the effects of FGF9 on myogenesis, C2C12 myoblasts were treated with 10 and 50ng/mL recombinant FGF9, and the effects on differentiation were measured 72h later by Fusion Index (FI). To study molecular mechanisms of FGF9 on myogenesis, the expression of myogenic regulator genes (MyoG, Mhc and Myostatin) were analyzed by RT‐qPCR. To investigate the effects of FGF9 on calcium homeostasis of myotubes, intracellular Ca2+ measurements were performed.

Compared to control, a significant decrease in FI was found in FGF9‐treated groups (p < 0.05) suggesting that FGF9 inhibits C2C12 myoblast differentiation. In FGF9‐treated groups, the expression of MyoG and Mhc decreased significantly, while the expression of Myostatin increased significantly (p < 0.05). These results suggest that FGF9 inhibition of C2C12 cell differentiation may be mediated by the upregulation of Myostatin. FGF9‐treated myotubes showed a lower Ca2+ peak and a shortened relaxation phase in response to caffeine‐induced Ca2+release (p < 0.05), suggesting FGF9 inhibits calcium homeostasis in myotubes. PCR array revealed that in 10ng/ml FGF9 treated C2C12 cells, compared to control, the expression of Acsl5 and Icam1 significantly increased, while Bamp4, Hey1, Wnt1 and Wnt6 significantly decreased.

FGF9 could be a mediator of bone to muscle signaling, likely an inhibitory osteokine via modulation of myostatin expression and intracellular Ca2+ signaling. This could have important applications to musculoskeletal diseases.

Support or Funding Information

This study was supported by: NIH‐NIA P01 AG039355 (SD, MB), the George W. and Hazel M. Jay Endowment (MB), and the UT System Science and Technology Acquisition and Retention Program (STARS) (MB).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Mechanistic comparison of human DNA Ligases I and III
Authors: O'Brien Pat
Published: June 2021

DNA ligation is an essential step to complete DNA repair, replication and recombination processes. The human genome encodes two divergent, yet structurally similar DNA ligases, LIG1 and LIG3. Each enzyme has unique amino and carboxy‐terminal extensions and the conserved catalytic core is only 22% identical. LIG3 is essential for mitochondrial genome maintenance, but is also expressed ubiquitously as a nuclear isoform. To understand the mechanism and fidelity of human DNA ligation we are working to characterize DNA LIG1 and LIG3 and their associated protein complexes. Transient and steady state kinetic approaches are used to elucidate distinct enzyme conformational changes and the role of the essential metal ions in substrate selection and catalysis. By determining the similarities and differences between these DNA ligase isoforms, we provide new insights into the clinical situations in which mutation or changes in gene expression are associated with diseases such as cancer and primary immune deficiency.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.