bluuebunny/crossref_metadata_2025 huggingface dataset:
Targeting The Laminin Receptor / PEDF Interface to Treat Prostate Cancer Cells
Authors: Umbaugh Charles , Diaz‐Quiñones Adriana , Zhou Jia , Cheng Xiaodong , Figueiredo Marxa
Published: June 2021

The non‐integrin laminin receptor (LAMR1) is a multifunctional protein localized in the cytosol and at the membrane. Recently, the potent antiangiogenic protein required for retinal development and maintenance, pigment epithelium derived factor (PEDF), was confirmed as a binding partner for LAMR1. LAMR1 is frequently overexpressed in cancer cells. Our lab and others have demonstrated that PEDF abolishes neo‐vasculature in prostate tumors, resulting in decreased tumor growth and burden and abatement of metastatic potential.

Using crystal structures for PEDF and LAMR1, we mapped the interaction interface and screened the Maybridge HitFinder Database in silico (>14,000 compounds) using Docking@UTMB and AutoDock Vina. We selected seven hits and conducted a medium throughput screen in multiple cell lines. IC50s were generated using a six point dose curve and a four parameter non‐linear regression. Orthogonal validation was performed using luciferase assays against cancer specific promoters. Finally, we identified the 67 kDa species of LAMR1 and successfully knocked endogenous protein down using siRNA duplexes.

Our phenotypic screen revealed three putative laminin antagonists, two agonists, and two ineffective compounds. We established that PEDF and the peptide P18 can mediate changes in gene expression. One compound significantly reduced Fas Ligand promoter activity and phenocopied PEDF, suggesting that these compounds may modify the tumor microenvironment. Next, we will confirm LamR specificity using cell‐surface binding assays and siRNA.

Funding: UTMB CAR P50DA033935 Pilot Grant and Startup Funds
Antibacterial Activity of Crude Methanolic Extract of Buchholzia Coriacae (Wonder Kola) on some Bacteria from Veterinary Teaching Hospital, Ahmadu Bello University, Zaria
Authors: Kwanashie Clara , Esonu Mary , Mamman Paul
Published: June 2021

Medicinal plants are plants which contain substances that could be used for therapeutic purposes. In recent years, there has been a gradual renewal of interest in the use of medicinal plants worldwide. As a result of this interest, crude methanolic extract of the plant wonder kola of botanical name Buchholzia coriacea, family Capparaceae was used on some bacteria organisms (Escherichia coli, Staphylococcus aureus, Salmonella Gallinarium 1, Salmonella Gallinarium 2 and Salmonella Gallinarium 3 Salmonella spp) to establish the antibiotic activity of wonder kola on these organisms. The result showed that wonder kola has antibiotic activity against these organisms at various concentrations as shown thus Escherichia coli at 500 gm/ml, Staphylococcus aureus at 5.6 mg/ml, Salmonella Gallinarum 1 at 200 mg/ml, Salmonella Gallinarium 2 at 31.25 mg/ml and Salmonella Gallinarium 3 at 31.25 mg/ml. It can be concluded that wonder kola (Buchhalzia coriacea) has significant antibiotic activity on these organisms.
Redox‐sensitive Sp‐3 Transcription Factor via Renal AT1 Receptor Contributes to Age‐related Hypertension
Authors: Saleem Mohammad , Asghar Mohammad
Published: June 2021

Renal angiotensin II type 1 receptor (AT1R) plays pivotal role in blood pressure control. We recently reported an exaggerated AT1 receptor function and hypertension in aged (21 month) Fisher 344 X Brown Norway (FBN) rats. Antioxidant tempol treatment reduced AT1R function and blood pressure in these rats suggesting a role of oxidative stress in this phenomenon. However, oxidative stress mechanism for higher AT1R function is not known. Therefore, we became interested in delineating the mechanism of oxidative stress induced up‐regulation of AT1R in aging kidney. mRNA levels of renal AT1R were measured by qRT‐PCR while nuclear Sp‐3 and AT1R protein levels were measured by western blot. To further examine whether oxidative stress increases nuclear translocation of Sp‐3 and up‐regulation of renal AT1R expression, we treated human kidney 2 (HK2) cells with pro‐oxidants diethyldithiocarbamate (DDC) and hydrogen peroxide (H2O2). Acute treatment of DDC (30min) and H2O2 (20min)increased nuclear levels of Sp‐3 which was attenuated with tempol treatment. DDC chronic treatment (2.5 hour) increased the expression of AT1R while tempol treatment attenuated this effect. Moreover, Sp‐3 plasmid increased and Sp‐3 siRNA decreased the levels of AT1R protein in HK2 cells. Our results suggest that redox‐sensitive Sp‐3 transcription factor up‐regulates renal AT1 receptor and may contribute to age‐related hypertension.

Source of research support: NIH/NIA AG039856
GPER/GPR30 and Components of Store‐operated Ca2+ entry
Authors: Terry Lara , VerMeer Mark , Francis Sarah , Tran Quang‐Kim
Published: June 2021

The novel G protein‐coupled estrogen receptor 1 (GPER/GPR30) has been found to participate in numerous cardiovascular functions. Store‐operated Ca2+ entry (SOCE) is an essential mechanism required for many cell activities. We found that GPER agonist G‐1 inhibits SOCE in endothelial cells. Heterologous expression of GPER causes a 40% decrease in the rate of SOCE. GPER antagonist G15 acutely increases rates of SOCE by 16% and when treated chronically, increases total Ca2+ signals by 50% in vascular smooth muscle. Consistently, GPER gene silencing increases SOCE by approximately 50%. In endothelial and ventricular tissues, GPER coimmunoprecipitates with the stromal interaction molecule 1 (Stim1) and the sarcoplasmic reticulum Ca2+‐ATPase (SERCA), two essential molecular switchers of SOCE. In addition, Stim1 colocalizes with GPER when heterologously expressed in HEK 293 cells. In addition, overexpression of GPER substantially reduces thapsigargin‐induced Ca2+ release from the endoplasmic reticulum in vascular endothelial cells, an indication of reduced SERCA activity. These data suggest that GPER may be an important regulatory input of store‐operated Ca2+ entry via its interactions with key components of store‐operated Ca2+ entry.
Cocaine sensitization is mediated by proteasome function in an activity‐dependent manner
Authors: Howell Kristin , Patrick Gentry , Anagnostaras Stephan
Published: June 2021

Exposure to addictive drugs can produce structural and functional modifications in neurons and synapses that entail the dynamic regulation of protein synthesis and degradation. Protein degradation is mediated by the ubiquitin proteasome system. Recently, it was discovered that CaMKIIa regulates the proteasome by phosphorylating the ATPase 19S regulatory particle subunit of the 26S proteasome, Rpt6, at serine 120 in an activity‐dependent fashion. Based on these findings, a line of phospho‐mimetic, S120D knock‐in mice was generated. Behavioral sensitization, an enhanced sensitivity to a drug's effects is characterized by increased psychomotor activation and is accompanied by major structural remodeling. Here, we investigated the effects of proteasomal function in sensitization. S120D and WT mice received three training sessions during which mice were injected with cocaine (15 mg/kg, i.p) and activity was assessed. Mice then underwent a post‐training sensitization challenge test, consisting of a final injection of cocaine (15 mg/kg). There were no acute activity differences between WT and S120D mice following the first injection. However, while robust sensitization developed in WT mice following repeated cocaine administration, locomotor activity in S120D mice remained stable across all sessions. Sensitization, assessed as the difference in activity between the challenge test and first injection, revealed a complete absence of sensitization in S120D mice. These data provide strong evidence that permanently mimicking Rpt6 S120 phosphorylation occludes cocaine‐induced sensitization.
Behavioral, Biochemical and Molecular modulation in Brain and Spinal cord : A potential antinociceptive‐antidepressant mechanism of Xanthophylls
Authors: Badgujar Lohit , Saraf Madhusudan
Published: June 2021

Aim

The present study was designed to investigate the antinociceptive‐antidepressant effects of xanthophylls, Lutein and in chronic pain‐depression dyed.

Methods

SD rats (230‐250 g) were anesthetized under ketamine‐xylazine (80 ‐ 20 mg/kg, intraperitoneally) and chronic constriction injury (CCI) was performed (Bennett and Xie, 1988). Vehicle‐olive oil (0.5 ml), Lutein (LT 20 & 40 mg/kg), Zeaxanthin (ZX 20 & 40 mg/kg) and Lutein‐Zeaxanthin (LZ 25 & 50 mg/kg) were administered from day 14 to day 38. Behavioural observations were recorded on 0, 7, 14, 21, 28 & 35th day. Animals were sacrificed on 36th day; sciatic nerve, spinal cord and brain were isolated for biochemical estimations (Brain monoamines, spinal GABA, total ROS in brain, spinal cord and sciatic nerve)and molecular expression.

Result

Pain, anxiety and depression (increased immobility at expenses of climbing behaviour) were significantly increased in CCI animals. Decreased brain noradrenalin and increased spinal ROS was observed in CCI animals compared to sham. LT 40, ZX 40 and LZ 25 and 50 decreased the mechanical allodynia, thermal hyperalgesia and allodynia. Similarly, LT (40 mg/kg), ZX (40 mg/kg) and their combination LZ (25 and 50 mg/kg) showed reduction in depression behaviour i.e. decreased time of immobility and increased climbing in FST. Modulation in brain monoamine levels and decreased spinal ROS were observed in LT 40, ZX 40, LZ 25 and LZ 50 treated animals. Similarly, modulation in protein and mRNA expression at Spinal cord were observed.

Conclusion

xanthophylls LT, ZX and LZ play their antinociceptive‐antidepressant role by modulating ROS & monoamines level at brain along with the protein and mRNA expression and ROS at spinal cord.
Knockout of the Ergothioneine Transporter ETT in Zebrafish Results in Increased 8‐Oxoguanine Levels
Authors: Krueger Johanna , Pfeiffer Carolin , Gruendemann Dirk
Published: June 2021

Ergothioneine (ET) is expected to provide antioxidative properties due to its double‐bonded sulfur. It is acquired by plants and vertebrates solely through dietary means. However, the precise physiological function of ET remains unclear. Case‐control studies suggest an association of polymorphisms in the human ET transporter (ETT) gene with chronic inflammatory diseases like rheumatoid arthritis [Nat Genet 35, 341‐348], Crohn's disease [Nat Genet 36, 471‐475] and diabetes [Biochem J 25, 171].

Methods

Here we have procured, based on retroviral insertion [Genes Dev 13:2713‐24], an ETT knock‐out zebrafish (Danio rerio). The expression profile of ETT in wild‐type zebrafish was analyzed by RT‐PCR and real‐time RT‐PCR and validated by measuring ET tissue content.

Results

Strong signals were obtained with kidney, intestine, skin, brain and eye. In whole‐fish lysates of homozygous knock‐out animals, ET content (0.06 ± 0.01 ng/g wet mass; n=3) was reduced more than 1000‐fold compared to wild‐type (138 ± 61 ng/g wet mass; n = 3). Similarly, all examined organs contained virtually no ET. Carnitine and acetylcarnitine were reduced across all examined tissues of knock‐out fish by a factor of 2.0 ± 0.2, and 1.7 ± 0.1, respectively. Incubation in sublethal concentrations of Pb2+ and Cu2+ led to an increase of the lipid peroxidation markers 4‐hydroxy‐2,3‐trans‐nonenal (HNE) and malondialdehyde (MDA) in knock‐out versus wild‐type zebrafish. Comparison of unstressed fish at the level of small molecules by LC‐MS difference shading [Proc. Natl. Acad. Sci. U.S.A. 102:5256‐61] revealed a 3.8‐fold increase of 8‐oxoguanine in the skin of ETT knockout fish.

Conclusion

Based on this phenotype we propose that the universal and specific purpose of ET is to eliminate singlet oxygen.
Characterization of Corning® HepatoCells for Drug Uptake Transport Assays
Authors: zuo Rongjun , Cooper Kirsten , Andes Mercyanne , Li Feng , Parikh Sweta , Li Na , Liu Jin , Faris Ronald , Crespi Charles , Patten Christopher
Published: June 2021

Increasing evidence shows that drug transporters play an important role in drug‐induced hepatotoxicity and adverse drug‐drug interactions. Regulatory agencies (USFDA and EMA) recommend in their drug interaction guidelines to investigate clinically relevant transporters for new molecular entities. Here we report characterization of Corning HepatoCells (derived from primary human hepatocytes) for uptake transporter study, specifically OCT1, OATP1B1/1B3, and NTCP. When cultured on Corning BioCoat™ Collagen I coated tissue culture plates for 3‐4 days, Corning HepatoCells demonstrated time‐dependent and concentration‐dependent uptake of prototypical substrates of OCT1, OATP1B1/1B3, and NTCP, with Km values comparable to literature data. Multiple lots of Corning HepatoCells generated similar kinetic profiles. Inhibition assays were also performed and the results showed concentration dependent inhibition with similar profiles among multiple lots of cells, and IC50 values were in line with literature data from other test systems. In addition to active expression of uptake transporters, Corning HepatoCells also demonstrated bile canaliculi formation suggesting active expression of efflux transporters. In conclusion, the study demonstrated that Corning HepatoCells actively express functional uptake transporters, and this model represents a renewable source of human hepatic cells with consistent performance for uptake transporter study.
Simple Method Provides Resolution of Albumin, Lipoprotein, Free Fraction, and Chylomicron to Enhance the Utility of Protein Binding Assays
Authors: Brockman Adam , Oller Haley , Moreau Benoit , Kriksciukaite Kristina , Bilodeau Mark
Published: June 2021

Medicinal chemists have been encouraged in recent years to embrace high speed protein binding assays. These methods employ dialysis membranes in 96 well format or in spin filters. Membrane‐based methods do not separate lipoprotein binding from albumin binding. While controls can account for membrane binding to an extent, membrane methods still introduce interference into the experiment. Ultracentrifugation methods, in contrast, do not introduce interference if density gradients can be avoided and they resolve lipoprotein from albumin. A new generation of compact, fast ultracentrifuges facilitate the rapid and fully‐informative separation of plasma into albumin, albumin/fatty‐acid complex, lipoprotein, protein‐free, and chylomicron fractions with no need of a salt or sugar density gradients. We present a simple and fast ultracentrifuge method here for two platinum compounds and a taxane that otherwise bound irreversibly to dialysis membranes and which exhibited distinctive lipoprotein binding behaviors. This new generation of ultracentrifugation methods underscores a need to further discuss protein binding assessments as they relate to medicinal chemistry efforts.
Prevention of Cardiovascular Disease at Outpatient Clinic Service in Kazakhstan
Authors: Turlybekova K.D. , Khismetova Z.A. , Rakhypbekov T.K.
Published: June 2021

High public and medical importance of diseases of the circulatory system (hereinafter ‐ SBR) due to their wide distribution, poor prognosis and significant social and economic losses. Components of the study for the prevention of coronary heart disease (CHD) were: detection and dynamic monitoring of individual groups (CHD) risk; training of patients with diseases of the circulatory system in the school, “Coronary heart disease”; adherence to the principles of continuity in the monitoring of patients with coronary artery disease between general practitioners (GPs), cardiologist and physician day hospital; empowerment of day hospitals; active health education outreach.

During the period from 2010‐2012 to in the medical preventive institution surveyed 1,852 people. Among examined patients 42.3% had a SBR requiring active interventions for secondary prevention, while 48% of them have never consulted a doctor, and the pathology was revealed during questioning and subsequently the active examination. 64.5% ‐ are at the ages of 50‐59 years. At risk group the largest share (30.5%) were in people aged 45‐54 years; 25.7% ‐ 44 years; 23.4% ‐ 50‐54 years; 17.8% of those at risk group in the age 55‐59 years and 21.6% ‐ 35‐40 years. Healthy people predominated in the age group 35‐39 years (55.3%).

Thus, considering that if hypertension is a strong risk of vascular events: stroke, myocardial infarction, treatment costs are significantly was higher (8‐12 times), than the costs of preventive measures for hypertension, it is necessary to develop ways of improving economic approach to the development of the above diseases.
Systemic and Microcirculatory Vasoactive Effects of a Third Generation Perfluorocarbon in a Rodent Top‐Load Model
Authors: Crossland Randy , Macko Antoni , Song Bjorn , Sheppard Forest
Published: June 2021

Background

Restoration of oxygen carrying capacity (OCC) is essential to adequately treat hemorrhagic shock. Multifunctional resuscitation fluids (MRFs) containing oxygen carrier (OC) molecules are being developed to improve OCC. However, adverse vasoactivity of previous generations of OCs has limited MRF development. Newer, third generation PFCs have garnered interest as potential OCs for use in MRFs. Here, as part of an evaluation as a potential OC for use in a MRF, we tested a third generation PFC for systemic and microvascular vasoactivity in a rodent model.

Methods

Male Sprague‐Dawley rats (250‐350 grams) were randomly assigned to receive four sequential intravenous infusions (1.5 mL/kg each) of either PFC (Oxycyte®; n=10) or 0.9% NaCl (control; n=10). At the conclusion of each study, phenylephrine (PE; 1 mg/kg) was infused to confirm physiological intactness of the animal. Heart rate (HR) and mean arterial pressure (MAP) were continuously monitored. Vasoactivity was assessed by intravital microscopy of spinotrapezius arcade arterioles. Statistical comparisons were made by ANOVA. Results reported as mean ± SEM.

Results

No observed differences were seen in MAP, HR, or microvascular arteriole diameters between groups at baseline or throughout the study (Table 1). In both PFC and control groups, PE constricted arterioles by 50% (p<0.001) and elevated MAP 2‐fold (p<0.001).

PFC (n=10)
Control (n=10)

Baseline
Final Dose
PE
Baseline
Final Dose
PE

MAP (mmHg)
94 ± 4
99 ± 5
198 ± 3
95 ± 2
90 ± 4
199 ± 5

HR (bpm)
362 ± 6
344 ± 36
326 ± 10
364 ± 9
359 ± 10
314 ± 11

Arteriole Diameter (µm)
58 ± 4
58 ± 4
26 ± 4
57 ± 2
62 ± 4
24 ± 4

Funding: DoD Hemorrhage and Resuscitation Research and Development Program (WUN G1018)

Conclusion

Although these data demonstrate no adverse systemic or microcirculatory vasoactive effects, further evaluation of the third generation PFC in a clinically relevant hemorrhage model is warranted.
It's Strong, It's Stable, It's Streptavidin
Authors: Futch Liane , Pham Tiffany , Adamson Bennett , Reilly Bianca , Kronenfeld Jason , Patterson Kiersten , Inostroza Martin , Fraijo Savanah , Montoya Joe , Chavez Ryan , Hazzard James
Published: June 2021

Many proteins bind to substrates, so what makes streptavidin binding biotin so amazing? Perhaps, the fact that its dissociation constant (Kd = 10‐15 M) is the lowest in nature. The bacterium Streptomyces avidinii has taken advantage of this incredibly low Kd to develop a defense mechanism with which it can withhold valuable biotin (vitamin B7) from competing bacteria. This high affinity binding is facilitated by the structure of tetrameric streptavidin, consisting of a dimer of dimers each formed by the interaction between monomers that involves a single amino acid, Trp120. Each subunit consists of an 8 antiparallel beta strand barrel forming the biotin binding site which is capped by a flexible loop between strands 3 and 4 which essentially acts as a lid over the binding site on the end of the barrel. Tight biotin binding results from multiple hydrogen bonding interactions, and a hydrophobic Trp‐lined binding site that allows for extensive van der Waal interactions. The strong streptavidin‐biotin interaction has been exploited in biotechnology for processes such as protein identification, purification, isolation, and enrichment; as well as, nucleic acid detection involving binding of biotinylated targets by immobilized proteins. The Ironwood Ridge High School SMART Team (Students Modeling a Research Topic) has modeled streptavidin bound with biotin and bound with a Strep‐tag using JMOL and 3‐D fabrication to speculate on the mechanism, and to highlight the residues, which facilitate streptavidin's record breaking dissociation constant.
Reviving an Impaired Cardiomyocyte
Authors: Kunkel George , Chaturvedi Pankaj , Tyagi Suresh
Published: June 2021

Although cardiac resuscitation can revive the whole body, the mechanisms are unclear. To this end we propose that reviving a dysfunctional cardiomyocyte will shed light on resuscitation mechanisms and pave the way to treat cardiac myopathies. Calpain is a cardiac malignant protease responsible for the formation of multiple cardiomyopathies resulting in heart failure. Calpain upregulation causes degradation of cytoskeletal and contractile proteins resulting in decreased cardiac output and force of contraction. We hypothesize that upregulation of calpain 1 due to high calcium harbor by the PCP4 (Purkinje cell protein) and CAM (Calmodulin) complex activates CAMKII (Calmodulin Kinase II), leading to its autophosphorylation and eventually death of the myocyte. These consequences are accompanied by increase in the levels of MMP9, fibrosis due to higher collagen deposition and decreased levels of PCP4, worsening the condition. To validate the hypothesis we isolated cardiomyocytes using the Radnoti perfusion apparatus and observed myocyte death within 5 minutes of adding calcium (2mM). Western blotting showed high levels of calpain 1 along with calmodulin kinase II and PCP4 in the dead myocyte as compared to the live myocyte. Our preliminary data suggests the involvement of calpain 1, CAMKII and PCP4 in myocyte death. Interestingly, we found decreased levels of calpain 1 and PCP4 in the hearts of exercised mice with diabetes (db/db), suggesting that exercise plays a crucial role in reviving an impaired myocyte. The study examining the effects of exercise on myocyte resuscitation via calpain inhibitors in diabetic mice is currently in progress.
Divergent relationship between arterial baroreflex sensitivity and mental stress‐induced blood pressure reactivity in women and men of Oman Family Study
Authors: Albarwani Sulayma , Millar Philip , Picton Peter , Jaju Deepalu , Al‐Yahyaee Said , Bayoumi Riyadh , Floras John , Hassan Mohamed
Published: June 2021

The arterial baroreflex is important for buffering blood pressure (BP) responses to stress. Whether the sensitivity of the arterial baroreflex (BRS) is associated with hemodynamic responses to mental stress is sex dependent is unclear. We tested the relationships between BRS, BP and heart rate (HR) responses to a modified Word Conflict Test (WCT) in a cohort of semi‐rural, healthy Omani Arab women (n=69; 25±7 years; BMI: 23±5 kg/m2 [mean±SD]; BP: 104±10/65±8mmHg) and men (n=55; 24±7 years; BMI: 23±5 kg/m2; BP: 120±18/76±15mmHg). Resting BRS was similar in women and men (16±10 vs. 16±8 ms/mmHg). In women, the WCT increased (All P<0.01) systolic (Δ10±17mmHg), diastolic (Δ6±13mmHg), mean arterial pressure (Δ8±15mmHg), and HR (Δ8±15bpm). In men, the WCT increased (All P<0.01) systolic (Δ10±18mmHg), diastolic (Δ7±16mmHg), mean arterial pressure (Δ9±17mmHg), and HR (Δ13±15bpm). Resting BRS was +ve associated with changes in systolic (r=0.40, P=0.001), diastolic (r=0.34, P=0.003), mean arterial pressure (r=0.33, P=0.005) and HR (r=0.27, P=0.03) in women but ‐ve correlated with systolic BP (r=‐0.30, P=0.03) and HR (r=‐0.24, P=0.08) in men. These findings demonstrate a sex‐based difference in the role of the arterial baroreflex in buffering BP where mental stress in women showed higher vagal modulations of HR that may play a larger role in determining BP. As exaggerated stress reactivity is predictive of future hypertension, high BRS in women and low BRS in men may be important factors to identify individuals at risk.
Repeatability of Reactive Hyperemia Flow‐Mediated Dilation in the Superficial Femoral Artery
Authors: Schmitter Sarah , King Trevor , Pyke Kyra
Published: June 2021

Reactive hyperemia (RH) is induced via the release of a temporary limb occlusion and causes a transient increase in artery shear stress. Flow‐mediated dilation stimulated by RH (RH‐FMD) is a standard index of endothelial function in the brachial artery however, relatively few studies have assessed RH‐FMD in the superficial femoral artery (SFA). The purpose of this study was to determine the repeatability of RH‐FMD in the SFA. SFA RH‐FMD was measured in 19 subjects (10 males and 9 females; age: 24 ± 2 years). Subjects performed four 5‐minute occlusion RH trials in a seated position with the cuff placed above the knee. Six minutes of recovery followed cuff release. SFA diameter and blood velocity were assessed using echo and Doppler ultrasound, respectively. Shear stress was estimated using shear rate (SR = mean blood velocity/SFA diameter). RH‐FMD was calculated as the % increase from baseline diameter to the peak 3 second diameter measurement post occlusion release. Results are expressed as mean ± SD. The RH SR area under the curve (AUC) for the 30 seconds following cuff release was higher in trial 1 versus trial 2 and 3 (p < 0.001 and p = 0.021). There were no differences in %RH‐FMD across trials (RH trial 1: 5.2 ± 2.6%, trial 2: 5.0 ± 2.3%, trial 3: 4.5 ± 2.2%, and trial 4: 5.0 ± 2.9%, p = 0.465). FMD normalized to the SR AUC also did not differ between trials (p = 0.879). In conclusion, despite some variability in the SR stimulus, there was no systematic change in %RH‐FMD over four repeated trials.

Funded by NSERC and CFI
Effects of Pamidronate in Acute Cardiotoxicity Induced by Doxorubicin in Rats
Authors: Polegato Bertha , Carvalho Paula , Bergamasco Carolina , Gonçalves Andrea , Azevedo Paula , Modesto Pamela , Roscani Meliza , Fernandes Ana , Paiva Sergio , Zornoff Leonardo , Matsubara Luiz , Okoshi Marina , Minicucci Marcos
Published: June 2021

Aim

Evaluate pamidronate (Pam) effects in ventricular function, matrix metalloproteinase‐2 (MMP‐2) activity, oxidative stress, and myocardial energetic metabolism during acute doxorubicin (Dox)‐induced cardiotoxicity.

Methods

64 Wistar rats were allocated in 4 groups: Control, Dox, Pam and Dox + Pam. Rats received Pam (3 mg/kg, IP) or saline. After 24 hours, rats received Dox (20 mg/kg, IP) or saline. Forty‐eight hours after Dox injection, rats were euthanized. Statistical analysis: two‐way ANOVA.

Results

Dox induced diastolic and systolic ventricular dysfunction and increased MMP‐2 activity. Dox increased myocardial oxidative stress (increased in lipid hydroperoxide, and decreased in catalase, superoxide dismutase and glutathione peroxidase myocardial levels) and impaired myocardial energetic metabolism (increased in phosphofructokinase and decreased in β‐hydroxyacyl dehydrogenase myocardial levels). Pam associated with Dox did not change ventricular function or MMP‐2 activity, but it decreased lipid hydroperoxide (p=0.042) and increased catalase (p<0.001) and superoxide dismutase levels (p<0.001). Also, Pam improved phosphofructokinase (p=0.021) and β‐hydroxyacyl dehydrogenase levels (p<0.001).

Conclusion

Pam attenuated acute Dox‐induced cardiotoxicity because it decreased oxidative stress and modulated myocardial energetic metabolism.

Acknowledgement: PROPe‐UNESP and FAPESP.
Effects of Aging on Wire Injury‐induced Neointimal Hyperplasia: An in vitro and in vivo Study
Authors: Wu Chieh‐Hsi , Pan Chun‐Hsu , Wu Chi‐Han
Published: June 2021

In the balloon angioplasty‐injured vascular tissues, abnormal cell proliferation and migration of smooth muscle cells have been believed to play an important role during the disease progression. It still lacks final conclusion and research evidences at present to clarify whether possessing the characteristics of advanced aging will become a susceptibility factor of restenosis. Therefore, the senescence accelerated mouse‐prone 8 (SAMP8) was applied as animal model to induce neointimal hyperplasia by wire injury in this project. The statistical results suggested that the ratio of neointimal‐to‐medial area (N/M ratio) was lower in the SAMP8 mice than in the wild‐type mice of the same age, which implicated that the aging factor may reduce the severity of neointimal hyperplasia. The immunohistochemistry staining was further used to examine the expression of β‐galactosidase, a senescence marker, in the vascular tissues. The level of β‐galactosidase was higher in SAMP8 mice than in the wild‐type mice of the same age. Hence, a cellular senescence model was induced by CoCl2 treatment in the vascular smooth muscle cells (VSMCs) to investigate the regulations and mechanisms of senescence in the cell proliferation and migration of VSMCs. We found that CoCl2‐induced senescent VSMCs to the serum‐triggered stimulations in the cell growth and migration were negatively correlated with treatment concentration of CoCl2. Similarly, this phenomenon was also found in the regulation of AKT and ERK 1/2. Thus, aging factor may reduce the sensitivity of VSMCs to the exogenous stimulatory factors with consistent results to the in vivo study.
Within Subject Variability and Duration of Brachial Artery Dilation in Response to GTN
Authors: McPhee Iain , Levac Brendan , Pyke Kyra
Published: June 2021

Glyceryl trinitrate (GTN) elicits an endothelium independent dilation by supplying exogenous nitric oxide to vascular smooth muscle. GTN tests are often used in conjunction with flow mediated dilation (FMD) testing to isolate impaired FMD to an endothelium dependent mechanism. The purpose of this investigation was to assess trial to trial variability in GTN induced dilation. A secondary purpose was to evaluate the duration of diameter elevation post GTN administration. A 400‐µg GTN dose was administered sublingually to 17 healthy male subjects (23±3 years) at two time points, trial 1‐ 0min, and trial 2‐ 160min. Pre GTN baseline (1min) and post GTN (10min continuously) artery diameters were measured via ultrasound. Diameter was also measured for 1min at 40min, 80min, and 140min post trial 1 GTN administration. The response to GTN was determined as the percent change from the pre GTN baseline diameter to the peak 30 second average diameter post GTN administration. The within subject trial to trial coefficient of variation (CV) (SD/Mean*100) was calculated from the percent change from baseline for each subject after GTN trial 1 and 2. Data are mean ±SD. Percent change in diameter was not different between trials (21.4± 5.7% vs. 22.4± 5.9%, p=0.153). Within subject trial to trial CV for the GTN response was 7.9± 6.7%. Brachial artery diameter remained elevated from the trial 1 baseline at 40min (p<0.001), and 80min (p<0.001), but not 140min (p=0.513) post GTN administration. These results suggest that there is a fairly low within subject variability in the response to GTN between trials, and that the dilatory response to GTN may have persisted for at least 80 min post GTN administration. The potential for prolonged dilation in response to GTN must be considered when planning assessments pre/post acute interventions. NSERC, CFI
Evidence that Shift Work Influences the Relationship Between Reactivity to Mental Stress and Indices of Vascular Health: A Pilot Study
Authors: Pyke Kyra , Tranmer Joan , Carson Ira , Batson Morgan
Published: June 2021

High cardiovascular reactivity to stressful tasks is a predictor of increased future cardiovascular risk and may play a mechanistic role in disease progression. The purpose of this investigation was to examine the relationship between acute cardiovascular stress reactivity and indicators of vascular health status in shift workers (SW) and non‐shift workers (NSW). Female hospital employees were recruited. 18 NSW and 19 SW (minimum 6 yrs experience) participated. Blood pressure (Finometer), central pulse wave velocity (PWV) (carotid to femoral; applanation tonometry) and common carotid artery intima media thickness (IMT) (echo ultrasound) were assessed at rest. With continued blood pressure assessment, participants then completed a 20 min mental stress task with speech and arithmetic components (based on the Trier Social Stress Test). Systolic blood pressure (SBP) reactivity was quantified as the difference between the resting baseline SBP and the SBP during the stress task. Data are mean ± SD. SW and NSW did not differ with respect to age (42 ± 11 yrs, p=0.563), central PWV (6.6 ± 1.3 m/s, p=0.416), IMT (0.47 ± 0.11 cm, p=0.207) or SBP reactivity (20.3 ± 11.2 mmHg, p=0.717). In NSW there was no relationship between SBP reactivity and either PWV (p=0.789; r=0.07) or IMT (p=0.876; r=0.06). In contrast in SW, both PWV (p=0.032; r=0.507) and IMT (p=0.062; r=0.435) increased with increasing SBP reactivity. These data suggest that shift work experience may alter the relationship between cardiovascular stress reactivity and vascular health. Garfield Kelly Cardiovascular Research and Development fund, KGH.
PDE8A Shapes The Profile of Cyclic AMP Signaling Events: Assessing The Importance of PKA‐dependent Feedback Regulation
Authors: Jiang Jason Yusheng , Abi‐Jaoude Joanne , Nichols Jonathan , Curci Silvana , Hofer Aldebaran
Published: June 2021

Phosphodiesterases (PDEs) function as scavengers to clear the cyclic nucleotide messengers cAMP and cGMP inside cells. PDE8 is a cAMP‐specific PDE participating in the control of many cell functions, including steroidogenesis, T cell adhesion, lymphocyte chemotaxis, and cardiac excitation–contraction coupling. However, PDE8 is not sensitive to IBMX (a universal inhibitor of the PDE family), and the lack of available pharmacological tools has hindered the study of PDE8 isoforms in living cells. Here we used FRET sensors Epac H30 and AKAR4 to image cAMP and PKA activity, respectively, in cultured colonic epithelial cells overexpressing either PDE8A or mCherry‐PDE8A. We report that overexpressed mCherry‐PDE8A is distributed throughout the cytoplasm but is generally excluded from the nucleus. The presence of this PDE isoform powerfully suppressed isoproterenol (ISO) stimulated cAMP signaling. Next, taking advantage of a newly synthesized high affinity PDE8 inhibitor, we observed that PDE8 was ordinarily quiescent in resting cells, however, following elevation of intracellular cAMP, PDE8 inhibition caused cAMP to increase by 2.5‐fold, suggesting dramatic cAMP‐dependent upregulation of PDE8 activity. Mirroring the effect of cAMP, we found that PKA activity (as measured by the AKAR4 ratio change) was increased 3.0‐fold under these conditions. Specifically eliminating PKA activity using PKI‐mCherry enhanced (by 1.9‐fold) ISO‐induced cAMP signals. Our data show that PDE8 is able to strongly influence the profile of the cAMP signal, and this is achieved through PKA‐dependent modification of PDE8 activity.
Airway exposure of e‐cigarette‐vapors impairs autophagy and induces aggresome‐formation
Authors: Shivalingappa Prashanth , Westphal Colin , Vij Neeraj
Published: June 2021

Objective

Electronic‐cigarettes (e‐cigarettes) are proposed to be a safer alternative to tobacco‐cigarettes. Hence, we evaluated if e‐cigarette‐vapors (eCV) impairs cellular proteostasis similar to cigarette‐smoke (CS)‐exposure.

Results

First, we evaluated the impact of eCV‐exposure (~2.5mg or 7.5mg) on Beas2b cells that showed significant increase in accumulation of ubiquitinated‐proteins (ub, insoluble‐fractions) with time‐dependent decrease in proteasomal‐activities from 1hr to 6hrs (p<0.01) of eCV‐exposure as compared to room‐air control. We verified that even minimal eCV‐exposure (1hr) induces valosin containing protein (VCP; p<0.01), sequestosome‐1/p62 (aberrant‐autophagy marker; p<0.05) and aggresome‐formation (ub‐accumulation) using immunoblotting (IB), fluorescence microscopy and immunopreciptation (IP). The inhibition of protein synthesis by 6hr cyclohexamide (50µg/ml) treatment significantly (p<0.01) alleviates eCV‐induced (1hr) aggresome‐bodies. We also observed that eCV (1hr) induced protein aggregation can activate apoptosis (caspase‐3/7) and senescence (SA‐β‐gal) activities (p<0.01) as compared to room‐air controls. We verified using carbamazepine (CBZ, 20µM, 6hrs) that eCV induced changes in ub‐accumulation, proteasomal activity, autophagy, apoptosis and senescence could all be controlled by autophagy induction by CBZ. We further confirmed the role of acute eCV‐exposure on autophagy impairment in murine lungs (C57BL/6 and CD1) by IB (ub, p62, VCP) and IP (VCP, p62) similar to in‐vitro experiments.

Conclusion

eCV induced autophagy impairment and aggresome‐formation activates apoptosis/senescence suggesting its potential role in COPD‐emphysema pathogenesis.
CFTR: Structural Mishap by Mutation
Authors: Link T , Arpornsukant P , Chareddy A , Alluri A , Chyong D , Eshleman C , Hu M , Jain S , Kahn J , Kemelmakher H , Kepner A , Krishnaswamy A , Raman V , Sanders K , Stock G
Published: June 2021

The CFTR (Cystic Fibrosis Transmembrane Conductance Regulator) protein is a member of the ABC (ATP‐binding cassette) family, which functions as a low conductance channel regulating the flow of certain anions (especially chloride and thiocyanate) across membranes of epithelial cells, allowing for osmosis‐based regulation of mucus consistency. The proper function of CFTR depends on the amino acid phenylalanine at position 508, which mediates a tertiary interaction between the surface of the N‐terminal nucleotide‐binding domain and cytoplasmic loop‐4 in the C‐terminal membrane‐spanning domain. Miscoding caused by the most common identified mutation, ΔF508, results in loss of phenylalanine by deletion of base pairs G, A, and A from the sequence “T‐A‐G” at position 507 and adjacent “A‐A‐A”, resulting in a codon made for isoleucine, but not for phenylalanine. Inheritance of two copies of the mutation causes part of the CFTR protein to misfold and degrade, not becoming fully processed in the endoplasmic reticulum. This mutation, among others, leads to a class II mutation of cystic fibrosis. The misfolded CFTR is either not transported to the surface of the membrane or is only transported in residual amounts, preventing the regulation of mucus consistency and causes cystic fibrosis. The mechanism of the ATP‐dependent gating mutation in CFTR protein can be further understood through modeling. Understanding the mutation that causes the misfolding of CFTR can help develop new treatments for those afflicted with cystic fibrosis. Walton High School SMART (Students Modeling A Research Topic) Team has designed a model of CFTR using 3D printing technology to investigate structure‐function relationships.
ATP Antagonism Attenuates the Reflex Cutaneous Vasoconstriction Response in Humans
Authors: Smaller Kevin , Lang James
Published: June 2021

Norepinephrine is responsible for ~60% of the reflex cutaneous VC response in young individuals, whereas the remainder is due to a coreleased sympathetic adrenergic neurotransmitter(s) that has yet to be identified. There is conflicting evidence supporting neuropeptide Y in this role; however, other putative cotransmitters have yet to be tested in human skin. We hypothesize that ATP contributes to the reflex cutaneous VC response to whole‐body cooling (Tsk = 30.5 oC). Three microdialysis (MD) fibers were placed in the forearm skin of 7 (25 ± 1 years; 4 males, 3 females) individuals for infusion of 1) lactated Ringer's solution (control), 2) 10 mM L‐NAME, and 3) purinergic receptor blockade with 1mM suramin + L‐NAME. Laser Doppler flux (LDF) was measured over each MD site and cutaneous vascular conductance (CVC) was calculated as CVC = LDF/mean arterial pressure and expressed as a percent change from baseline (%ΔCVC). L‐NAME was used to block the vasodilatory influence of ATP and unmask the P2X‐mediated VC response to exogenous ATP infusion (‐18 ± 2 %ΔCVC). This response was blocked in the suramin‐pretreated site (‐2 ± 1 %ΔCVC). During whole body cooling (Tsk = 30.5 oC), the VC response in the suramin site was attenuated (‐24 ± 3 %ΔCVC; p < 0.05) compared to the control site (‐35 ± 4 %ΔCVC). These data suggest that ATP has a contributory role in the reflex cutaneous VC response in humans during whole‐body cooling.

**Funded by Iowa Osteopathic & Education Research Grant (IOER #03‐14‐01)
Disadvantages and complications of laparoscopic operations
Authors: Fursov A.B. , Ismailov A.S. , Kuspaev E.N.
Published: June 2021

Analyzed 5000 laparoscopic cholecystectomy performed various clinics and complications during surgery. The main difficulties and errors in the treatment of patients.

Found that many complications during the operation associated with the method of laparoscopic surgery. This is primarily facilitated by Visual inspection (in 35% of cases), in the zone of operation of several instruments of cops, working part of each of them can be a conductor of electrical current one. That resulted in significant currents with burns the internal organs (10%). in addition, our use of irrigation during the operation contributed to the electrical conduction along the dièlek‐electric facilities on top of the metal tools. This led to the emergence of assessmenttion ways current through liquid and electric sparks (0,5%).

A complication in the postoperative pancreatitis we observed only one (0.2%) case. Wound suppuration and formation of inflammatory infiltrations of the anterior abdominal wall were 0.4% of patients (paraumbilikal‐infection of the wound). Pneumonia is diagnosed in the postoperative period, only 0.2% of patients.

4,7% of patients have been simultaneous operational intervention. In the second halfofthe cases is cholecystectomy and hernia repair (paraumbili‐locally or umbilical hernia). Overall postoperative lethality was 0.66%. Overall‐a single case of death of the patient after through injury of abdominal aorta trochar.Severe intraoperative complications (biliary injuries Pu Tei) in 3 times more likely to occur under emergency laparoscopic holecistèktomiâh about the destructive forms of cholecystitis than routine operations.
Analysis of the reasons of development of lethal cases of surgical patients
Authors: Ismailov A.S. , Kuspaev E.N. , Fursov A.B.
Published: June 2021

The retrospective analysis of stories of diseases of the died patients of office of the general surgery is carried out to the period from 2011 to 2013. Among 121 died sick offices of the general surgery 51 patients in age group were more senior than 60 years (42,1%). So at 31 patients the heavy course of the main disease was aggravated with existence of accompanying pathology. 29 patients were hospitalized in a hospital in a critical condition.

Discussion

In 75% of cases it is necessary to state not preventability of development of a lethal outcome in view of weight of a condition of the patient. In 17% of cases development of a lethal outcome could be prevented at the timely request of the patient for medical care, at timely establishment of the correct diagnosis and at purpose of adequate treatment. In the analysis of the main diseases which have led to development of a lethal outcome of patients of office of the general surgery it is possible to divide into 3 groups.

The I group – patients with the diagnosis sharp pancreatitis, pancreatonecrosis (14,9%), and patients with the diagnosis sharp destructive cholecystitis (10,7%). These nosological forms are the most widespread in structure of the hospitalized patients therefore at arrival of patients with the complicated forms of a course of a disease at this category of patients the lethal outcome developed.

The II group – patients with diagnoses at which the high rate of a lethality (the cirrhosis complicated by a syndrome of portal hypertensia and bleeding from varicose and expanded veins of a gullet) is noted.

The III group – patients with not surgical diagnoses which were hospitalized because of a nontransportable serious condition of the patient (a myocardial infarction, pyelonephritis, disseminirovanny tuberculosis of lungs, purulentsalpingooophoritis).
PICK1 is Required for ASIC1‐Mediated Activation of NFATc3 in Pulmonary Arterial Smooth Muscle Cells
Authors: Yellowhair Tracylyn , Browning Carly , Herbert Lindsay , Giermakowska Wieslawa , Gonzalez‐Bosc Laura , Jernigan Nikki
Published: June 2021

The development of chronic hypoxia (CH)‐induced pulmonary hypertension is associated with increased pulmonary arterial smooth muscle cell (PASMC) Ca2+ influx through the acid sensing ion channel 1 (ASIC1) and subsequent ASIC1‐dependent activation of the Ca2+/calcineurin‐dependent transcription factor, nuclear factor of activated T‐cells isoform c3 (NFATc3). Both ASIC1 and calcineurin B have been shown to interact with protein interacting with C kinase 1 (PICK1) in neurons. Therefore, we hypothesize that PICK1 is a required scaffolding component facilitating the ASIC1‐dependent activation and nuclear translocation of NFATc3 in PASMC following CH. To test this hypothesis, we examined the interaction between PICK1/ASIC1/calcineurin B using the Duolink in situ proximity ligation assay. We found PICK1 interacts with both ASIC1 and calcineurin B in mouse PASMCs. We next determined the effect of CH (1 wk @ 380 mmHg) on PICK1 expression and found that CH increases the expression of PICK1 in mouse pulmonary arterial homogenates. We then examined the effect of PICK1 inhibition on endothelin‐1 (ET‐1; 100 nM) stimulated NFATc3 nuclear translocation. The PICK1 inhibitor, FSC231 (50 µM) abolished ET‐1 mediated nuclear translocation of NFATc3 in mouse PASMC. Together, these data show PICK1 is required for the ASIC1‐dependent activation and nuclear translocation of NFATc3 in PASMC. This ASIC1/PICK1/NFATc3 signaling complex may contribute to the vascular remodeling and increased vascular contractility associated with CH‐induced pulmonary hypertension.
Intramyocellular Triacylglycerol is Associated with Peroxisomal Biogenesis in Skeletal Muscle from Lean and Obese Humans
Authors: Huang Tai‐Yu , Zheng Donghai , Park Sanghee , Houmard Joseph , Hickner Robert , Cortright Ronald
Published: June 2021

Obesity is associated with elevated levels of lipids (intramyocellular triacylglycerols; IMTG) and reductions in mitochondrial fatty acid oxidation in skeletal muscle, both which are associated with insulin resistance. Known predominately for their actions in liver, peroxisomes are subcellular compartments essential for lipid disposal by chain‐shortening very long‐ and long‐chain fatty acids to acyl‐carnitines, permitting CPT‐1 independent entry into the mitochondria for oxidation. We hypothesized that peroxisome proliferation in human skeletal muscle occurs as an adaptive response to elevated lipid supply and accumulation (IMTG) which could facilitate mitochondrial fatty acid oxidation. Assays for the peroxisomal biogenesis marker PEX19 protein content and IMTG were assessed using vastus lateralis from obese and lean humans. Peroxisomal membrane protein 70 (PMP70) was measured after lipid incubation of human skeletal muscle primary myotubes. Results: 1. A significant relationship between tissue IMTG and PEX19 was identified in human skeletal muscle tissue (P<0.05) but not % body fat or BMI 2) PEX19 protein content was elevated (P<0.05) in tissues from obese individuals with high IMTG and 3) An increase in peroxisomal PMP70 was observed (P<0.05) with high fat incubation of human primary myotubes.

Conclusion

This is the first report associating lipid oversupply, IMTG accumulation, and peroxisomal biogenesis in human skeletal muscle suggesting a functional‐adaptive role for peroxisomes in mitochondrial lipid oxidation and obesity.