bluuebunny/crossref_metadata_2025 huggingface dataset:
The Effects of Dietary Cinnamic Acid and p‐coumaric Acid on Glucose and Lipid Metabolism in Normal Rats Fed a High‐Sucrose Diet
Authors: Kishida Kunihiro , Takeshima Ken , Ozaki Yoshihiko , Ihara Hayato
Published: June 2021

Cinnamic acid (CIN) and p‐coumaric acid (COU) are widely distributed in edible plants and ingested by human in normal diet. CIN and COU are highly absorbed via the monocarboxylic acid transporter in gastrointestinal tract. The hypoglycemic effect of CIN on diabetic mice and rats has been observed while we reported the hypolipidemic effect of COU on normal rats fed a high‐fat, high‐cholesterol diet. In this study, we tested whether CIN and COU improved glucose and lipid metabolism in normal rats fed a high‐sucrose diet. Male SD rats were fed a high‐sucrose (50 %) diet containing 0.2% CIN or COU for 120 days. No significant differences were observed in total food intake, body, liver and visceral fat weight, serum glucose, lipid profile, aspartate transaminase (AST) and alanine transaminase (ALT) levels, hepatic triacylglycerol and cholesterol levels among all groups. Dietary CIN but not COU significantly decreased the hepatic mRNA expression of liver X receptor α (LXRα), ATP‐binding cassette, sub‐family A, member 1 (ABCA1), low density lipoprotein receptor (LDLr) and sterol regulatory element binding protein 2 (SREBP2) compared to control group. There were no significant differences among all groups concerning the other hepatic mRNA expression involved with lipid metabolism (ACC1, FAS, ME, G6PDH, SCD1, SREBP1c, ChREBP, CPT1a, PPARα, MTP, CYP7A1) and glucose metabolism (GK, G6Pase, F1,6BP, PEPCK1, PEPCK2). These results suggest CIN and COU did not improve glucose and lipid metabolism in normal rats fed a high‐sucrose diet although dietary CIN induced some variations in gene expression involved with lipid metabolism.
D‐xylose as a sugar complement exerts antidiabetic effects in streptozotocin‐nicotinamide‐induced diabetic rats
Authors: Kim Eunju , Kim Yoo‐Sun , Kim Kyung‐Mi , Jung Sangwon , Yoo Sang‐Ho , Kim Yuri
Published: June 2021

Diabetes mellitus is a complex metabolic disease. Type 2 diabetes is more frequent and characterized by insulin resistance. D‐xylose is a sucrose inhibitor and may be useful as a functional sugar complement to inhibit increase of blood glucose level. The aim of study was to evaluate the antidiabetic potential of D‐xylose in streptozotocin‐nicotinamide‐induced diabetic rats. The rats were divided into following groups; I ‐nondiabetic, II ‐ diabetic control (DB), III ‐ DB + sucrose mixed with 5% D‐xylose (Xylo 5), IV‐ DB + sucrose mixed with 10% D‐xylose (Xylo 10). Xylo 10 supplementation significantly reduced fasting serum glucose level (P < 0.05). Xylo 10 tended to exert lower area under the glucose curve (AUG) and increase the level of insulin compared to DB, but they were not statistically significant. Xylo 10 tended to down‐regulate the level of phosphoenolphyruvate carboxylase (PEPCK). In blood analysis, the number of red blood cells and the level of hematocrit were significantly increased in Xylo 10 compared to DB. Histopathologically, whereas focal necrosis and dilated acini were observed in pancreas of rats in DB, both Xylo 5 and Xylo supplemented rat showed minimal pathological changes. In conclusion, D‐xylose may be useful for improving abnormalities of diabetic conditions.
Ethanol extract of Pinus eldarica needle reduces atherosclerosis in ApoE‐deficient mice
Authors: Jang Yeon Jeong , Jung Yun Joo
Published: June 2021

Atherosclerosis is a chronic inflammatory disease initiated by vascular inflammation and monocyte recruitment in part. Pinus eldarica needle have been widely used in traditional medicine for the treatment of various diseases. However, the effect of ethanolic extract of Pine eldarica needle on inflammatory diseases like atherosclerosis is not known yet. Herein we evaluated whether ethanolic extract of Pine eldarica needle has a protective effect against atherosclerosis in experimental mice. 6 male ApoE –/– mice and 6 male control (C57BL/6J) mice were fed with 30% high‐cholesterol diet for 21 weeks. 6 of the ApoE –/– mice were orally administered pine needle extract(10 mg/kg and 20 mg/kg body weight, every two days for 21 weeks). Pine needle extractshowed the suppressive effects on the expression of endothelial intercellular adhesion molecule 1 (ICAM‐1) and vascular cell adhesion molecule 1 (VCAM‐1) and the serum levels of CCL2, CCL5, IL‐6 and TNF‐α in ApoE –/– mice. In addition,administration of pine needle extractattenuated the serum levels of oxLDL, HDL, LDH, triglyceride and total cholesterol. Furthermore, the serum levels of SREBP‐2, LOX‐1 and LXRs were inhibited by the pine needle extract. Collectively, these results suggest that of ethanolic extract of Pine eldarica needle might play a protective role against atherosclerosis.
Hemostasis in Epileptic Seizures
Authors: Sanyk Oleksandr , Zaporozhets Tetiana , Sanyk Lidia , Bodulev Alexei
Published: June 2021

Convulsive seizures can be accompanied by serious changes in the body. The purpose of this study is to investigate the platelet aggregation using the Born method and the hemolysate aggregation test, coagulation hemostasis (plasma recalcification time, prothrombin time (PT), thrombin time (TT), activated partial thromboplastin time, antithrombin III, fibrinogen, beta‐naphthol test, soluble fibrin monomer complex (SFMK), euglobulin clot lysis time (ECLT) and histological changes of the brain in 20 rats after an epileptic seizure caused by parenteral administration of Nikethamide. All animal experimentation protocols were approved by the UMSA Animal Ethical Committee. Immediately after the convulsive seizure the platelet count, the height of aggregatogram and index of platelet aggregation have been reduced. Shortening of the prothrombin and thrombin time in intact fibrinogen concentration, decreasing of SFMC without changing of ECLT was revealed. Blockade of microcirculatory, injury of the vascular wall, perivascular changes (edema, hemorrhage) were observed in the brain. These results suggest that we have identified laboratory signs of disseminated intravascular coagulation (DIC) (hypercoagulation phenomena, thrombocytopenia, and thrombocytopathia), which were confirmed by morphological features. These data can be used in the management of patients with seizures.
Dilute Russell's Viper Venom Clotting Time (DRVVT) is Useful for the Routine Monitoring of Newer Non‐Vitamin K Anticoagulants
Authors: Brayne Mark , Kola Jake , Hoppensteadt Debra , Iqbal Omer , Fareed Jawed
Published: June 2021

The newer non‐vitamin K anticoagulants such as rivaroxaban (Bayer Healthcare) (R), apixaban (BMS/Pfizer) (A) and dabigatran (Boehringer Ingleheim) (D) have been approved to manage thrombotic and cardiovascular disorders. This study was designed to determine which of the available clotting assays could be used to monitor these new oral anticoagulants in plasma.

Materials

Citrated blood was drawn from ten donors and platelet poor plasma (PPP) was obtained. The PPP was supplemented with A, R and D in a concentration range of 0‐1.0 ug/ml. In addition, plasma samples, containing varying concentrations of either A, R or D were also blindly analyzed. The plasma samples were analyzed using PT/INR (Innovin, Seimens, Deerfield,IL), APTT reagent (Platelin, TCoag, Ireland) dilute Russell's viper venom time (DRVVT) and DRVVT confirm (HemosIL, Instrumentation Laboratories (Bedford, MA).

Results

In the PT and APTT assays a concentration dependent modest increase in clotting time was observed. In contrast, both the DRVVT and DRVVT confirm assays the concentration‐response curves were straight lines with a steep slope. Therefore, it was possible to extrapolate the concentrations of the unknown samples using both of these assays. In addition , there was a good correlation between the calculated concentration and the actual concentration in the range of 50 to 1000 ng/ml (r2=0.82) with both assays.

Conclusions

These results suggest the dRVVT and DRVVT confirm assays are sensitive to A, D and R at concentrations > 50 ng/ml. Therefore, A,D and R can be monitored by these assays. Both of these assays are commercially available and can be readily adapted to any automated instrument.
IL‐6 Production in Human Bronchial Epithelial Cells in Response to House Dust Mite Allergen
Authors: Lewis Christina , Fischer Horst
Published: June 2021

Airway epithelial cells lie at the interface between host and environment, representing the first line of defense against pathogens. In addition to its role as protective barrier, the airway epithelium can activate underlying dendritic cells, thereby initiating and prolonging airway inflammation. Furthermore, alterations in epithelial ion transport mechanisms can modify many aspects of its signaling, mucus properties, bacterial defenses, and induction of immunity. IL‐6 is a proinflammatory cytokine produced mainly by the innate immune system and is increased in many pulmonary diseases, but its role(s) remain unclear. The objective of our research is to determine the mechanism(s) by which the airway epithelium drives the immunopathology of allergic asthma and to clarify the relationship(s) between the epithelium and IL‐6 in asthma pathogenesis. Normal Human Bronchial Epithelial (Lonza) were grown on collagen‐coated transwell inserts and cultured at ALI until full differentiation (~20 d, TER ~500 Ω), and subsequently treated with house dust mite allergen (HDM, 50 or 100 ug/mL; Greer) for 24 h. IL‐6 production was measured via ELISA (R&D), and increased in dose‐dependent manner after HDM treatment (50 ug/ml HDM: 878±235 pg/mL, or 142% increase, and for 100 ug/mL of HDM:1325±18.7 pg/mL, or 215% increase, p=0.04) compared to sham treatment (618±433 pg/mL). Our data indicate that bronchial epithelial cells may drive allergic asthma via IL‐6 production. Our ongoing studies seek to determine the relationship(s) between the airway epithelium, IL‐6 production, and other aspects of epithelial dysfunction in asthma pathogenesis, including dysregulated mucus production and epithelial ion transport.
Visualization and Objective Quantification of Autonomic Circuitry in Mouse Area Postrema Using a Genetic Method
Authors: Gokozan Hamza , Corcoran Sarah , Czeisler Catherine , Gygli Patrick , Catacutan Fay , Baig Faisal , Otero Jose
Published: June 2021

The transcription factor PHOX2B is a master regulator of autonomic nervous system function as well as the specification of noradrenergic neurons. Incomplete maturity and/or absence of noradrenergic neuron networks are a proposed mechanism of infant breathing disorders including apnea of prematurity, sudden infant death syndrome, and congenital central hypoventilation syndrome (CCHS).

PHOX2B mutations result in the “chemosensation” loss characteristic of CCHS, resulting in poor respiratory drive secondary to hypercarbia (high pCO2). The objective of our work is to identify PHOX2B derived structures in the developing brain and generate new animal models capable of tracing the neuronal connections between neuronal nuclei during embryonic and postnatal development.

We utilized a PHOX2B‐CRE X ROSA‐GFP mouse to trace the appearance of PHOX2B‐derived cells in nuclei of the hindbrain. In order to trace the connections between these nuclei other hindbrain nuclei, we utilized a transgenic mouse in which targeted neurons display traceable processes and synapses. This transgenic mouse reagent (Jackson Labs stock number 010590) shows CRE‐mediated expression of tdTomato in cell bodies and GFP expression at synapses. Using these conditional gene targeting approaches we were able to conclude that neurons derived from PHOX2B progenitor domains innervate certain hindbrain structures.

By using our transgenic approach, we were able to (1) identify PHOX2B derived progenitors generate both neurons and glia, (2) show PHOX2B derived neuronal synapses in Ventral Medulla and Area Postrema. We also present an unbiased stereology method which can be used to determine the complexity of PHOX2B neuronal network.
Protein Expression of Alzheimer's disease‐ and Reduced Hippocampal Volume‐ Risk Loci in Human Hippocampus
Authors: Adams Stephanie , Norman Tim , Tilton Kathy , Seshadri Sudha , Delalle Ivana
Published: June 2021

We examined the protein expression of novel genome‐wide association studies (GWAS) identified risk alleles implicated in Alzheimer's disease (AD) and low hippocampal volume using postmortem hippocampi with no or mild‐to‐moderate AD‐associated pathological changes. We found BIN1, the most significant late‐onset AD susceptibility locus after APOE, expressed in glia of all the examined cases while the punctate signal in neuropil varied substantially. EPHA1, another confirmed AD risk locus, is an ephrin receptor implicated in neuronal apoptosis. We found EphA1 in the neuronal cytoplasm, with a trend of decreasing signal intensity as AD progressed. Methionine sulfoxide reductase B3 (MSRB3) locus, critical for oxidative stress, was identified in GWAS on hippocampal volume. We found MSRB3 signal mostly as cytoplasmic puncta in pyramidal neurons from all of the examined cases, albeit in various amounts. Finally, LEMD3, a possible causal gene at the MSRB3 locus, was expressed in neurons as well as astrocytes. Neuronal expression of LEMD3 may increase with AD progression. Our data provide first insight into the human hippocampal cell type‐ and cell compartment‐specific protein expression of AD‐ and hippocampal volume reduction‐ risk alleles. We are currently examining the correlation of these proteins' expression with the cognitive status reflected in CDR (Clinical Dementia Rating) scores.
The onset and decay of heterologous tolerance to morphine after subcutaneous (sc) injection for varying lengths of time
Authors: Thompson Benjamin , McKeel Jackie , Taylor David
Published: June 2021

The development and decay of heterologous tolerance following chronic morphine exposure via pellet implantation is well documented in the guinea pig longitudinal muscle/myenteric plexus (LM/MP). However, this method of administration only permits qualitative analysis of the decay of tolerance because the time at which drug exposure ends is unknown. Therefore, we assessed tolerance development in animals treated with twice daily sc injections of morphine for 1, 2, 4, 7 or 10 days. The analgesic effect of morphine was assessed in vivo using the paw pressure test and the characteristics of tolerance evaluated by comparing the neurogenic twitch inhibition by DAMGO and 2‐CADO in LM/MP from the same animals. Decay of tolerance was determined by evaluating animals or tissues obtained from them at 0, 1, 2 or 4 days after treatment cessation. A significant level of tolerance was generated by 2 days after treatment, which peaked at 4 days and was maintained at 7 and 10 days of treatment. The data indicated that the time for return to baseline correlated to the magnitude of tolerance in all animal groups. By 2 days after treatment tolerance began to decline and by 4 days it was no longer significant. The fact that the decay of tolerance is a function of both the length of treatment and magnitude of tolerance developed reinforces the idea that tolerance following chronic treatment with morphine is an adaptive process produced by alterations in one or more of several cellular signaling proteins. Protein analysis of the LM/MP from these animals via western blot revealed no change in the amount of the µ, A1, or A2a receptors. These data indicate that the observed tolerance cannot be explained by a decrease in the number of receptors.
Meta‐Analysis of General Anesthesia on Total Knee Angioplasty and Total Hip Angioplasty
Authors: TSAI JEFFREY
Published: June 2021

Both total knee angioplasty (TKA) and total hip angioplasty (THA) may be carried out under general anesthesia, however, many surgeons prefer to perform the surgery under regional anesthesia due to the profound preemptive analgesic effects which are provided by the regional aesthetic techniques. Both centroneuraxis which includes spinal and epidural as well as peripheral nerve blocks are used for TKA and THA. Even though the former is used less frequently mainly in both TKA and THA due to the increased administration of low molecular heparins mainly with the resultant epidural hematoma formation, the latter is enjoyed more popularly. In this paper, we will discuss the use of peripheral nerve blocks for both TKA and THA. In addition, the surgery also includes use of femoral/sciatic nerve blocks as well as intra‐articular injection of the local anesthetics or even opioids at the knee and hip joints. They are mostly devoid of any extensive sympathectomy and helps in providing sufficient surgical analgesia along with a motor nerve block. Conclusively, this meta‐study is to analyze the fundamental pros and cons of perioperative anesthesia for both TKA and THA.
Anti‐hyperalgesic Effects of Imidazoline I2 Receptor Ligands and their Interactions with Opioids in a Rat Model of Inflammatory Pain
Authors: Thorn David , Zhang Yanan , Li Jun‐Xu
Published: June 2021

Pain affects > 30% of the U.S. population and currently available analgesics are not adequate to meet the clinical needs. Opioids are effective for treating moderate to severe pain; however, their use is limited due to the unwanted effects. We recently found that imidazoline I2 receptor ligands produce antinociceptive effects in rat models of chronic pain and also enhance the antinociceptive effects of morphine. However, little is known of their interaction with other opioids. This study examined the anti‐hyperalgesic effects of selective I2 receptor ligands (2‐BFI and phenyzoline) alone and in combination with oxycodone using the von Frey filament test in rats with complete Freund's adjuvant (CFA)‐induced inflammatory pain. In CFA‐treated rats, 2‐BFI (1‐10 mg/kg, i.p.) and phenyzoline (17.8‐56 mg/kg, i.p.) dose‐dependently produced significant antinociceptive effects. When studied as combinations, 2‐BFI and oxycodone produced additive interactions, while phenyzoline and oxycodone produced synergistic interactions for their effects against mechanical hyperalgesia in CFA‐treated rats. Taken together, these studies suggest that imidazoline I2 receptor ligands alone have anti‐hyperalgesic effects and also enhance the effects of opioids in a rat model of chronic pain. Therefore, drugs acting on the imidazoline I2 receptor may represent a novel class of pharmacotherapy for the management of chronic pain.
Subcellular Localization and Oligomerization of R7 RGS Proteins: a Potential Mechanism of Regulating GPCR Signaling
Authors: Tayou Junior , Wang Qiang , Slepak Vladlen
Published: June 2021

Regulators of G‐protein signaling (RGS proteins) accelerate the GTPase activity of Gi/o family heterotrimeric G‐proteins. Our work is focused on R7 family RGS proteins, which include RGS6, 7, 9 and 11. These proteins exist as obligatory dimers with the atypical Gβ subunit, Gβ5 and are involved in regulation of sensory transduction, addiction, memory and learning, and metabolism. We previously showed that RGS7 localizes not only to the plasma membrane but also to distinct cytoplasmic puncta, both in primary neurons and transfected cells. Here we show that cytoplasmic RGS7 colocalizes with vesicles containing neurotrophic factors such as BDNF. We also used chemical crosslinking, immunoprecipiatation and mass spectrometry to identify binding partners of vesicular RGS7. This approach identified a few candidate proteins. However, our results unexpectedly showed that RGS7 exist as an oligomer ((Gβ5‐RGS7)n , where n=2 or 3) in the vesicles. When R7 binding protein (R7BP) recruits RGS7 to the plasma membrane, it causes the departure of RGS7 complex from the vesicles and dissociation of the oligomeric form. The DEP (but not RGS or DHEX) domain of RGS7 is required for oligomerization. Gβ5 is essential for RGS7 stability but not for oligomerization. We also found that RGS7 hetero‐oligomerize with RGS6 and 11 but not with RGS9. Our on‐going studies are aimed on understanding the role oligomerization and vesicular localization play in signal transduction.
Systematic Analysis of Yeast F‐box Proteins Reveals a New Role of Ubiquitination in Polarity Establishment
Authors: Sheetz Joshua , Kelley Joshua , Miller Samantha , Hurst Jillian , Young Everett , Dohlman Henrik
Published: June 2021

Ubiquitination is a post‐translational modification that regulates protein activity by tagging specific substrates for degradation. Regulation of proteins by ubiquitination has been demonstrated to help terminate the response to mating pheromone in budding yeast. The yeast pheromone response is comprised of two branches, one leading to MAPK activation and gene transcription, the other leading to cellular polarization and shmoo formation. The Skp1/Cdc53/F‐box (SCF) ubiquitin ligase employs different F‐box subunits to recruit specific substrates for ubiquitination. In yeast, F‐box proteins have been identified as interchangeable components of the SCF complex. To identify pathway regulators, we conducted a screen in which we measured activation of the pheromone response in 16 yeast strains ‐‐ each lacking one of the candidate F‐box genes. We characterized the abilities of these deletion mutants to affect either the MAPK or the polarity branch of the pathway. Our study reveals that one F‐box protein, Pfu1, selectively regulates the polarity branch. Using live cell imaging in a microfluidic chamber, we demonstrate that Pfu1 is required to restrict the cell to a single polarity site. In contrast, Pfu1 is has no effect on MAP kinase activation or transcriptional induction. While polar axis formation is a fundamental and well characterized feature of budding, the mechanisms used to regulate cell polarity components during mating are largely unknown. The existence of an F‐box protein exclusively involved in mating projection development reveals ubiquitination as a means to independently regulate branches of a signal transduction pathway.
NSAIDs Cause Significant Changes in Gene Expression in Small Intestinal Epithelial Cells
Authors: Silver Kris , Lillich James
Published: June 2021

Non‐steroidal anti‐inflammatory drugs (NSAIDs) are among the most widely used drugs for the amelioration of pain and inflammation. Unfortunately, NSAIDs are commonly associated with a wide range of toxic side effects, particularly in the gastrointestinal tract (GI). Though indiscriminant inhibition of COX enzymes has been suggested as a cause of adverse GI side‐effects, mounting evidence indicates that COX inhibition and loss of prostaglandin production do not entirely explain NSAID‐induced GI effects, and that other mechanisms also contribute to NSAID toxicity. Currently lacking from the body of NSAID activity/toxicity data is a genomic level examination of the effects of NSAIDs in vivo on gene expression in intestinal epithelial cells. Therefore, the current project was undertaken to examine the effects of oral NSAID treatment on gene expression in rat duodenal mucosa. Rats were treated with control, indomethacin (non‐specific), or NS‐398 (COX‐2 specific) for 72 h. Rats were then euthanized and mucosal scrapings were collected from rat duodenum. RNA was isolated then subjected to Illumina sequencing and comparisons of gene expression in duodenal mucosa revealed 698 or 17 genes that had significantly altered expression following oral treatment with indomethacin or NS‐398, respectively. A total of eight genes (one up and seven down) were affected by both indomethacin and NS‐398 treatment, including stomatin (up), amylase 2A3, carboxyl ester lipase, RGD1561381, Tmem238, ENSRNOG00000015847, Lsm5, and Tceal8. Our results provide new insight into the mechanisms involved in NSAID‐induced GI toxicity and provide a basis for future experiments aimed at understanding the complex interactions between NSAIDs, intestinal epithelia, and the intestinal lumen.
Regulated Expression of the I Prostanoid Receptor (IP) Gene by Androgens: Combined role for Serum Cholesterol
Authors: Eivers Sarah , Kinsella B
Published: June 2021

The prostanoid prostacyclin plays a key cardioprotective role within the vasculature, acting as an inhibitor of platelet aggregation, vasodilator and promotes vascular repair in response to injury. Previous studies established that both estrogen and low‐cholesterol increase expression of the human prostacyclin receptor (hIP) through a transcriptional mechanism, potentially accounting for some of the cardioprotective effects of prostacyclin, estrogen and low‐cholesterol within the vasculature. There is increasing evidence that androgens may also confer cardioprotective effects, such as in men, but through unknown mechanisms. Herein, it was aimed to investigate whether androgens either alone or in various conditions of serum‐cholesterol may regulate expression of the hIP gene (PTGIR) within the vasculature. In normal serum conditions, the androgen dihydrotestosterone (DHT) was found to increase expression of the hIP and without any involvement of the functional cholesterol‐responsive sterol response element (SRE). The DHT‐responsive region was localized to an androgen response element (ARE), lying within close proximity to the functional SRE within the core promoter of the PTGIR. However, under conditions of low‐cholesterol, DHT led to substantial increases in hIP expression that occurred through a mechanism involving both the androgen‐responsive ARE and the cholesterol‐responsive SRE. Collectively, these data establish that the PTGIR is under the transcriptional regulation of androgens but that this regulation may be influenced by serum‐cholesterol levels. Funded by the Programme for Research in Third Level Institutions (PRTLI; MolCellBiol) and the European Regional Development Fund.
Role of SLMAP Genetic Variants in Susceptibility of Diabetes and Diabetic Retinopathy in Qatari Population
Authors: Upadhyay Rohit , Crystal Ronald , Ding Hong
Published: June 2021

Objective

Overexpression of SLMAP gene has been associated with diabetes and endothelial dysfunction of macro‐ and micro‐blood vessels. In this study, our primary objective is to explore the role of SLMAP gene polymorphisms and genetic variants of SLMAP in the susceptibility of type 2 diabetes, with or without type 2 diabetes associated retinopathy (DR), in the Qatari population.

Research design and methods

A total of 341 Qatari subjects (non‐diabetic controls and type‐2 diabetic patients with or without diabetic retinopathy) were genotyped for SLMAP gene polymorphisms (rs17058639 C>T; rs1043045 C>T and rs1057719 A>G) usingTaqman SNP genotyping assay.

Results

SLMAP rs17058639 C>T polymorphism was associated with the presence of diabetic retinopathy among Qataris with type 2 diabetes. One‐way ANOVA and multiple logistic regression analysis showed SLMAP SNP rs17058639 C>T as an independent risk factor for DR development. SLMAP rs17058639 C>T polymorphism also had predictive role for severity of DR. Haplotype Crs17058639Trs1043045Ars1057719 was associated with the increased risk for DR among Qataris with type 2 diabetes.

Conclusions

The data suggests the potential role of SLMAP SNPs as a risk factor for the susceptibility of DR among type 2 diabetes patients in Qatari population.
Expression of GDNF Throughout Rat Brain After Intranasal Delivery of hGDNF Plasmid Nanoparticles
Authors: Aly Amirah , Harmon Brendan , Shah Parin , Sesenoglu‐Laird Ozge , Padegimas Linas , Cooper Mark , Waszczak Barbara
Published: June 2021

Glial cell line‐derived neurotrophic factor (GDNF) gene therapy is a promising approach for CNS disorders such as Parkinson's disease (PD). We are examining intranasal administration of PEG‐CK30 DNA nanoparticles (NPs) encoding hGDNF, developed by Copernicus Therapeutics, Inc. We have shown that intranasal NPs incorporating hGDNF DNA (pGDNF), or a hGDNF‐eGFP fusion plasmid (pUGG), transfect brain cells in vivo, and that intranasal pGDNF NPs protect substantia nigra (SN) dopamine neurons in a rat model of PD.

Here, we examined transfection in rat brain 7 days after intranasal delivery of these NPs. ELISA showed increased GDNF levels throughout the brain of rats given pGDNF relative to saline controls. Immunohistochemistry (IHC) revealed that the number of eGFP positive cells was significantly higher in rats given intranasal pUGG NPs compared to background fluorescence in controls, with the highest numbers in midbrain. Double label IHC showed that most eGFP positive cells were immediately adjacent to capillary endothelial cells staining for RECA‐1, and they were often contiguous to GFAP positive astrocytic endfeet enwrapping capillaries. When eGFP cells were observed adjacent to dopamine neurons in SN, or NeuN labeled cells in any region, these neurons were also found close to a capillary. This pattern suggests that pericytes are transfected after intranasal administration of DNA NPs, consistent with distribution by perivascular transport. These results confirm delivery and transfection of cells in rat brain after intranasal administration of Copernicus' DNA NPs and support use of intranasal hGDNF NPs as a non‐invasive gene therapy for PD.
Cerium Oxide Nanoparticles (CeO2 NP) Treatment Ameliorates Sepsis Induced Diaphragm Dysfunction
Authors: Asano Shinichi , Ravikumar Arvapalli , Manne Nandini , Nandyala Geeta , Ma Bing , Maheshwari Mani , Nepal Niraj , Selvaraj Vellaisamy , Rice Kevin , Blough Eric
Published: June 2021

CeO2 nanoparticles have been posited to exhibit anti‐inflammatory and anti‐oxidative activities suggesting that these particles may be of potential use for the treatment of disorders characterized by increased inflammatory/oxidative stress. Severe inflammatory disorders such as that seen during sepsis are associated with diaphragm dysfunction and poor patient prognosis. We hypothesized that CeO2 nanoparticle treatment would ameliorate diaphragm dysfunction in sepsis by improving the immune response and oxidative stress levels. Sprague Dawley rats were randomly assigned into the following groups: Sham control, CeO2 nanoparticle treatment only (0.5mg/kg i.v.), sepsis (cecal inoculum method) and sepsis + CeO2 nanoparticles. Serum cytokine (TNF‐ α, IFN‐γ and IL‐6) levels were elevated at 3 and 18 h after sepsis induction and significantly lowered with CeO2 nanoparticle treatment (~60 % of Sepsis: P<0.05). Nanoparticle treatment decreased sepsis associated impairments in diaphragmatic contractile (Po) function (Sham:25.6 ±1.6 N/cm2 vs. CeO2:23.4 ± 0.8 N/cm2, vs. Sep:15.9 ± 1.0 N/cm2 vs. Sep + CeO2:20.0 ± 1.0 N/cm2, P<0.05). These improvements in contractile function were accompanied by a normalization of protein translation signaling (AKT, S6 and 4EBP), diminished proteolysis (caspase 8 activity and ubiquitin), and decreased inflammatory signaling (Stat3). Histological analysis suggested that nanoparticle treatment was associated with diminished sarcolemma damage and diminished inflammatory cell infiltration. These data indicate CeO2 nanoparticles may improve diaphragmatic function in the septic laboratory rat.
Antithrombin Perfluorocarbon Nanoparticle Deposition Prevents Stent Thrombosis
Authors: Palekar Rohun , Vemuri Chandu , Marsh Jon , Arif Batool , Wickline Samuel
Published: June 2021

Background

Despite pharmacologic modification of vascular implants, stent thrombosis remains clinically significant. The purpose of this research was to demonstrate the utility of antithrombin nanoparticles to prevent stent thrombosis.

Methods: Perfluorocarbon nanoparticles conjugated to multiple copies of the direct thrombin inhibitor D‐phenylalanyl‐L‐prolyl‐L‐arginyl‐chloromethyl ketone (PPACK‐NP) were tested in static and in flow‐loop in vitro models for their ability to prevent thrombosis on bare metal stents. Thrombin‐adsorbed stents were treated with saline, free PPACK, control NP or PPACK‐NP, followed by exposure to platelet poor plasma (PPP) or platelet rich plasma (PRP) for 60 minutes. In PRP‐exposed groups, subsets were pretreated with either clopidogrel or vorapaxar.

Results: Treatment of thrombin‐adsorbed stents with PPACK‐NP abrogated clot formation following PPP (p<0.05) or PRP (p<0.0005) exposure. Scanning electron microscopy of PRP exposed stents treated with PPACK‐NP revealed accumulation of adsorbed nanoparticles (Fig. A, arrow) on the surface with little fibrin or platelet deposition compared to control NP‐treated stents (Fig. B). Furthermore, PPACK‐NP treatment in combination with platelet inhibitors limited clot formation over that of clopidogrel (p<0.05) and vorapaxar (p<0.005) alone. Application of PPACK‐NP in an in vitro flow model confirmed the ability of PPACK‐NP to prevent the formation of an occlusive thrombus on stents compared to controls (Fig. C&D).

Conclusion

In summary, our results demonstrate the ability of PPACK‐NP to prevent stent thrombosis and suggest the potential for enhancing stents with nanoparticle coating to improve patency.

Research Support: NIH HL112303

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Methylene Blue Protects the Liver from Steatohepatitis via AMPK Activation
Authors: WU HONGMIN , Shin Seo Young , Kim Tae Hyun , Kim Sang Geon
Published: June 2021

Methylene blue (MB) has been under clinical trials for expanded therapeutic applications. This study investigated the effects of MB on mitochondrial dysfunction and steatohepatitis caused by toxicant or high‐fat diet feeding, and the underlying basis. MB treatment enabled HepG2 cells to survive against oxidative stress elicited by severe oxidative stress, restore GSH content, and decrease H2O2 production. Consistently, mitochondria dysfunction was abrogated. MB treatment enhanced hepatic mitochondrial DNA contents and oxygen consumption rates. In an animal model, MB treatment protected the liver from injury against toxicant. In mice fed on a high‐fat diet, MB treatment inhibited hepatic fat accumulation and steatohepatitis. MB activated AMP‐activated protein kinase (AMPK)both in vivo and in vitro. MB activation of AMPK caused to inhibition of glycogen synthase kinase‐3b(GSK3b), contributing to protecting mitochondria and hepatocytes. Consistently, the activation of AMPK enhanced the expression of carnitine palmitoyltrasnferase‐1 (CPT‐1) and peroxisome proliferator‐activated receptor a (PPARa), which promoted mitochondrial biogenesis. These results show that MB has the ability to protect the liver from the insult of toxicant or metabolic dysfunction in association with AMPK activation, implying its potential therapeutic impact on energy metabolism.
Incretins Increase the Tissue Level of Endogenous Antioxidants in Experimental Diabetes Mellitus
Authors: Adeghate Ernest , Lotfy Mohamed , Singh Jaipaul
Published: June 2021

Diabetes‐induced angiopathy is caused by chronic hyperglycemia via increased tissue level of free radicals. A decrease in the tissue levels of free radicals or elevation of the tissue concentration of endogenous antioxidant defense system can prevent or delay diabetes‐induced complications. The aim of the study was to examine the effect of incretins on selected markers of the antioxidant system (catalase, dismutase and glutathione reductase) in streptozotocin‐induced diabetic rats using immunohistochemical techniques. The extent of glutathione peroxidase gene expression was also determined in the pancreas of control and GLP‐1 or exenatide‐treated rats. The administration of GLP‐1 and exenatide caused large and significant (p<0.05) increases in the number of catalase‐immunopositive cells in incretin‐treated rats when compared to untreated diabetic controls. The percentage number of glutathione reductase‐ and dismutase‐immunoreactive cells was significantly increased (p<0.05) in diabetic animals treated with either GLP‐1 or exenatide compared to those of untreated diabetic rats. Morphological studies showed that catalase, dismutase and glutathione reductase co‐localized with insulin in pancreatic beta cells. GLP‐1 and exenatide caused marked (p<0.05) elevations in the tissue level of glutathione peroxidase gene expression when compared to that of untreated control. In conclusion, incretin‐induced increase in the tissue level of endogenous antioxidants may play an important role in the protection of the pancreatic beta cell.
In silico and in vitro evaluation of benzothiazole derivatives with inhibitory activity on 11‐beta‐hydroxysteroid dehydrogenase type 1
Authors: C. Cabrera Perez Laura , Padilla Martinez Itzia , Cruz Alejandro , Correa Basurto Jose , Rosales Hernandez Martha , Loera Pineda Juan
Published: June 2021

11‐beta‐hydroxysteroid dehydrogenase 1 (11‐βHED1) plays an important role in obesity and metabolic syndrome, due to it regulates cortisol levels. High cortisol levels produce gain of adipose tissue, release of proinflammatory substances, and free radicals (FR)1,2. Then the inhibition of 11‐βHED1 by thiazole derivatives has been reported. However, nowadays there are not a drug to inhibit the enzyme. For instance, inhibitors developed by Amgen/Biovitrum have not yet been approved due to it is not concluded, because, their clinical trials was halted3. For this reason, in present work, eleven benzothiazole derivatives were studied by in silico methods. Studies by blind docking allowed to determine that compound 1 (C1) has better affinity for 11‐βHED1 (PDB:4BB6) than compound 2 (C2), because interacts by hydrogen bond with Y177 in 11‐βHED1 dimer, suggesting that C1 could be used in obesity treatment, besides that in previous studies it demonstrated antioxidant properties that can counteract the FR generated during obesity.

Additionally, the results of the in silico studies of C1 and C2 will be supported in vitro by NADPH production, which is proportional to inhibitory effect on 11‐βHED1.1Jan H et al., 2014; 2Celso E, 2011; 3Rob M., et al 2012.
In Vitro Acetylation of Seven Anti‐glaucoma Agents in Rat, Rabbit and Human Ocular and Liver S9 Fractions
Authors: Bushee Jennifer , Dunne Christine , Argikar Upendra
Published: June 2021

Ocular N‐acetyltransferase(NAT) research has been limited to probe substrate activity in specific tissues from rat or rabbit, but not for a model that represents the whole eye. The comparison of acetylation as a biotransformation pathway between preclinical species and human has not been well documented. We aimed to understand functional ocular NAT activity across rat, rabbit, and human ocular S9 fractions in comparison with liver S9 fractions for seven clinically approved anti‐glaucoma compounds. Specific probe substrates were examined in human ocular, liver, and kidney S9 fractions to asses human NAT1 and 2 activities. High resolution mass spectrometry was utilized to characterize the acetyl conjugate(s) for each anti‐glaucoma agent in ocular and liver S9 fractions. Direct acetyl conjugates were detected across all species in ocular and liver fractions for betaxolol, carteolol and levobunolol, indicating that these compounds are non‐specific NAT substrates. Oxidation and O‐acetylation of levobunolol was detected in rat and rabbit liver S9 only. Species‐ and organ‐specific metabolic differences were observed for metoprolol, metipranolol, propranolol, and timolol. Neither ocular nor liver fractions from rat or rabbit were useful to predict human ocular acetylation in vitro. Ocular and renal S9 fractions did not catalyze the acetylation of sulfamethazine, a NAT2 probe substrate, contrary to the observation in liver S9. The data suggests a strong role of NAT1 in human ocular S9 fractions. This is the first report of direct acetyl conjugates for the b‐adrenergic receptor antagonist class of anti‐glaucoma agents in rat, rabbit and human ocular S9s.
sGC Stimulation Results in Potent Anti‐hypertensive Effects as a Stand‐alone or Combination Therapy in the Rat
Authors: Tobin Jenny , Jefferson Robert , Shea Courtney , Solberg Erik , Long Kimberly , Nakai Takashi , Moore Joel , Sheppeck James , Germano Peter , Wakefield James , Sykes Kristie , Rivers Samuel , Masferrer Jaime
Published: June 2021

Soluble guanylate cyclase (sGC) stimulators have the potential to treat a variety of cardiovascular diseases via the nitric oxide (NO)‐sGC‐cyclic guanosine monophosphate (cGMP) pathway. Ironwood Pharmaceuticals has synthesized a number of novel sGC stimulators with the objective of creating a once‐a‐day oral anti‐hypertensive.

A screening paradigm that evaluated sGC enzyme stimulation, pharmacokinetics (PK), and pharmacodynamics (PD) identified three novel sGC stimulators that induce sustained cardiovascular effects after oral dosing. Potency against sGC was evaluated using an in vitro cell based assay that measured cGMP production. Compounds were subsequently evaluated in PK and blood pressure PD in vivo rat models. All compounds were potent stimulators of sGC in vitro (EC50 = 40‐220nM), had sustained compound exposure in vivo (Thalf 7‐12 hours) after oral dosing, and significantly decreased mean arterial blood pressure (MABP) in vivo (蠅 10mmHg) after oral dosing.

The potential for sGC stimulators to work in combination with reference anti‐hypertensive therapies was assessed in our in vivo PD assay. Doses of Losartan, Atenolol, Amlodipine or each of our sGC stimulators that induced a decrease in MABP (蠅 10mmHg) when dosed alone were chosen. Combining sGC stimulators with anti‐hypertensives resulted in an additional 5‐50% decrease in MABP over either therapy given alone. These data indicate that IWP sGC stimulators possess potent anti‐hypertensive effects in the rat and that their efficacy increases when dosed in combination with standard anti‐hypertensive agents.

This research was funded by Ironwood Pharmaceuticals.
The Cardiovascular Effects Produced by Intracerebroventricular Microinjection of N/OFQ in Angiotensin II/High Salt Diet Rats.
Authors: Nguyen Tin , Hollister Bernadette , Franklin Cynthia , Gottlieb Helmut
Published: June 2021

Intracerebroventricular (ICV) administration of the opioid‐like peptide, nociceptin/Orphanin (N/OFQ), produces a free water diuresis, in addition to marked effects on cardiovascular function and renal sympathetic nerve activity (RSNA). The present study utilized an angiotensin II high salt diet (ANGII) hypertension model to determine the central‐mediated cardiovascular effects of N/OFQ in anesthetized rats. Rats were chronically instrumented with cannulas in the femoral artery, femoral vein and urinary bladder and implanted with a renal nerve electrode. Cardiovascular and renal parameters were measured during control (C, two 10‐minutes) and for 60 minutes after ICV injection of saline vehicle or N/OFQ (10 mg/5 ml) in normotensive or ANGII rats. ICV N/OFQ produced a marked decrease in heart rate (HR), mean arterial pressure (MAP) and RSNA in both normotensive and ANGII rats. However, N/OFQ produced greater percentage decreases in HR, MAP and RSNA in ANGII rats than normotensive rats. N/OFQ also produced a significant increase in urine flow rate (V) in both groups, but ANGII blunted this diuretic effect. ICV saline did not produce any significant changes in any of the measured parameters. These findings suggest that, in urethane‐alpha‐chloralose anesthetized rats, N/OFQ appears to have greater effect on the cardiovascular system than on the renal. As such, drugs targeting the central N/OFQ receptors could facilitate the lowering of blood pressure in diseases, such as salt‐sensitive hypertension. Supported by NIH 5SC2 HL104639
Neddylation Promotes Vascular Endothelial Dysfunction: A Role For Histone Deacetylases 2 and Arginase 2.
Authors: PANDEY DEEPESH , Hori Daijiro , Kim Jae , Berkowitz Dan , Romer Lewis
Published: June 2021

Emerging evidence strongly supports a role for HDAC2 in the transcriptional regulation of endothelial genes and vascular function. We have recently demonstrated that HDAC2 reciprocally regulates the transcription of Arginase2, which is itself a critical modulator of endothelial function via endothelial NOS. Moreover the abundance of HDAC2 is decreased in response to the atherogenic stimulus, OxLDL via a mechanism that is apparently dependent upon proteasomal degradation. Neddylation is a post‐translational protein modification that is tightly linked to ubiquitination and thereby protein degradation. We propose that changes in Neddylation may modulate vascular endothelial function in part through alterations in the proteasomal degradation of HDAC2. In HAEC, OxLDL stimulation resulted in globally increased Neddylation of cellular proteins. Pre‐incubation of mouse aortic rings with the Neddylation activating enzyme inhibitor, MLN4924, prevented OxLDL‐induced endothelial dysfunction. MLN also enhanced HDAC2 abundance, and decreased expression and activity of Arg2 in HAEC. Furthermore MLN blocked OxLDL‐mediated reduction of HDAC2 levels. Additionally, co‐transfections of HAEC with the de‐neddylase SENP8 increased HDAC2 levels in a dose‐dependent fashion that was also consistent with the results of Neddylation activating enzyme inhibition. Our findings indicate that the observed improvement in endothelial dysfunction with Neddylation activating enzyme inhibition is likely mediated by an HDAC2‐dependent decrease in Arginase2. Neddylation activating enzyme may therefore be a novel target in endothelial dysfunction and atherogenesis.
Evaluation of the Cardiovascular Responses in Nicotinamide+Streptozotocin Diabetic Pithed Rats
Authors: Centurion David , Torres‐Gutierrez Miriam , Gutierrez‐Lara Erika , Sanchez‐Lopez Araceli
Published: June 2021

In vitro studies have demonstrated that during diabetes a vascular dysfunction is observed. However, very few studies in vivo have confirmed these findings in the systemic vasculature. Thus, this study evaluated the vasopressor responses induced by sympathetic stimulation or by i.v. bolus injections of the agonists noradrenaline (α1/2), methoxamine (α1), UK 14,304 (α2), and the vasodepressor responses induced by acetylcholine and sodium nitroprusside in rats pretreated with: (i) nicotinamide + streptozotocin (NAD+STZ), (ii) saline + buffer, (iii) saline + streptozotocin (SAL+STZ); and (iv) nicotinamide + buffer. For this purpose, rats were treated with a single dose of nicotinamide (230 mg/kg, i.p.) or its vehicle (saline, 1 ml/kg) and 15 min after streptozotocin (65 mg/kg, i.p.) or its vehicle (citrate buffer, 1 ml/kg) was administered. 12 weeks latter, a tolerance glucose curve was determined after administration of glucose (1 g/kg, p.o.). Then, under anesthesia, the animals were pithed and prepared to measure blood pressure and heart rate. In animals treated with NAD+STZ intolerance to glucose was observed. Moreover, the vasopressor responses to sympathetic stimulation or noradrenaline, methoxamine and UK 14,304 remained unchanged. In contrast, in animals treated with SAL+STZ, the vasopressor responses induced by sympathetic stimulation or by methoxamine and UK 14,304 were blunted. Furthermore, the vasodepressor responses induced by acetylcholine, but not those induced by sodium nitroprusside, were significantly decreased in rats treated with NAD+STZ or SAL+STZ. These results suggest that in NAD‐STZ diabetic rats an impairment on the vasodepressor responses induced by acetylcholine was observed probably by an endothelial mechanism.