bluuebunny/crossref_metadata_2025 huggingface dataset:
Tectonics form a transition zone complex of ciliopathy proteins that regulate ciliary composition
Authors: Reiter Jeremy , Garcia-Gonzalo Francesc , Corbit Kevin , Dowdle William , Yee Laura
Published: June 2021

Primary cilia are projections found on many human cells. These cilia do not move, but function as cellular antennae to receive and interpret signals. For example, olfactory cilia sense signals from the environment, such as odorants. Other cilia sense cues from other cells, such as Hedgehog proteins, to coordinate tissue patterning and growth. We have identified mechanisms of ciliary signaling, cilium formation, and the regulation of ciliary composition. Defects in ciliary signaling cause some forms of birth defects, cancer, polycystic kidney disease, and other disorders collectively called ciliopathies. Investigating ciliopathies is illuminating how disturbing the function of a single organelle can give rise to diverse human diseases.

We have found that two ciliary proteins of the Tectonic family, Tctn1 and Tctn2, form part of the transition zone, a region at the base of the cilium. Tctn1 is an extracytosolic glycoprotein that interacts with transmembrane components of the transition zone such as Tmem67 to connect to a cytosolic transition zone complex comprised of most known Joubert‐ and Meckel‐associated proteins. Loss of components of the Tectonic transition zone complex in mice compromise ciliogenesis in some tissues, and deregulate ciliary protein composition in others. We hypothesize that transition zone defects disrupt intercellular signaling, causing the multiorgan disease observed in ciliopathies, including polycystic kidney disease and birth defects such as polydactyly.
Knocking E. coli off of their pedestals: Understanding the strategies microbes exploit to generate morphological structures during their disease processes
Authors: Guttman Julian Andrew
Published: June 2021

When the extracellular attaching and effacing pathogens [enteropathogenic E. coli, enterohaemorrhagic E. coli and Citrobacter rodentium] exploit their host cells, morphologically distinct actin‐rich structures are generated beneath the attached bacteria. These structures, called pedestals, protrude from the cell surface, enable the bacteria to “surf” atop infected cells and are hallmarks of the infections. Contained within these structures are numerous actin‐associated components that would be predicted to be at dynamic actin‐rich structures including the Arp2/3 complex, cortactin, profilin and cofilin. Recently my lab has discovered that unexpected proteins are also present within pedestals. These include clathrin‐mediated endocytic proteins and protein components of the spectrin cytoskeleton. These proteins are crucial for pedestal generation as their alteration blocks pedestal formation and often also inhibits attachment of the bacteria to their target cells, thus halting the infections. By using E. coli pedestals as a model system to also study general cell motility we have shown novel roles for a variety of cellular proteins. Taken together our examination of E. coli pedestals has already provided potential targets for pharmaceutical intervention as well as novel insights into general cell biological processes.
Mechanisms of oxLDL on Endothelial Biomechanics
Authors: Oh Myung-Jin , Shentu Tzu-Pin , Levitan Irena
Published: June 2021

Oxidized low density lipoprotein (oxLDL) is a major factor in the initiation and development of atherosclerosis. Our lab has shown that oxLDL increases endothelial stiffness suggesting that changes in endothelial biomechanics are important for endothelial dysfunction. The mechanisms, however, by which oxLDL alters endothelial biomechanics are not understood. Our goal is to elucidate the mechanisms of oxLDL‐induced cell stiffening. Since cell stiffness is determined by the cytoskeleton, in particular the actin cytoskeleton, we tested whether oxLDL‐induced cell stiffening is mediated by the Rho‐GTPases, the major regulators of actin cytoskeleton dynamics. We targeted RhoA, that induces stress fiber formation. We show that oxLDL induced cell stiffness is abrogated by inhibiting RhoA with C3 transferase, suggesting that RhoA and its downstream targets are involved in oxLDL induced cell stiffening. We next explored the role of Rho kinase (ROCK), an immediate downstream effector of RhoA. Inhibition of ROCK by Y27632 also inhibited oxLDL induced cell stiffening. In contrast, inhibiting myosin light chain kinase (MLCK) by ML7 had no effect on oxLDL induced cell stiffness. We propose, therefore, that oxLDL‐induced endothelial stiffening is mediated by the RhoA/ROCK pathway and plays an important role in dyslipidemic conditions in vivo. (Supported by HL083298)
Angiopoietin like‐2 stimulates leukocytes adhesion to the native aortic endothelium in LDLr−/−; hApoB100+/+ mice
Authors: Farhat Nada , Thorin-Trescases Nathalie , Villeneuve Louis , Allen Bruce G. , Thorin Eric
Published: June 2021

Angiopoietin like‐2 (Angptl2), which circulating levels are increased in patients with coronary artery disease, could contribute to chronic inflammation. We hypothesized that the pro‐inflammatory effect of Angptl2 contributes to atherogenesis by stimulating leukocyte adhesion in atherosclerotic mice (LDLr−/−; hApoB100 +/+, ATX).

In C57Bl/6 control mice (WT), Angptl2 levels increased with age (P<0.05) in the plasma and the aorta. In ATX mice, levels were higher (P<0.05) than in WT mice both in plasma (3 folds) and the aorta (2 folds), and they further rose with age (P<0.05). In vitro, basal endothelial levels of TNF‐α and IL‐6 gene expression were higher in ATX compared to WT mice; stimulation with Angptl2 (100 nM) further raised their expression in both WT and ATX mice. Ex vivo, Angptl2 stimulated the adhesion of leukocytes on the native endothelium of the aorta of ATX mice (P<0.05), but not WT mice. Leukocytes adhesion on the endothelium was due to a rise (P<0.05) in the abundance of adhesion molecules (mRNA and protein) and the surface of endothelial cells (ICAM‐1) and leukocytes (Mac‐1, L‐selectin and PSGL‐1). Antibodies against P‐selectin or ICAM‐1 prevented (P<0.05) leukocyte adhesion in ATX mice.

Our data suggest that Angptl2 could significantly contribute to the atherosclerotic process by stimulating pro‐inflammatory cytokines secretion and leukocytes adhesion.
Neuroprotection of sulfuretin is mediated by inhibition of microglial inflammatory activation in BV‐2 cells
Authors: Hong Sa-Ik , Kwon Seung-Hwan , You In-Jee , Jung Yang-Hee , Kim Min-Jung , Ma Shi-Xun , Sohn Uy-Dong , Lee Seok-Yong , Jang Choon-Gon
Published: June 2021

In the present study, we investigated the effects of sulfuretin on inflammatory responses induced by lipopolysaccharide (LPS) in murine microglial BV‐2 cells. BV‐2 cells were treated with sulfuretin prior to LPS exposure, and the effects of such treatment production of NO, inducible nitric oxide synthese (iNOS), cyclooxygenase‐2 (COX‐2), prostaglandin E2 (PGE2), interleukin‐1β (IL‐1β), and tumour necrosis factor‐α (TNF‐α) were analysed by Griess reaction, ELISA, western blotting. Moreover, we investigated the effects of sulfuretin on LPS‐induced nuclear factor‐κB (NF‐ κB), heme oxygenase‐1 (HO‐1) activation, phosphorylation of mitogen‐activated protein kinases (MAPKs) such as 1/2 c‐Jun N‐terminal kinase (JNK), p38 MAPK, extracellular signal‐regulated kinase 1/2 (ERK 1/2), as well as phosphatidylinositol 3‐kinases (PI3K)/Akt. Our studies showed that sulfuretin markedly suppressed the production of NO and PGE2, as well as expression of IL‐1β, TNF‐α, iNOS, COX‐2 in a dose‐dependent manner. Moreover, sulfuretin significantly suppressed NF‐ κB and HO‐1 activation, phosphorylation of JNK, p38 MAPK, ERK 1/2, and PI3K/Akt. These results indicated that sulfuretin has a strong anti‐inflammatory activity in brain microglia, and could be a potential therapeutic agent for the treatment of neuroinflammatory diseases such as Alzheimer's disease, stroke, and Parkinson's disease.
Nasal Spray of Bioactive Polyphenol Metabolites as a Novel Therapy for Alzheimer's Disease and Other Forms of Dementia
Authors: Knable Lindsay Alexis , Wang Jun , Ferruzzi Mario G , Vempati Prashant , Freire Daniele , Gong Bing , Pasinetti Giulio Maria
Published: June 2021

While polyphenolic compounds have many health benefits, the potential development of polyphenols for the prevention/treatment of neurological disorders is largely hindered by their complexity and limited knowledge regarding their bioavailability, metabolism and bioactivity in the brain. We recently demonstrated that dietary supplementation with a grape‐derived polyphenolic preparation, namely a monomeric‐enriched catechin and epicatechin fraction (Mo), significantly improves cognitive function in a mouse model of Alzheimer's disease (AD). We also found that Mo treatment resulted in the accumulation of proanthocyanidin metabolites in the brain at a concentration of ~400 nM. One of the metabolites identified in the brain following Mo treatment, Metaphenol‐A1, was shown to promote basal synaptic transmission and long term potentiation (LTP) at physiologically relevant concentrations in hippocampal slices through mechanisms associated with cAMP‐response‐element‐binding‐protein signaling. In this study, we tested whether application of Metaphenol‐A1 via a novel, non‐invasive intranasal delivery apparatus can effectively improve LTP and cognitive function in the AD mice. Our study will provide insights into developing a novel, safe approach to directly deliver a bioactive therapeutic agent to the central nervous system for AD prevention/treatment.

Supported by Discretional Funds to GMP.
ageLOC Recharge improves memory and learning ability in memory impairment models
Authors: Tan Ningzhi , Yao Yisang , Yang Guang , Wu Zimei , Zhu Jia-Shi
Published: June 2021

Based on the differentiated gene expressions in skeletal muscle and neocortex between the young (5 mo) and old (25 mo) mice, ageLOC Vitality was formulated with ingredients (Cordyceps sinensis Cs‐4 and extracts from ginseng and pomegranate) that oppose the age associated changes. In addition to its effect in improving physical endurance and sexual functions, we examined its effect in memory improvement at a dose of 200, 400 or 800 mg/kg in mice in this study. Shuttle box avoidance test was used to examine the markers: active avoidance latency (AAL) and active avoidance response rate (AARR) in 3 models: memory acquisition impairment (MAI) by scopolamine; memory consolidation impairment (MCI) by sodium nitrite; memory reproduction impairment (MRI) by ethanol. Recharge shortened AAL and increased AARR in (1) MAI model: AAL −47 to −70% (p<0.001) and AARR +109 to +209% (p=0.007); (2) in MCI model: AAL +9.5 to −44% (p=0.024) and AARR +59 to +149% (p=0.002); and (3) in MRI model: AAL +10 to −52% (p=0.014) and AARR +17 to +203% (p<0.001). Our results indicate ageLOC Recharge is effective in improving memory and learning ability in mice with memory impairment, supporting the potential anti‐aging effect of ageLOC Recharge.
The effect of oral creatine monohydrate supplementation on cognitive function in young women
Authors: Skelton Michele , Dellis-Leeper Erica
Published: June 2021

Clinical studies have demonstrated Creatine (Cr) monohydrate's impact on cognitive function. Increased memory and intelligence have been reported in Cr supplementation studies involving elderly individuals and vegetarians. Recent studies suggest that early administration of oral Cr‐monohydrate may reduce the risk of neurological deficiencies. The purpose of this study is to determine if short term supplementation with oral Cr‐monohydrate improves cognitive function among young women (26.7 ± 4.4 years). Ten women completed 4‐weeks of oral Cr‐monohydrate supplementation (5g per day). The women completed a battery of computerized cognitive function tests prior to and after supplementation. Post‐test scores on cognitive function tests involving central executive tasks and short term memory were significantly higher than pre‐test scores (p ≤ 0.05). There were no differences in scores on long term memory tests or reaction time. Results from this study suggest that acute supplementation of Cr‐monohydrate may positively impact cognitive function in young women.
High‐throughput screening and evaluation of anti‐cancer compounds
Authors: Carey Colleen Marie , Chen Bo , Jiang Na , Sharma Ashok , She Jin-Xiong
Published: June 2021

Early drug discovery for cancer treatment was based on targeting known proliferation mechanisms and the disruption of essential cellular functions. These therapies, although specific, were shown to have a wide range of toxicities. In contrast, therapies targeted to non essential functions were less toxic and less effective. Currently, the discovery phase has shifted to identify targeted therapies without specific molecular targets in mind. This approach allows for the identification of compounds against multiple targets or cancer pathway specific molecules. Furthermore, the identification of compounds that are efficacious and safe, particularly at the level of the immune system, will result in overall better therapies being developed. In this proposal various genomic and proteomic tools will be used to advance the identification of novel therapeutic compounds. A 150,000 molecule chemical library has been pre‐screened at various levels and narrowed down to 8 lead compounds. These compounds have minimal toxic effects on the immune system and will be further evaluated to identify the genomic and proteomic signatures for safety, efficacy and dose. We will also elucidate the molecular pathways that are targeted as they relate to cancer. This research is supported by funding to JX She.
Endothelium‐dependent modulation of cerebral artery myogenic tone by cyclopiazonic acid
Authors: Tam Raymond , Kerr Paul M , Ondrusova Katarina , Plane Frances
Published: June 2021

The myogenic response, the ability of cerebral arteries to constrict in response to increases in intra‐vascular pressure, is a key autoregulatory mechanism by which cerebral blood flow is maintained within narrow limits despite large changes in pressure. The endothelium modulates arterial diameter via the release of factors such as nitric oxide (NO) and by electrical coupling with smooth muscle cells. We hypothesized that disruption of endothelial intracellular Ca2+ stores will affect the cerebral artery myogenic response. In endothelium‐intact middle cerebral arteries, cyclopiazonic acid (CPA; 10 μM) and thapsigargin (500 nM), inhibitors of the sarcoplasmic Ca2+‐ATPase (SERCA), each inhibited development of myogenic tone, e.g. 80mmHg, tone development was reduced by 65.5 ± 6.0% (n=6). In endothelium‐denuded arteries, CPA and thapsigarin had no effect the myogenic response. In endothelium‐intact arteries, L‐NG‐nitroarginine methyl ester (100 mM), an inhibitor of NO synthase, did not affect the CPA‐induced loss of myogenic response but addition of apamin (50nM) and Tram‐34 (1 μM), inhibitors of endothelial small and intermediate conductance Ca2+‐activated potassium (KCa) channels, prevented the effect of CPA. These data indicate that inhibition of endothelial SERCA leads to the activation of KCa channels and loss of cerebral myogenic tone. Supported by HSFC.
Viability of two sub‐strains of C57BL/6 mice in response to cardiac pressure‐overload
Authors: Parikh Jay , Wang Sophia , Azizi Khalid , Bird Elizabeth , Spellman Matthew , Lai N. Chin
Published: June 2021

The inbred mouse strain C57BL/6 is highly suitable and commonly used for the generation of transgenic mice as tools in dissecting biological processes and diseases. However, this parent line is available from a variety of sources with different sub‐strains. We examined male C57BL/6J mice from the Jackson Laboratory (n=29) and male C57BL/6NHsd from the Harlan Laboratories (n=36) in the setting of transverse aortic constriction (TAC), which is a common interventional procedure for studying pressure‐overload in hearts. Under anesthesia, the aortic arch between the innominate and the left carotid artery was constricted to 0.413 mm (27 ga needle) with a 7‐0 silk suture. Survival was monitored for 3 wks, and subsequently left ventricular function was assessed with a 1.4F Millar pressure transducer. Three wks after TAC, the survival rate was 50% for the C57BL/6NHsd and 79% survived for the C57BL/6J (Top Figure). A significant decline of 1058 mmHg/s (21%) in cardiac contractility (Bottom Figure) and 1228 mmHg/s (22%) in the rate of decay was found in C57BL/NHsd mice.

Conclusion

This study demonstrates phenotypic differences in viability and cardiac performance in response to TAC for these two sub‐strains. Therefore, it is imperative to carefully consider the appropriate line for the design of transgenic mice.
Acute effects of intermittent pneumatic compression induced hemodynamics on vascular function in humans
Authors: Sheldon Ryan D. , Roseguini Bruno T. , Laughlin M. Harold , Newcomer Sean C.
Published: June 2021

Intermittent pneumatic leg compression (IPC) therapy is commonly used to treat symptoms associated with peripheral artery disease. One hypothesis is that IPC increases shear rate (SR) in the arteries and enhances endothelial cell (EC) function. To test this hypothesis we characterized the hemodynamic profile and used flow mediated dilation (FMD) to assess EC function in response to a single session of high frequency (HF) and low frequency (LF) IPC. Eleven male subjects (age: 23.5 ± 4.7 years) underwent an hour of HF (2s inflation/3s deflation) and LF (4s inflation/16s deflation) IPC treatment on separate days. Doppler/ultrasonography of the popliteal artery (PA) was used to measure diameter and blood velocity at the 5th and 45th minute of treatment and to assess FMD pre and post treatment. IPC had no effect on PA FMD (HF: pre: 2.45±.50%, post: 2.76 ±.32%; LF: pre: 3.16±.36%, post: 3.56±.52%). Cuff deflation elevated blood flow (BF) and SR and decreased oscillatory shear index (OSI) from baseline at the onset of both treatments (p<.01). This effect was no longer evident by the 45th minute of HF treatment only. When compared to baseline both treatments significantly reduced BF and SR and increased OSI during cuff inflation (p<.01). In conclusion, a single session of IPC does not acutely impact PA FMD despite significant alterations to the hemodynamic profile during treatment. HL35088 (MHL, BTR); ACSM Research Endowment (SCN)
Ginkgolide B inhibits renal cyst development in in vitro and in vivo cyst models
Authors: Yang Baoxue , Zhou Hong , Gao Jinsheng , Xia Yin
Published: June 2021

Autosomal dominant polycystic kidney disease (ADPKD) is a common inherited disease characterized by massive enlargement of fluid‐filled cysts in the kidney. To examine whether ginkgolide B, a natural compound, inhibits cyst development, a MDCK cyst model, an embryonic kidney cyst model and a PKD mouse model were used. Interestingly, ginkgolide B significantly inhibited MDCK cyst formation dose‐dependently, with up to 69% reduction by 2 μM ginkgolide B. Ginkgolide B also significantly inhibited cyst enlargement in the MDCK cyst model, embryonic kidney cyst model and PKD mouse model. To determine the underlying mechanisms, the effect of ginkgolide B on the MDCK cell viability, proliferation, apoptosis, chloride transporter CFTR activity and intracellular signaling pathways were also studied. Ginkgolide B did not affect cell viability, proliferation, and expression and activity of the chloride transporter CFTR that mediates cyst fluid secretion. Ginkgolide B induced cyst cell differentiation and altered Ras/MAPK signaling pathway. Taken together, our results demonstrate that ginkgolide B inhibits renal cyst formation and enlargement, suggesting that ginkgolide B might be developed into a novel candidate drug for ADPKD.
Possible mechanism of neutrophil death in Diabetes mellitus
Authors: Kuwabara Wilson Mitsuo Tatagiba , Curi Rui , Alba-Loureiro Tatiana Carolina
Published: June 2021

Diabetes mellitus was induced by streptozotocin (65 mg/Kg, i.v.) two weeks before. Neutrophils were obtained from the peritoneal cavity 4 hours after injection of 1% oyster glycogen solution. Cells were maintained in culture for 0, 60, 120 and 180 minutes with or without PMA stimulus (20nM). Neutrophils were then analyzed by flow cytometer for membrane integrity, DNA fragmentation and mithocondrial transmambrane potential. Caspases 1 and 3 activity was evaluated by spectrofluorimetry. The content of mithocondrial proteins released during cellular death (Cytochrome C, Smac/Diablo, Endonuclease G (Endo G), Apoptosis Inducing Factor (AIF), High temperature requirement protein A2 (HtrA2)/Omi) was evaluated by Western Blotting. Our study showed that neutrophils from the diabetic rats presented a decrease in membrane integrity (p<0,001), increase in DNA fragmentation (p<0,01) and an increase in the mitochondrial depolarization (p<0,0001), after 60 min of PMA stimulus when compared to non‐stimulated cells from the same group. However, at time 0, neutrophils from diabetic rats already presented a decrease in the mitochondrial transmembrane potential (p<0,001) when compared to cells from the group control. Caspases 1 and 3 activity was not observed. It was not observed difference in the content of the mitochondrial proteins cytochrome C, Smac/Diablo and Endo G. However, content of AIF and HtrA2/Omi was increased 60 minutes after PMA stimulus. The death process in neutrophils from diabetic rats seems to be caspase 1 and 3 independent and dependent on alterations of mitochondrial transmembrane potential presented at time 0 without stimulus. FAPESP, CNPq and CAPES.
Sex‐related differences in monoamine oxidase activity
Authors: Salom Lorena P , Evanson Kirk W , Stone Audrey J , Kluess Heidi A
Published: June 2021

Monoamine oxidase is an enzyme that breaks down biological amines, such as norepinephrine and serotonin and renders them inert for producing vasoconstriction. The purpose of this study was to characterize monoamine oxidase activity in skeletal muscle arteries and to determine if estrogen contributes to sex‐related differences in monoamine oxidase activity. Five male and 20 female F344 rats were anesthetized. Twelve females were ovariectomized and a subset of six received an estrogen pellet (17β‐estradiol (4μg/day)). In all animals, the white gastrocnemius 1A arteriole (WG1A) was removed. The arterioles were homogenized in enzyme buffer. Luminometric and spectrophotometric assays were used to quantify monoamine oxidase activity and total protein content of the vessel, respectively. Relative luminescence units (RLU) were proportionate to monoamine oxidase activity. Monoamine oxidase activity in the WG1A was 29.85±18.71 RLU·μg−1 protein for males, but only 11.42±5.13 RLU·μg−1 protein in females (p<0.05). Estrogen replacement had no effect on monoamine oxidase activity (Ovariectomized: 16.63±10.64 RLU·μg−1 protein; Ovariectomized+estrogen: 16.93±10.38 RLU·μg−1 protein). These results suggest that there are sex‐related differences in monoamine oxidase activity, but, estrogen does not appear to be the hormone that influences sex‐related differences in monoamine oxidase activity.
Hypoxia attenuates the ability of cyclic guanine‐dependent protein kinase (PKG) to increase BK channel activity
Authors: Thorpe Richard Burdette , Williams James M , Pearce William Julian
Published: June 2021

OBJECTIVE

Chronic hypoxia attenuates endothelium‐dependent vasodilatation, more in fetal than adult arteries. Here we examine the hypothesis that these effects involve inhibition of PKG activity and reduced BK channel activity.

METHODS

Pregnant and Non‐Pregnant ewes were held at sea level (normoxic) or at 3820m (hypoxic) for the final 110 days of gestation. Carotid arteries from non‐pregnant adults and term fetal sheep were prepared for 5HT concentration‐response determinations with ±8‐pCPT‐cGMP and ±iberiotoxin. Abundances of α/β isoforms of PKG‐I were determined by Western blot. PKG activity was determined by enzyme assay.

RESULTS

PKG activation depressed 5HT‐induced contractility in normoxic but not hypoxic arteries. Hypoxia did not alter PKG abundance or specific activity but reduced iberiotoxin's ability to block PKG's effect on 5HT‐induced contractions, more in fetal than adult arteries.

CONCLUSION

Activation of BK channels accounts for a significant portion of PKG‐induced vasorelaxation in ovine carotid arteries. Chronic hypoxia attenuates the ability of PKG to activate BK channels without effects on PKG activity or abundance.

Supported by PHS Grants P01‐HD31226 and R01‐HL64867.
An imaging apparatus for simultaneous measurement of isometric contraction and Ca2+ fluorescence in large blood vessels of the rat
Authors: Tykocki Nathan R. , Wiseman Robert W. , Jackson William F. , Watts Stephanie W.
Published: June 2021

While simultaneous measurement of contraction and Ca2+ transients can be measured in small vessels (< 3 mm diameter), few strain gauges are available small and sensitive enough for concurrent measurement of Ca2+ transients and contraction in large vessels (> 3 mm diameter). We constructed a highly sensitive low‐noise strain gauge for the simultaneous measurement of vessel tension development and Ca2+ transients using fluorescent Ca2+ indicators. The force transducer was fabricated from aluminum and produced a sensitivity of 0.26 μV/mg over a broad range of applied load (0.05 to 5 g) and a rapid (102.8 Hz) frequency response. Blood vessels mount to the transducer with two stainless steel pins submerged above a coverslip in a 5 ml preparation bath. We utilized this apparatus for concurrent measurement of global Ca2+ and force production in rat aorta (RA) and vena cava (RVC) using the Ca2+ indicator Fura‐2. ET‐1 (100 nM) caused a transient spike in global Ca2+ that peaked when RA and RVC reached 25% of maximal contraction. After exposure to ET‐1, RVC Ca2+ peaked within 76±9 sec and reached maximum contraction within 8±3 min. In RA, Ca2+ peaked within 128±23 sec and reached maximum contraction within 18±4 min. These findings show that global Ca2+ increases more rapidly in response to ET‐1 in RVC than in RA, and that this difference correlates to differences in the time to maximal force development. Supported by NIH P01HL70687.
Stimulation of thromboxane A2 receptor induced the constriction and remodeling of the ductus arteriosus in the rat
Authors: Yokota Tomohiro , Aida Takashi , Shiraishi Ryosuke , Ichikawa Yasuhiro , Fujita Takayuki , Yokoyama Utako
Published: June 2021

The ductus arteriosus (DA) is an essential vascular shunt that connects the pulmonary artery to the aorta in the fetus. After birth, the DA immediately closes. DA closure is occurred by its constriction and remodeling. Patency of the DA (PDA) is a common life‐threatening complication of premature infants. Although cyclooxygenase inhibitors are frequently used for PDA, they often show little effect with severe side‐effects. Thromboxane A2 receptor (TP) is related to the vasoconstriction and the proliferation and migration of vascular smooth muscle cells (SMC) in the atherosclerosis. Hence, we hypothesized that TP stimulation induced closure of the DA by inducing the constriction and remodeling.

To assess the constrictive effect of the TP agonist U46619 on the DA, adjunct arteries, and peripheral vessels in vivo, we injected U46619 into the rat neonates and two distinct PDA models we generated. We found that U46619 induced DA constriction without constriction of other arteries in vivo. In addition, U46619 didn't form thrombosis in the lung. To investigate the role of TP stimulation in the DA remodeling, we examined the U46619‐induced intimal thickening (IT), proliferation, and migration using DA explant and cultured DA SMC. We found that U46619 induced IT by promoting the proliferation and migration of SMC. We conclude that TP agonists could be an alternative therapy for PDA.
AKAP150 is required for NFATc3‐induced vascular BKCa channel suppression during diabetic hypertension
Authors: Nystoriak Matthew A. , Nieves-Cintrón Madeline , Hinke Simon A. , Scott John D. , Santana Luis F. , Navedo Manuel F.
Published: June 2021

Activation of large conductance Ca2+‐activated (BKCa) K+ channels hyperpolarizes arterial smooth muscle, thus opposing vasoconstriction and contributing to lower blood pressure. Reduced expression of smooth muscle BKCa channels contributes to vascular dysfunction and hypertension in diabetes mellitus. However, the molecular mechanisms involved in this are unknown. We tested the hypothesis that hyperglycemia (HG) leads to BKCa suppression via activation of the calcineurin/NFATc3 signaling pathway in smooth muscle. Single BKCa records in inside‐out membrane patches (−40 mV) revealed reduced open probability (PO) in arterial myocytes under hyperglycemic conditions. Decreased BKCa PO in myocytes in HG was associated with downregulated mRNA and protein expression of BKCa pore‐forming α and accessory β1 subunits. Conversely, no effect of HG on BKCa subunit expression or activity was observed in myocytes isolated from NFATc3 null mice. Furthermore, HG‐induced BKCa suppression required Ca2+ influx via L‐type voltage‐gated Ca2+ channels, activation of calcineurin, and subcellular targeting of this phosphatase by the anchoring protein AKAP150. Overall, these results support the concept that hyperglycemia leads to vascular dysfunction via local activation of AKAP150‐bound calcineurin, NFATc3 nuclear translocation and reduced vascular BKCa expression.
Hsp90 inhibition prevents monocrotaline‐induced pulmonary hypertension in rats, as revealed by high resolution echocardiography
Authors: Dimitropoulou Christiana , Meadows Louise , Su Yunchao , Litwin Sheldon , Catravas John D
Published: June 2021

Four weeks after a single sc injection of monocrotaline (MCT; 60mg/kg), rats develop pathologic changes resembling those of human pulmonary arterial hypertension (PAH). Using high resolution echocardiography, we investigated non‐invasively, the ability of the hsp90 inhibitor 17‐AAG (10 mg, 3x weekly) to prevent the effects of MCT. PA flow velocity time integral (VTI), PA acceleration time (PAT), cardiac index (CI) and cardiac output (CO) values were reduced in rats treated with MCT alone, indicative of PAH. 4‐ or 2‐week (starting at week 3 after MCT) treatment with 17‐AAG prevented the decrease in VTI and PAT, but not in CO and CI. Right ventricle (RV) free wall (RVFW) thickness also increased with MCT, but not in the 4‐week 17‐AAG treatment group. Invasive assessment of right ventricular pressure (RVP) at the end of the 4‐week treatments confirmed increased RVP in the MCT, but not in the 17‐AAG treated animals. We conclude that treatment with hsp90 inhibitor for all or part of the 4‐week exposure to MCT effectively prevents the initiation/progression of PAH and that high resolution echocardiography is a useful tool for the non‐invasive, real‐time, repeated evaluation of cardiopulmonary changes in this model of PAH. (Supported by the AHA).
Treatment of mice with delta‐aminolevulinic acid, a generator of the guanylate cyclase activator protoporphyrin IX, prevents the development of hypoxia‐induced pulmonary hypertension
Authors: Alhawaj Raed , Patel Dhara , Ahmad Mansoor , Rémond Mathieu C , Eisenberg Leonard M , Wolin Michael S
Published: June 2021

Since our previous studies found that delta‐aminolevulinic acid (ALA) could generate protoporphyrin IX in amounts which activated soluble guanylate cyclase in pulmonary arteries, we investigated if treatment of mice with ALA (50 mg/kg/d) could prevent the development of pulmonary hypertension. Mice exposed to hypoxia (10% oxygen) for 21 days showed a significant increase in pulmonary artery pressure measured by echo Doppler pulmonary artery flow acceleration time/ejection time and right ventricle/left ventricle+septum weight ratios. These changes were attenuated in ALA‐treated mice. The previously observed 21d hypoxia‐elicited suppression of contraction to 10nM phenylephrine (PE) in isolated ~180μm pulmonary arteries by endogenously generated extracellular peroxide (reversed by 1μM catalase) and loss of the hypoxic pulmonary vasoconstriction response were prevented by treatment of mice with ALA. Thus, treatment of mice with ALA prevented the development of pulmonary hypertension and peroxide‐associated changes pulmonary artery reactivity caused by exposure of mice to 21d hypoxia. Supported by NIH grants HL31069, HL43023 and HL66331.
Pulmonary hypertension induced right heart failure: species comparison of global and local inflammatory responses
Authors: Walker Lori A. , Brown R. Dale , Bull Todd M. , Stenmark Kurt R. , Buttrick Peter M.
Published: June 2021

Pulmonary artery hypertension (PAH) is, in part, an inflammatory disease and local inflammation in both the proximal pulmonary vasculature and the right ventricle (RV) is correlated with RV failure in both animal models and in human disease. Despite this, the precise linkages between local inflammation and RV dysfunction are ill defined. It is also unclear whether data generated in rodent models have fidelity with larger animals and humans. Therefore, we measured inflammatory markers in several distinct contexts: 1) the hypoxic rat, 2) the hypobaric hypoxic calf, and 3) in human systemic scleroderma (SSd) (with PAH). The profile of inflammatory changes across these models and species was surprisingly concordant. In both rat and bovine RV tissue, increases in IL1‐ra, IL‐1β, IL‐2, IL‐6, TNF‐α, GM‐CSF, and fractalkine were seen, all of which were elevated in serum samples from patients with SSd. VEGF and IFN‐γ increased in tissues but were unchanged in human serum samples. MCP‐1 was increased in SSd serum but was not examined in rat or bovine tissues. These results suggest that inflammatory changes associated with pulmonary artery hypertension are conserved across species and may contribute to PAH associated RV dysfunction. Supported by NIH Grants R21HL104352 (LW&PB), HL084923‐05 (KS) and HL014985‐39 (KS)
The genetic association of polymorphisms rs599839, rs646776 and rs4970834, in chromosomal locus 1p13.3 with coronary artery disease
Authors: Rizk Nasser , El-Menyar Ayman , Wais Idil Souleman , Egue Huda , Baluli Hissa Mohamed , Al Suwaidi Jassim
Published: June 2021

Background

Abnormalities in lipids with increased serum concentration of LDL‐C are risk factors for coronary artery disease (CAD), and atherosclerosis. The aim of this study was to investigate the association between serum LDL‐cholesterol, and significant coronary artery stenosis with well‐known genetic variants in chromosomal locus 1p13.3 among CAD patients with and without significant coronary artery stenosis [case and control groups] respectively.

Methods

Blood was drawn from a total of 178 patients. Genotyping for the variants rs599839, rs646776 and rs4970834, were performed by the 5’ nuclease test using TaqMan assay.

Results

The HDL‐C level of rs599839 was [12.01± 3.9 mM vs. 3.46± 1.45 mM] for GG vs. AA+AG using the recessive genetic model with p value=0.006. Using the additive model, rs64776 variant had a significantly higher triglycerides levels [7.76±2.25 vs. 1.74±0.15 mM] for AA vs. GG respectively with p value=0.031. No significant difference was observed for LDL‐C and cholesterol among genotypes of all variants using different genetic models. The rs599839 variant had a significant association with the severity of coronary stenosis using additive model. The OR was 2.7 times among patients with significant stenosis (p =0.04). Stepwise regression analysis revealed that the following independent variables have a statistically significant effect on LDL‐C level; BMI, diabetes, TC, TG, HDL‐C, and anti‐lipidemic treatment in presence of rs59839 and rs646776 and rs4970834.

Conclusion

In summary, polymorphism rs646776 and of rs599839 were significantly associated with HLD‐C and triglycerides levels, but not with LDL‐C. In addition the variant rs599839 had a significant association with atherosclerotic lesion in coronary vessels.
Deletion of the Antioxidant Enzyme Methionine Sulfoxide Reductase‐A Impairs Autonomic Regulation and Exacerbates Angiotensin‐induced Hypertension and End‐Organ Damage
Authors: Sabharwal Rasna , El Accaoui Ramzi N , Davis Melissa K , Goeken J Adam , Weiss Robert M , Abboud Francois M , Meyerholz David K , Chapleau Mark W
Published: June 2021

Methionine sulfoxide reductase‐A (MsrA) reverses methionine oxidation. We hypothesized that MsrA protects against angiotensin II (AngII)‐induced autonomic and cardiovascular dysfunction. Blood pressure (BP) and heart rate (HR) were measured in MsrA−/− (n=13) and control C57BL/6 (n=7) mice by telemetry, before and during AngII infusion (1000 ng/kg/min). Baroreflex sensitivity (BRS, sequence technique), and cardiac vagal and sympathetic tone (HR responses to methylatropine and propranolol) were measured. In control mice, AngII increased mean BP and sympathetic tone while decreasing BRS and vagal tone (Table). The deleterious effects of AngII were exaggerated in MsrA−/− mice (Table). The antioxidant tempol (T, 1mM drinking water) completely reversed AngII‐induced hypertension and autonomic dysregulation in MsrA−/− mice (Table). Left ventricular (LV) dysfunction (decreased ejection fraction, increased volume/mass ratio), increased ascending aorta diameter, and aneurysms, assessed by echocardiography and post‐mortem histopathology, were present only in AngII‐infused MsrA−/− mice. In summary, MsrA deficiency impairs autonomic regulation and exacerbates AngII‐induced hypertension, LV dysfunction, and arterial pathologies. These results identify MsrA as a novel therapeutic target in hypertension. (HL14388, VA)
Body weight (BW) and body fat (BF) gain due to ovarian hormone loss is attenuated by inhibiting angiotensin converting enzyme (ACE) or angiotensin type 1 receptors (AT1R) in Dahl salt‐sensitive (DS) female rats
Authors: Ji Hong , Zheng Wei , Bajaj Bilkish , Wu Xie , Liu Jun , Sandberg Kathryn
Published: June 2021

Objective

Ovarian hormone loss increases BW and BF gain in women. We and others have shown that ovariectomy (Ovx) increases BW while 17β‐estradiol replacement prevents this effect in DS rats and other animal models. Previous studies implicate the renin angiotensin system (RAS) in BF regulation. Here, we studied the effect of captopril (C), an ACE inhibitor and losartan (L), an AT1R antagonist (0.5g/L each, drinking water) on BW and BF gain induced by Ovx.

Method

Female DS rats were sham‐operated (Sh; n=6) or ovariectomized and treated with vehicle (Ovx), C (Ovx+C) or L (Ovx+L) and maintained on a low sodium (0.01% Na+) diet for 3 months.

Results

Analysis of lean and fat mass by DXA revealed that while Ovx had no effect on the % lean mass gain, Ovx increased the accumulation of BF (p<0.01 vs Sh) and BW (p<0.0001 vs Sh), while both C & L attenuated these effects [BF (% increase from Sh): Ovx, 139; *Ovx+C, 113; *Ovx+L, 115; *p<0.05 vs Ovx; BW (final‐initial)gm/day: Ovx, 0.40; *Ovx+C, 0.27; *Ovx+L, 0.22; *p<0.01 vs Ovx]. The glucose dysregulation induced by Ovx was also markedly improved by C or L treatment.

Conclusions

These findings suggest that 17β‐estradiol loss causes dysregulation of the adipose RAS and thereby contributes to Ovxinduced BF and BW gain and glucose insensitivity. Supported by AG19291 & HL57502.
Effects of the cacao flavanol (‐)‐epicatechin on a rat model of metabolic syndrome
Authors: Ceballos Guillermo , Chamorro German , Ortiz Pilar , Gutierrez Gabriela , Villarreal Francisco
Published: June 2021

Metabolic syndrome (MS) is a cluster of conditions that include increased blood pressure, elevated insulin levels, excess body fat and abnormal lipid levels that when present together, increasing the risk of heart disease, stroke and diabetes. An underlying defect of MS is detrimental changes in mitochondrial structure/function which adversely affects cellular bioenergetics (i.e. metabolism). Flavonoids are natural occurring compounds that are known for having beneficial effects on health and are recognized as safe and non‐toxic. Cacao contains by weight the greatest amounts of flavanols in particular, (‐)‐epicatechin (EPI). We have demonstrated that EPI increases mitochondrial volume and cristae density. In this study, we examined the effects that low dose EPI (1 mg/kg/day by gavage) has on a rat model of MS. MS was induced in male Sprague‐Dawley rats of 8 weeks of age (~200 g weight) by the use of a high cholesterol and high fructose diet given for 7 weeks. Two treatment schemes were tested. In a preventive scheme, EPI was given every day together with the MS diet. In a therapeutic scheme EPI was given for 10 days after the 7 week MS diet implementation. Animals had body weight recorded, blood chemistry and nitrate levels as well as blood pressure. The use of EPI as preventive led to notable and significant improvements in all measured parameters. The use of EPI as a therapeutic intervention also led to improvements in MS endpoints. In conclusion, results indicate that the use of EPI at low doses leads to significant improvements in MS endpoints when used under a preventive or therapeutic scheme.
Effect of Castration on Cardiac Index in the Spontaneously Hypertensive Rat
Authors: Martin Doug , Kjellsen Barton , Messenger Tami
Published: June 2021

Androgens amplify the development of hypertension in the spontaneously hypertensive rat (SHR). We showed that androgens increase venous tone, an important contributor to cardiac filling in SHR. The current study tested the hypothesis that castration of SHR would alter the hemodynamic profile during hypertension development in the SHR.

SHR were subjected to sham operation, castration or castration+testosterone replacement at 4 weeks of age. Cardiac output was tracked longitudinally using echocardiography. At 6 weeks of age the rats were implanted with radiotelemetry transmitters to monitor arterial pressure and heart rate. These variables were monitored to 13 weeks of age.

At 5 weeks of age cardiac index was slightly higher in sham operated (51±5 ml/min/100g) compared to the castrated SHR (40±4 ml/min/100g). Testosterone treatment of castrated SHR partially restored cardiac index (49±8 ml/min/100g). Cardiac index decreased rapidly as the rats aged. At 13 weeks of age there were no differences in cardiac index amongst the groups (Sham: 25±1 ml/min/100g; Castrated 24±2 ml/min/100g; Castrated+testosterone 27±4 ml/min/100g)

Collectively, these data are consistent with the possibility that androgens amplify the development of hypertension, at least in part, by increasing cardiac index during the very early stages of hypertension. Supported by NIH #2RO1HL63053‐ARRA (DM).
Expression and functions of inositol monophosphatase in seawater adapted euryhaline teleosts such as the European eel (Anguilla anguilla) and tilapia (Oreochromis mossambicus)
Authors: Kalujnaia Svetlana , Gellatly Steven , Cramb Gordon
Published: June 2021

Myo‐inositol is a major osmolyte, which plays an important role in cyto‐protection in many organisms exposed to a variety of environmental stresses. Myo‐inositol monophosphatase (IMPA) de‐phosphorylates any myo‐inositol 1‐phosphate recycled from the membrane phospholipid pool or synthesised de novo from glucose‐6 phosphate to generate myo‐inositol. Studies have identified changes in expression of a number of Impa isoforms when euryhaline fish move between freshwater (FW) and seawater (SW), however these changes in gene expression are isoform‐ and species‐specific. The Impa1 isoform is primarily expressed in epithelial tissues exposed to the aquatic environment. In eels and tilapia, Impa1 expression was located throughout the basal/germinal cell layers within the stratified epithelia of the oesophagus, skin, fin and epithelia of the branchial arch and primary filaments, the interstitial cells and/or glomerulae between renal tubules and, specifically in eels, also in branchial and ray fin chondrocytes. In all tissues examined from eels and tilapia, Impa1 expression and inositol contents were markedly up‐regulated after transfer of fish to SW. These results suggest that increased inositol concentrations promotes renal water retention and maintains the cell volume of epithelial and epidermal tissues when fish are exposed to the dehydrating effects of the SW environment.
PTG (Protein Targeting to Glycogen) regulates the osmoprotective transcription factor NFAT5 via SHP‐1 phosphatase
Authors: Zhou Xiaoming , Burg Maurice B. , Ferraris Joan D.
Published: June 2021

Hypertonicity alters phosphorylation of NFAT5, which regulates its activity. PTG is a regulatory subunit of protein phosphatase1 (PP1). By screening a genome‐wide siRNA library, we previously found that PTG inhibits NFAT5 transcriptional activity. The present study addresses the mechanism involved. siRNA‐mediated knock‐down of PTG increases high NaCl‐induced NFAT5 transactivating activity and protein abundance, and over expression of PTG reduces high NaCl‐dependent NFAT5 nuclear localization. We examined the effect of PTG siRNAs on phosphorylation of regulatory sites in p38 and ERK kinases and SHP‐1 phosphatase, previously known to contribute to regulation of NFAT5. SHP‐1 was affected, but not p38 or ERK. PTG siRNAs increase high NaCl‐induced phosphorylation of SHP‐1‐S591, which inactivates it. Mutation of SHP‐1‐S591 to A increases the inhibitory effect of SHP‐1 on NFAT5 transcriptional activity. Further, siRNA‐mediated knock‐down of the catalytic subunit of PP1, PP1gamma, also increases NFAT5 transcriptional activity. High NaCl reduces PP1 activity. We conclude: 1) that reduction by high NaCl of the inhibitory effect of protein phosphatase 1 on NFAT5 activity contributes to high NaCl‐induced activation of NFAT5 by increasing NFAT5 transactivating activity, nuclear localization and protein abundance, and 2) this effect is mediated through SHP‐1.
Renal Collecting Duct Specific GSK 3 alpha Regulates Cellular Distribution and Lithium‐Induced NDI
Authors: Rao Reena , Nilsson Line , Tao Shixin , Woodgett James , Norregaard Rikke
Published: June 2021

Lithium (Li) induced nephrogenic diabetes insipidus is associated with reduced Aquaporin 2 (AQP2) levels and increased proliferation of collecting duct cells (CD) in rodent models. Li is a potent inhibitor of glycogen synthase kinase 3 (GSK3). GSK3 inhibition is known to increase cell proliferation in non‐renal cells. In the current study we examined isoform specific roles of GSK3α and GSK3β in CD structure and function in normal and Li treated mice. We found that gene deletion of GSK3 alpha (3 alpha −/−) caused urinary concentrating defect and polyuria at basal conditions. In these mice, AQP2 protein levels were lower and H‐ATPase, a marker for type A intercalated cells were significantly higher, compared to WT or 3β‐CDnull mice. LiCl treatment (4mmol/Kg, daily IP injection) for 6 days increased urine output and reduced AQP2 levels in WT, but not in 3 alpha −/− mice. Immunostaining for PCNA, marker for dividing cells, AQP4 for principal cells and H‐ATPase demonstrated increase in proliferation of principal cells of cortical and medullary CD cells in WT and 3beta‐CDnull mice, but not in 3 alpha −/− mice. These studies showed that gene deletion of GSK3 alpha changed normal cell distribution in renal CD, reduced AQP2 expression and caused polyuria and that, GSK3 alpha, rather than GSK3beta, is a critical target of Li in renal CDs.