bluuebunny/crossref_metadata_2025_split huggingface dataset:
Inhibition of Angiotensin II Type I Receptor Agonistic Autoantibody Reduces Mitochondrial Mediated Oxidative Stress and Hypertension in Reduced Uterine Perfusion Pressure Rats
Authors: Vaka V. Ramana , Cunningham Mark W , Shirey Kristin , Ibrahim Tarek , Hosler Jonathan , LaMarca Babbette 
Published: November 2023

IntroductionPreeclampsia is characterized by new onset hypertension that usually occurs in the 3rd trimester of pregnancy, and is associated with oxidative stress and increased plasma levels of angiotensin II type 1 receptor agonistic autoantibodies (AT1‐AA). Inhibition of the AT1‐AA in the reduced uterine perfusion pressure (RUPP) rats, a model of preeclampsia, decreases the blood pressure and oxidative stress. Although there is a decrease in overall oxidative stress, the changes in mitochondrial mediated oxidative stress has not been explored with AT1‐AA inhibition in the RUPP rat. We hypothesize that AT1‐AA inhibition prevents mitochondrial dysfunction and decreases mitochondrial mediated reactive oxygen species (ROS) in the RUPP rat.MethodsFemale Sprague Dawley rats were dived into three groups; normal pregnant (NP), RUPP, and RUPP + AT1‐AA inhibition (RUPP+7AA). AT1AA inhibition was achieved by administration of a 7 amino acid epitope binding peptide that inhibits the AT1‐AA from binding to the AT1 receptor. On day 14 of gestation, the RUPP surgery was performed with or without the 7AA peptide (2μg/ml saline) administered via mini‐osmotic pumps. On day 19 of gestation, conscious mean arterial blood pressure (MAP) was measured in RUPP and normal pregnant (NP) rats, placentas were collected for histology and isolation of mitochondria. Dihydroethidium (DHE, 10 μmol/L) staining was performed to determine placental superoxide levels. Isolated mitochondria from placentas were used to measure ROS and enzyme activities of Complex I and IV of the electron transport chain. All rats were age matched to control.ResultsMean arterial blood pressure was higher in RUPPs (n=8; 125± 6 mmHg) vs NPs (n=7; 98±3 mmHg). RUPPs treated with 7AA (n=8; 103± 6 mmHg) showed a significant reduction in blood pressure vs RUPP rats. Superoxide levels were elevated in the placenta of RUPP vs NP rats, and was lowered in RUPP+7AA treated rats. The enzyme activities of Complex I (160 nmol/e‐/min/mg) and Complex IV (501 nmol/e‐/min/mg) in RUPP rats were reduced compared to NPs (198 and 686 nmol/e‐/min/mg respectively). AT1‐AA inhibition improved Complex I and IV activities (198 and 756 nmol/e‐/min/mg respectively) vs RUPPs. ROS levels were higher in the placental mitochondria of RUPP vs NP (15× 10−3 vs 5× 10−3, relative units normalized to Complex IV activity). RUPPs that received 7AA peptide showed a decreased ROS levels (13× 10−3, relative units, normalized to complex IV activity).ConclusionInhibition of AT1AA reduces the blood pressure, placental superoxide levels, mitochondrial electron transport chain activity, and mitochondrial ROS production in RUPP rats. Our data suggest that AT1‐AA inhibition could serve as a novel therapy to reduce mitochondrial mediated oxidative stress in preeclampsia.Support or Funding InformationResearch Supported by RO1HD067541
Effects of NBCn1 deletion on behavioral actions of ethanol in mice
Authors: Choi Inyeong , Jones Jayre , Chung Jaegwon , Lee Jae‐Kyung , Schank Jesse 
Published: November 2023

Genomic studies have been directed towards screening the gene maps of a large number of drug abusers and non‐abusers for genetic differences that might explain the propensity towards addiction. Studies show that the human SLC4A7 gene has genome‐wise association with multidrug abusers in the US population. The SLC4A7 gene encodes the sodium‐bicarbonate cotransporter NBCn1, the membrane protein that moves HCO3− into the cytosol and keeps intracellular pH from falling below normal. In the central nervous system, NBCn1 is particularly important for maintaining pH at synapses, where a small pH change in the synaptic cleft exerts significant effects on membrane excitability and complex firing patterns. In this study, we examined the effect of NBCn1 deletion on behavioral actions of ethanol in mice. Slc4a7 knockout mice lack the functional NBCn1 and develop abnormal pH in neurons and these animals are useful to gain knowledge of pH effects on alcoholism features. Knockout mice and wildtype mice were injected with 3.5 g/kg ethanol and the loss of righting reflex was examined to determine alcohol sedation. Knockout mice showed a 33% increase in the sleep time relative to the wildtype. The sleep latency (onset after injection) was not significantly different between wildtypes and knockout mice. In two bottle choice drinking experiment, mice were given to 3–12% alcohol, 4 days at each concentration. The mutant mice had alcohol consumption by 50% more than the controls at every concentration. Thus, the NBCn1 knockout mice drink more alcohol and also show increased sensitivity to alcohol‐induced sedation. To examine the effects of alcohol on glutamate signaling in knockout mice, we performed extracellular recordings of the CA3 region in hippocampal slices and found decreased NMDA receptor activity in knockout mice. Knockout mice have low NMDA receptor expressions determined by immunoblots with NR1 and NR2 antibodies. In adult wild type mice that were chronically administered with ethanol (>1 month), NBCn1 mRNA was downregulated in the hippocampus and midbrain compared to untreated control mice determined by qPCR. These results are consistent with our hypothesis that pH regulated by NBCn1 contributes to the susceptibility to ethanol. The lack of NBCn1 in KO mice lowers cell pH and makes mice more susceptible to ethanol, and a similar effect might occur when NBCn1‐dependent pH regulation is altered by an abnormal NBCn1.Support or Funding InformationAmerican Heart Association; NIH; Emory URC grant
Microfluidic Models of the Microvasculature for Red Blood Cell Metrology in Sickle Cell Disease
Authors: Betz Jordan , LaVan David , Ackerman Hans 
Published: November 2023

The microvasculature plays an important role in the pathology of sickle cell disease. As erythrocytes deliver oxygen while traversing the arterial and capillary networks, deoxygenated sickle hemoglobin begins to polymerize into fibers, causing changes in erythrocyte mechanical and adhesive properties that contribute to the obstruction of capillaries and postcapillary venules. Micropipette aspiration, atomic force microscopy, optical tweezers, and other related experimental techniques have been used to measure mechanical and adhesive properties of erythrocytes, but these approaches require static conditions that ignore the effects of shear stress. More recently, microfluidic devices have been introduced that are capable of providing realistic shear rates, but employ channel diameters that are much larger than those of human capillaries. We used photolithography and soft lithography techniques to fabricate microfluidic devices from (poly)dimethylsiloxane that recapitulate the internal dimensions of human capillaries. The physical constraints imposed by these channels allow us to measure the dynamics of deformation and “reformation” ‐ how quickly a red cell recovers its biconcave disc shape upon exit from a capillary channel.In our devices, the initial inlet channel bifurcates five times, resulting in 32 capillaries that are 4 μm wide and 1 mm long. The channels are 2.5 μm tall, which orients the erythrocytes flat and keeps them within the focal plane of a microscope objective lens. We used high speed phase contrast video microscopy to record and analyze the cells under physiologically representative flow conditions at1% hematocrit. We measured the differences in erythrocyte elongation (the ratio of the maximum and minimum Feret diameter) and circularity (4π*Area/Perimeter2), as well as characterizing the time to return from the constrained shape in the capillaries to the more relaxed discoid shape, in sickle cell patients and healthy controls under both oxygenated and deoxygenated conditions. These measurements provide quantitative assessment of erythrocyte deformability and shape recovery that we will use to study vascular occlusion that occurs in sickle cell disease.Support or Funding InformationAll samples were drawn from volunteers with informed consent under protocol 03‐H‐0015 as reviewed and approved by the National Heart, Lung, and Blood Institute's Institutional Review Board. This research was supported by the Division of Intramural Research of the National Heart, Lung, and Blood Institute and the Material Measurement Laboratory of the National Institute of Standards and Technology.
A Diet High in Fat and Cholesterol Reduces Flow‐induced Dilations of Small Visceral Arteries and Involves Kir Channel Impairment
Authors: Fancher Ibra S , Ahn Sang Joon , Bian Jing‐Tan , Phillips Shane A , Levitan Irena 
Published: November 2023

Distinct mechanisms regarding the progression of vascular endothelial dysfunction in diet‐induced obesity remain largely unknown. A diet high in saturated fat and cholesterol promotes vascular dysfunction prior to more severe disease states. Vascular K+ channels have also been implicated in the progression of vascular disease with obesity and our group established a definitive effect of dyslipidemia on Kir channel function. In this study we investigated the role of flow‐induced Kir2 channel activation and suggest Kir2 channel impairment in 1) a mouse model of diet‐induced obesity and 2) morbidly obese humans. We expand our investigation to spatially distinct arteries isolated from visceral and subcutaneous adipose. We show that a high fat, high cholesterol diet reduces flow‐induced vasodialtion in visceral arteries of mice and humans while subcutaneous endothelium‐dependent function remains intact. Importantly, barium reduces flow‐induced vasodilations in normal fed mice and in subcutaneous arteries, but has no effect on visceral arteries of obese mice or humans, implicating spatially distinct Kir channel dysfunction in obesity. Interestingly, visceral endothelial‐dysfunction correlates with elevated serum LDL. Our future direction will focus on the molecular mechanisms of Kir channel dysfunction in diet‐induced obesity.Support or Funding Information5 T32 DK080674 andNIH R01 HL073965
Lysophosphatidic Acid Promotes a Switch from Nitric Oxide to Hydrogen Peroxide as the Mediator of Flow‐Induced Dilation in Human Arterioles
Authors: Chabowski Dawid S. , Kadlec Andrew O. , Ait‐Aissa Karima , Hockenberry Joseph C. , Beyer Andreas M. , Gutterman David D. 
Published: November 2023

IntroductionMicrovascular dysfunction is strongly predictive of cardiovascular pathology. Previous studies in our laboratory have shown that flow‐induced dilation (FID) in patients with coronary artery disease (CAD) is dependent on hydrogen peroxide (H2O2) as opposed to the more vasculoprotective nitric oxide (NO)‐mediated mechanism observed in arterioles from patients without CAD. Lipid phosphate phosphatase 3 (LPP3) is an integral membrane protein that dephosphorylates lysophosphatidic acid (LPA). Large genome‐wide association studies have identified a single nucleotide polymorphism in PPAP2B, a gene encoding LPP3, as a predictor of CAD independent of traditional risk factors. A pro‐inflammatory endothelial phenotype with decreased levels of endothelial nitric oxide synthase (eNOS) and atherosclerosis occurs with reduced LPP3 expression and an increase in LPA. We hypothesize that treatment of healthy microvessels with LPA induces a switch in mediator of FID from NO to H2O2, recapitulating the disease state.MethodsHuman arterioles were isolated from healthy adipose and prepared for video‐ and fluorescence microscopy. LPA (10 μmol/L) was added to the bath for 30 minutes prior to constriction with Endothelin‐1. Internal diameters were measured at steady‐state during graded increases in intraluminal pressure gradients (flow) in the presence/absence of a NOS inhibitor (L‐NAME; 10−4 M) or H2O2 scavenger (PEG‐catalase; 500 units/mL). MitoPY1 fluorescent probe was used to detect mitochondrial H2O2 in response to flow in the presence and absence of LPA and PEG‐catalase.ResultsAcute exposure to LPA reduced FID compared to control vessels (% max dilation, LPA: 61.7±6.0, n=4 vs. Control: 77.7±3, n=2). PEG‐catalase largely suppressed the remaining dilation (LPA + PEG‐catalase: 17.2±6.8, n=3). Interestingly, treatment with L‐NAME improved FID compared to LPA alone (LPA+LNAME: 83.7±6.8, n=3 vs. LPA: 61.6±6.0, n=4). LPA increased flow‐induced mitochondrial H2O2 production (% increase to flow, Control: −1±25, n=3 vs. LPA: 154±40, n=4). PEG‐catalase suppressed the increase in LPA‐induced, flow‐stimulated H2O2 production (PEG‐cat: 37±19, n=2). In summary, LPA is capable of shifting the mediator of dilation from NO to H2O2 in healthy arterioles. In addition, in the presence of LPA, blocking NOS paradoxically improves FID, raising the question whether NOS is uncoupled in this situation. Mitochondria appear to be the source of LPA‐induced increase in H2O2. We conclude that LPA may play a role in microvascular dysfunction in CAD, invoking changes in NOS and ROS formation.
The microtubule‐associated End Binding proteins regulate the ER exporting and trafficking of the TRPM4 channels
Authors: Romero Anibal , Blanco Constanza , Morales Danna , Mogollones Ignacio , Álvarez Alhejandra , Varela Diego , Cáceres Mónica , Cerda Oscar 
Published: November 2023

Trafficking and localization of ion channels are critical events for their proper function. These processes are relevant in several cellular functions, such as developmental programmes, cell polarity and local membrane potential. TRPM4 is a Ca2+‐activated non‐selective cationic channel expressed in different tissues and cell types, and is involved in a wide variety of physiological processes, such as insulin secretion and mechanotransduction. Altered activity, expression and localization of these channels might trigger several pathophysiological conditions. Indeed, TRPM4 plasma membrane overexpression has been related to cardiovascular and neurodegenerative diseases and cancer. Therefore, the mechanisms involved in the regulation of TRPM4 activity constitute an important area of biomedical research. Protein‐Protein Interactions (PPIs) control the expression, trafficking, localization and biophysical properties of ion channels. Thus, the identification of novel TRPM4‐related PPIs and their characterization might contribute to dissect the regulatory mechanisms of this channel. Interestingly, bioinformatics analyses of the primary sequence of TRPM4 allowed us to identify a putative interacting motif to End Binding (EB) proteins, novel members of the microtubule plus‐end tracking proteins (+TIPs). These proteins bind a consensus motif (SxIP) in their substrates and are involved in a plethora of cellular processes, including growing dynamics of the microtubule cytoskeleton, focal adhesion dynamics, cell migration and protein trafficking, targeting and localization. Here, we provide novel data suggesting that TRPM4 interacts with EB proteins. Moreover, we show that the mutation (TRPM4ΔSWIP and TRPM4SWNN) of the putative EB‐binding motif abolishes the TRPM4‐EB interaction. We also found that these mutant variants show a reduced expression of the mature population of the channel and display an endoplasmic reticulum (ER)‐like distribution. Furthermore, these mutant variants present a decreased expression in the plasma membrane, consistent with a reduction in the whole‐cell electrophysiological recordings. Conversely, we demonstrated that the expression of a soluble fragment containing the wild type N‐terminal region of TRPM4 (EGFP‐N‐TRPM4WT) reduces the membrane expression of TRPM4. These results suggest that EB interaction is necessary for the proper trafficking and activity of TRPM4. These findings might contribute to the understanding and characterization of novel mechanisms involved in TRP channel trafficking/localization, and could be applied to the design of therapeutic strategies against TRPM4 gain‐of‐function‐associated pathologies.Support or Funding InformationThis research was funded by Fondecyt 11121239 to O.C. and Fondecyt 11140064 to M.C. A.R. is a Conicyt Doctoral Fellowship recipient.
Salidroside Inhibits Clinorotation‐induced Apoptosis in Pulmonary Microvascular Endothelial Cells by regulating signaling pathway of Endoplasmic reticulum stress
Authors: Kang Chunyan , YUAN Ming , WANG Jingyu , YUAN Min , LIU Changting 
Published: November 2023

ObjectiveMicrogravity or simulated microgravity induces dysfunction of endothelium. We have previously demonstrated that clinorotation induces apoptosis in microvascular endothelial cells. Salidroside(Sal), one of main active ingredients extracted from the root of Rhodiola rosea, has been documented to have potent antiapoptotic effects on some cells. The present study investigated whether salidroside inhibited clinorotation‐ induced apoptosis in microvascular endothelial cells by regulating the signaling pathway of endoplasmic reticulum(ER) stress.MethodsHuman pulmonary microvascular endothelial cells(HPMECs) were divided into control group, clinorotation group, clinorotation+Sal and clinorotation+Sal+ LY294002 pretreatment groups. Microgravity was simulated by clinorotation. After 72h clinorotation, the apoptotic rate of HPMECs was detected by TUENL assay and JC‐1 staining. The protein expressions of PI3K, Akt, Bcl‐2, Bax, caspase‐3 and caspase‐9 were determined by Western blot. Meanwhile, the ER stress related proteins such as GRP78, CHOP, IRE1a and ATF6 were measured.ResultsTUNEL assay, together with JC‐1 staining indicated that 72h clinorotation significantly induced apoptosis in HPMECs, which was reversed by salidroside treatment. Clinorotation also decreased the protein expression of PI3K, Bcl‐2 and phospho‐Akt level, increased the apoptosis related protein expression of Bax, caspase 3 and caspase 9. Meanwhile, Clinorotation increased the ER sensors protein expression of GRP78, CHOP, IRE1a and ATF6. Pretreatment with salidroside reversed the changes of these protein expression induced by 72h clinorotation, however, the protective effects of salidroside were reversed by inhibitor of PI3K/Akt, LY294002.ConclusionOur results suggested that the apoptosis of HPMECs induced by 72h clinorotation can be alleviated by pretreatment of salidroside. The protective effects of salidroside on HPMECs apoptosis may come from the upregulation of PI3K/Akt signal pathway and downregulation of ER stress pathway.Support or Funding InformationFunded by the Defense Medical Fund of China (Grant No. 06Z048) and State key lab of space medicine fundamentals and application, China Astronaut Research and Training Center (grant No. SMFA14B01).
Tolfenamic acid downregulates β‐catenin in colon cancer
Authors: Lee Seong‐Ho , Ha Taekyu , Lou Zhiyuan 
Published: November 2023

Tolfenamic acid, a nonsteroidal anti‐inflammatory drug (NSAIDs) has been shown to induce apoptosis and suppress cancer cell growth and tumorigenesis in colon cancer. Wnt/β‐catenin pathway is highly activated in colon cancer and considered as target for cancer prevention and therapy. The current study was performed to study if tolfenamic acid affects β‐catenin pathway in colon cancer. Tolfenamic acid resulted in an inhibition of cell growth and downregulation of β‐catenin in dose‐ and time‐dependent manner in different types of human colon cancer cells. Reduction of β‐catenin was associated with ubiquitin‐mediated proteasomal degradation. Treatment of tolfenamic acid to colon cancer cells downregulated expression of Smad2 and Smad3. Overexpression of Smad2, but not Smad3 blocked tolfenamic acid‐induced downregulation of β‐catenin. Decreased expression of β‐catenin, Smad2, Smad3 and VEGF were observed in tumor tissues of mice treated with tolfenamic acid at dose of 50 mg/kg body weight. Our findings suggest that tolfenamic acid inhibited cancer cell growth by inducing β‐catenin degradation via ubiquitin‐proteasome pathway and Smad2 mediated tolfenamic acid‐induced downregulation of β‐catenin.Support or Funding InformationAmerican Cancer Society grant (RSG‐11‐133‐01‐CCE) to S‐H Lee
Paks integrate polarity and vesicle transport signaling in ovarian epithelial cancers
Authors: Radu Maria , Mumber Hannah , Chernoff Jonathan 
Published: November 2023

P21 activated kinases (Paks) are effectors for small RhoGTPases and have been widely implicated in regulation of cytoskeleton arrangements and cell architecture. Paks gained a lot of momentum when several members of the family have been shown to be over‐expressed in a variety of cancers, including ovarian, breast, prostate and colon. These discoveries led to augmented efforts to discover new and improved Pak inhibitors to be used in clinic. Recent data shows however that the absence or complete inhibition of Pak2 leads to lethality in adult animals. It is thus imperative that we have a better understanding of Pak regulated pathways and their direct substrates.In an effort to unveil new molecular efforts in Pak signaling, we employed a modified kinase assay in conjuncture with peptide identification by LC/MS. A high percentage of the newly discovered Pak substrates fell in the category of molecules that control cell communication. Cell communication relates to all modes of inter and intra cell messaging, including cytoskeleton arrangement and vesicle transport. Importantly, these are also key elements that when deregulated lead to epithelial cancer initiation and progression.Among the newly identified Pak direct targets that regulate cell communication CDK16 and NSF have been previously shown to regulate each other and together to control polarity and vesicle transport respectively. We found that Pak1 and Pak2 phosphorylate CDK16 at S119 and NSF at S207. Our data has shown that unphosphorylatable CDK16 mutants are unable to bind Pak1 and 2 and that inactive Paks do not bind CDK16. This suggests that functional protein complexes are formed between CDK16 and Pak1 and 2. Furthermore, we found that in an ovarian carcinoma cell line overexpression of an unphosphorylatable CDK16 mutants can induce stress fiber and focal adhesion disassembly (link to deregulated polarity), while a CDK16 mutant that mimics a constitutively phosphorylated state can partially rescue stress fiber disassembly induced by Pak inhibition.These data strengthen our hypothesis that Paks regulate epithelial cancer progression by phosphorylating CDK16 and NSF, proteins that integrate polarity and vesicle trafficking pathways. To confirm this hypothesis we will first investigate the temporal and spatial regulatory events that take place between Pak, CDK16 and NSF and how these events influence polarity and vesicle transport. Secondly, we will investigate how the dis regulation of Pak/CDK16/NSF pathway leads to epithelial cancer initiation and spread. Thirdly, we will consider the possibility of combining Pak and CDK16 inhibition to arrest the growth various epithelial carcinomas.Our results address the possibility to target Pak signaling in Pak overexpressing cancers by inhibiting CDK16, a downstream kinase effector and to override some of the critical effects of current Pak inhibitors.Support or Funding InformationNIH T32 Training grant; NCI CA06927 to Fox Chase Cancer Center