bluuebunny/crossref_metadata_2025_split huggingface dataset:
Contribution of kv7 channels to acetylcholine‐induced dilation in human adipose arterioles
Authors: Nishijima Yoshinori , Zhang David 
Published: November 2023

Voltage‐gated K+ (KV) channels, especially shaker‐related KV1, serve as important regulators of vascular tone and reactivity. Emerging evidence suggests that KCNQ or KV7 channels may represent another subfamily of KV channels contributing to vascular tone regulation. Functional KV7 channels have been detected in vascular smooth muscle and endothelial cells of several vascular beds, although their significance in mediating physiological responses remains largely unclear. We investigated the contribution of KV7 channels to acetylcholine (ACh)‐induced dilation in ex vivo human adipose arterioles. Fresh adipose tissues were obtained as discarded surgical specimens from subjects without and with coronary artery disease (non‐CAD and CAD, respectively), and arterioles (100–200 μm, internal diameter) were dissected and cannulated under 60 mmHg and examined for diameter changes using video microscopy. In arterioles from non‐CAD subjects, ACh (10−9–10−5 M) induced potent vasodilation in a concentration‐dependent manner (% maximal dilation at 10−7 and 10−5 M, 62±6 and 93±2, respectively, n=6). KV7 channel blocker linopirdine (10 μM) significantly reduced the dilation to 10−7 M ACh (% dilation 18±6 vs. 62±6 in control), whereas blockade of KV1.5 (a major KV1 channel in human adipose arterioles) by DPO‐1 (1 μM) or Psora‐4 (30 nM) was without effect. In arterioles pretreated with L‐NAME (100 μM, a nitric oxide synthase inhibitor) and indomethacin (10 μM, a prostacyclin inhibitor), linopirdine also reduced ACh‐induced dilation (% dilation to 10−7 ACh, 10±4 vs. 30±6 in control with L‐NAME and indomethacin, and % dilation to 10−6 M ACh, 45±8 vs. 80±4 in control with L‐NAME and indomethacin). In contrast to arterioles from non‐CAD subjects, ACh‐elicited dilation in CAD arterioles was not affected by linopirdine (% dilation at 10−7 M, 57±4 vs. 66±7 in control). These results indicate that KV7 channels are involved in ACh‐induced dilation in human adipose arterioles from non‐CAD subjects and part of this effect is related to an endothelium‐dependent hyperpolarization (EDH) mechanism of dilation. In addition, the involvement of KV7 in ACh‐induced arteriolar dilation seems to be reduced in subjects with disease (CAD). This study provides further support for the recruitment of KV7 to the functional K+ channel repertoire in vascular cells.Support or Funding InformationNational Heart, Lung and Blood Institute Grant R01‐HL 096647
Vasodilatory Function in Human Skeletal Muscle Arteries with Advancing Age: Role of Adropin
Authors: Kwon Oh Sung , Andtbacka Robert , Hyngstrom John , Richardson Russell 
Published: November 2023

BackgroundAging is associated with impaired endothelium‐dependent dilation in human skeletal muscle feed arteries (SMFAs). However, the specific mechanisms underlying the age‐related impairment in the regulation of blood flow have yet to be clarified. Recent evidence has suggested that adropin, a peptide secreted by the liver, may play a role in cardiovascular health. Therefore, this study sought to determine if treatment with adropin, in vitro, can restore the age‐related endothelial dysfunction in human SMFAs.MethodsAdropin protein expression was assessed in 3 groups of subjects, young (28±4 yrs, n=10), middle aged (50±6 yrs, n=10) and old (75±7 yrs, n=16) with Western blot analysis. Using pressure myography, vasodilation in SMFAs with and without adropin incubation, from these young, middle aged, and old subjects was assessed in response to three stimuli 1) flow‐induced shear stress, 2) acetylcholine (ACh), and 3) sodium nitroprusside (SNP).ResultsProgressive age‐related reductions in endothelial‐dependent vasodilatory function were evident in response to both flow (Young: 97 ± 13; Middle‐age: 62 ± 12; Old: 32 ± 14%, P < 0.05) and ACh (Young: 98 ± 15, Middle‐age: 68 ± 13 %; Old: 40 ± 10 %, P < 0.05). Adropin treatment in the middle aged and old SMFAs restored the vasodilatory response to flow (Middle‐age + Adropin: 88 ± 10; Old + Adropin: 78 ± 11 %) and ACh (Middle‐age + Adropin: 94 ± 16%; Old + Adropin: 83 ± 10 %, P < 0.05) (p<0.05). Endothelium‐independent vasodilation (SNP) was unaffected by age and adropin, also, had no effect. Interestingly, adropin upregulated eNOS protein expression in the middle aged and old SMFAS and there was a significant positive correlation between adropin protein expression and both flow‐mediated and ACh‐dependent vasodilation.ConclusionsOverall, these findings suggest that adropin may play a critical role in age‐associated endothelial dysfunction. Furthermore, adropin could be a novel therapeutic approach to restore endothelial function and NO bioavailability in the elderly.Support or Funding InformationNIH Heart, Lung, Blood Institute grants (HL‐ 091830) and Veterans Affairs Rehabilitation Research and Development Merit (E6910‐R)
Enhanced myogenic response of cerebral arteries induced by pre‐existing hypertension is intact after traumatic brain injury
Authors: Szarka Nikolett , Amrein Krisztina , Czeiter Endre , Buki Andras , Koller Akos , Toth Peter J 
Published: November 2023

Traumatic brain injury (TBI) impairs pressure‐induced myogenic autoregulation of cerebral blood flow (CBF), which is associated with increased mortality and bad functional outcome of TBI patients. When blood pressure drops lack of myogenic autoregulatory function of cerebral vessels exposes brain tissue to hypoperfusion and ischemia; in case of high blood pressure it allows high blood volume to penetrate the brain exacerbating disruption of the blood brain barrier, development of hemorrhages and elevation of intracranial pressure. Hypertension, the most important cardiovascular risk factor in TBI patients, enhances myogenic tone of cerebral arteries via increased vascular production of 20‐hydroxyeicosatrienoic acid (HETE), which protects brain tissue form pressure/volume overload but leads to increased susceptibility to cerebral ischemia. Although both effects may potentiate the vascular consequences of TBI, it is not known how hypertension modulates the effect of TBI on myogenic responses of cerebral vessels. To establish the effect of TBI on cerebral myogenic responses in hypertension we studied isolated middle cerebral arteries (MCA) of normotensive and spontaneously hypertensive rats (SHR) after severe impact acceleration diffuse brain injury. We found that TBI diminished myogenic constriction of MCAs in normotensive animals, but the 20‐HETE mediated enhanced myogenic tone in SHRs was not affected by traumatic brain injury. These results suggest that following brain trauma hypertensive patients may be protected from high blood pressure, but are likely to be more prone to cerebral ischemia when blood pressure drops, and therefore require different therapeutic strategies than normotensive patients.Support or Funding InformationThis work was supported by grants from the Marie Curie Actions SMARTER 7th Framework Program of the European Union 606998 (to NSz and AK), the Hungarian Academy of Sciences (the Bolyai Research Scholarship BO/00634/15 to PT), the Hungarian Brain Research Program (Grant No. KTIA_13_NAP‐A‐II/8 to PT, ECz and AB) and the Hungarian Scientific Research Fund (K108444 to AK).
CRISPR/Cas9 creation of a mouse lacking a unique motif on alpha globin demonstrates its critical role in regulating nitric oxide in vivo
Authors: Filho Gilson Bras Broseghini , Butcher Joshua T , Keller Steven TC , Poonam Sharma , Kwangseok Hong , DeLalio Leon J , Wenhao Xu , Padilha Alessandra Simão , Columbus Linda , Sonkusare Swapnil , Isakson Brant E 
Published: November 2023

Alpha globin and eNOS form molecular complexes in microvascular endothelial cells that can regulate nitric oxide (NO) availability. On the alpha globin protein we have identified the amino acids 35–44 (FASFPTTKTY) as being essential for eNOS interaction. Using in silico prediction, we identified the 37–40 amino acid motif (SFPT) as being especially crucial for eNOS interaction, which we confirmed with purified protein binding assays. To confirm the importance of this motif in vivo, we used CRISPR/Cas9 to create a mouse with alpha globin missing amino acids 37–40 in the Hba1 gene. Whole genome sequencing confirmed the mutation and demonstrated a lack of any off‐target effects. We found that homozygous Hba1Δ37–40 mice were not viable (0 mice born from 8 different heterozygous pairs; 16 litters) and were re‐absorbed in utero by approximately day E12. The lethal homozygous Hba1Δ37–40 embryos had significantly dilated arteries and cardiac ventricles. For this reason we utilized Hba1WT/Δ37–40 heterozygous animals; these mice had no difference in total alpha globin protein expression in arterioles and were viable. These mice had decreased blood pressure and significantly increased heart rates. Angiotensin II infusion only had a mild effect on blood pressure that was significantly lower than controls with Angiotensin II. Blood NOX levels were significantly higher than controls. In line with this observation, third‐order mesenteric arterioles from heterozygous Hba1WT/Δ37–40 mice had significant increases in NO release when stimulated with the endothelial muscarinic agonists carbachol. We also detected a significant decrease in the amount of cGC present in smooth muscle cells. Our results demonstrate the importance of discreet motifs on alpha globin in regulating NO in endothelium in a unique CRISPR‐derived in vivo mouse model.Support or Funding InformationFunding: HL088554,HL120840.
Maintenance of Nerves in the Rat Mesentery Culture Model
Authors: Hodges Nicholas 
Published: November 2023

Neurovascular patterning represents an emerging area of microvascular research and is critical for tissue remodeling. While overlapping molecular signals highlight links between angiogenesis and neurogenesis, advancing our understanding of their spatial and temporal coordination is limited by a lack of in vitro models that contain both systems. One such potential model is the rat mesentery culture model, which our laboratory has recently introduced as an ex vivo tool to investigate cellular dynamics during angiogenesis and lymphangiogenesis in an intact microvascular network scenario. The objective of this study was to evaluate the effect of media type on nerve maintenance in the rat mesentery culture model. Adult male Wistar rat mesenteric tissue windows were harvested, rinsed in sterile DPBS and media and then cultured per media group: 1) minimum essential media (MEM) and 2) nerve basal medium (NBM) with NGF and 20% fetal bovine serum (FBS). On day 3 tissues were labeled for PECAM and class III β‐tubulin or PECAM and NG2. Unstimulated (not cultured) tissues were also labeled as a control group. Nerve presence at the arteriole level was quantified by the ratio of network feeding arterioles with nerve alignment to the total number of arterioles. Nerve presence at the capillary level was quantified by the length of nerve alignment per vascular area (μm/mm^2). NG2 and class III β‐tubulin positive nerve presence along feeding arterioles was decreased for MEM only group compared to control levels (NG2: Control = 0.875 ± 0.06, MEM = 0.6 ± 0.09, p < 0.05; Class III β‐tubulin: Control = 0.937 ± 0.06, MEM = 0.412 ± 0.10, p < 0.05). In contrast, nerve presence along arterioles identified by either label was not statistically different between the control and NBM+NGF+FBS groups (NG2: Control = 0.875 ± 0.06, NBM+NGF+FBS = 0.917 ± 0.05; Class III β‐tubulin: Control = 0.937 ± 0.06, NBM+NGF+FBS = 0.937 ± 0.06). NBM+NGF+FBS also maintained NG2 and class III β‐tubulin positive nerve length per vascular area compared to the control group (NG2: Control = 1930 ± 134, NBM+NGF+FBS = 1846 ± 214 μm/mm^2; Class III β‐tubulin: Control = 1134 ± 124, NBM+NGF+FBS = 1589 ± 299 μm/mm^2). Nerve presence at the capillary level was not quantified for the MEM group because of the lack of consistent capillary regions with observable nerves. The results of this study supports the use of specific media types to maintain nerve presence across cultured microvascular networks and implicates the rat mesentery culture model as a novel tool to investigate neurovascular patterning ex vivo.Support or Funding InformationNIH R01AG049821, NIH P20GM103629
Functional microdomain in the cardiac pacemaking in mouse
Authors: Lang Di , Warden Aleah , Kamp Timothy , Glukhov Alexey 
Published: November 2023

BackgroundHeart rhythm imitates from the leading pacemaker called sinoatrial node (SAN). SAN cell contains a number of unique ion channels and transporters responsible for cardiac pacemaking which were prevailingly believed freely mobile in the cell membrane. Recently, it has been reported that Caveolin‐3 (Cav3), a myocyte‐specific scaffolding protein, compartmentalizes these ion channels and transporters within caveolar membranes. These proteins are attributed to a surface “membrane clock” which is spatiotemporally coupled with subcellular calcium handling machinery (i.e. “Ca2+‐clock”) forming a coupled‐clock pacemaker cell system. Such coupling is achieved via extremely complex crosstalk between the two clocks and is hypothesized to rely on functionality of a macro‐molecular pacemaker signaling complex targeted to caveolae.Methods and ResultsIn vivo ECG telemetry, ex vivo high‐resolution calcium/voltage optical mapping, and in vitro confocal imaging of [Ca2+]i were performed in wild type (WT, n=17) and conditional cardiac‐specific Cav3 knockout (Cav3−/−, n=14) mice. Cav3−/− mice exhibited SAN pacemaking abnormalities characterized by alternating periods of tachycardia‐bradycardia rhythm originated from distinct anatomical locations, including SAN (during tachycardia) and ectopic foci located within the intercaval region outside of the SAN (during bradycardia). The ectopic foci areas consisted of non‐tubulated myocytes and had prolonged Ca release time and Ca transient duration due to desynchronized sarcoplasmic reticulum Ca release. This was reproduced in non‐tubulated (but not in tubulated) WT atrial myocytes by disruption of caveolae by methyl‐β‐cyclodextrin. To investigate unstable SAN pacemaking, we characterized subsarcolemmal local Ca2+ releases (LCRs) from ryanodine receptors (RyR), a critical component of Na+/Ca2+ exchanger (NCX)‐mediated late diastolic depolarization of SAN action potential (AP). In Cav3−/− SAN myocytes, regular LCRs were recorded; however, a half of them failed to initiate APs resulting in irregular SAN rate. We found significantly prolonged LCR‐to‐AP period (73.9±6.7 ms in WT vs. 130.6±15.0 ms in Cav3−/−, p<0.01) and increased LCR amplitude (normalized value: 0.54±0.046 %/μm in WT vs. 0.90±0.13 %/μm in Cav3−/−, p<0.01). A longer time and a higher driving force needed to activate NCX and boost AP may indicate redistribution of NCX from their canonical location in caveolae to non‐native membrane regions where their functional coupling with RyRs is lost.ConclusionOur findings demonstrate that Cav3 plays a crucial role in supporting functional integrity of the SAN by synchronizing calcium‐voltage coupling and regulating rhythmic LCRs. Loss of Cav3 disrupts caveolae‐associated macro‐molecular pacemaker signaling complex which results in functional uncoupling of NCX from RyRs and affects calcium‐voltage synchronization leading to SAN dysfunction.
Ipragliflozin Prevents the Worsening of Ischemic Heart Failure in Diabetic Rats Independent of its Diuretic Effect
Authors: Kamada Kazuhiro , Saku Keita , Kishi Takuya , Akashi Takuya , Takehara Takako , Tsutsui Hiroyuki , Sunagawa Kenji 
Published: November 2023

IntroductionAlthough diabetes mellitus (DM) increases cardiovascular events, no clinical trials had ever shown the benefit of glucose‐lowering therapy on heart failure (HF). Recently, sodium‐glucose cotransporter 2 (SGLT2) inhibitor has shown to lower cardiovascular events and hospitalization for HF in DM (EMPA‐REG OUTCOME). To clarify the beneficial mechanisms of SGLT2‐inhibitor for HF, we investigated the impact of SGLT2‐inhibitor, ipragliflozin, on hemodynamics and cardiac function in a rat model of HF with DM.MethodsWe allocated Sprague‐Dawley rats into streptozotocin (60 mg/kg)‐induced DM or non‐DM groups. We created myocardial infarction (MI) in both groups and administered ipragliflozin (3 mg/kg/day) or vehicle from two weeks after MI. Four weeks after the initiation of treatment, we compared hemodynamic parameters among 4 groups (DM‐MI+ipragliflozin, n=9; DM‐MI+vehicle, n=6; MI+ipragliflozin, n=7; MI+vehicle, n=7).ResultsDaily urine volume nearly quadrupled in DM rats in comparison with non‐DM rats. In DM rats, ipragliflozin significantly lowered blood glucose (309±36 vs. 478±107 mg/dL, p<0.01), prevented the worsening of left ventricular (LV) ejection fraction (−1.4±4.8 vs. −8.8±5.1%, p<0.05), increased max LVdP/dt (6193±667 vs. 5015±456 mmHg/sec, p<0.01) and tended to decrease LV end‐diastolic pressure (24.8±5.3 vs. 30.5±5.7 mmHg, p=0.069). However, ipragliflozin did not affect urine volume. In non‐DM rats, the beneficial effects of ipragliflozin on HF were limited.ConclusionsIpragliflozin attenuated the worsening of HF in DM rats, while its beneficial effects were obscured in non‐DM rats. These results indicate that ipragliflozin has direct cardioprotective effects on HF in DM independent of its diuretic effects.
Diaphragm Muscle Function and Antioxidant Levels in Chronic Heart Failure
Authors: Hackfort Bryan T , Lewis Robert , Gao Lie , Zucker Irving H , Schultz Harold D 
Published: November 2023

Nearly 5.7 million adults in the United States have chronic heart failure (CHF). CHF is associated with dyspnea and impaired diaphragm function. Shortness of breath in CHF patients may lead to decreased exercise and mobility, further exacerbating skeletal muscle atrophy and the effects of heart failure. Oxidative stress contributes to limb skeletal muscle weakness in heart failure, however the role of oxidative stress in the diaphragm is poorly characterized. Nuclear factor (erythroid derived 2)‐like 2 (Nrf2) is a master regulator of antioxidant enzymes and is decreased in limb skeletal muscles following heart failure. We hypothesize that Nrf2 is decreased and oxidative stress is increased in diaphragm muscle following heart failure resulting in impaired diaphragm muscle function. To test this hypothesis, 8–9 week old, male mice were infarcted by left coronary artery ligation and aged for 10 weeks post MI. Diaphragm function was measured using plethysmography in sham and heart failure conditions. After collection of the diaphragm, protein levels of antioxidant enzymes were measured. Preliminary results (N=2–3) show Nrf2 is not changed (1.1 fold relative to sham) in the diaphragm following CHF. Further, the antioxidant enzymes glutathione peroxidase (2.0 fold), catalase (1.3 fold), and SOD2 (1.9 fold) all tended to increase with CHF compared to sham. Breathing studies showed CHF mice (N=4) tended to have increased respiratory rates (RR = 265 breaths per minute) compared to sham (RR = 208) and decreased tidal volumes. While preliminary, these results suggest that altered breathing dynamics may be associated with altered diaphragm workload in heart failure, with an increase in antioxidant enzymes independent of Nrf2. The increase in antioxidant enzymes in CHF may be related to more oxidative stress from increased workload. Further studies will increase sample size and look at oxidative stress in diaphragm via DHE stain, diaphragm muscle function via muscle tension studies, and mitochondrial respiration.Support or Funding InformationThis work was supported by funds from the National Institutes of Health 5P01HL062222‐17
Identification of HuR‐dependent gene expression in cardiac hypertrophy
Authors: Slone Samuel , Anthony Sarah R , Nieman Michelle L , Lorenz John N , Rubinstein Jack , Benoit Joshua B , Tranter Michael 
Published: November 2023

HuR is a widely expressed RNA binding protein that interacts with specific AU‐rich domains in target mRNAs and regulates their expression by modulating stability and/or translation. RNA binding proteins such as HuR are becoming recognized as potential mediators of cardiac physiology and pathology but the role of HuR in cardiac hypertrophy is largely unknown. Our results show that HuR undergoes cytoplasmic translocation, indicative of its activation, in hypertrophic cardiac myocytes and is necessary for hypertrophic cell growth. Using an inducible cardiomyocyte‐specific HuR deletion mouse (iCM‐HuR−/−), we have shown that cardiac specific deletion of HuR significantly reduces hypertrophy compared to wild‐type mice at 8 weeks post‐transverse aortic constriction (TAC). More importantly, the iCM‐HuR−/− mice exhibited significantly preserved ejection fraction, reduced chamber dilation, and less fibrosis compared to WT at 8 weeks post‐TAC. To elucidate the downstream target genes of of HuR in the heart and identify potential mechanisms by which HuR deletion reduces pathological cardiac hypertrophy, we used RNA‐seq to identify HuR‐dependent gene expression in the setting of cardiac hypertrophy. Using the NIH DAVID Functional Annotation Tool, our results indicate a significant increase in genes promoting anti‐apoptotic Akt signaling in hearts from iCM‐HuR−/− mice following TAC. However, since HuR binding typically stabilizes target mRNA, we observed a much larger number of significantly down‐regulated mRNA transcripts in iCM‐HuR−/− mice. Among these were genes involved in key cellular growth processes such as muscle cell differentiation, cell proliferation, cell cycle, and positive regulation of transcription. Thus, our results suggest that HuR deletion protects the heart from pathological cardiac hypertrophy by promoting pro‐survival Akt signaling while decreasing the expression of genes involved in cell growth and differentiation.Support or Funding InformationThis work was partially funded by an American Heart Association Scientist Development Grant and NIH R01 HL132111 (MT).
Alterations in the Balance of Cardiomyocyte Glucocorticoid and Mineralocorticoid Receptor Signaling Lead to Heart Failure
Authors: Oakley Robert , Cruz‐Topete Diana , He Bo , Foley Julie , Myers Page , Murphy Elizabeth , Chambon Pierre , Willis Monte , Cidlowski John 
Published: November 2023

Stress is associated with adverse cardiac outcomes; however, the mechanisms underlying these effects are poorly understood. Glucocorticoids are stress hormones that regulate cell and tissue homeostasis through two closely related nuclear receptors, the glucocorticoid receptor (GR) and mineralocorticoid receptor (MR). Cardiomyocytes express both these receptors but little is known concerning their coordinated actions in heart physiology and pathology. Here, we show that patients with failing hearts have a disparity in the relative levels of GR and MR compared to non‐failing donor hearts. To examine the in vivo function of glucocorticoid signaling in the heart, we generated mice with cardiomyocyte‐specific deletion of GR (cardioGRKO), MR (cardioMRKO), or both GR and MR (cardioGRMRdKO). The cardioMRKO mice exhibited normal heart function whereas the cardioGRKO mice spontaneously developed cardiac hypertrophy and left ventricular systolic dysfunction. Surprisingly, the cardioGRMRdKO mice were protected from cardiac disease, despite having many of the same pathological gene changes that were present in the GR deficient hearts. Microarray analysis revealed a set of cardioprotective gene changes that occurred specifically in the cardioGRMRdKO hearts. Re‐installation of MR into the cardioGRMRdKO hearts by adeno‐associated virus gene delivery reversed these cardioprotective gene changes and resulted in cardiac dysfunction. These findings reveal critical gene‐regulatory roles for both GR and MR in the heart and suggest that an imbalance in these two signaling pathways leads to heart disease. Therapies designed to modulate cardiomyocyte glucocorticoid signaling to favor more GR and less MR activity may provide new approaches for treating the failing heart.Support or Funding InformationNIH