bluuebunny/crossref_metadata_2025_split huggingface dataset:
In Mice, KCNQ Channel Modulators Affect Afferent Nerve Activity and not Urinary Bladder Smooth Muscle Contractility Directly
Authors: Tykocki Nathan R. , Heppner Thomas J. , Bonev Adrian D. , Nelson Mark T. 
Published: November 2023

KCNQ channels are a family of voltage‐dependent K+ channels, which open in response to membrane depolarization to regulate cell excitability. KCNQ activators, such as retigabine, have been used successfully to treat epilepsy; interestingly, these drugs also caused urinary retention. Using ex vivo measurements of bladder pressure and afferent nerve activity, isometric contractility of bladder strips, and electrophysiological recordings from freshly isolated urinary bladder smooth muscle (UBSM) cells, we explored the role of KCNQ channels as regulators of murine urinary bladder function. In the ex vivo bladder, both the KCNQ activator retigabine and the voltage‐gated Ca2+ channel inhibitor diltiazem blocked transient contractions (TCs) and associated bursts of afferent nerve activity. In addition, retigabine reduced baseline afferent nerve activity in the absence of TCs. Simulating TCs by mechanical deformation of the bladder wall also caused bursts of afferent activity; retigabine, but not diltiazem, inhibited these afferent bursts as well. This suggests that the effects of retigabine were not limited to detrusor muscle, and also included inhibition of afferent nerve activity. In bladder strips, neither retigabine (10 μM) nor the KCNQ antagonist XE‐991 (10 μM) altered contractions in response to electrical field stimulation (0.5 – 50 Hz). Retigabine did, however, significantly reduce 60 mM KCl‐induced contractions, possibly indicative of voltage‐dependent Ca2+ channel inhibition and not KCNQ activation. Consistent with contractility data, retigabine inhibited currents through voltage‐dependent Ca2+ channels and had no effect on K+ currents, as measured in UBSM cells using the patch‐clamp technique. Lastly, retigabine had no effect on resting membrane potential in UBSM cells, using the current‐clamp configuration of the patch clamp technique. Taken together, these data indicate that UBSM cells lack functional KCNQ channels, and the effects of retigabine on detrusor contractility may be due instead to non‐specific inhibition of voltage‐gated Ca2+ channels. Alternatively, urinary retention caused by retigabine may be related to inhibition of afferent nerve activity and not diminished detrusor contractility.Support or Funding InformationSupported by NIH K01DK103840 to NRT and NIH R37DK053832 to MTN.
High fat diet induces hypertension but not vascular and cardiac remodeling in female Dahl sensitive rats.
Authors: Fernandes Roxanne , Bonilla Patricia A Perez , Garver Hannah , Galligan James J , Fink Gregory D , Xu Hui 
Published: November 2023

Mechanisms responsible for obesity associated hypertension may be different in males and females. Altered vascular reactivity to sympathetic neurotransmitters and inflammation‐induced vascular remodeling/fibrosis contribute to obesity associated hypertension in males, but recently elevated leptin and aldosterone levels have been proposed to cause obesity associated hypertension in females. We and others have found that high fat (HF) feeding significantly increases mean arterial blood pressure (MAP) in male Dahl salt sensitive (SS) rats. Here we sought to test whether HF diet increases MAP in female Dahl SS rats, and if the mechanisms responsible are similar in males and females. Female and male Dahl SS rats were fed either normal fat diet (NF, 10% kcal from fat with 0.24% NaCl, 0.4% K+) or HF diet (60% kcal from fat with 0.3% NaCl, 1% K+) for 24–26 weeks starting at weaning (3 weeks of age). At the end of the study MAP was significantly and similarly higher in HF versus NF males (n=9–10, 158±1 mmHg vs 130±1 mmHg) and HF versus NF females (n=4–5, 157±3 mmHg vs 125±7 mmHg) (P<0.05). Neither HF males nor females had larger final body weights than their respective NF groups. Hyperlipidemia and increased visceral fat were found in HF males, but not females, compared with NF rats. No differences in plasma levels of fasting glucose, insulin, leptin, aldosterone, or BUN were found between HF versus NF males or females. Male HF rats showed a greater acute depressor response to ganglion blockade than did male NF rats, but this was not seen in female rats. HF feeding did not cause alterations in mesenteric arterial (MA) contractile responses to sympathetic neurotransmitters (norepinephrine and ATP) in males or females. HF feeding significantly increased MA wall‐thickness and cardiac mass in males, but the changes were much less dramatic in females. Morphological studies did not reveal greater fibrosis in adventitia of mesenteric, intrarenal and coronary arteries from HF fed males or females. Our studies show that HF diet promotes hypertension in Dahl SS males and females without causing overt metabolic dysfunction in either sex. Females, however, are less prone to develop hypertension associated vascular and cardiac remodeling, and only males exhibit a neurogenic component to their hypertension.Support or Funding InformationNIH2P01HL070687‐11A1
Microvascular endothelial function in lean versus obese women with polycystic ovary syndrome: role of the endothelin B receptor
Authors: Usselman Charlotte W , Taylor Hugh S , Stachenfeld Nina S 
Published: November 2023

Polycystic ovary syndrome (PCOS) affects 10% of women and is associated with myriad cardiovascular risk factors, including obesity and endothelial dysfunction. PCOS is also associated with elevated levels of plasma endothelin‐1 (ET‐1), one of several circulating indicators of endothelial injury and dysfunction. Women with PCOS have reduced vasodilatory responses to ET‐1 as mediated through the endothelin receptor subtype B (ETBR). We hypothesized that impairments in ETBR‐dependent and independent vasodilation are more severe in obese than in lean women with PCOS. We tested 3 lean (age 30 ± 6 y; BMI 23 ± 1 kg/m2) and 3 obese (33 ± 5 y; 36 ± 4 kg/m2) women diagnosed with PCOS according to the Rotterdam criteria. We assessed cutaneous vascular conductance (CVC) with laser Doppler flowmetry during intradermal microdialysis perfusions of ET‐1 (1, 3, 4, 5, and 7 pmol) with either lactated Ringer's solution, ETBR inhibition (BQ‐788), or nitric oxide inhibition (L‐NMMA). Dose‐response curves (%CVCmax) as a function of the log concentration of ET‐1 demonstrated reduced vasodilatory responses in lean PCOS (logEC50 0.61 ± 0.07) relative to obese PCOS (logEC50 0.52 ± 0.47). Inhibition of ETBR decreased the ET‐1‐induced vasodilation in both lean (i.e., logEC50 0.66 ± 0.11) and obese (logEC50 0.62 ± 0.21) groups. Contrary to our hypothesis, this inhibition was two‐fold greater in obese PCOS women, indicating a greater role for ETBR‐induced vasodilation under basal conditions relative to lean PCOS. Conversely, nitric oxide inhibition had little effect in either lean (logEC50 0.61 ± 0.32) or obese PCOS women (logEC50 0.54 ± 0.18) compared to ET‐1 alone. Our data suggest that impairments to endothelial function are more severe in lean than obese women with PCOS, and that ETBR plays an important role in this potentially adverse cardiovascular outcome.Support or Funding InformationSupported by The John B. Pierce Laboratory and the American Heart Association (15POST24990003).
Sex differences in Mesenteric Arterial Function at Pre‐Diabetic and Diabetic Stage of UC Davis Type 2 Diabetic Mellitus Rats
Authors: Shaligram Sonali , Akther Farjana , Graham James L , Stanhope Kimber L , Havel Peter J , Rahimian Roshanak 
Published: November 2023

UC Davis type 2 diabetes mellitus (UCD‐T2DM) rat is a validated model of type 2 diabetes exhibiting etiology very similar to clinical T2DM seen in humans compared to other rodent models. It is characterized by polygenic adult‐onset obesity and diabetes observed in both sexes along with development of insulin resistance, impaired glucose tolerance, and subsequent β‐cell decompensation. The objectives of this study were to investigate whether 1) the mesenteric arterial function is impaired and 2) sex differences exist in the development of abnormal vascular responses in UCD‐T2DM rats at pre‐ diabetic and diabetic stage.Endothelium‐dependent vasodilation (EDV) to acetylcholine (ACh, 10−8 to 10−5 M) was measured in mesenteric arteries (MA) pre‐contracted with phenylephrine (PE, 2 μM). Concentration response curves (CRC) to sodium nitroprusside (SNP) (Nitric oxide (NO) donor, 10−9 to 10−5 M) and contractile agents such as PE (10−7 to 3×10−5 M) or endothelin‐1 (ET‐1, 10−10 to 10−7 M) were also determined.ACh‐induced relaxation was significantly impaired in MA of pre‐diabetic and diabetic rats, regardless of sex. However, in pre‐diabetic stage, the extent of impairment was greater in males compared to that of female rats. When animals become diabetic, MA from female rats demonstrated a greater suppression of EDV than that of males. Similar to ACh responses, the relaxation responses to NO donor (SNP) were also impaired in MA of pre‐diabetic and diabetic rats, in both sexes. Furthermore, female diabetic rats exhibited more impairment compared to male diabetic. In addition, arteries from female diabetic but not male diabetic showed a larger shift of SNP CRC to the right compared to their respective pre‐diabetic animals. Finally, the sensitivity of MA to contractile agents such as PE and ET‐1 was significantly enhanced in female diabetic rats compared to other groups.Our data, for the first time, show that the mesenteric arterial function of UCD‐T2DM rats is impaired in both sexes as early as pre‐diabetic stage. Moreover, we demonstrated a greater predisposition of MA to vascular injury in female diabetic rats, might be due to an altered sensitivity of arteries to contractile agents and NO.Support or Funding InformationSupported by NIH/NIHLBI
Increases in Cytochrome P450 Expression and 20‐HETE Production Reduce Mean Arterial Pressure in an Animal Model of Preeclampsia
Authors: Cotton Joshua , Wu Wenjie , Murphy Sydney 
Published: November 2023

Recent data suggests cytochrome P450 (CYP450) vasoactive eicosanoids in the pathophysiology of preeclampsia. Thus, we investigated whether alterations in CYP450 expression and 20‐HETE production contribute to maternal hypertension during pregnancy. In order to address this hypothesis, we utilized a newly characterized model of hypertension during pregnancy also documented to have a genetic deficiency in 20‐HETE production, the Dahl S rat. In addition, we utilized a CYP4A1 overexpression transgenic rat. Mean arterial pressure, measured via telemetry, and proteinuria were measured in 12 week old Spraque Dawley (SD), Dahl SSJr, and CYP4A1 Tg rats at baseline, prior to pregnancy, and gestational day (GD) 5, 12, and 19. Consistent with previous reports, a reduction in MAP at term during normal pregnancy was not seen in the Dahl S rat (1.5±6 mmHg, p<0.05; SD, 14±1 mmHg; CYP4A1 Tg, 18±2 mmHg). Proteinuria was elevated in both Dahl SSJr (107±45 mg/day, p<0.05) and CYP4A1 Tg (53±5 mg/day, p<0.05) rats relative to SD (10±1 mg/day) pregnant rats on GD19. Pup number was reduced in the Dahl S (9±1, p<0.05) and increased in the CYP4A1 Tg (11±1, ns) rat compared to the SD (12±1), with no differences noted in placenta to pup weight. CYP450 enzymatic activity, as measured by production of 20‐HETE, within the placenta and kidney were significantly increased in the placenta (0.002±0.0006 vs 0.4±0.2 pmol/min/mg protein) and kidney of CYP4A1 Tg rats (4.4±0.5 vs 1.9±0.7 pmol/min/mg protein). Therefore, alterations in CYP450 enzyme activity and 20‐HETE may contribute to the increases in MAP during pregnancy and may be a potential therapeutic target in the treatment of preeclampsia.Support or Funding Information14SDG20160020; HL121042
Expression of MEF2 transcription factors in human placenta and involvement in trophoblast invasion and differentiation
Authors: Li Lucy , Gong Xiaoming , Rubin Lewis P 
Published: November 2023

IntroductionThe important and common adverse pregnancy outcome, preeclampsia, is characterized by placental trophoblast dysfunction and insufficient (shallow) trophoblast uterine invasion/remodeling during early pregnancy. Myocyte enhancer factor 2 (MEF2) family transcription factors play an important role in cell differentiation and invasion in a variety of cell types and tissues. The objective of this study is to explore the role of MEF2s in trophoblast differentiation and uterine invasion in normal placental development and preeclampsia.MethodsHuman cytotrophoblasts (CTBs) were isolated from term placentas by mechanical and enzymatic digestion and purified using Percoll gradient centrifugation and differential adhesion. Highly purified (>95%) CTBs in acute cultures (1–4 d) were used to analyze expression of MEF2 isoforms and MEF2‐dependent cell signaling pathways. The effects of MEF2 on human trophoblast invasion and differentiation and cellular pathways were examined principally in two relevant human trophoblast cell lines: HTR8/SVneo (first trimester human extravillous trophoblast phenotype) and BeWo (choriocarcinoma CTB‐like) cells. Invasion/migration was evaluated by Transwell (including chemotactic) invasion and “scratch” migration 2D assays; differentiation was assessed by trophoblast syncytial formation and expression of trophoblast differentiation genes.ResultsMEF2 isoforms are differentially expressed in first and third trimester placental trophoblast. In particular, MEF2D and 2B are highly expressed during first trimester placental development. In other experiments, transient overexpression of MEF2D in HTR8/SVneo cells promotes trophoblast invasion and migration. This invasive property is blocked by exogenous expression of a dominant negative MEF2. In contrast, MEF2A is the major isoform expressed in term trophoblast. MEF2A expression is associated with CTB differentiation into syncytiotrophoblast; transient overexpression of MEF2A enhances cytodifferentiation/syncytia formation. Based on p38 MAPK null experimental animals, p38 is the principal upstream regulator of MEF2. During in vitro differentiation of primary human CTB cultures, we found the course of p38 activity (phosphorylation) parallels MEF2A expression. Placental tissues obtained from pre‐eclamptic and normal (non‐pre‐eclamptic) pregnancies exhibit differences in MEF2 isoform expression.ConclusionThese findings suggest individual MEF2 family members differentially regulate trophoblast invasion and differentiation. Dysregulation of MEF2 expression or signaling in early pregnancy may be associated with placenta‐related pregnancy disorders, including preeclampsia.Support or Funding InformationSeed grant from Laura W. Bush Institute for Women's Health, Texas Tech University Health Sciences Center
NMDA receptor‐mediated synaptic transmission in the spinal dorsal horn and spinal nerve ligation‐induced nociceptive behavior are enhanced in serine racemase knockout mice
Authors: Fukushima Teruyuki , Kato Eiko , Konno Ryuuichi , Hori Yuuichi 
Published: November 2023

N‐methyl‐D‐aspartate (NMDA) receptors are reported to play an important role in synaptic plasticity that may underlie not only learning and memory but also persistent pain. Peripheral nerve injury or tissue inflammation often induces an abnormal pain state, so‐called neuropathic pain. The activation of NMDA receptors mediates the development and maintenance of neuropathic pain. D‐serine co‐localizes with NMDA receptors to a high degree and acts as an endogenous coagonist of the NMDA receptor “glycine site”. D‐serine is synthesized from L‐serine by serine racemase (SR), an enzyme that is abundant in the central nervous system (CNS). To elucidate the contribution of SR to synaptic transmission in the CNS, we generated SR knockout (KO) mice.Animal experimental procedures were reviewed and approved by the institutional animal care and use committees at Dokkyo Medical University. The care and use of the animals were in accordance with the guidelines of the International Association for the Study of Pain. In the present study, we investigated NMDA receptor‐mediated synaptic transmission in the superficial dorsal horn (SDH) of spinal cord slices by means of tight‐seal whole‐cell recordings in SR‐KO mice and wild type (WT) mice. We also analyzed nociceptive behavior, which is mediated by NMDA receptors in the SDH, induced by spinal nerve ligation (SNL). The amplitude of the NMDA receptor‐mediated component of excitatory postsynaptic currents (EPSCs), normalized to that of the non‐NMDA receptor‐mediated component of EPSCs, was smaller in SR‐KO mice than in WT mice. In contrast, the time decay course and the synaptic charge transfer of the NMDA component were slower and larger in SR‐KO mice than in WT mice, respectively. Mechanical allodynia induced by the SNL was augmented in SR‐KO mice compared to WT mice. It is speculated that SR‐KO mice have a different NMDA receptor subunit composition compared to WT mice, which might explain the differences in synaptic transmission and nociceptive behaviors between the SR‐KO mice and WT mice. Real‐time RT‐PCR analysis indicated that the expression level of NR2B subunit of NMDA receptors was slightly but distinctly increased in SR‐KO mice.Finally, the present observations could suggest the possibility that the SR activity modulates the neuropathic nociception by changing the subunit composition of NMDA receptors. SR‐KO mice may provide a unique tool to investigate the synaptic action of D‐serine in the CNS.
Inactivation of the PLA2 Activity of Prdx6 Ameliorates Sepsis‐Induced Acute Lung Injury
Authors: Vazquez‐Medina Jose Pablo , Tao Jian‐Qin , Patel Priyal , Dodia Chandra , Sorokina Elena , Feinstein Sheldon I , Chatterjee Shampa , Fisher Aron B 
Published: November 2023

Prdx6 is a multi‐functional protein that expresses peroxidase, phospholipase A2 (PLA2) and LPC acyltransferase activities. While the three activities of Prdx6 contribute to either the scavenging of lipid hydroperoxides or to the repair of peroxidized membranes, the PLA2 activity of Prdx6 (aiPLA2) is also required for NOX2 activation in pulmonary microvascular cells (PMVECs), alveolar macrophages and polymorphonuclear neutrophils. During sepsis‐induced acute lung injury (ALI), NOX2 is activated generating superoxide radicals that contribute to vascular inflammation. To test whether inactivation of aiPLA2 ameliorates sepsis‐induced ALI, we used a knock‐in mouse model that lacks aiPLA2 activity due to a mutation of a key member of the PLA2 catalytic triad (Prdx6‐D140A). Prdx6‐D140A mice treated IP with LPS (10 mg/Kg) exhibited increased survival rates compared to WT mice (75% vs 25% 4 days after LPS). Total nucleated cells in BALF, lung VCAM‐1 expression, and lung permeability increased significantly (p<0.05) after 24 h treatment with LPS in WT but not in Prdx6‐D140A mice. In PMVECs, LPS treatment (1 μg/mL for 8h) increased Prdx6 phosphorylation and aiPLA2 activity by two‐fold (p<0.05). Furthermore, phosphorylated Prdx6 was localized at the plasma membrane after LPS treatment. LPS also increased extracellular hydrogen peroxide generation (Amplex Red oxidation) by 70% in WT but not in NOX2 null or Prdx6‐D140A PMVECs (p<0.05). Similarly, LPS increased intracellular hydrogen peroxide generation by 90% in WT PMVECs stably‐expressing the pHyPer‐Cyto sensor. Increased intracellular hydrogen peroxide generation was prevented by pre‐treatment with MJ33, a pharmacological inhibitor of aiPLA2. These results demonstrate that aiPLA2 is needed for LPS‐induced, NOX2‐driven oxidant generation in PMVECs. Results also show that genetic inactivation of aiPLA2 reduces LPS‐induced vascular inflammation, increased vascular permeability, and mortality in an animal model. Furthermore, these studies suggest that aiPLA2 could be a target for prevention or treatment of sepsis‐induced ALI.Support or Funding InformationResearch funded by NHLBI L105509 and HL102016.
Acute High Fat Diet Exposure Results in Discrete Shifts in the Adipose Tissue Inflammation Profile in Mice
Authors: Hetrick Byron , Huffard Adrian , McCurdy Carrie 
Published: November 2023

Obese adipose tissue is characterized by chronic, low grade inflammation that is currently recognized to play a direct role in adipocyte insulin resistance. Lean adipose tissue is characterized by a variety of leukocytes that maintain tissue homeostasis, for example, anti‐inflammatory M2 macrophages which are supported by IL‐4 secretion by Eosinophils. A pro‐inflammatory set of leukocytes, for example, M1 macrophages, neutrophils, and CD8+ T‐cells are elevated in obese adipose tissue. While the inflammation profile of lean and obese adipose tissue has been characterized to a large extent, little is known about the temporal changes and relative importance of inflammatory factors during the transition from lean to obese state. We have characterized changes to the inflammatory phenotype of mouse epididymal and subcutaneous adipose depots upon exposure to an acute (3 day) high fat diet in order to understand early changes to adipose tissue inflammation that may contribute to insulin resistance prior to the full onset of obesity. C57BL/6J mice were fed either a high fat diet (45% calories from fat) or control diet. After three days, the mice were euthanized and the epidiymal and subcutaneous adipose tissue was removed, and digested into a cell suspension. The stromal vascular fraction was isolated by differential centrifugation, and the leukocyte content of the tissue was measured by multi‐color flow cytometry. We observed a decrease in anti‐inflammatory leukocytes, such as eosinophils and regulatory T‐cells in the epididymal fat depot. While we did not see an increase in the amount of M1 polarized macrophages, the number of TNFα expressing macrophages was increased, which may indicate early stages of a polarization shift in the macrophage subset from M2 to M1. We also observed an increase in B‐cells and Natural Killer T‐cells indicating they play a role in the initiation of a high fat diet induced pro‐inflammatory phenotype. Surprisingly, we saw very different effects in the subcutaneous adipose depot. Eosinophils and IL‐10 expressing macrophages were increased upon high fat diet exposure indicating that this fat depot responds to prevent or delay the pro‐inflammatory phenotype associated with insulin resistance. In summary, only a subset of inflammatory factors are affected by acute exposure to high fat diet, indicating a complex network of signaling orchestrates the transition of adipose tissue from a lean state to a pro‐inflammatory, obese state. From our observations, we predict that high fat diet exposure initially causes a reduction in anti‐inflammatory factors, causing proliferation, infiltration, or polarization of pro‐inflammatory factors that block the insulin signal transduction cascade in adipocytes, resulting in insulin resistance.Support or Funding InformationNIH R01 DK095926