bluuebunny/crossref_metadata_2025_split huggingface dataset:
IL1β Disrupts Barrier Integrity and Elicits Formation of Focal Adhesions in Lymphatic Endothelial Cells
Authors: Adderley Shaquria P , Zhang Xun E , Breslin Jerome W 
Published: November 2023

A functional role of changes in lymphatic vessel permeability for the optimization of immune and inflammatory responses has recently been reported. The precise mechanisms within lymphatic endothelial cells that promote changes in barrier integrity remain incompletely understood. In the current study we investigated the effects of two different inflammatory mediators on lymphatic endothelial barrier function: histamine, which produces local inflammatory responses; and IL1β, an inflammatory cytokine commonly elevated in a variety of diseases. Human adult lymphatic endothelial cell (HALEC) and human juvenile lymphatic lymphatic endothelial cell (HJLEC) monolayers were used in order to examine endothelial‐specific mechanisms. Transendothelial electrical resistance (TER) was used as an index of barrier function. Immunofluorescence microscopy was used to examine localization of vinculin and VE‐cadherin. The roles of MLCK and p38 MAPK kinase, both previously reported to be involved in inflammation‐induced elevations in microvascular permeability, were tested using the inhibitors ML7 (1 μM) and SB203580 (10 μM), respectively. The results showed that histamine (10 μM) produces a rapid decrease in resistance, which recovers within 5 mins, whereas IL1β (25 ng/ml) causes an initial increase in resistance followed by a decrease starting at 3 hours and continuing until approximately 4 hours after treatment. The recovery of the IL1β‐induced decrease in barrier function begins approximately 7 hours into the time course. Treatment with IL1β or histamine also increased the number of focal adhesions containing vinculin, 5.5 h after treatment. Surprisingly, inhibition of p38 MAP kinase with SB203580 potentiated the response of IL1β. Inhibition of MLCK with ML7 had no effect on the IL1β response. The data suggest that the effect of IL1β on endothelial barrier function has different dynamics than that of histamine. However, in both cases the cells appear to be responding to stress, with elevated focal adhesions 5.5 h after treatment. The p38 MAPK kinase appears to have a protective role for barrier function in lymphatic endothelium.Support or Funding InformationSupported by NIH grant R01HL098215.
Disruption of interleukin 12 stimulates and enhances angiogenesis in type 2 diabetes
Authors: Ali Maha Ali Essameldeen , Mali Vishal , Haddox Samuel , Abdelghani Soad , Eldeek Sahar , Abulfadl Atif , Belmadani Souad 
Published: November 2023

IntroductionPrevious studies reported the involvement of interleukin (IL)‐12 in cardiovascular complications in type 2 diabetes. We previously showed that ischemia‐induced angiogenesis was compromised in type 2 diabetic mice. In this study, we determined whether IL‐12 disruption would rescue angiogenesis in type 2 diabetic mice.MethodsType 2 diabetes was induced by feeding control and IL‐12 deficient mice with high fat diet (60% high fat diet “HFD”) for 12 weeks. Body weight, glucose test tolerance and insulin test tolerance were assessed. After 12 weeks of HFD, femoral artery was ligated and then blood flow recovery was assessed once a week for 4 weeks.ResultsControl (C57/Bl6) and IL12−/− mice fed with HFD became markedly obese after 12 weeks and exhibit glucose intolerance and insulin resistance. Using laser Doppler technique, data revealed that blood flow recovery was fully restored in IL12 −/− and in C57/Bl6 mice without HFD 3 weeks after femoral ligation. After 12 weeks of HFD, blood flow recovery was compromised and only reached 50% in control mice. However, in IL‐12−/− fed with HFD, the blood flow recovery was fully restored and reached 100% indicating that the disruption of IL‐12 in type 2 diabetes protects angiogenesis in response to ischemia. The recovery was associated with an enhancement in eNOS/Akt/VEGFR2 phosphorylation, and the reduction in oxidative stress.ConclusionThe disruption of IL‐12 increases angiogenesis and blood flow recovery after hind limb femoral ligation in obese type 2 diabetic mice independently of obesity and type 2 diabetes.Support or Funding InformationEmbassy of Arab Republic of Egypt Culture and Educational bureau
Smooth Muscle Specific Expression of a Dominant Negative Cullin 3 Mutant (Cul3Δ9) Causes Vascular Dysfunction in Mice Mediated by RhoA/Rho‐kinase
Authors: Agbor Larry N , Ibeawuchi Stella‐Rita C , Hu Chunyan , Quelle Frederick W , Sigmund Curt D 
Published: November 2023

Dominant de novo mutations in Cullin 3 (Cul3) resulting in deletion of exon 9 (Cul3Δ9) cause severe early onset human hypertension mediated in part by impairing the degradation of WNK4 in the kidneys. However, the extra renal mechanisms remain uninvestigated. Previously, we showed that knockout of Cul3 in HEK293T cells induced by clustered regularly interspersed short palindromic repeats (CRISPR)‐Cas9 resulted in accumulation of the Cul3 substrate RhoA. We hypothesized that smooth muscle (SMC) expression of Cul3Δ9 protein in mice impairs endogenous Cul3 wildtype (WT) function and causes vascular dysfunction via elevated RhoA/Rho‐kinase signaling. We generated a novel transgenic mouse model inducibly expressing Cul3Δ9 protein specifically in smooth muscle (termed S‐Cul3Δ9) and assessed vascular responses in the cerebral basilar artery and aorta using a pressurized and a wire myograph respectively. Basilar artery from S‐Cul3Δ9 transgenic mice exhibited significantly impaired relaxation responses to acetylcholine (ACh) (at 100 μmol/L: 15±4% S‐Cul3Δ9 vs 65±5% WT, p<0.0001) and to the nitric oxide donor sodium nitroprusside (SNP) (at 100 μmol/L: 59±2% S‐Cul3Δ9 vs 90±5% WT, p<0.05). Vasoconstriction responses to angiotensin II (Ang II) and to phenylephrine (PE) were significantly elevated in S‐Cul3Δ9 transgenic mice compared to WT. Consistent with data from basilar artery, aorta from S‐Cul3Δ9 transgenic mice exhibited impaired ACh‐mediated relaxation (at 100 μmol/L: 55±2% S‐Cul3Δ9 vs 71±7% WT, p<0.0001). RhoA protein expression was significant elevated in aorta of S‐Cul3Δ9 transgenic mice (1.6±0.2 S‐Cul3Δ9 vs 1.0±0.1 WT, P<0.05). Interestingly, incubation with the Rho‐kinase inhibitor (Y27632) completely restores ACh relaxation responses to WT levels in both the basilar artery and aorta of S‐Cul3Δ9 mice. Taken together, these findings suggest a novel vascular specific mechanism whereby Cul3Δ9 protein interferes with endogenous Cul3 and impairs the degradation of Cul3 substrates such as RhoA, contributing, at least in part, to vascular dysfunction in mice.Support or Funding InformationThis work was supported by National Institutes of Health Grants HL048058, HL062984, and HL084207 (to C. D. S.).
Impaired myogenic response of MCA elevates transmission of pressure to penetrating arterioles and contributes to cerebral vascular disease in aging hypertensive FHH rats
Authors: Fan Fan , Pabbidi Mallikarjuna , Lin Rick C.S. , Ge Ying , Gomez‐Sanchez Elise P , Rajkowska Grazyna K , Moulana Mohadetheh , Gonzalez‐fernandez Ezekiel , Sims Jerrell , Elliott Matthew R , Paul Ian A , Alexander Alexander P , Mosley Thomas H , Harder David R , Roman Richard J 
Published: November 2023

We recently identified a K572Q mutation in Add3 in FHH rats. The present study examined whether it contributes to cerebral vascular dysfunction in aging and after the development of hypertension. The expression of Add3 is lower in FHH than in FHH.1BN rats. The inner diameter of MCA decreased by 80 ± 2 % and 71 ± 4% in FHH.1BN and FHH.Add3 transgenic rats, respectively, when perfusion pressure was increased from 40 to 140 mmHg. The pressure in terminal pial arteries was greatly increased from 37 ± 1 to 63 ± 4 mmHg in FHH rats when MAP was increased from 100 to 160 mmHg but was unchanged (34 ± 1 to 36 ± 1 mmHg) in FHH.1BN rats. The inner diameters of capillaries in the deep cortex were markedly larger (8 ± 2 vs. 22 ± 1 μm) and capillary density was reduced (15 ± 0.4% vs. 8 ± 2%) in FHH than in FHH.1BN rats. The expression of phosphorylated‐ERK1/2 and phosphorylated‐PKC was reduced in the brain of FHH than FHH.1BN rats. CBF rose by 99 ± 7%, 64 ± 5% and 42 ± 4% in FHH, FHH.1BN and FHH.Add3 rats, respectively, when MAP was increased from 100 to 190 mmHg, demonstrating that the impaired autoregulation of CBF in FHH rats was rescued by introduction of wildtype Add3. BBB leakage was greater in FHH rats than FHH.1BN and FHH.Add3 rats after induction of DOCA/salt hypertension. Hypertensive FHH rats exhibited marked neurodegeneration and vascular remodeling of the neocortex and hippocampus with a 33% reduction in the density and size of neurons relative to FHH.1BN and FHH.Add3 rats. Gap junctions between endothelial cells were larger in hypertensive FHH rats with neuronal, mitochondrial and myelin degeneration in the vacuolated area near leaky capillaries. The hypertensive FHH rats took 2.5 times longer to escape from an eight‐arm water maze than hypertensive FHH.1BN rats suggesting spatial learning and memory dysfunction. These findings suggest that the K572Q mutation in FHH rats decreases the expression and activity of Add3 which is involved in phosphorylation of PKC and ERK1/2. This contributes to the impaired myogenic response and autoregulation of CBF, therefore elevating the transmission of pressure in penetrating arterioles that ultimately promotes loss of cerebral capillaries, BBB leakage and neurodegeneration, which may contribute to vascular cognitive impairments with hypertension and age.Support or Funding InformationThis study was supported by grants HL36279 and DK104184 (RJR) from the National Institutes of Health.
Influence of Acute Lower Limb Heating on Clinical Measures and Functional Capacity in Peripheral Artery Disease
Authors: Pellinger Thomas K. , Pearce Catherine B. , Simmons Grant H. 
Published: November 2023

Acute local heating of human limbs produces increases in conduit artery blood flow and shear stress, and these increases are comparable in magnitude to those achieved during both whole body and single limb exercise. Numerous studies have shown that acute exposure to periods of high shear stress; whether by exercise or heating, results in increased conduit artery flow‐mediated vasodilation immediately following exposure. However, it is unknown if these transient increases in conduit artery vascular function are sufficient to improve the matching of oxygen supply and demand distal to the site of stenosis in patients with peripheral artery disease (PAD). Thus, the purpose of this study was to test the hypothesis that a single bout of lower limb heating (for 15 or 45 minutes) increases resting popliteal artery blood flow and 6 minute walk distance (6MWD) in patients with PAD. One 71 year old male patient with PAD (Intermittent Claudication, Fontaine stage II) attended the laboratory for a screening visit and two experimental visits during which bilateral lower limb heating was carried out via water bath immersion (42C; 40 cm depth) for either 15 or 45 minutes. Clinical and cardiovascular measures including resting single leg popliteal artery blood flow (Doppler ultrasound), ABI, and toe oxygen saturation (pulse oximetry) were carried out prior to and beginning 10 minutes following limb heating. Popliteal vascular conductance (PVC) was calculated as flow/mean arterial pressure. 6MWD and time/distance to pain onset were measured on the screening day and each experimental day 45 minutes post‐leg immersion. The 15 minute leg heating trial increased popliteal artery blood flow from 113.7 to 169.1 mL min−1 (48.6%), PVC from 1.41 to 2.15 mL min−1 mmHg−1 (52.4%), and 6MWD from 660 to 800 feet (21.2%). The 45 minute leg heating trial increased popliteal artery blood flow from 91.8 to 159.3 mL min−1 (73.5%), PVC from 1.17 to 1.99 mL min−1 mmHg−1 (70.6%), and 6MWD from 660 to 770 feet (16.7%). Furthermore, the subject reported onset of leg pain at 2:45 and 325 feet of the screening visit 6MW, which was delayed until 3:00 and 500 feet after the 15 minute leg heating trial and until 4:00 and 550 feet after the 45 minute leg heating trial. Neither of the heating trials evoked changes in ABI or toe oxygen saturation. Although data collection is ongoing, these initial findings suggest that acute lower limb heating in patients with PAD may increase exercise tolerance which, if undertaken routinely, could increase either patient adherence to or efficacy of exercise prescription in this population.Support or Funding InformationSupported by the Salisbury University Faculty Mini‐Grant
Endothelial calreticulin and its role in vascular function
Authors: Biwer Lauren , Good Miranda , Sonkusare Swapnil , Isakson Brant 
Published: November 2023

Myoendothelial junctions (MEJ) are projections from endothelial cells (EC) that make contact with neighboring smooth muscle cells (SMC) and are the anatomical hallmarks of resistance arteries. MEJ also exhibit presence of endoplasmic reticulum (ER), as evidenced from the ER retention signal –KDEL immunostaining and the live ER tracker (glibenclamide) in holes of the internal elastic lamina. Within the ER, calreticulin is a calcium binding protein that can bind approximately 50% of ER calcium and we found it was significantly enriched in the non lipid‐raft fraction of the MEJ in vitro. Due to the importance of ER calcium signals at the MEJ in controlling vascular tone, we investigated the in vivo role of calreticulin in the endothelium and the MEJ. We generated a tamoxifen inducible, endothelial specific calreticulin floxed mouse. To confirm knockdown of calreticulin in the EC, we digested diaphragm skeletal muscle to obtain microvascular EC from mice injected with peanut oil or tamoxifen. Live cells were stained with CD31 antibody (to mark EC), CD45 (to exclude leukocytes), fixed and permeabilized, and stained with calreticulin. We saw a significant decrease in the percentage of CD31+, Calreticulin+ cells after tamoxifen injection, indicating less EC were expressing calreticulin in our knockout mouse. In calreticulin mice with and without tamoxifen, we could not identify any basal change in blood pressure as measured by radiotelemetry. However, after acute injection of 0.1mg/kg PE, the rapid and significant increase in blood pressure seen in control mice was significantly blunted in mice lacking EC calreticulin. Because of this, we used pressure myography on isolated resistance arteries from mice (both third order mesenteric arteries and thoracodorsal arteries) lacking EC calreticulin and found them to have less constriction to PE dose response curves. In order to determine if this blunted response is due to less agonist‐induced ER calcium release, we also measured calcium transients at MEJs of intact resistance arteries. In summary, calreticulin in the EC appears to play an important role in the regulation of vascular tone and blood pressure, possibly through its actions at the ER in MEJs. We hypothesize calreticulin's presence at the MEJ may be to mediate ER calcium release and provide the negative feedback that occurs upon SMC stimulation with PE.Support or Funding InformationAmerican Heart AssociationNational Institute of Health
Effects of arginine vasopressin (AVP) deficiency on long term arterial blood pressure in normal and spontaneously hypertensive rats (SHR)
Authors: Villanueva‐Rodriguez Gloria Marcela , Organista‐Esparza Alejandro , Biñuela‐Berni Veronica , Herrera‐Carreon Paulina , Tinajero‐Ruelas Manuel , Rivera‐Cerecero Claudia Veronica , Chavira‐Ramirez Roberto Veronica , Kovacs Kalman , Quintanar‐Stephano Andrés 
Published: November 2023

The role of AVP controlling the arterial blood pressure (BP) in basal conditions remains unclear. Previously, we described that neurointermediate pituitary lobectomy (NIL) in rats, induced an early but transient diabetes insipidus and permanent low mean BP (MBP), without changes in aldosterone, Na and K serum levels (1). In order to obtain a deeper insight in the temporal relationship between NIL and MBP, here we evaluated the effects of NIL on AVP and oxytocin (OT) serum levels and MBP from time 0 to 6 hours and at 3, 15, 45 and 90 days post NIL. Water intake and urine output, urine sodium and potassium concentrations and sodium and potassium serum levels, were measured at these same times, whereas the renin‐angiotensin II‐aldosterone system (RAAS) was assessed at 3 and 45 days post NIL. All values were compared with the respective sham operated (SHAM) groups. Values are expressed as mean ± SEM.ResultsNIL caused significant and permanent low AVP and OT serum levels (10.6 ± 0.8 versus (vs) 2.4 ± 0.16 (pg/ml), SHAM vs NIL respectively), transient diabetes insipidus of 15–30 days of duration and a significant and permanent decrease of the MBP in 70% of the NIL animals (106.8 ± 3.15 vs 73.8 ± 3.2 mmHg respectively). Hypotension was apparent 30 minutes after NIL. Indirect systolic blood pressure (SBP) (cuff tail) were measured in SHR and NIL‐SHR rats at 15 and 3 days before and 8, 20, 35 and 45 days after surgeries. In all times SBP was significantly decrease in NIL animals. The last measured was 166.5 ± 16.8 vs 121.4 ± 12.16 mmHg, SHR vs NIL‐SHR). As in previously studies, no significant differences in the RAAS and serum electrolytes were evident, whereas urine electrolytes were lower in NIL rats (Na; 33.08 ± 6.177 vs 58.99 ± 15.061 and K; 67.148 ± 12.515 vs 126.9 ± 26.97 respectively). We concluded that: 1) the permanent deficiency of AVP (NIL animals) is responsible of the long‐term low BP levels. 2) At the present we suppose that in NIL animals, the normal Na and K serum levels are due to the remnant basal concentrations of AVP (2.4 ± 0.16) which well may be enough to maintain the hydro‐electrolytic balance in the body fluids. 3) The decreased Na and K levels in the urine of the NIL animals may be explained partially for the kidney adaptations of the several segments of the nephron to the low levels of AVP. 4) Further studies are needed to explain why 30% of the NIL animals did not develop hypotension, and to explain how heart, blood vessels and kidney mechanisms adapt to these new physiological conditions.Support or Funding InformationSupported by UAA‐PIFF 14‐1 and CONACYT‐221262 grants (AQS) and CONACYT (GMV) PhD scholarship, México, and the Jarislowsky and Lloyd Carr‐Harris Foundations.
Detrimental Changes in Perivascular Adipose Gene Expression in Metabolic Syndrome
Authors: Seldomridge Ashlee , DeVallance Evan , Lemaster Kent , Branyan Kayla , Frisbee Jefferson , Setola Vincent , Chantler Paul 
Published: November 2023

Metabolic syndrome (MetS) is a clustering of major risk factors that increases the risk of cardiovascular mortality. Perivascular adipose tissue (PVAT) is an important regulator of vascular function through release of adipocytokines. Although PVAT is typically beneficial in a healthy individual, an obese individual exhibits greater production of pro‐inflammatory adipocytokines. PVAT has the capability of altering vascular tone through release of nitric oxide (NO) by endothelium nitric oxide synthase (eNOS). Tetrahydrobiopterin (BH4) is a critical cofactor for eNOS, oxidative stress can reduce BH4 production catalyzed by GTP‐cyclohydrolase‐1 (Gch1). This leads to uncoupled eNOS, which can produce superoxide adding to the oxidative burden. The increased oxidative load is additionally problematical as decrease in the antioxidant system is believed to accompany MetS. These suspected alterations in MetS PVAT may further contribute to diminish NO production from endothelium.PURPOSETo demonstrate MetS disrupts eNOS and Gch1 expression and therefore decreases PVAT derived NO production.METHODSPVAT tissue surrounding the aortic was removed from 17‐week old male obese Zucker (model of MetS) rats and their lean controls (LZR). Levels of eNOS, Gch1, and SOD‐1 expression were measured by qPCR in the PVAT. NO bioavailability was directly assessed using a DAF‐FM diacetate assay of PVAT exudate.RESULTSOZR PVAT had reduced NO production by 36% (p≤0.01). Further, OZR caused an up‐regulation of eNOS, specifically a 1.8 fold increase in OZR vs. LZR (p≤0.001). However, Gch1 expression decreased 1.6 fold (p≤0.001) in OZR vs. LZR. SOD‐1 expression showed only a slight decrease in OZR.CONCLUSIONSThe PVAT is ill equipped to deal with the increased oxidative load of MetS. Consequently, despite increased eNOS expression in MetS, NO levels are decreased, likely due to a decreased Gch1 expression, and less production of BH4. The possible uncoupling of eNOS may further increase the oxidative load. Not only did these changes reduce PVAT derived NO but also reduced endothelium release of NO. Thus alteration to PVAT in MetS greatly reduces the global bioavailability of NO, which likely impairs aortic reactivity.Support or Funding InformationNIHAHA
Glutaminase Stimulates the Proliferation of Human Endothelial Cells
Authors: Durante William , Liu Xiao‐ming , Yates Benjamin , Yu Yajie , Peyton Kelly J. 
Published: November 2023

Glutaminase (GLS) is a mitochondrial enzyme that metabolizes glutamine to glutamate and ammonia. Two distinct isoforms of GLS, GLS1 and GLS2, have been identified in mammals that exhibit distinct tissue distribution, enzyme kinetics, structural properties and molecular regulation. Although abundant GLS activity has been reported in endothelial cells, its functional role remains largely unknown. In the present study, we characterized the expression of GLS in endothelial cells and investigated the effect of GLS on the proliferation of endothelial cells. Human umbilical vein endothelial cells (HUVEC) expressed high levels of GLS1, but GLS2 was not detected in these cells. Treatment of HUVEC with the non‐selective GLS inhibitor 6‐diazo‐5‐oxo‐L‐ornithine (DON) or the selective GLS1 inhibitor bis‐2‐(5‐phenylacetamido‐1,3,4‐thiadiazol‐2‐yl)ethyl sulfide (BPTES) resulted in a concentration‐dependent inhibition of endothelial cell growth. Both GLS inhibitors also blocked HUVEC DNA synthesis and this was paralleled by a significant decrease in the phosphorylation of retinoblastoma protein. In addition, knockdown of GLS1 expression using a siRNA approach inhibited the mitogenic response of HUVEC. Flow cytometry experiments revealed that DON or BPTES arrested HUVEC in the G0/G1 phase of the cell cycle, as demonstrated by an increase in the percentage of cells in G0/G1 with a corresponding decline in the portion of cells in S and G2/M phases. Cell cycle arrest by the GLS inhibitors was associated with a dramatic reduction in cyclin A expression without any change in the expression of the other G1 cyclins, cyclin D1 and E, the cyclin‐dependent kinase inhibitors, p21 or p27, or p53. In conclusion, this study demonstrates that endothelial cells selectively express GLS1, and that GLS1 plays a fundamental role in stimulating human endothelial cell cyclin A expression, cell cycle progression, and proliferation.Support or Funding InformationSupported by American Heart Association Grants 15GRNT25250015.