bluuebunny/crossref_metadata_2025 huggingface dataset:
L‐2‐oxothiazolidine‐4‐carboxylic acid: a novel antioxidant agent that protects cadmium‐induced thromboembolic events in mice (671.2)
Authors: Fahim Mohamed , Nemmar Abderrahim , Shafiullah Mohamed , Javed Mohamed , Hasan Mohamed
Published: June 2021

Cadmium (Cd) has been demonstrated to induce brain edema and hemorrhage, and blood‐brain barrier disruption. Cd may contribute towards endothelial damage either directly by interacting with the endothelium or by inducing the generation of reactive oxygen species in endothelial and inflammatory cells. However, the underlying mechanisms are not fully understood. Therefore, the aim of this study is to examine the influence of Cd exposure on the in vivo platelet aggregation, and to assess the possible protective effects of L‐2‐Oxothiazolidine‐4‐Carboxylic acid (OTC) against cerebral microvessels injury induced by Cd in mice. Mice received cadmium chloride at a dose of 1 mg Cd/kg/day, i.p., for 3 weeks. OTC pretreatment (80mg/kg/day, i.p.) was started one hour before Cd administration. Cd compared with control significantly reduced both the time(in seconds) for the first aggregate(48.00+3.74 vs 79.33+20.73) and the time until flow stops(152+17.24 vs 179.00+13.62). OTC either by itself or concurrently with Cd significantly increased both the time for the first aggregate(156.67+10.98 vs 135.33+13.60) and the time until flow stops(239.00+20.22 vs 201.17+2.93). OTC, through its antioxidant and anti‐inflammatory actions, appeared to effectively protect Cd induced thrombosis in mice. Hence, OTC represents a potential candidate to protect the cerebral microvessels from the detrimental effect of Cd toxicity.
The effect of EPA and DHA on platelets treated with multiple agonists found in the thrombus core (671.3)
Authors: McCue Betsy , Shaull Lance , Kollasch Anna , Ye Shaojing , Whiteheart Sidney , Larson Mark
Published: June 2021

The omega‐3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been consistently associated with reduced platelet function, and more recently, with impaired thrombin generation. However, almost all studies to date have focused on how EPA and DHA alter platelet physiology in response to monotypic agonist stimulation. This reductionist approach does not recapitulate the complex chemical environment found in a developing thrombus. Therefore, we asked if platelets treated with multiple agonists like those found in the thrombus core would also display EPA and DHA‐mediated inhibition. We found that optical aggregometry and dense granule secretion was diminished after EPA and DHA incorporation in platelets stimulated with the core clot agonists. We also noted a reduction in both the protein content of platelet lipid rafts and thrombin generation after core agonist stimulation, consistent with our earlier observations. Transmission electron micrographic imaging also established that EPA and DHA‐treated platelets showed moderate ultrastructural malformations after core agonist treatment. These results all suggest that EPA and DHA can abrogate platelet signaling responses within the core of a developing thrombus, further illustrating the potential health benefits of increased EPA and DHA intake.

Grant Funding Source: Supported by an Institutional Development Award (IDeA) from the NIH/NIGMS, #P20GM103443
Increase in cerebrovascular permeability leads to fibrinogen‐cellular prion protein complex formation after traumatic brain injury (672.1)
Authors: Muradashvili Nino , Tyagi Reeta , Benton Richard , Tyagi Suresh , Lominadze David
Published: June 2021

Traumatic brain injury (TBI) is the most common cause of death and disability. It is accompanied with development of inflammation and thus linked to increased content of inflammatory mediators including fibrinogen (Fg). It is known that TBI is associated with loss of memory that can be affected by the level of cellular prion protein (PrPC).

We tested the hypothesis that TBI‐induced an increase in cerebrovascular permeability leads to Fg exstravasation in pericontusional area through activation of MMP‐9. Deposited Fg contributes to formation of Fg‐PrPC complex and can be related to loss of short‐term memory.

Pial venular leakage was observed by intravital fluorescence microscopy in male wild‐type (WT, C57BL/6J) and MMP‐9 gene knockout (MMP9‐/‐) mice after TBI. Fg deposition and formation of Fg‐PrPC complex were assessed by immunohistochemistry. Short‐term memory of mice was estimated by a novel object recognition test.

Cerebrovascular leakage, Fg deposition, and it’s co‐localization with PrPC were increased after TBI in each experimental group compared to those in respective sham‐operated mice. However, they were greater in WT than in MMP9‐/‐ mice.

Thus, our data suggest that TBI‐induced increased cerebrovascular permeability and the resultant Fg deposition leads to Fg‐PrPC complex formation. The latter can be involved in loss of memory. These results indicate a novel role of Fg in post‐TBI pathology.

Grant Funding Source: NIH P30 GM103507 and NS084823
Atrial natriuretic peptide attenuates thrombin‐induced inflammation (672.4)
Authors: Curry Fitz‐Roy , Jiang Yan , Kim Min‐Ho , Clark Joyce , Adamson Roger , Simon Scott
Published: June 2021

One microcirculatory homeostatic action of atrial natriuretic peptide (ANP) in the absence of inflammation is to increase barrier permeability to plasma proteins (Ps) when plasma volume is increased. However several investigations, particularly in cultured endothelial cells, suggest that high concentrations of ANP may also attenuate Ps. We tested the hypothesis that this action of ANP was characteristic of endothelium expressing a pro‐inflammatory phenotype. In individually perfused rat mesentery microvessels with no prior exposure to inflammation, pretreatment for 30 min with ANP (100 ng/ml) did not attenuate the increased permeability after exposure to bradykinin (10 nM). However in mouse skin where thrombin increased Ps 24 hours after an aseptic wound (AJP Heart, 296: 848, 2009), ANP added to the wound (50 ng/min/kg bw) significantly attenuated the thrombin induced transvascular albumin. At the same time, ANP attenuated thrombin induced neutrophil accumulation in the wound, and ANP alone increased Ps. Our results confirm that ANP acts to attenuate transvascular exchange in an intact microvasculature but only under inflammatory conditions. The mechanism of action appears to be independent of the normal ANP signaling pathways to increase Ps, and may involve reduced production of pro‐inflammatory agents from activated endothelium, neutrophils, or other inflammatory cells.

Grant Funding Source: Supported by NIH HL28607
Variable roles of the H1‐H3 receptors and PKC in histamine‐induced barrier dysfunction in cultured endothelial cells from different sources (672.6)
Authors: Adderley Shaquria , Zhang Xun , Sarangan Sandhya , Breslin Jerome
Published: June 2021

We tested the hypothesis that histamine‐induced endothelial barrier dysfunction requires different receptors and signaling pathways depending upon endothelial cell (EC) source. Changes in barrier function elicited by 10 μM histamine were compared between human umbilical vein EC (HUVEC) and human dermal microvascular EC (HMVEC‐D) monolayers, using determination of transendothelial electrical resistance (TER). H1, H2, and H3 receptors were blocked with 10 μM mepyramine, 10 μM cimetidine, and 0.01‐1 μM ciproxifan, respectively. Inhibitors of PKC (GFX109203X), PI3K (PI828) and phospholipase C (U73122) were used to evaluate downstream signals. Western blots and immunofluorescence microscopy analysis demonstrated H1, H2, and H3 expression in both EC types. Blockade of H1, H2, or H3 significantly inhibited the histamine‐induced decrease in TER in HMVEC‐D. In contrast, only blockade of H1 significantly attenuated the histamine‐induced drop in TER in HUVEC. Inhibition of either PKC or PI3K significantly attenuated histamine‐induced barrier dysfunction in HUVEC, but combined inhibition of both was required to attain significant attenuation in HMVEC‐D. Inhibition of PLC also inhibited the histamine‐induced decrease in TER in HMVEC‐D, but interestingly, combined PLC and PKC inhibition did not. This study suggests a common role for H1, but differential roles for H2 and H3 in histamine‐induced endothelial barrier dysfunction among different endothelial cell types. In addition, PKC appears to have either a promoting or inhibitory role depending on the endothelial cell source. Supported by NIH R01HL098215.

Grant Funding Source: NIH R01HL098215
Functional changes in cardiovascular responses with exposure to oxidative insult in a zebrafish embryo model (674.12)
Authors: Frame Mary , Gunter Geoffrey , Thompson Holly
Published: June 2021

Our goals were to design an observation chamber to observe the cardiovascular responses of anesthetized (0.1% MS222) zebrafish embryos (N=50) at 4‐5 dpf (days post fertilization), and to determine if oxidative insult altered these responses. The chamber was molded out of a silicone elastomer, and consisted of a shallow depression to hold the fish, surrounded by a larger moat that is maintained at 28.5C. Without restraining the fish, this system permits volume exchange (to test different doses/agents) while keeping the fish continuously within the field of view. Heart rate (HR), and diameter (D) changes for the aorta and caudal artery were obtained in response to acetylcholine (ACH 1e‐4M), nitroprusside (NP 1e‐4M), adenosine (ADO 1e‐4M) and phenylephrine (PE 1e‐6M). All solutions were made in instant ocean without methylene blue. The baseline D were ~50 um (aorta) and ~30 um (caudal a.). Baseline HR was 96 b/m. Maximal dilation values were 25% ACH, 30% NP, 20% ADO; maximal constriction to PE was ‐50%. Changes in HR were ‐74% ACH, ‐40% NP, ‐50% ADO, and +20% PE. Oxidative stress was induced by exposure to 1e‐4M peroxide, or 0.25 ug/ml nanometal oxide (silica, ceria, alumina) for 24 hours at 28.5C. Following oxidative stress, dilation and decreased HR to ACH was attenuated, consistent with endothelial dysfunction. Responses to NP or ADO were unchanged. Responses to PE were an accentuated constriction, but unchanged HR. This chamber provides a means to continuously observe vasoactive responses within the zebrafish embryo. The zebrafish model has the potential to test for oxidative insult impact on the cardiovascular system.
Computational analyses of oxygen transport suggest that perfusion heterogeneity, not overall oxygen delivery, limits muscle performance in metabolic syndrome (674.7)
Authors: Frisbee Jefferson , Wu Fan
Published: June 2021

The ability of in situ skeletal muscle to perform is compromised with development of metabolic syndrome in obese Zucker rats (OZR); a model of vascular disease without atherosclerosis. At 17 weeks, skeletal muscle function appears to be normal, implicating the microcirculation as a major contributor to poor performance; yet actual contributors to this are unclear. We performed computational analyses on single capillary O2 transport in LZR and OZR, using the Krogh model to compute axial PO2 distribution given metabolic rate and capillary diameter. The target variable is ΔPO2 between the capillary center and outer wall of a tissue cylinder. Initial analyses suggest that, assuming uniform microvascular perfusion, the maximum axial ΔPO2 is 13 mmHg in LZR and 10 mmHg in OZR at fatiguing contractions, suggesting that oxygenation in both is sufficient to support metabolism and that O2 consumption is not limited by bulk perfusion in LZR or OZR. Combined with our previous findings on capillary‐level flow distributions [Wu et al., 2011], this implies that increased muscle fatigue in OZR is associated with heterogeneous flow distribution, causing local mismatch between O2 supply and metabolic demand.

Grant Funding Source: Supported by National Institutes of Health and American Heart Association
Comparison of plasma and 5% albumin resuscitation on preserving endothelial glycocalyx and microvascular permeability in vivo after severe hemorrhagic shock in rats (676.12)
Authors: Torres Luciana , Sondeen Jill , Salgado Christi , Dubick Michael , Torres Filho Ivo
Published: June 2021

Plasma expanders have been used to promote volume replacement and improve hemodynamics and tissue perfusion after HS. We have investigated the effects of high viscosity plasma expanders on the EG of cremaster muscle venules from anesthetized rats subjected to 40% HS for 1h followed by 1h resuscitation with 5% albumin (ALB, Buminate), fresh frozen plasma (FFP) or 6% Hetastarch (HEX, Hextend). EG thickness, blood flow, and permeability were studied using intravital microscopy. Hemodynamics and coagulation tests (ROTEM) were measured. Mean arterial pressure was significantly lower than baseline for all groups (hypotensive resuscitation). HEX group showed significantly lower EG thickness, higher blood flow and permeability than the FFP. ALB partially restored EG thickness (80% of baseline), but showed higher blood flow and shear rate than the FFP group (p<0.05). Permeability was 20% higher in the ALB group than baseline. EXTEM and FIBTEM showed higher clotting time and lower clot strength in the ALB and HEX groups compared to FFP. In conclusion, resuscitation with ALB alone is not sufficient to return the EG thickness, permeability and coagulation to normal after HS possibly due to the lack of other plasma components beyond that of albumin. Resuscitation with FFP is superior to high viscosity plasma expanders, including the main FFP protein constituent alone.

Supported by US Army Medical Research & Materiel Command.
In vivo effects of blood products on the endothelial glycocalyx, microvascular permeability and coagulation in severe hemorrhagic shock (676.15)
Authors: Torres Filho Ivo , Torres Luciana , Sondeen Jill , Salgado Christi , Dubick Michael
Published: June 2021

Transfusion with blood and blood products are used to treat hemorrhage and improve tissue oxygen delivery. However, since the effects of products such as packed red blood cells (PRBC) on the EG are unknown, we compared the effects of resuscitation with blood products to fresh whole blood (FWB) on cremaster muscle venules. Anesthetized rats subjected to 1h of HS were resuscitated with 1:1 PRBC:LR (Lactated Ringer's), 1:1 wPRBC (washed PRBC):LR or FWB. EG thickness, blood flow and permeability were studied using intravital microscopy. Hemodynamics and coagulation tests (ROTEM) were performed. EG thickness and permeability were restored in the PRBC:LR compared to FWB and SHAM groups, but not in wPRBC:LR (p<0.05). FWB restored coagulation function. EXTEM and FIBTEM clotting times were longer in PRBC:LR and wPRBC:LR. EXTEM clot strength fell >30% in wPRBC: LR. Resuscitation with FWB was superior in reverting coagulopathy, restoring EG, permeability and blood flow. PRBC:LR improved EG and permeability (due to the presence of plasma proteins), but coagulation and blood flow were not recovered. wPRBC:LR did not improve EG or permeability (due to plasma protein removal after washing). Data suggest that removal of residual plasma proteins from PRBC and its association with a protein‐free solution (LR) impair microcirculation and coagulation function in this HS model.Supported by US Army Medical Research & Materiel Command.
Vasopressin receptor 2 stimulation increases permeability in human microvascular endothelial cells (676.16)
Authors: Lopez Ernesto , Fujiwara Osamu , Zhu Yong , Enkhbaatar Perenlei
Published: June 2021

Introduction: Arginine vasopressin (AVP) is administered in sepsis to increase arterial pressure by activating AVP receptor 1 (V1R). Limited attention has been given to effects of AVP receptor 2 (V2R) on endothelial cells. Previously, in our ovine model of sepsis, we have reported that treatment of severe sepsis with V1R selective agonist prevents vascular hyper‐permeability as compared to animals treated with non‐selective AVP. We hypothesize that V2R activation is critically involved in the pathogenesis of vascular hyper‐permeability, which is a severe complication of sepsis.

Methods: An electrical impedance assay was adapted to assess acute changes of connectivity in confluent Human Lung Microvascular Endothelial Cells (HMVECs). Confluent cells were exposed to different doses of V2R specific agonist, Desmopressin (DDAVP), Vascular Endothelial Growth Factor (VEGF) or vehicle. The intercellular connectivity (electrical impedance) was measured every 15 min during 24 hours (h). A HMVECs permeability assay was also performed measuring the amount of FITC‐dextran passing through the endothelial monolayer after 18 h of treatment with DDAVP or VEGF.

Results: V2R expression in HMVECs was verified with RT‐PCR. DDAVP caused a decrease of HMVECs connectivity with a maximum difference at 0.5 h of treatment. Its clear dose dependent effect was also seen at 0.5 h with 0.3nM, 1nM, 3nM (p蠄0.05), 10nM (p蠄0.0001), and 30nM (p蠄0.0001). The effect was similar to changes seen with 3 ng/ml VEGF (p蠄0.0001), but milder than 10 ng/ml VEGF. The changes in HMVECs connectivity remained during the rest of the experiment. The FITC‐dextran permeability assay also showed a trend of increased permeability with 10 nM DDAVP and 50 ng/ml VEGF.

Conclusions: These results suggest that the activation of V2R disrupts the endothelial barrier within minutes, thus causing hyper‐permeability. Additional studies are being performed in our in vivo model to elucidate the role of V2R in sepsis induced vascular hyper‐permeability.

Grant Funding Source: Supported by NIH RO1 GM097480, SHC85500, SHC84050 and CONACYT‐Mexico
Pre‐diabetes promotes sympathetically‐mediated arteriolar dysregulation in response to skeletal muscle contraction (676.20)
Authors: Novielli Nicole , Medeiros Philip , Jackson Dwayne
Published: June 2021

Augmented sympathetic vascular regulation is associated with pre‐diabetes, thus it may contribute to impaired contraction‐evoked arteriolar dilation in skeletal muscle, previously demonstrated by our group. Thus we tested the hypothesis that blunted contraction‐evoked arteriolar dilation in pre‐diabetes occurs as a result of augmented sympathetic neuropeptide Y1 receptor (NPY Y1R) and α1 adrenergic receptor (α1R) control. Using intravital video microscopy, second‐ third‐ and fourth‐order arteriolar diameter changes were measured in response to electrical field stimulation of the gluteus maximus muscle in pre‐diabetic (PD, Pound Mouse, n=6‐13) and control (CTRL, c57bl6, CTRL, n=5‐12) mice. Baseline diameter was similar between groups, however vasodilatory responses to single tetanic (100Hz; 400 and 800ms) and sustained rhythmic (2 and 8Hz; 30s) contractions were blunted by 50% or greater in PD versus CTRL (p<0.05). In line with our hypothesis, decrements in contraction‐evoked dilation in PD were restored upon Y1R and α1R blockade (BIBP3226 and prazosin, 100nM). Arteriolar reactivity (% change from baseline diameter) to muscle contraction was greatest at distal arterioles in CTRL. Such spatial differences in arteriolar reactivity were absent in PD, but were restored with Y1R and α1R blockade. Finally, arteriolar vasoconstrictor responses to increasing doses of NPY (10‐13 ‐ 10‐8 M) or PE (10‐9 M ‐ 10‐5 M) were greater in PD versus CTRL, an observation that was consistent with greater Y1R and α1R expression in isolated arterioles and vascular smooth muscle cells in PD. Deficits in contraction‐evoked arteriolar dilation in this model of pre‐diabetes are a result of elevated sympathetic arteriolar Y1R and α1R expression and activation.

Grant Funding Source: NSERC
Potassium and calcium channel gene expression in small arteries in porcine and rat models of diet‐induced obesity (676.4)
Authors: Jensen Lars , Salomonson Max , Sorensen Charlotte , Holstein‐Rathlou Niels‐Henrik , Pedersen Henrik , Christoffersen Berit , Olsen Lisbeth , Ludvigsen Trine
Published: June 2021

Obesity is an increasing problem worldwide leading to cardiovascular morbidity. Only limited information exists on the transcriptional regulation of arterial K+ and Ca2+ channels in obesity. We quantified, by real‐time PCR, mRNA expression of K+ channels and L‐type Ca2+ channels (LTCC) in small mesenteric (MA), middle cerebral (MCA), and left coronary arteries (LCA) of lean vs. obese rats and minipigs. Male Sprague Dawley rats were fed a high‐fat (FAT; N=5), high‐fructose (FRUC; N=7), high‐fat/high‐fructose (FAT/FRUC; N=7) or standard diet (STD; N=7‐11) for 28 Weeks. FAT and FAT/FRUC became obese, whereas FRUC and STD were lean. Systolic blood pressure (SBP) averaged over 14 weeks was increased (P<0.001) in FRUC and FAT/FRUC but not in FAT. Kir, Kv1.2, Kv1.5, and Kv7.4 channels were up‐regulated in FAT, whereas SKca and IKca channels were down‐regulated in FAT/FRUC (P<0.05). There were no transcriptional changes in FRUC. Castrated male Göttingen minipigs with or without diabetes were fed a diet rich in fat, cholesterol and fructose (OB+DIAB; N=3 vs. OB; N=2, respectively) or a standard diet (STD; N=3) for 22‐45 weeks. Body weight, total body fat content and plasma triglyceride levels were increased in OB and OB+DIAB. BKca, IKca, SKca and/or LTCC mRNA was up‐regulated in LCA from OB and OB+DIAB (n.s.). Expression of BKca mRNA was increased, whereas IKca mRNA decreased in MCA from OB (n.s.). SKca mRNA was decreased in MA from OB (n.s.). Diet‐induced obesity in rats and minipigs lead to complex changes in K+ and Ca2+ channel gene expression, which, in rats, did not seem to be linked with changes in SBP. These transcriptional changes may lead to disturbances in microvascular flow patterns in obesity.

Grant Funding Source: Supported by: Danish Medical Research Council; The Foundation for the Advancement of Medical Science
Sildenafil improves cardiovascular function in experimental hypertension (676.5)
Authors: Balarini Camille , Dias Ananda , Fahning Bernah , Cintra Amanda , Frossard Jessica , Gomes Isabele , Gava Agata , Meyrelles Silvana , Vasquez Elisardo
Published: June 2021

The aim of the present study was to evaluate the beneficial cardiovascular effects of sildenafil treatment on experimental hypertension.

Male C57BL/6 mice (n=5‐10) were subjected to 2K1C hypertension. After 14 days, sildenafil (40 mg/kg/day) or vehicle was administrated, during a period of 14 days. At the end of experimental period, animals were anesthetized and catheterized to blood pressure recording and had the mesenteric arteriolar bed removed to vascular function studies (acetylcholine ‐ ACh and norephrine ‐ Nor curves) (Protocol 02/2013‐ CEUA‐EMESCAM). Data are mean ± SEM. One‐way ANOVA was performed, followed by Tukey's post hoc test.*p<0.05 and **p<0.01 vs. 2K1C + vehicle. Sildenafil treatment reduced mean arterial blood pressure (124±2 vs. 112±3* mmHg) and heart rate (577±32 vs. 472±19* bpm) in hypertensive‐treated animals. Endothelial function was impaired in hypertensive animals and was successfully restored by sildenafil (Rmax ACh: 50.5±2% vs. 71.6±3%**). Blockage of nitric oxide production with L‐NAME revealed that the participation of this molecule in ACh‐induced vasodilation was augmented by sildenafil (AUC: 60.6±8 vs. 126.8±24*). Similarly, blockage with indomethacin showed the increased participation of COX‐derived prostanoids in vasodilation of treated animals (AUC: 37.9±9 vs. 95.9±17*). The participation of reactive oxygen species was evaluated by blockage of NADPH oxidase with apocinin and revealed that sildenafil was efficient in reducing oxidative stress (AUC: 94.±7.3 vs. 47.4±10.4*). Furthermore, sildenafil was also successful in reducing hyperactivity to Nor in 2K1C animals (Rmax: 167±8 vs. 117±9** mmHg). Thus, sildenafil can restore endothelial function and reduce blood pressure in experimental hypertension and could be suggested as a novel therapeutic approach while treating hypertensive patients.

Grant Funding Source: Supported by: Fapes 54498465; CNPq 476525/2012‐8, 473082/2013‐6; FAPES/CNPq/PRONEX Edital 012/2009
The 3D structure of the myoendothelial projections: intracellular organelles, protein trafficking and biological function (677.12)
Authors: Maarouf Nadia , Welsh Donald
Published: June 2021

The 3‐D structure of the myoendothelial projections: intracellular organelles, protein trafficking and biological Function Nadia Maarouf and Donald G. Welsh Department of physiology and pharmacology, University of Calgary, Calgary, AB, Canada Endothelial and smooth muscle cells communicate with one another in order to regulate vessel diameter. A key element of heterocellular communication is the myoendothelial projection, a thin endothelial extension that crosses the internal elastic lamina to make contact with the smooth muscle. The aim of this study was to ascertain the structural composition of the myoendothelial projection using electron tomography, a technique that enables the generation of 3‐D models. Anatomical models from mesenteric arteries reveal that myoendothelial projections are generally devoid of membranous structures, contrary to current thought. Intriguingly, key organelles such as endoplasmic reticulum with ribosomes, caveolae and vesicles were found in abundance at the base of the projection. The presence and positioning of the ribosomal endoplasmic reticulum suggests that in addition to governing Ca2+ regulation, this organelle contributes to protein packaging and trafficking to the plasma membrane which may include ion channels. In summary, the electron tomography approach provides valuable structural insight into the detailed composition of myoendothelial projections and their potential role in arterial tone regulation.

Grant Funding Source: CIHR
Far red/near infrared light induces vasodilation and an increase in activity of large conductance potassium channel currents in murine femoral arterial muscle (677.13)
Authors: Lohr Nicole , Lindemer Brian , Harder David , Gebremendhin Debebe
Published: June 2021

Over 8 million Americans are treated for symptoms related to peripheral artery disease (PAD), such as intermittent claudication, impaired wound healing, and critical limb ischemia. We have shown light irradiation at the far red/near infrared region (670 nm (R/NIR)) can stimulate angiogenesis by increasing NO independent of NOS through release of NO from nitrosyl‐heme stores. We hypothesized R/NIR can stimulate vasodilation through increases in large conductance potassium channel (BKCa) activity. To test vasodilation, facial arteries from C57Bl/6 mice were isolated, pressurized to 60mmHg, and pre‐constricted with U46619 in the dark.

The vessels were irradiated with 670 nm light (10mW/cm2) up to 5 min at a time with 10 min dark period between irradiation. Vessel diameter increased up to 17.8% (±3.0, p<0.05) after 5 min of R/NIR exposure. The effect of red light on single channel BKCa current was determined in enzymatically dissociated murine femoral arterial muscle cells, using the patch clamp technique and symmetrical KCl [145mM] recording solution at a patch potential of +60 mV (RT). Exposure of cell attached patches to light ( λ =670 nm, 10 mW/cm2) markedly increased the opening frequency of the large conductance potassium channels (BKCa) single channel current. This effect of light on BKCa single channel current activity could be the underlying mechanism for the vasodilatory action of red light in isolated murine femoral arterial segments. Conclusion: R/NIR exposure acts to stimulate vasodilation, by increasing the opening frequency of BKCa single channel current recorded in cell attached patches of femoral arterial smooth muscle cells.
Acute effects of dietary bisphenol A on endothelial function in humans (677.14)
Authors: Chantler Paul , Casto Erica , DeVallance Evan , Fournier Sara , Fancher Ibra , Dick Gregory
Published: June 2021

BPA is present in canned goods and detectable in the urine of >95% of US adults. Studies suggest a link between BPA and cardiovascular disease, but mechanistic information in human subjects is extremely limited. We investigated whether consuming 12‐oz of canned soup daily for 5 days would impair flow‐mediated dilation (FMD) of the brachial artery. FMD is an assay of endothelial function that provides insight into cardiovascular health. Using a double‐blinded crossover design, we compared the effects of 5 days of canned vs. fresh soup consumption in 7 healthy adults. The fresh soup recipes were matched for sodium and fat content, but contained no canned ingredients. After each week of soup, urinary BPA and FMD (with B‐mode ultrasound using published guidelines) were measured. Data were compared using paired t tests. BPA was undetectable in fresh soup (<0.01 ug/ml), but was 0.98 ± 0.72 ug/ml in canned soup. Thus a 12‐oz portion of canned soup could be expected to contain approximately 350 ug of BPA (representing a concentration of 4.4 uM). After a week of fresh soup, urinary BPA was 1.61 ± 0.48 ug/L. In contrast, after a week of canned soup, urinary BPA increased to 5.85 ± 4.87 ug/L (p < 0.05). However, FMD did not differ following a week of canned or fresh soup (10 ± 4 vs. 8 ± 2%, p = NS). These preliminary findings indicate that acute BPA consumption has little or no effect on endothelial function in healthy young individuals.

Grant Funding Source: Supported by T32HL090610 and AHA 11CRP7370056
The vasoconstrictor agent phenylephrine selectively modulates endothelium‐ vs. smooth muscle‐mediated inhibition of myogenic tone in rat cremaster arteries (677.5)
Authors: Mishra Ramesh , Rahman Mohammad , Wulff Heike , Hill Michael , Braun Andrew
Published: June 2021

The α1‐adrenergic agonist phenylephrine (PE) is reported to impact endothelium‐mediated vasorelaxation by influencing vasoactive signals transferred from endothelium to vascular smooth muscle (VSM), however, it is unclear whether this putative process occurs in myogenically active vessels at physiologic pressures. In the present study, we have examined the impact of PE and the TP receptor agonist U46619 on endothelium‐dependent and independent vasorelaxation of cannulated rat cremaster arterioles pressurized to 70 mmHg in the absence of luminal flow. Vasorelaxant agents acting via the endothelium (i.e. 0.3 μM acetylcholine (ACh) and 3 μM SKA‐31) or directly on VSM (i.e. 10 μM adenosine (ADO), 10 μM SNP, 5 μM pinacidil) reversibly inhibited myogenic tone by 50‐80%. Bath addition of 25 nM PE evoked a modest constriction on top of the existing myogenic tone, but did not alter the magnitude of vasorelaxant responses to ACh and SKA‐31. However, ADO‐mediated vasorelaxation was somewhat depressed and the response to pinacidil was enhanced. In the presence of U46619 (5 nM), no clear effects on either endothelium‐dependent or ‐independent vasodilatory stimuli were noted. In summary, the cellular pathways activated by PE and U46619 in VSM do not appear to influence endothelium‐mediated inhibition of myogenic tone in cremaster arterioles, suggesting that myo‐endothelial communication is unaffected by signaling events occurring downstream of α1‐adrenoceptor or TP receptor activation in these vessels.

Grant Funding Source: Supported by CIHR
Evidence that flow mediated dilation dynamics differ in smokers versus non‐smokers: a pilot study (677.6)
Authors: King Trevor , Findlay Briar , Gupta Parnika , Szijgyarto Ingrid , Pyke Kyra
Published: June 2021

Endothelial function is impaired in smokers as measured via handgrip exercise (HGEX) induced flow mediated dilation (FMD). The goal of this pilot study was to determine whether the dilation and reconstriction dynamics of FMD were impaired in young male smokers. 8 smokers and 14 non smokers (age 21 ± 2) performed 10 minutes of rhythmic HGEX bracketed by 4 minutes of arterial compression to ensure rapid shear stress onset and return to baseline. Brachial artery diameter and blood velocity assessed via echo and Doppler ultrasound. Shear stress estimated as shear rate (SR = brachial artery blood velocity/diameter). Dynamic parameters: time to onset of dilation and reconstriction (TD1); rate of dilation and reconstriction(Tau1). Alpha set at P<0.1, data ± SD. SR stimulus did not differ between groups(p=0.60). TD1 of dilation was longer in smokers (29.7 ± 17.0 s) vs. non smokers (17.5 ± 11.9 s)(p=0.06), and Tau1 was faster among smokers (27.5 ± 9.5 s vs. 48.6 ± 32.4 s, p = 0.09). Constriction dynamics were not different between groups (p>0.1). Among smokers, dilation tau1 was faster than reconstriction tau1(27.5 ± 9.5 s vs. 72.3 ± 56.4 s, p=0.06), whereas there were no differences between onset and constriction dynamics among non‐smokers (p=0.43). These results suggest that smoking impacts the dynamics of the vascular response to shear stress with a preferential impact on dilation vs. reconstriction.

Grant Funding Source: Supported by NSERC and CFI
Myogenic characteristics of the afferent arteriole in papillectomized and in Cx40 deficient mice (678.1)
Authors: Jacobsen Jens Christian , Sørensen Charlotte
Published: June 2021

Glomerular flow is regulated through diameter changes in the afferent arteriole (AA). In turn diameter of the AA is regulated primarily by the myogenic response and by the tubuloglomerular feedback (TGF) mechanism constituting renal autoregulation. Connexin 40 (Cx40) is the main connexin of endothelial gap junctions in AA and it has been shown previously that Cx40 knock‐out (Cx40KO) mice lack TGF. In the present study we investigated whether absence of functional Cx40 in the endothelium also affects the properties of the myogenic response in the AA. We compared the diameter changes of the AA to an increase in renal perfusion pressure, which elicits an autoregulatory AA contraction induced by the myogenic response and TGF. We compared papillectomized wildtype mice (lacking TGF, but with intact intracellular communication in the endothelial layer) versus Cx40KO mice (lacking TGF and with deficient intracellular communication in the endothelium). We found no differences between the two groups indicating that the level of myogenic tone is largely determined by the local pressure, independent of longitudinal endothelial communication. In addition we modeled the observed myogenic response and estimated the characteristic relation between circumferential stress and vessel wall activation leading to the observed myogenic behavior.
Quantification of microvascular hemodynamic using cross correlation analysis of intravital microscopy videos (678.11)
Authors: Ortiz Daniel , Briceno Juan , Cabrales Pedro
Published: June 2021

Blood flow patterns and endothelium interactions are of great importance for the progression of cardiovascular diseases, thus quantitative analysis of blood flow dynamics at the microcirculation level of great relevance. Regulatory mechanisms mediated by blow flow have been studied in detail using in vitro approaches. However, these mechanisms have not been completely validated in vivo due to technical limitations involved in the quantification of flow patterns in the mcirocirculation with the required level of detail. Intravital microscopy combined with high‐speed video recordings has been used for the analysis of blood flow in small blood vessels of chronic and acute experimental animal tissue preparations. This tool is used to study the interaction between the flowing blood and the vessel walls of arterioles and venules with great temporal and spatial resolution. Our objective is to develop a simple and robust cross‐correlation algorithm for the automatic analysis of high‐speed video recordings of microcirculatory blood flow. The algorithm has been validated using in vitro (glass microchannels) and in vivo (window chamber models) systems. Results indicate that the algorithm’s ability to estimate the velocity of local red blood cells (RBCs) as a function of blood vessel radius and time is highly accurate.The algorithm could be used to explore dynamic changes in blood flow under different experimental conditions including a wide range of flow rates and hematocrit levels. The algorithm can also be used to measure volumetric flow rates, radial velocity profiles, wall shear rate (WSR), and wall shear stress (WSS). Several applications are presently explored, including the analysis of velocity profiles in the branches of arterial bifurcations. Our work demonstrates the robustness of the image cross‐correlation technique in various flow conditions and elucidates its potential application for in vivo determination of blood flow dynamics in the microcirculation.

Grant Funding Source: Supported by: NIH R01 HL52684, R01 HL064395 and R01 HL062318 and ARMY: W81XWH‐11‐2‐0012
Cutaneous thermal injury increases leukocyte adherence in rat mesenteric venules by a mast cell‐independent mechanism (678.15)
Authors: Howard James , Lucero Raulee , Moncure Michael , Holloway Naomi , Wood John
Published: June 2021

Background: The inflammatory response following major burns is known to cause remote organ dysfunction, although the underlying mechanisms are not yet known. Purpose: Our goal was to determine if thermal injury to the skin of rats would promote microvascular inflammation in the mesenteric microcirculation, and if the mechanism involved degranulation of mast cells. Methods: Intravital microscopy was used to assess mast cell degranulation and leukocyte adherence in mesenteric venules of anesthetized rats. Thermal injury was induced by exposing skin on the back of animals to steam for 1 min. The area of injury was ~ 40% of the total surface area. Control animals were not exposed to steam. Results: Leukocyte adherence significantly increased during 5 h after burn, but not in control animals. Mast cell degranulation was not increased by burns, and the mast cell stabilizer cromolyn did not attenuate leukocyte adherence in burned rats. Conclusions: Thermal injury to the skin of rats resulted in microvascular inflammation in the mesenteric microcirculation of rats by a mechanism independent of mast cell activation.
Age‐related progression of bone marrow blood vessel ossification in rat and, potentially human, long bone (678.19)
Authors: Prisby Rhonda
Published: June 2021

BACKGROUD: Bone perfusion and bone vessel location, density and function are associated with bone mass. We recently demonstrated ossification of bone vessels in rats. The objective of this study was to assess ossification with advancing age and to confirm ossification in human patients. METHODS: Femora from young (n=8) and old (n=8) male Fischer‐344 rats were examined to quantify patent vs. ossified and calcified vessels. Femora from additional young (n=5) and old (n=5) rats were analyzed for vessel ossification via µCT. Femoral vessels from a patient with ASVD and from rats were examined via microscopy. RESULTS: Isolated vessels (human and rat) had osteocyte lacunae on the surface. Patent vessel number was reduced (p<0.05) with age (0.47±0.05/mm2 vs. 0.29±0.05/mm2). Higher (p<0.05) volumes of ossified vessels were evident with age (0.01±0.01% vs. 0.49±0.11%). Calcified vessel volume/tissue volume (0.32 ±0.08% vs. 1.16±0.31%) and /patent vessel volume (17.2±5.9% vs. 62.4±15.5%) were augmented (p<0.05) in old rats. Ossified vessel number was higher (p<0.05) in old rats (0.07±0.01/mm2 vs. 0.19±0.04/mm2). CONCLUSIONS: For the first time, ossification of human bone marrow blood vessels was demonstrated. Ossification begins in youth and progresses with age, corresponding with a reduced number of patent vessels. Such pathology is presumably associated with bone disease.

Grant Funding Source: Supported by NIH (NIAMS) Grant 1R15AR062882‐01
CD34+ stem cell treatment for acute ischemic stroke (678.20)
Authors: Gavins Felicity , Smith Helen , Granger D. Neil
Published: June 2021

Stroke is the most common cause of significant acquired disability in adults for which only limited therapeutic strategies exist, limited by a brief therapeutic window (<4 h). No pharmacological compound to date has been found to increase recovery reliably once brain injury has occurred. Stem cell transplantation techniques offer novel approaches that may enhance endogenous repair mechanisms, promote brain recovery and regeneration, and may ultimately reduce neurological deficits in stroke patients. While showing positive results, current stem cell therapies are highly inefficient, with a tiny percentage of injected cells arriving at the intended injured site.

Following on from our previous open label phase 1 study in stroke patients, to determine the safety and feasibility of treatment with immunoselected CD34+, we have used a murine model of ischemic stroke (the middle cerebral artery occlusion model) to investigate the migration of CD34+ stem cells to the infarcted area. Using both intravital and confocal intravital microscopy we have been able to explore both leukocyte and stem cell trafficking in the murine brain pre and post stroke. Both leukocyte rolling and adherence were increased in mice subjected to stroke vs. sham mice. Stem cell treatment reduced these leukocyte interactions, with effects observed upto two weeks post stroke. These novel data suggest a therapeutic potential role for the use of CD34+ stem cells for acute ischemic stroke
Gender‐based protection against microvascular dysfunction with depressive symptoms in mice (678.3)
Authors: Skaff Paulina , Stanley Shyla , Brooks Steven , Butcher Joshua , Frisbee Jefferson
Published: June 2021

Human subjects suffering from clinical depression frequently experience these symptoms as a result of the presence of chronic, unresolved stresses in their environment. This can have significant impacts on a poor outcome for cardiovascular health, and one of the central markers of dysfunction is a blunted vasodilator response to imposed stimuli. There is increasing evidence that this may be due to a reduction in the bioavailability of signaling molecules contributing to the dilator response. To investigate this in an animal model of chronic stress and depressive symptoms, we used the unpredictable chronic mild stress (UCMS) model to simulate human clinical depression in mice. In response to the UCMS protocol, all mice exhibited significant depressive symptoms, although this was increased in female mice vs. males. Endothelium‐dependent dilator responses in ex vivo aortic rings in response to increasing concentrations of methacholine were reduced by UCMS, and this effect was more extensive in male than in female mice. Treatment of rings with the antioxidant TEMPOL improved dilator responses in vessels from both genders. Inhibition of NOS (L‐NAME) had no impact on dilator responses in rings from UCMS males, but reduced responses in females. Inhibition of cyclooxygenase (indomethacin) blunted dilation in UCMS males but had a smaller effect on responses in UCMS females. These results suggest that, while female mice are more susceptible to depressive symptoms, they are somewhat protected from the impaired vascular function. While male mice are less susceptible to the depressive symptoms their vascular function is impaired to a greater extent.

Grant Funding Source: Supported by National Institutes of Health and American Heart Association
Cutaneous microvascular endothelium‐dependent vasodilation is impaired in young obese subjects (678.4)
Authors: Patik Jordan , Christmas Kevin , Brothers R.
Published: June 2021

Cutaneous microvascular endothelium dependent vasodilation is impaired in young obese subjects Jordan C. Patik, Kevin M. Christmas, and R. Matthew Brothers Environmental and Autonomic Physiology Laboratory, Department of Kinesiology and Health Education, University of Texas at Austin Microvascular dysfunction is a precursor to a variety of cardiovascular diseases and contributes to insulin resistance and type II diabetes. Obese individuals are at a greater risk for the development of these aforementioned conditions relative to their lean counterparts. The purpose of this study was to asses endothelium dependent microvascular function in young, otherwise healthy, obese individuals. Specifically, we tested the hypothesis that the cutaneous microvascular response to intradermal administration of methacholine (MCh) would be blunted in obese relative to lean individuals. 6 young obese and 5 young lean individuals (Age: 24±2.2 vs. 25±1.18 years P>0.05; BMI: 34.96±1.89 vs. 22.24±0.84 kg/m2, P<0.001) participated in this study. Microvascular function was assessed using laser Doppler flowmetry to calculate cutaneous vascular conductance over a range of MCh concentrations infused through microdialysis membranes. Dose response curves were constructed to determine the effective concentration that elicits 50% of maximal conductance (EC50). Young obese subjects had significantly blunted EC50 responses to MCh infusion relative to the lean individuals (Obese: ‐3.173±0.526 vs. Lean: ‐4.875 ±0.199 log molar concentration of MCh, P = 0.02). Our preliminary results support the hypothesis that impaired endothelium dependent vasodilation is present in relatively young obese individuals prior to the development of obesity related disease.
Contribution of Kv1 channel activity to myogenic responses in skeletal muscle arterioles with aging and exercise training (678.7)
Authors: Ghosh Payal , Solis Fredy , Dominguez Jimmy , Delp Michael , Muller‐Delp Judy
Published: June 2021

The impaired myogenic constriction of skeletal muscle arterioles observed with advancing age may be the result of increased Kv channel activity. Exercise training has been shown to restore myogenic responses. The purpose of this study was to determine the role of Kv1 channel activity in these adaptations in reactivity.

Young and old male Fischer 344 rats were randomly assigned to either a sedentary (YSED and OSED) or exercise trained (YET and OET) group. Training consisted of treadmill running (15m/min, 15°incline, 1hr/d, 5d/wk for 10 wks). Soleus muscle first order arterioles were isolated and cannulated for in vitro study. Myogenic responsiveness was determined by exposing arterioles to incremental changes in intraluminal pressure from 0 to 140 cmH20, either in the presence or absence of Kv1 channel blocker correolide.

Exercise enhanced myogenic tone in arterioles from both old and young rats and restored myogenic responsiveness in old rats to that of YSED rats. Kv1 channel blockade abolished the training effect in young rats, and resulted in greater constriction at lower pressures in YSED, but had no effect in YET rats. In both OSED and OET rats, Kv1 channel blockade led to increased myogenic constriction; however, the training effects persisted.

These results suggest that Kv1 channel activity may play a role in modulating myogenic tone in young and old rats. Increased myogenic constriction after training in young rats may be mediated through a reduction in Kv1 channel activity. However, improved myogenic responsiveness with training in old rats appears to be mediated through a different pathway.

Grant Funding Source: Supported by NIH R01HL090937
Bcl‐2 expression in endothelial cells impacts postnatal retinal vascular development without affecting retinal neovascularization (679.2)
Authors: Sorenson Christine , Zaitoun Ismail , Wang Shoujian , Sheibani Nader
Published: June 2021

Bcl‐2 plays an important role in retinal vascular homeostasis. Global lack of Bcl‐2 expression results in attenuation of postnatal retinal vascular development and neovascularization during Oxygen‐induced Ischemic Retinopathy (OIR).To determine the impact lack of Bcl‐2 expression in endothelial cells has during retinal vascular development, remodeling, and neovascularization, we generated mice carrying a conditional Bcl‐2 allele (Bcl‐2Flox/Flox) and VE‐cadherin‐cre called Bcl‐2EC mice. Bcl‐2EC mice displayed decreased numbers of endothelial cells at both 3 and 6 weeks of age compared to wild‐type controls. However, the remodeling of the retinal vasculature proceeded with similar percentage loss of endothelial cells in both wild‐type and Bcl‐2EC mice. In addition, Bcl‐2EC mice had increased pericyte numbers at 6 weeks of age contributing to a lower EC/PC ratio. Retinal arteriogenesis was also decreased in Bcl‐2EC mice similar to what we observed in global Bcl‐2 knockouts. However, unlike the global knockouts the migrating front of the retinal vasculature proceeded normally and neovascularization during OIR occurred at levels similar to wild‐type mice. Therefore, our data suggests that Bcl‐2 expression modulates endothelial cell survival impacting postnatal retinal vascular development with minimal impact on retinal neovascularization during OIR.

Grant Funding Source: Supported by R21EY023024