bluuebunny/crossref_metadata_2025 huggingface dataset:
Anti vascular inflammatory effect of Samul‐tang in HUVECs
Authors: Choi Eun Sik , Lee Yun Jung , Yoon Jung Joo , Ahn You Mee , Park Ji Hun , Han Byung Hyuk , Namgung Seung , Jin Xian Jun , Kang Dae Gill , Lee Ho Sub
Published: November 2023

Samultang (, SMT) is a well known herbal prescription treating overall haematological disorders defined as blood deficiency, blood stasis and blood loss in traditional korean medicine. SMT is recorded in several formulary including ‘Treasured Mirror of Eastern Medicine’ (, Donguibogam) and consists of 4 herbs : Angelicae Radix, Cnidii Rhizmoa, Rhemanniae Radix Preparata and Paeonia Radix. Major cause of atherosclerosis and other vascular diseases is inflammation and vascular endothelium is emphasized as central spot of vascular inflammatory process. Thus, we investigated the effects of SMT water extracts on vascular inflammation in HUVECs (Human Umbilical Vein Endothelial Cells).

Expression of CAMs (cell adhesion molecules) such as VCAM‐1 (vascular cell adhesion molecule‐1), ICAM‐1 (intracellular adhesion molecule‐1), E‐selectin (endothelial‐selectin) and HL‐60 monocyte adhesion were induced by TNF‐α stimulation. However, SMT pretreatment (10–50 μg/ml) inhibited CAMs expression and monocyte adhesion significantly. TNF‐α stimulation led HUVECs to produce ROS (reactive oxygen species) and translocate NF‐κB (nuclear factor‐κB) into nucleus, but SMT suppressed ROS production and NF‐κB nuclear localization significantly. Furthermore, activation of NF‐κB was also suppressed by SMT in dose dependent manner.

HO‐1 (heme oxygenase‐1) protein level was significantly upregulated by SMT treatment (10–50 μg/ml) as HO‐1 inducer CoPP (cobalt protoporphyrin) did. Besides, nuclear translocation of Nrf‐2 (nuclear factor erythroid 2‐related factor 2), which transcripts antioxidative genes including HO‐1, was increased by SMT dose dependently. Intracellular NO (nitric oxide) and nitrite products were also increased by SMT treatment.

Taken together, these results suggests that SMT might exerts vascular protective effects by suppressing vascular inflammatory process triggered from damaging stimulus including TNF‐α and by promoting synthesis of beneficial bioactive substances including HO‐1.
Effect of Hwangryunhaedoktang on TNF‐α‐induced vascular inflammation in human umbilical vein endothelial cells
Authors: Han Byung Hyuk , Lee Yun Jung , Yoon Jung Joo , Choi Eun Sik , Namgung Seung , Kim Hye Yoom , Ahn You Mee , Jin Xian Jun , Lee Ho Sub , Kang Dae Gill
Published: November 2023

Hwangryunhaedoktang (HHT) originally recorded in an ancient Korea medicinal book named “Donguibogam” and has been used for the treatment of inflammatory hemorrhage relevance to vascular inflammation which can be cause an atherosclerosis. This study was designed to demonstrate whether HHT has an inhibitory effect on vascular inflammation induced by TNF‐α in human umbilical vein endothelial cells (HUVECs).

Pretreatment with HHT decreased the adhesion of HL‐60 cells to TNF‐α‐induced HUVEC. HHT suppressed TNF‐α‐induced expression level of cell adhesion molecules such as intracellular adhesion molecule‐1 (ICAM‐1), vascular cell adhesion molecule‐1 (VCAM‐1), and endothelial cell selectin (E‐selectin), as well as dose dependently inhibited TNF‐α‐induced matrix metalloproteinase‐2/‐9 activity. HHT significantly decreased TNF‐α‐induced intracellular reactive oxygen species (ROS) production. Western blot and immunofluorescence analysis showed that HHT inhibited the translocation of p65 NF‐κB to the nucleus. Phosphorylation of IκB‐α also was inhibited in cytoplasm by pre‐incubating with HHT, on the other hand increased expression of IκB‐α.

These data showed that treatment with HHT regulate the ROS/NF‐κB signaling pathway. Therefore a traditional herbal formula HHT might be potential therapeutic agent of atherosclerosis.
Purinergic Signaling In Rat Mesenteric Arteries In Vivo
Authors: Aalkjaer Christian , Mazur Aleksandra , Matchkov Vladimir
Published: November 2023

The role of ATP for vascular function is well documented. In this study we wanted to assess the role of purinergic signaling for rat small mesenteric arteries in vivo. Rats were anesthetized with Ketamin/Xylazin and the intestine exteriorized through a laparotomy. A small mesenteric artery segment was positioned in a 200 μl reservoir and the diameter monitored. The mean diameter of the arteries (μm) was 241±15 (n=12). Application of l‐NAME, indomethacin, TRAM‐34 and apamin resulted in significant constriction (diameter 180±12 (n=12)). Addition of the broad purinergic receptor blocker suramine under these conditions resulted in dilation (diameter 180±19 and 276±19, without and with suramine, respectively (n=6)). During TTX sensitive electrical field stimulation a frequency dependent constriction was seen. At 16 Hz stimulation the response was inhibited by the a1‐receptor blocker prazosin but also by suramin plus the P2X1 inhibitor NF944. The prazosin sensitive constriction to exogenous norepinephrine (NE) was also inhibited by suramin plus NF944. We conclude that rat mesenteric arteries have vasodilator tone from the endothelium and ATP receptor dependent vasoconstrictor tone in vivo and that tone development to both endogenous and exogenous NE is dependent on purinergic receptors.
H+ Induces Arteriolar Vasodilation but Does Not Initiate a Conducted Response in Skeletal Muscle Microvasculature
Authors: Charter Mackenzie E , Murrant Coral L
Published: November 2023

Active hyperemia is the process where blood flow is recruited to a tissue to meet increased metabolic demand. Conducted responses are important to this process in order to direct blood flow to capillaries to feed active skeletal muscle fibres. A conducted response is a coordinated vasodilatory signal that travels from low to high branch order arterioles resulting in an increase in capillary perfusion. Conducted responses are hypothesized to be initiated by metabolic vasodilators ‐ byproducts of skeletal muscle metabolism‐ however, exactly which metabolites are involved remains unclear. Two vasodilatory metabolites of skeletal muscle metabolism are hydrogen ion (H+) and carbon dioxide (CO2). Both H+ and CO2 are interrelated through the bicarbonate (HCO3−) buffer (HCO3− + H+ <‐‐> H2CO3 <‐‐> CO2 + H2O) whereby H+ reacts with HCO3− to produce CO2 and CO2 reacts in reverse to produce H+. Thus, it is difficult to study the individual effects of these metabolites. In order to separate them we used a Tris (amine) buffer to superfuse the preparation, to allow us to apply H+ without changing CO2, thus allowing us to investigate the vasodilatory actions of H+ independently. Using an in situ, intact hamster cremaster muscle model, H+ was applied to the whole preparation through the superfusate or to a small region of a 2A arteriole (max. diameter approx. 40μm) via micropipette. Arteriolar diameter was recorded and analyzed offline. A range of H+ concentrations (63.1 – 251.2 nMol; pH 7.2 – 6.6) applied to the entire preparation induced arteriolar vasodilation. The maximum change in diameter was 9.9±2.1μm, which was not a maximal dilation in the arteriole (only 71.1% of maximal diameter). Micropipette application of 251.2 nMol H+ (pH 6.6), induced a local arteriolar vasodilation of 3.5±0.4μm however it did not elicit a significant vasodilation 1000μm upstream of the micropipette application site. These novel data demonstrate that H+ is a local arteriolar vasodilator with a limited capacity to cause vasodilation and does not cause a conducted response, which is important in directing blood flow to active skeletal muscle cells during active hyperaemia. These data bring into question the importance of H+ and changes in pH in the active hyperaemic response.

Support or Funding Information

NSERC, Canada
Vascular Function Assessed by Passive Leg Movement and Flow‐Mediated Dilation: Evidence of Construct Validity
Authors: Rossman Matthew J , Groot H. Jonathan , Garten Ryan S , Witman Melissa , Richardson Russell
Published: November 2023

The vasodilatory response to passive leg movement (PLM), often performed in either the upright seated or supine posture, appears to provide a novel, non‐invasive assessment of vascular function. However, PLM has yet to be compared to flow mediated vasodilation (FMD), a test recognized to have prognostic value in terms of vascular health. Therefore, to evaluate the construct validity of PLM, brachial (BA) and superficial femoral (SFA) artery FMDs (Doppler ultrasound) and upright seated and supine PLM were performed in 10 young (22±1) and 10 old (72±2) subjects. During PLM, leg blood flow (LBF, Doppler ultrasound), mean arterial pressure (MAP, Finapres), and leg vascular conductance (LVC, LBF/MAP) were measured, with peak change from baseline LBF (ΔLBF) and LVC (ΔLVC) characterizing the PLM‐induced hyperemic and vasodilatory responses. Age‐associated reductions in vascular function were evident with both PLM and FMD testing. During upright seated PLM, ΔLBF and ΔLVC were significantly correlated with BA (r=0.58 and r=0.69) and SFA (r=0.68 and r=0.64, for ΔLBF and ΔLVC, respectively) FMD. For supine PLM, there was some evidence of a positive relationship between both ΔLBF and ΔLVC and BA FMD (r=0.28 and r=0.3), but ΔLBF and ΔLVC were significantly related to SFA FMD (r=0.53 and r=0.55). Collectively, these data indicate that PLM, particularly upright seated PLM, likely provides a prognostic index of vascular health analogous to the traditional FMD test. Given the relative ease of PLM implementation compared to FMD testing, these data have important implications for the potential clinical use of PLM to assess vascular health.

Support or Funding Information

National Institutes of Health grant PO1 HL 09830, HL‐103786 and HL‐116579 and a VA Merit grant E6910R (RSR)
Effect of mantidis ootheca on the mechanism of vasorelaxation in thoracic aorta
Authors: Kim Hye Yoom , Lee Yun Jung , Ahn You Mee , Tan Rui , Park Ji Hun , Choi Eun Sik , Lee So Heun , Lee Han Sol , Kang Dae Gill , Lee Ho Sub
Published: November 2023

The aim of the present study was to define the effects of aqueous extract of mantidis ootheca (AMO) on the vascular tension and its responsible mechanisms in rat thoracic aortic rings. The eggs of a mantis are enclosed in a foamy pouch. The extracts of AMO‐induced dose‐dependent relaxation of phenylephrine‐precontracted aorta, which was abolished by removal of functional endothelium. Endothelium‐denudation abolished the AMO‐induced vasorelaxation. Pretreatment of the endothelium‐intact aortic rings with NG‐nitro‐L‐arginine methylester (L‐NAME) and 1H‐[1,2,4]‐oxadiazolo‐[4,3‐alpha]‐quinoxalin‐1‐one (ODQ) inhibited the AMO‐induced vasorelaxation. Similarly, wortmannin and LY‐294002, an inhibitors of the phosphatidylinositol 3‐kinase (PI3K), an upstream signaling molecule of eNOS, attenuated the AMO‐induced vasorelaxation. Furthermore, K+ channel inhibition with tetraethylammonium, 4‐AP and glibenclamide had inhibitory effects on the AMO‐induced vasorelaxation. AMO‐induced vascular relaxations were also markedly attenuated by addition of inhibitory muscarinic receptor, atropine and methoctramine. Taken together, the present study suggests that AMO relaxes vascular smooth muscle via endothelium‐dependent activation of NO‐cGMP signaling through the PI3K/Akt‐, possible involvement of K+ channel.
Sprague Dawley Severe Hemorrhagic Shock Model Assessed by Systemic and Microcirculatory Parameters
Authors: Nugent William H , Song Bjorn K
Published: November 2023

Traumatic injury with hemorrhage is a primary cause of death in young, healthy adults with a significant portion of mortality occurring post‐resuscitation. Three factors contribute to outcomes: severity of hemorrhage, duration of hemorrhagic shock (HS), and resuscitation. To evaluate the efficacy of resuscitative intervention, we have developed a severe HS model sensitive to various techniques and solutions of resuscitation. Male Sprague Dawley rats were cannulated with arterial and venous lines to facilitate blood withdrawal, reperfusate infusion, continuous infusion of anesthesia, and monitoring of systemic variables (heart rate, mean arterial pressure [MAP]) with a pressure transducer (MP‐150; BIOPAC Systems Inc., Goleta, CA). The thin spinotrapezius muscle was exteriorized and secured in situ to a thermoregulated platform, which fit onto the stage of an upright microscope (Axio Imager.A2m, Carl Zeiss, Germany) for intravital microscopy of the microcirculation. Phosphorescence quenching microscopy assessed tissue oxygenation (ISF PO2). A 45% estimated blood volume withdrawal rate of 3.5 ml × kg−1 × min−1 produced a rapidly lethal (~1 h) model in sham (no treatment) animals with a good extension of survival times in animals resuscitated with either a non‐oxygen carrying colloid (Hextend™; Hospira, Inc, Lake Forest, IL) or whole blood. Hemorrhage produced a MAP of 20–25 mmHg and a non‐detectable ISF PO2. Following the 30 min HS phase, reperfusion of 20% estimated blood volume with Hextend™ elevated MAP to near baseline and increased survival times to 3 hours. However, Hextend™ did not change ISF PO2, which remained non‐detectable until the animals expired. Reperfusion of 20% estimated blood volume with whole blood elevated MAP back to baseline and elevated tissue ISF PO2 to normoxic levels. Additionally, whole blood extended survival times to 8 hours – the time of experiment termination. In summary, we have developed an in vivo model of severe hemorrhagic shock that allows for simultaneous characterization of systemic and microcirculatory metrics. The model is sufficiently severe to ensure consistency in post‐hemorrhage ischemia and mortality, while providing outcome sensitivity to reperfusion solutions and techniques.

Support or Funding Information

Song Biotechnologies
Effects of Novel Oxygen Therapeutic on Systemic and Microcirculatory Variables in Sprague‐Dawley Rat Isovolemic Hemodilution Model
Authors: Song Bjorn K , Nugent William H , Sayes Christie M , Bruce Erica D
Published: November 2023

Oxygen therapeutics (OTs), sometimes referred to as blood substitutes, have been investigated for almost half a century, and remain actively researched. OT's have traditionally been categorized as either hemoglobin‐based oxygen carriers (HBOCs) or perfluorocarbons (PFCs). Unlike blood, HBOCs and PFCs do not require blood typing, have a long shelf life, do not transmit blood borne diseases, and in most cases do not need refrigeration. Despite the promising attributes of HBOCs and PFCs, there are concerns regarding arteriolar constriction that leads to increased blood pressure and increased white blood cell content for HBOCs and PFCs, respectively. This study investigated the systemic (heart rate, mean arterial pressure) and microcirculatory (vasoactivity, interstitial fluid oxygenation ‐ISF PO2) impact of a novel OT named OX66 (Hemotek LLC, Plano, TX). OX66 is different from HBOCs and PFCs in that it does not contain hemoglobin or long carbon chains; rather, it is a poly‐oxygenated aluminum hydroxide that contains 66.20% oxygen. OX66 is a true clathrate (lattice‐like) structure having large areas within the structure able to capture and hold oxygen. In this study, male Sprague‐Dawley rats underwent a 50% blood volume isovolemic hemodilution exchange with either OX66 or phosphate buffered saline (PBS; volume control), since OX66 was suspended in PBS. Isovolemic hemodilution is the reduction of red blood cells (hematocrit) with an equal volume of hemodiluent, i.e., crystalloids, colloids or OTs. The withdrawal/infusion rate was 2.0 ml × min−1 × kg−1 and performed through a cannulated carotid artery and jugular vein. Systemic measurements were recorded via a cannulated femoral artery that was connected to a pressure transducer (MP150; Biopac Systems, Inc, Goleta, CA), while microcirculatory parameters were collected through phosphorescence quenching and intravital microscopic examination of the exteriorized spinotrapezius muscle. Compared to baseline, a 50% blood volume exchange with either hemodiluent produced a significant reduction in heart rate, blood pressure, arterial diameter and ISF PO2 in all animals. However, OX66 animals demonstrated an improvement in ISF PO2 compared to PBS animals. This finding demonstrates that OX66 both transports and releases oxygen to the peripheral microcirculation.

Support or Funding Information

Support: Baylor University‐Hemotek Medical Applications Project: 32370181
A Dynamical Computational Model of Circulating Blood Gases, Hemoglobin Binding, and pH Regulation in the Microcirculation
Authors: Bassingthwaighte James B , Dash Ranjan K.
Published: November 2023

A new computational model is developed to characterize the dynamical aspects of simultaneous blood‐tissue transport and exchange of O2, CO2, HCO3− and H+ in the coronary microcirculation by accurately simulating the rapidly changing intra‐capillary concentration profiles as the blood gets warmed, acidified, deoxygenated, and carbonated while flowing through the myocardial capillaries. It is based on our 2006 steady‐state, nonlinear, convection‐diffusion‐permeation‐reaction computational model of circulating blood gases, hemoglobin (Hb) binding, and acid‐base buffering in the blood which is available for download from our JSim model repository (http://www.physiome.org/jsim/models/webmodel/NSR/Exchange_O2_CO2_HCO3_and_H+). The kinetics are complicated by: (i) intravascular axial diffusion, (ii) RBC/plasma velocity ratio > 1, (iii) gas and ion permeation through RBC, endothelial and cell membranes, (iv) carbonic anhydrase‐catalyzed CO2 hydration reaction and HCO3‐ buffering in blood and tissue, (v) nonlinear competitive temperature‐dependent binding of Hb, O2, CO2 and H+ inside RBCs, (vi) myoglobin‐facilitated transport of O2 in cardiac cells, and (vii) O2 consumption and CO2 production inside mitochondria, with CO2 production and the respiratory quotient dependent upon the substrates, gas concentrations, cardiac work levels, and the rate of mitochondrial oxidative phosphorylation. These interactions smooth the axial spatial gradations in fluxes of O2 from blood into tissue and of CO2 out into the lung air. The simplest version of this extended model focuses on intracapillary RBC‐plasma interactions and the conditions for alveolar‐arterial differences in oxygen saturation. The cell‐tissue versions incorporate a new kinetic model of cytochrome c oxidase (CcO) that accurately characterizes the kinetics of O2 consumption by CcO as a function of cytochrome c redox fraction, mitochondrial inner membrane electrical and proton gradients (i.e. proton motive force; energy state), O2, and pH. The model serves as the dynamical exchange component of whole body gas exchange and acid‐base regulation. With augmentation to bind 15O2 or 17O2, this model can be used for the analysis of dynamic PET image sequences or NMR studies for estimating the regional tissue O2 consumption.

Support or Funding Information

Supported by NIH grant U01‐HL122199
Rho‐Kinase Inhibition Restores ATP Release from Deoxygenated Red Blood Cells of Older Adult Humans
Authors: Racine Matthew L. , Joyner Michael J. , Dinenno Frank A.
Published: November 2023

Substantial evidence indicates that red blood cells (RBCs) can act as a ‘sensor’ for oxygen demand in addition to their traditional role as an oxygen ‘carrier’, in that they are able to stimulate vasodilation and increase oxygen delivery to the tissue through the release of ATP in direct proportion to the degree of hemoglobin (Hb) deoxygenation. However, RBCs from older adults have an impaired ability to release ATP in response to Hb deoxygenation compared to RBCs from young adults, yet the underlying cause of this remains unknown. RBC membrane deformability has been shown to decrease with advancing donor age, thus we hypothesized that increasing membrane deformability (via the Rho‐kinase inhibitor Y‐27632) would restore ATP release from RBCs of older adults, while decreasing membrane deformability (via the cell‐stiffening agent diamide) would impair ATP release from RBCs of young adults. Isolated RBC ATP release in response to normoxic (PO2 ~110 mmHg) and hypoxic (PO2 ~20 mmHg) stimuli was quantified in RBCs from young (23 ± 3 years; n = 11 for Y‐27632 and diamide) and older (68 ± 7 years; n = 11 and 6 for Y‐27632 and diamide, respectively) adults using the luciferin‐luciferase technique following RBC incubation with either saline (control), Y‐27632, or diamide. The relative change in ATP release from normoxia to hypoxia in saline control conditions was impaired in RBCs from older (41.5 ± 12.1% and 63.5 ± 18.5%) vs. young (110.4 ±12.7% and 146.0 ± 29.3%) adults (P < 0.05; control trials for Y‐27632 and diamide, respectively). Y‐27632 improved RBC ATP release in older adults (98.8 ± 19.8%) such that it was not different from the young control (P > 0.05), whereas diamide decreased RBC ATP release in young adults (57.4 ± 8.4%) such that it was not different from the older control (P > 0.05). Preliminary data from our laboratory indicate that these drugs affected RBC membrane deformability as expected based on other reports of the effects of Y‐27632 and diamide on RBC membrane deformability. We conclude that decreased membrane deformability contributes to the impaired ability of RBCs from older adults to release ATP in response to Hb deoxygenation, and that this may have implications for impaired vascular control with advancing age.

Support or Funding Information

HL095573, HL119337, F31HL126377
Mechanistic Investigation of Reversal of Endothelial Dysfunction in hyperglycemia
Authors: Patel Hemang , Chen Juan , Kavdia Mahendra
Published: November 2023

Progression of endothelial dysfunction has emerged as a significant risk factor for diabetes related complications such as retinopathy, nephropathy, neuropathy and vascular dementia. At elemental level, endothelial dysfunction results from and into disproportionate changes in vasoactive reactive oxygen species (ROS). Superoxide (O2−), nitric oxide (NO) and hydrogen peroxide (H2O2) regulates the redox state of dysfunctional endothelium. Increased O2− levels rapidly react away with NO to reduce its bioavailability, which sets the stage for endothelial dysfunction in hyperglycemia. Simultaneous regulation in H2O2 levels supports either vasoconstrictive or vasorelaxative processes. Several therapeutic strategies had limited success to manage endothelial dysfunction and results from these strategies have shown a lack of understanding of the redox interplay. We studied the regulation of overall redox state and endothelial function during hyperglycemia. We measured O2−, NO, and H2O2 levels using dihydroethidium (DHE), DAF‐FM diacetate and Amplex Red assay, respectively, during sodium nitrite (NaNO2), superoxide dismutase (SOD) and catalase treatments in hyperglycemia treated HUVECs. Hyperglycemia significantly increased O2− and H2O2 levels and decreased NO levels in HUVECs. Treatments to exogenous SOD and NaNO2 decreased O2− levels and significantly increased NO levels in hyperglycemic‐HUVECs. Similarly, treatment with catalase also induced similar changes in O2− and NO levels. Our results suggest that besides the O2− and NO, regulation of H2O2 levels also serves a viable therapeutic alternative for improving the redox state of hyperglycemic‐HUVECs. These results also suggest that production of NO, O2− and H2O2 is tightly regulated in hyperglycemia‐induced endothelial dysfunction. The indirect manipulation of the fate of O2− may hold a key to reverse and manage the hyperglycemic‐endothelial dysfunction.

Support or Funding Information

This study was funded by NIHR01 HL084337
MICROVASCULAR FUNCTION IS IMPAIRED IN ADULTS WHO REPORTED ADVERSE CHILDHOOD EVENTS
Authors: Miguelez Paula Rodriguez , Thomas Jeffrey , Seigler Nichole , Pollock Jennifer S , Harris Ryan A
Published: November 2023

INTRODUCTION

Microvascular function, assessed using the cutaneous microcirculation, has been proposed as a prognostic marker of cardiovascular diseases (CVDs). Adversity early in life, referred to as adverse childhood events (ACEs), has been associated with the development of CVDs during adulthood. Whether or not microvascular dysfunction is present in adults who report ACEs has yet to be investigated.

PURPOSE

This study sought to test the hypothesis that microvascular function is lower in adults who reported ACEs compared to adults who did not.

METHODS

Nineteen young adults (33 ± 2 yrs), with no evidence of overt CVDs, were separated into two groups based on their self‐reported ACE study questionnaire score; HiA: ACE score ≥ 4 (n=9) or NoA: ACE score = 0 (n=10). Augmentation index at 75 bpm (AIx75) was determined by applanation tonometry (SphygmoCor) as an independent predictor of CVD risk. Laser Doppler flowmetry (MoorVMS‐LDF) was used to evaluate microvascular function non‐invasively through the assessment of cutaneous flux (expressed as a perfusion unit (PU)) during baseline (BL), post‐occlusive reactive hyperemia (PORH), local thermal hyperemia (Ht), and iontophoresis of acetylcholine (Ach).

RESULTS

No differences in BL cutaneous flux were observed between groups (HiA: 12.1 ± 15.4 vs. NoA: 7.6 ± 4.8 PU), p=0.387). The PORH response was significantly lower (p=0.039) in HiA (25.1 ± 9.7 PU) compared with NoA (37.0 ± 13.1 PU). Additionally, endothelium‐dependent vasodilation, assessed by the iontophoresis of Ach peak flux relative to maximal Ht flux was significantly lower (p=0.009) in HiA compared to NoA (64 ± 24 vs. 119 ± 52 %, respectively). Further, strong correlations were identified between maximal Ach response and AIx75 (r=−0.475; p=0.040) and between ACE score and both PORH (r=−0.430; p=0.066) and maximal Ach response (r=−0.501: p=0.029).

CONCLUSION

For the first time, we have documented an impaired microvascular response and a reduced endothelium‐dependent cutaneous vasodilation in adults who have reported ACEs compared with adults who did not report any ACE. These findings suggest that the contribution of ACEs to CVDs later in life may be through a reduction in microvascular function.

Support or Funding Information

Support in part by NIH/NHLBI P01HL06999.
The Role of Pericytes for Angiogenesis in Skeletal Muscle Tissue
Authors: Hoier Birgitte , Hellsten Ylva
Published: November 2023

We and others have previously shown that pericytes are invested on most capillaries in skeletal muscle but the physiological role of pericytes in this tissue remains unclear. The aim of the present study was to elucidate whether pericytes contribute to angiogenesis in skeletal muscle by secretion of VEGF and promotion of endothelial cell proliferation.

Pericytes, endothelial cells and skeletal muscle cells were isolated from rat skeletal muscle tissue by use of Dynabeads coupled to specific antibodies and the cells were either characterized for the expression of angiogenic factors or maintained in culture for experiments. Pericytes were found to contain a number of angiogenic factors, e.g. Vascular endothelial growth factor (VEGF), endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor receptor‐2 (VEGFR‐2), Thrombospondin‐1 (TSP‐1), Angiopoietin 1 and 2, (Ang 1 and 2) and platelet‐derived growth factor receptor beta (PDGFR‐β). The expression of VEGF, VEGFR‐2, eNOS, and PDGFR‐β was higher in pericytes conditioned with skeletal muscle media. The muscle conditioned pericytes also released a 7‐fold greater amount of VEGF to the media than untreated control cells. Endothelial cells conditioned with media from pericytes, that first had been conditioned by muscle media, showed an improved rate of proliferation compared to non‐conditioned endothelial cells. Experiments with VEGF neutralizing antibodies added to the conditioned media suggested that VEGF was not responsible for the increased proliferation of endothelial cells.

In conclusion, we have identified a complex pattern of cellular communication in cultures of pericytes, endothelial and skeletal muscle cells whereby skeletal muscle cells activate pericytes and induce an upregulation of several angiogenic factors including VEGF. The activated pericytes release VEGF and other proangiogenic compounds and thereby induce proliferation of endothelial cells and capillary growth. The study strongly suggests that pericytes are important in capillary growth.

Support or Funding Information

The study was supported by The Danish Council for Independent Research‐ Medical Sciences.
Sigma‐1 Receptor Agonists Compromise Barrier Function of Endothelial Cell Monolayers with Differential Impact on Cells Derived from Cardiac or Dermal Microcirculation and Dermal Lymphatics
Authors: Schecker Caitlin , Adderley Shaquria , Breslin Jerome W
Published: November 2023

Sigma‐1 receptors are chaperone proteins in the endoplasmic reticulum known to modulate calcium signaling through the IP3 receptor. We recently demonstrated the presence of these receptors in the lymphatic vessels. In the current study we tested the hypothesis that Sigma‐1 receptors are present and functional in human cardiac microvascular endothelial cells (HCMEC), dermal microvascular endothelial cells (HDMEC) and human dermal lymphatic endothelial cells (HDLEC). Western blotting and immunofluorescence labeling were performed to verify expression of the Sigma‐1 receptor in these cell types. The cells were treated with the selective sigma‐1 receptor agonist, afobazole, or dehydroepiandrosterone (DHEA), which also can activate sigma‐1 receptors. Transendothelial electrical resistance (TER) was used as an index of barrier function. Western blot analysis confirmed expression of the Sigma‐1 receptor in HCMEC, HDMEC, and HDLEC. Immunofluorescence confocal microscopy studies showed that Sigma‐1 localization was largely punctate and perinuclear, with a similar pattern seen for endoplasmic reticulum. In HCMEC or HDLEC, DHEA and afobazole both decreased TER. However, in HDMEC, neither agonist caused a significant decrease in barrier function. The data demonstrates that Sigma‐1 receptors are indeed present in HCEC, HDEC and ADLEC, but functional responses to Sigma‐1 receptor agonists vary between the different endothelial cell sources.

Support or Funding Information

Supported by NIH grant R01HL098215 and APS Frontiers Fellowship.
Sphingosine‐1‐phosphate and 8‐pCPT‐2′‐O‐Me‐cAMP Can Restore Alcohol‐induced Junctional Disorganization and Barrier Dysfunction of Human Brain Microvascular Endothelial Cells
Authors: Alves Natascha Guimaraes , Adderley Shaquria , Yuan Sarah , Breslin Jerome
Published: November 2023

The blood‐brain‐barrier (BBB) is a highly selective endothelial barrier that protects the brain from blood‐born cytotoxins and ensures proper homeostasis of the central nervous system. Alcohol has previously been shown to compromise the BBB, however the mechanisms involved are incompletely understood. We hypothesized that alcohol increases BBB permeability by activating the p38 MAP kinase, disrupting paracellular junction proteins, such as VE‐Cadherin and Claudin‐5, and that microvascular barrier stabilizers such as sphingosine‐1‐phosphate (S1P) and 8‐pCPT‐2′‐O‐Me‐cAMP (8‐CPT) can resolve alcohol‐induced BBB dysfunction. Cultured primary human brain microvascular endothelial cell (HBMEC) monolayers were used to model BBB function so that endothelial‐specific mechanisms could be studied. Trans‐endothelial electrical resistance (TER) was used as an index of barrier function. Localization of VE‐cadherin and Claudin‐5 was determined by immunofluorescence microscopy. S1P (1 μM) or 8‐CPT (100 μM) were applied following treatment with alcohol to test their efficacies to rescue barrier function. The results show that treatment of HBMEC with alcohol (50–100 mM) causes a biphasic hyperpermeability response, characterized by an initial transient decrease in TER (10 min) that recovers, followed by a prolonged decrease in TER that lasts several hours. Surprisingly, we observed that pretreatment with the selective p38 MAPK inhibitor SB208530 (20 μM) does not block the alcohol‐induced decrease in HBMEC monolayer TER. Predictably, treatment of HBMEC monolayers with either S1P or 8‐CPT significantly enhances barrier function. However, pretreatment with S1P or 8‐CPT does not prevent the disruption caused by 75 mM alcohol, and S1P pretreatment can even exacerbate the alcohol‐induced decrease in TER. In contrast, application of S1P or 8‐CPT, after alcohol has already evoked a decrease in TER, causes a hastened recovery to normal HBMEC monolayer barrier function. Our Immunocytochemistry studies demonstrate colocalization of VE‐Cadherin and Claudin‐5 at intercellular junctions, and alcohol disrupts this organization. Importantly, treatment with S1P (after alcohol) can restore junctional integrity. Taken together, our results show that alcohol causes a biphasic decrease in HBMEC monolayer barrier function that does not require p38 MAP kinase activation, but does involve reorganization of junctional protein complexes. In addition, S1P and 8‐CPT can enhance barrier function and may be useful for restoring BBB function in the presence of alcohol intoxication.

Support or Funding Information

Supported by a grant from the USF MCOM Office of Research.
Mitigation of nitrosative stress and inflammation by Vitamin C in cardiomyopathy
Authors: Akolkar Gauri , Bagchi Ashim K , da Silva Dias Danielle , de Angelis Katia , Jassal Davinder , Singal Pawan K
Published: November 2023

Background

Oxidative/nitrosative stress is being linked as a key event in development of various cardiovascular diseases. Increase in the generation of reactive oxygen and nitrogen species is considered as a major trigger for cellular and tissue injury in the heart leading to myocardial dysfunction in general and doxorubicin (Dox) ‐ induced heart failure in particular. Previously we have shown that Vitamin C (Vit C), a very potent antioxidant, reduced Dox‐induced oxidative stress in isolated cardiomyocytes. However the role of Vit C in mitigation of cardiac nitrosative stress and inflammation needs to be understood. The aim of present study is to understand role of Vit C on cardiac nitrosative stress and inflammation.

Methods

Cardiomyocytes isolated from adult Sprague‐Dawley rats were treated with predetermined doses of Vit C (25 μM), Dox (10 μM) or Vit C (25 μM) + Dox (10 μM) for in vitro studies. To understand the effect of Vit C in vivo, adult male Wistar rats were treated with Vit C (50 mg/kg bw orally daily) for 6 weeks or Dox (15mg/kg bw given intraperitoneally in six equal doses for three weeks) or combination treatment with Vit C + Dox. At the end of 6 weeks, animals were euthanized and hearts and blood were collected for further analysis.

Results

Dox treatment increased levels of peroxynitrite in isolated cardiomyocytes and this effect was significantly reduced by Vit C co‐treatment. Both invitro and invivo, Dox induced an increase in levels of Nitric oxide, Nitric oxide synthase (NOS) activity and protein nitrosylation. These in vitro and invivo effects were blocked by Vit C. Dox caused an increased protein expression of inducible NOS which was prevented by Vit C, whereas expression of endothelial NOS was reduced. Treatment with Dox showed increase in levels of inflammatory cytokine TNFα and a decrease in the anti‐inflammatory cytokine IL10. Vit C co‐treatment prevented deleterious effects of Dox by reduction in levels of TNFα and increased levels of IL10.

Conclusion

Based on these results, it is suggested that an attenuation of major triggers of Dox‐induced cardiotoxicity by Vit C would be beneficial in protecting the heart from cardiac side effect of chemotherapy.

Support or Funding Information

CIHR and GETS, University of Manitoba.
Pharmacological Inhibition of NAD(P)H Oxidase with the Peptide gp91ds‐tat During Ischemic Stroke Reduces Infarct Volume and Improves Vascular Reactivity in Obese Zucker Rats
Authors: Brooks Steven D , Branyan Kayla , Asano Shinichi , Hu Hang , Ren Xuefang , Brock Robert W , Frisbee Jefferson C , Chantler Paul D
Published: November 2023

Production of reactive oxygen species (ROS) during ischemic stroke and subsequent reperfusion has been linked to increased infarct size and poor outcomes in multiple rodent models. NAD(P)H) oxidase (NOX) has been identified as a principal source of ROS generation during ischemia/reperfusion, and inhibition of NOX during ischemic stroke results in decreased infarct size and improved outcomes in preclinical models, although underlying mechanisms are unclear. Elevated blood glucose increases ROS production by NOX during ischemia/reperfusion, and vascular dysfunction can impair collateral flow; a stroke model that incorporates hyperglycemia and vascular dysfunction may therefore amplify ROS production and lead to larger infarcts and worsened outcomes. As such, the aim of this study was to determine the effects of gp91ds‐tat, a peptide inhibitor of NOX‐2, on infarct size and vascular reactivity in both the ipsilateral (IL) and contralateral (CL) middle cerebral arteries (MCAs) of lean and obese Zucker rats (LZR, OZR) two weeks after ischemic stroke. Strokes were induced in 17 week old rats for 60 min by transient MCA occlusion (tMCAO); 30 minutes into tMCAO, OZR received either gp91ds‐tat (10mg/kg body weight) or vehicle. Changes in cerebral blood flow were monitored by laser Doppler. 15 days after tMCAO, animals were sacrificed and the IL and CL MCAs were isolated and cannulated in an ex vivo microvessel preparation; MCA reactivity was assessed in response to acetylcholine (ACh). Infarct size was measured by TTC stain. Infarct size and mortality rates were increased at 15 days post‐tMCAO in control treated OZR vs. LZR. Vascular dysfunction was worsened in both CL and IL MCA; ACh‐induced vasodilation was reduced 35% in CL‐MCA compared to non‐stroke OZR MCA, while in IL‐MCA, dilation was completely ablated, likely due to reperfusion injury. In contrast, OZR treated with gp91ds‐tat demonstrated reduced infarct size and mortality, as well as improved recovery of cerebral blood flow after reperfusion vs. control OZR. Vascular reactivity improved significantly, as ACh‐induced dilation in CL‐MCA and IL‐MCA increased 12% and 67% respectively vs. control treated OZR, indicating gp91ds‐tat acutely reduced damage from reperfusion injury. Our findings suggest that preexisting metabolic syndrome can worsen neurological and cerebrovascular outcomes following ischemic stroke though a mechanism at least partially mediated by NOX. Targeted inhibition of NOX during ischemia/reperfusion may improve stroke outcomes, especially in patients with history of metabolic syndrome, and warrants further consideration as a treatment avenue for ischemic stroke.

Support or Funding Information

NIH COBRE GrantAHA Predoctoral Fellowship
Treatment with the Mitochondrial‐targeted Antioxidant Peptide SS‐31 Improves Cerebromicrovascular Function in Aged Mice
Authors: Tarantini Stefano , Hertelendy Peter , Valcarcel‐Ares Marta Noa , Gautam Tripti , Tucsek Zsuzsanna , Csiszar Anna , Ungvari Zoltan
Published: November 2023

Moment‐to‐moment adjustment of cerebral blood flow (CBF) via neurovascular coupling has a critical role in maintenance of healthy cognitive function. In advanced age increased oxidative stress and cerebromicrovascular endothelial dysfunction impair neurovascular coupling, likely contributing to age‐related decline of higher cortical functions. There is increasing evidence showing that mitochondrial oxidative stress plays a critical role in a range of age‐related cellular impairments, but its role in neurovascular uncoupling remains unexplored. The present study was designed to test the hypothesis that attenuation of mitochondrial oxidative stress may exert beneficial effects on neurovascular coupling in aging. To test this hypothesis 24 month old C57BL/6 mice were treated with a cell permeable, mitochondria‐targeted antioxidant peptide (SS‐31; 10 mg/kg/day, s.c.) or vehicle for 2 weeks. Neurovascular coupling was assessed by measuring CBF responses (laser Doppler flowmetry) and evoked field potentials in the somatosensory cortex evoked by contralateral electric whisker pad stimulation. We found that neurovascular coupling responses and ATP‐induced endothelium‐dependent CBF responses were significantly impaired in aged mice. Treatment with SS‐31 significantly improved neurovascular coupling responses, at least in part, by increasing NO‐mediated cerebromicrovascular dilation. These findings are paralleled by the protective effects of SS‐31 on mitochondrial production of reactive oxygen species in cultured cerebromicrovascular endothelial cells derived from aged animals. Thus, mitochondrial oxidative stress contributes to age‐related cerebromicovascular dysfunction. We propose that the antioxidant peptide SS‐31 should be considered for pharmacological microvascular protection for the prevention/treatments of age‐related diseases of the CNS.

Support or Funding Information

This work was supported by grants from the American Heart Association and the National Institute on Aging.
Critical role of telomerase in regulating cerebral vascular function and redox environment
Authors: Ait‐Aissa Karima , Hockenberry Joseph , Gutterman David D , Geurts Aron , Beyer Andreas M
Published: November 2023

Rationale

Cerebrovascular dysfunction has been directly been linked to cerebral micro bleeds and cognitive decline in clinical settings and rodent models of stroke and dementias. Recent clinical studies suggest that oxidative stress may facilitate the development and progression of impaired Flow‐mediated dilation (FMD) in patients with cerebrovascular disease. FMD is the most physiological relevant form of endothelial‐mediated vasodilation. Our laboratory has previously shown that telomerase, a ribo‐nucleoprotein that counteracts telomere shortening, has a protective effect on endothelial function under conditions of oxidative stress in the human microcirculation. For example, in the presence of coronary artery disease, decreased telomerase activity contributes to a shift in the mediator of FMD from atheroprotective nitric oxide (NO) to pro‐inflammatory and atherogenic hydrogen peroxide (H2O2). Endothelial dysfunction in the cerebral vasculature has been directly linked to cerebral microbleeds and cognitive decline in models of stroke and dementia, thus we hypothesized that TERT plays a critical role in maintaining normal NO‐mediated vasodilation in the cerebral vasculature.

Methods

Using Crisp/Cas9 we generated a novel deletion model for TERT (the catalytic subunit of telomerase) in rats on the WKY background. Middle cerebral arteries (MCA) were isolated from wild type (WT) and TERT−/− rats, prepared for videomicroscopy, and FMD was measured. The mediator of FMD was determined using a pharmacological inhibitor of NO synthase (L‐NAME) and a scavenger of H2O2 (PEG‐catalase).

Results

No changes in the magnitude of FMD were observed in MCA from TERT−/− compared to WT rats (max dilation: TERT−/− 71.97±14.7% vs. WT 80.9±10.6%; n≥3). In WT animals, pre‐incubation with L‐NAME abolished FMD while PEG‐catalase had no effect on FMD (max dilation: L‐NAME −5.8±7%*; Peg‐catalase 76.6±8.6%; n≥3; p<0.05). Conversely, in TERT−/− animals FMD was not affected in the presence of L‐NAME but abolished in presence of Peg‐catalase (max dilation: L‐NAME 57.8.0±16.5%; Peg‐catalase −4.3±2.4*%; n≥3; p>0.05 two way ANOVA RM).

Conclusions

Telomerase deficiency causes a switch in the vasoactive mediator of FMD from NO to H2O2, creating a pro‐oxidative environment in the cerebral vasculature. Understanding the regulatory role of telomerase in mediating this mechanistic switch may provide a novel therapeutic strategy for the treatment or prevention of cerebrovascular dysfunction.

Support or Funding Information

R01 HL113612/R21 OD018306
Exercise Mitigates Calpain 1 Induced Cardiac Dysfunction Through PCP4 in Diabetes
Authors: Chaturvedi Pankaj , Kunkel George H , Familtseva Anastasia , Kalani Anuradha , Tyagi Suresh C
Published: November 2023

Calpain 1 is a ubiquitous calcium dependent cysteine protease and found in cytoplasm as well as mitochondria. We have reported earlier that active calpain 1 is translocated from cytosol to mitochondria and activates MMP9. Although studies have reported that activation of calpain 1 is deleterious to the heart in various aspects but there are no reports whether calpain 1 can degrade purkinje cell protein (PCP4) and affect cardiac rhythm in diabetes. Therefore, we hypothesize that calpain 1 degrades PCP4 in diabetes to cause cardiac dysfunction and exercise can mitigate this. To test this hypothesis we recruited four groups of mice 1)db/+ control, 2)db/+ control with exercise, 3)db/db, 4)db/db with exercise. The mice were exercised on treadmill at 7m/min for 300m for 8 weeks and in agreement with the American Veterinary Research Guidelines. HL‐1 atrial cell line for in vitro studies and isolated cardiomyocytes for ex‐vivo studies were used. To evaluate whether activation of calpain can degrade PCP4 we treated the isolated cardiomyocytes with high calcium (40mM) till they lose their shape and evaluated the expression of PCP4, calmodulin, calmodulin kinase and calpastatin with Western blots and real time PCR. The induction of high levels of calpain 1 significantly downregulated PCP4 along with upregulated levels of calmodulin and calmodulin kinase. We used the CRISPR/Cas9 plasmid to knock down calpain in HL‐1 myocytes which restored the levels of PCP4 along with calmodulin and calmodulin kinase. In vivo, we found upregulated levels of calpain 1 in db/db mice as compared to controls which were mitigated in the exercised mice. Exercise also mitigated the levels of PCP4, calmodulin and calmodulin kinase in db/db mice. Conclusively our data strongly suggests that in diabetes there is high induction of calpain 1 with degrades PCP4, a protein important for cardiac rhythm and exercise can mitigate arrhythmias.

Support or Funding Information

Acknowledgements: This work was supported by NIH grants HL‐74185 and HL‐108621 and AHA grant 15POST23110021
Aging increases Reactive Oxygen Species and Interleukin‐17 concentrations in Peroxisome Proliferator Activated Receptor –alpha (PPAR‐α) knockout mice during Angiotensin II Hypertension through a NADPH Oxidase Subunit‐2 mechanism
Authors: Ananaba Ugoeze , Williams Nia C , Allard Joanne S , Lee Dexter L
Published: November 2023

Aging decreases myocardial peroxisome proliferator activated receptor –alpha (PPAR‐α) expression. Activation of PPAR‐α reduces hypertension, cellular oxidative damage and inflammatory cytokine production. We hypothesize that the absence of PPAR‐α would increase mean arterial pressure (MAP) during Angiotensin (Ang II) hypertension in aged mice hearts by increasing the expression of NADPH (NOX) subunit‐2, reactive oxygen species and interleukin‐17 (IL‐17) production. Male (40 weeks old) PPAR‐α knockout (KO) mice and wild‐type (WT) mice were implanted with biotelemetry devices and infused with a slow pressor dose of Ang II (400 ng/kg/min) for 12 days. Separate groups of Ang II treated mice were given the PPAR‐α agonist, fenofibrate (Fen) (145 mg/kg/day). On day 12 of Ang II, MAP was increased in KO compared to WT, 163 ± 6 and 148 ± 3 mmHg, respectively. Heart expression of NOX‐2 was increased in Ang II‐treated KO (90 ± 10%) and WT (80 ± 8%) mice, when compared to no treatment controls. WT + Ang II + Fen NOX‐2 expression was decreased (27 ± 6%) when compared to WT + Ang II alone. TBARS concentration was increased in KO + Ang II (12.4 ± 1 mM), when compared to WT + Ang II (10 ± 1 mM). IL‐17 concentrations were increased in KO + Ang II (1.7 ± 0.5 ng/mL) and WT + Ang II (1.2 ± 0.2 ng/mL), when compared to their respective controls. Our results suggest increased myocardial NOX‐2 expression, TBARS and increased IL‐17 concentrations in 40 week old PPAR‐α KO mice are potential mechanisms for the increased Ang II‐induced blood pressure responses.

Support or Funding Information

#1R25AGO47843‐01