Endothelin-1 Levels Research Articles

Endothelin-1 Correlates with Blood Pressure and Pulse Pressure in Females with Lupus Associated Hypertension

Systemic Lupus Erythematosus (SLE) is a common autoimmune disease that is classified by IgG complex deposition in end organs including the kidney, heart, and vasculature. SLE disproportionally affects women to men (9:1 ratio), with a high prevalence in African American women. Importantly, patients with an SLE diagnosis have common cardiovascular complications including hypertension. Importantly, numerous studies have reported that plasma and urinary endothelin-1 (ET-1) levels are elevated in patients with SLE or hypertension. However, few studies have investigated the role of the synergistic effects of SLE and hypertension on ET-1. Therefore, we hypothesized that women with SLE and hypertension would have elevated plasma ET-1 levels compared to SLE or hypertension alone. To investigate this, we recruited non-SLE non-HTN controls (n=8), non-SLE HTN (n=7), SLE non-HTN (n=5), and SLE-HTN (n=6) female human subjects and analyzed clinical and demographic characteristics, plasma ET-1, c-reactive protein (CRP), soluble vascular adhesion molecule-1 (sVCAM-1), and anti-Smith autoantibodies. We found that plasma ET-1 levels are significantly higher in African American compared to European American patients independent of SLE or blood pressure status (p=0.0182). When plasma ET-1 levels were correlated with age and body mass index (BMI), ET-1 had a positive association with age (r=0.4861;p=0.0118), but no correlation with BMI (r=0.2137;p=0.2945). When subjects we characterized by SLE diagnosis, we found that female subjects with SLE had elevated plasma ET-1 levels compared to non-SLE controls (p=0.0141). Next, we stratified subjects by blood pressure status and found that only SLE-HTN subjects had significantly elevated plasma ET-1 levels compared to non-SLE non-HTN controls (p=0.0005), and a trend for elevated ET-1 levels over SLE non-HTN (p=0.0799). Lastly, we correlated plasma ET-1 levels to offce systolic blood pressure, diastolic blood pressure and calculated pulse pressure. We found that plasma ET-1 positively associated with systolic blood pressure (r=0.5621;p=0.0035) and pulse pressure (r=0.5810;p=0.0023) but not diastolic blood pressure (r=0.3177;p=0.1217). These data demonstrate that plasma ET-1 levels associate with SLE diagnosis and blood pressure status in our cohort of female SLE human subjects. Further investigation of ET signaling should be done in SLE and SLE-HTN human subjects. Veterans Affairs Biomedical Laboratory Research & Development Career Development Award (CDA-2) 1IK2BX005605-01 to J.P.V.B, Medical University of South Carolina College of Medicine Program Project Grant to J.C.O and J.P.V.B., and Medical University of South Carolina Core Centers for Clinical Research Project Award P30AR072582 to M.A.C and J.P.V.B. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

A novel dual inhibitor of the angiotensin and endothelin systems for the treatment of hypertension

Hypertension, affecting nearly half of the US adult population, is a primary cause of morbidity and mortality worldwide. Despite the availability of several classes of antihypertensive medications, only 25% of patients have their blood pressure under control, highlighting the need for novel antihypertensive therapeutics with better effcacy. Among hypertensive cases, salt-sensitive and drug-resistant hypertension are particularly concerning, as they are unresponsive to a combined regimen of three or more classes of medications, including angiotensin II (Ang II) receptor blockers (ARBs), angiotensin-converting enzyme inhibitors (ACEis), and diuretics. Salt-sensitive, observed in 50% of the general and 73% of black hypertensive patients, disproportionately affects the latter, leading to significantly higher mortality. Salt-sensitive hypertension in humans, as well as in various experimental models of hypertension (e.g., Ang II-induced, Dahl-salt sensitive [DSS] rats, deoxycorticosterone acetate [DOCA]-salt hypertensive rats), is characterized by significantly elevated levels of plasma endothelin-1 (ET-1), oxidative stress, and kidney dysfunction. ET-1 is often considered a final common pathway in orchestrating cardiovascular dysfunction through a range of proinflammatory mediators such as Ang II, cytokines, hypoxia, thrombin, reactive oxygen species (ROS), oxidized LDL, and vasopressin, all of which stimulate ET-1 production. In this capacity, we report the discovery of a gut microbiota-derived metabolite, indole-3-acetic acid (IAA), that simultaneously blocks ET-1 receptors (ETAR and ETBR) and the ET-biosynthetic enzyme ECE-1. Our data demonstrate that IAA inhibits ET-1-induced constriction of resistance mesenteric arteries, ROS production in human aortic smooth muscle cells, activation of cultured endothelial cells, adhesion molecule expression, and leukocyte adhesion. IAA inhibited purified human ECE-1 and suppressed Ang-II-induced and hypoxia-elicited ET-1 production from endothelial cells in vitro. Administration of a once-daily, extended-release formulation of IAA inhibits blood pressure elevation and kidney dysfunction in both DSS and Ang II-induced hypertension. IAA treatment preserved endothelial nitric oxide (NO) production and maintained smooth muscle sensitivity to a NO donor, sodium nitroprusside. Furthermore, IAA inhibited the expression of ROS-associated proteins such as Nox2 and Nox4, reduced oxidative damages, and decreased the production of ET-1 in vivo. Overall, IAA is the first compound that, through novel dual targeting of both the angiotensin and endothelin systems, demonstrates profound antihypertensive action in both salt-sensitive and angiotensin II-induced hypertension. Thus, IAA has the potential to effectively treat hypertension that does not respond to conventional therapies. American Heart Association. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Endothelin-1 stimulated human preadipocyte growth through the AMPK, ERK, cJUN, PKC, and STAT3 pathways

High level of plasma endothelin (ET)-1 was noticed in obesity, and it was found to regulate adipogenesis and the endocrine activity of fat cells. However, relatively little is known about the ET-1 signaling pathway in preadipocyte growth. Our previous reports have shown that ET-1 and ET-3 stimulated murine 3T3-L1 preadipocyte growth through different kinase signaling cascades with ET type A receptor (ETAR) but not ETBR dependency. In this study, the hypothesis arose that ET-1 may regulate human visceral white preadipocyte growth through the AMPK, ERK, cJUN, PKC, and STAT3 pathways with ET receptor-specific dependency. As indicated by an increased number of cells and greater conversion of 3-4,5-dimethylthiazol-2-yl-2,5 diphenyl tetrazolium bromide (MTT) to formazan crystals by living cells, the stimulation of human preadipocyte growth by ET-1 depended on concentration. Neither ET-2 nor ET-3 stimulated human preadipocyte growth. When signaling pathways were examined, ET-1 signaling stimulated phosphorylation of AMPK, ERK, cJUN, PKC, and STAT3, proteins but not AKT, JNK, or p38 MAPK. Treatment with an ETAR antagonist, such as BQ610, or an ETBR antagonist, such as BQ788, blocked the ET-1-induced increase in cell growth and phosphorylated levels of AMPK, PKC, and STAT3 proteins. However, treatment with BQ788 but not BQ610 blocked the ET-1-increased levels of ERK and c-JUN protein phosphorylation. In addition, pretreatment with specific inhibitors of AMPK (compound C), ERK1/2 (U0126), JNK (SP600125), JAK2/STAT3 (AG490), or PKC (Ro318220) prevented the ET-1-induced increase in cell growth and reduced the ET-1-stimulated phosphorylation of AMPK, ERK1/2, c-JUN, STAT3, and PKC. However, neither the p38 MAPK antagonist nor the ceramide synthase inhibitor altered the effect of ET-1. The results indicate that the target molecules of ETAR and ETBR signaling pathways transducing ET-1 growth signals in human visceral preadipocytes can be similar (e.g., AMPK, JAK2/STAT3, and PKC) or different (e.g., ERK1/2 and JNK/c-JUN). This work was supported in part by the National Science and Technology Council, Taiwan, Grant NSTC 112-2320-B-008-001 to YHK and the Taoyuan Armed Forces General Hospital, Taiwan, Grant AFTGH to PJH, CCK, LJS, and YHK. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Role of local vasoconstrictors in diving performance of two pinniped species

Diving mammals extend submergence time through bradycardia and reduced peripheral perfusion to certain tissues. Thus, local blood flow regulation under hypoxia may be an essential physiological strategy that fine-tunes metabolic expenditures during breath-hold. We focused on a potent local vasoconstrictor, Endothelin-1 (ET1) and its receptors ETR-A and ETR-B, to determine their potential role in mammalian diving. We hypothesized that less perfused organs (kidney) would have higher signatures of local vasoconstriction (i.e. higher ET1 and receptors), compared to organs known to have higher relative perfusion underwater (heart/left ventricle). Additionally, we compared pinniped species with higher (Weddell seal) and lower (California sea lion) relative dive capabilities. Differences in mRNA expression of vasoconstrictive elements were assessed by 2-way ANOVA to compare tissue and species. In adult animals (n=7 both species) there were significant differences in expression of ET1 between species (F1,16=9.2, p=0.08) and tissues (F1,16=18.4, p=0.0006), with sea lions having consistently higher levels of ET1. ET1 expression was highest in the kidney, with sea lion kidney expressing 25x more ET1 mRNA than heart (posthoc p=0.0007). The expression ratio of ET A and B receptors also differed across tissue (F1,16=19.83 p=0.0004) with B/A highest in the kidney in both species. This is counter to expectations that submergence capability would relate to vasoconstriction potential across species, but supports the hypothesis that within a species, the less-perfused kidney should demonstrate higher potential for local vasoconstriction compared to the heart. Relative expression of ET receptors between tissues also runs counter to our hypothesis. In both divers, the heart expresses relatively more ETR-A, which has a critical role in vasoconstrictive signaling. On the other hand, ETR-B can signal for either vasoconstriction or vasodilation depending on its location within the endothelium or smooth muscle. Kidney in both species expresses relatively more ETR-B, indicating potential for nuanced vasoregulatory control through the endothelin pathway in this tissue. Receptor localization will be determined by immuno-staining to investigate functional differences between tissues. We also investigated the development of diving by comparing the same biomarkers in adults versus pups (n=10 seals, n=3 sea lions) using a separate 2-way ANOVA for each species. We expected that signatures of vasoregulatory potential would be less evident in the novice diving pups. Concordantly, ET1 is lower in sea lion pups than adults (F1,12=5.097, p=0.043); ET1 mRNA is 23x lower in the pup kidney compared to adult (posthoc p=0.02). Receptor expression followed the same patterns in both species, with the highest expression of ETR-A noted in the pup heart, indicating high vasoconstrictive potential in pups, but through tissue specific mechanisms. These results point to differences in the control of local vasoconstriction that occur with development and highlight the physiological components essential to the development of diving. Funding by NSF #2020706. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Profound Sex Differences in the Kidney Expression of Mitochondria-Related Genes in Endothelin-1 Receptor B (ETB) Deficient Rats Fed a High Salt Diet

Elevated levels of endothelin-1 (ET-1), overactivation of the ETA receptor and oxidative stress are critical in the development of hypertensive kidney disease. Females are more resistant than males to kidney damage; however, it is unclear if overactivation of the ETA receptor disrupts mitochondrial homeostasis similarly in both sexes. With these studies we aimed to determine if overactivation of the ET-1/ETA axis has a differential effect on mitochondrial homeostasis in the male and female kidney during high salt consumption. Male and female ETB deficient (ETB def) and transgenic (TG) control rats were placed on a high salt diet (4.0% NaCl) for 3 weeks. Kidneys were collected and the expression of 84 mitochondria-related genes in renal cortex and outer medulla was assessed with qRT-PCR arrays. In response to high salt, male ETB def rats displayed cortical overexpression of 13 genes when compared to male TG controls, while only 7 genes were upregulated in cortex of female ETB def rats compared to female TG controls. Genes upregulated in male ETB def rats are involved in apoptosis, mitochondrial biogenesis and transport, energy metabolism and ROS handling (selected genes, male ETB def vs. TG controls, n=3/group: Ucp1 (19-fold increase, p=0.03), Bnip3 and Scl25a21 (6-fold, p=0.003 and p=0.004, respectively), and Mfn1 (5-fold, p=0.03). Genes upregulated in females are involved in apoptosis, ROS handling and mitochondrial transport (selected examples, female ETB def vs. TG control, n=3/group: 2-fold increase in Sod1 (p=0.03), Akt1 (p=0.003) and Grpel1 (p=0.019). No common genes were changed between the sexes. When comparisons were made in the outer medulla, we found that deficiency of the ETB receptor in males resulted in a ≥30% reduction in the expression of 29 genes involved in apoptosis, mitochondrial biogenesis and transport, energy metabolism, ROS handling and other mitochondrial functions (p<0.05). On the other hand, the expression of 8 genes involved in mitochondrial transport and apoptosis was reduced by ≥80% in the female ETB def rat outer medulla vs. TG controls, while 3 genes involved in mitochondrial transport and energy metabolism had a 2-fold overexpression (p<0.05). Altogether, our results demonstrate that salt-sensitive hypertension affects mitochondrial homeostasis differentially in distinct regions of the kidney, and that the male and female kidney respond to salt in a disparate manner: with overactivation of the ET-1/ETA axis resulting in greater dysregulation of mitochondria-related genes in the male versus the female kidney during a high salt diet. Funded by UAB-UCSD O’Brien Center Summer Research Pathway for Students P30 DK079337 and KURE R25 DK115353 to BSM, NIGMS UAB CORD BBRT to GJC, and NIH K01HL145324 and Sapphire North America Return to Science Research grant to CDM. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Sex-specific role for the clock protein PER1 in renal ET-1 regulation in salt-sensitive hypertension

Global knockout (KO) of the clock protein PERIOD1 (PER1) in male, but not female, C57BL/6J mice displayed both blunted blood pressure and urinary endothelin-1 (ET-1) rhythms. More recently, we showed that Dahl salt-sensitive (SS) Per1 KO male rats had exacerbated SS hypertension, alongside increased renal injury, renal ET-1 gene ( Edn1) expression, and ET-1 peptide expression from kidneys collected during the rat’s inactive period (2 PM). Additionally, we see increased long non-coding RNA EDN1-AS, which has been suggested to modulate ET-1 production. However, it is unknown whether PER1 regulates rhythms of ET-1 in SS rats and whether PER1 regulates ET-1 in a sex-dependent way. The study objective was to test the hypothesis that PER1 negatively regulates renal EDN1-AS and Edn1 to modulate ET-1 in a sex- and time-dependent manner. To test this hypothesis, kidneys were collected during rat’s active period (2 AM) from male and female SS Per1 KO and SS control rats on a normal salt (0.4% NaCl) or 3 weeks high salt (4% NaCl) diet ( N=5-6). Renal ET-1 protein levels were measured by ELISA and semi- and quantitative PCR was performed to quantify the levels of EDN1-AS and Edn1. Renal ET-1 levels were significantly higher at 2 AM than our previously reported data at 2 PM in both SS Per1 KO and control male rats ( p<0.0001). Renal ET-1 levels were significantly increased in male SS Per1 KO rats following a high salt diet ( p=0.0362) but not in females ( p=0.9522). This was also reflected in the Edn1 mRNA levels (male, p=0.0005; female, p=0.0814). EDN1-AS levels were also increased in males ( p=0.0123) but not in females. Interestingly, significant sex differences were seen in EDN1-AS levels, where EDN1-AS levels were higher in males than females following both low and high salt diets, suggesting a potential novel sex-dependent mechanism for ET-1 regulation. Together, these data suggest sex-specific action of PER1 in the regulation of ET-1 in a model of salt-sensitive hypertension. Future work aims to investigate blood pressure and renal injury in female SS Per1 KO rats and whether this apparent negative regulation of ET-1 by PER1 contributes to the SS hypertensive and renal injury phenotype seen in male SS Per1 KO rats. This work was supported by the National Institutes of Health grants R35 HL135749 (to A.S.), R01 342 DK109570 (to M.G. and A.S.), and Department of Veteran Affairs grants I01 BX004024 (to A.S.). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

RENAL PROTECTIVE EFFECT AND CLINICAL ANALYSIS OF VITAMIN B 6 IN PATIENTS WITH SEPSIS.

Objective: To investigate the protective effect and possible mechanisms of vitamin B 6 against renal injury in patients with sepsis. Methods: A total of 128 patients with sepsis who met the entry criteria in multiple centers were randomly divided into experimental (intravenous vitamin B 6 therapy) and control (intravenous 0.9% sodium chloride therapy) groups based on usual care. Clinical data, the inflammatory response indicators interleukin 6 (IL-6), interleukin 8 (IL-8), tumor necrosis factor (TNF-α), and endothelin-1 (ET-1), the oxidative stress response indicators superoxide dismutase, glutathione and malondialdehyde, and renal function (assessed by blood urea nitrogen, serum creatinine, and renal resistance index monitored by ultrasound) were compared between the two groups. Results: After 7 d of treatment, the IL-6, IL-8, TNF-α, and ET-1 levels in the experimental group were significantly lower than those in the control group, the oxidative stress response indicators were significantly improved in the experimental group and the blood urea nitrogen, serum creatinine, and renal resistance index values in the experimental group were significantly lower than those in the control group ( P < 0.05). There was no statistical difference between the two groups in the rate of renal replacement therapy and 28 d mortality ( P > 0.05). However, the intensive care unit length of stay and the total hospitalization expenses in the experimental group were significantly lower than those in the control group ( P < 0.05). Conclusion: The administration of vitamin B 6 in the treatment of patients with sepsis attenuates renal injury, and the mechanism may be related to pyridoxine decreasing the levels of inflammatory mediators and their regulation by redox stress.

Association of endothelial dysfunction and peripheral arterial disease with sarcopenia in chronic kidney disease.

Endothelial dysfunction and peripheral arterial disease (PAD), which disturb skeletal muscle microperfusion, are highly prevalent in patients with chronic kidney disease (CKD). We evaluated the association of endothelial dysfunction and PAD with sarcopenia in patients with non-dialysis CKD. This cross-sectional study included 420 patients with stages 3-5 non-dialysis CKD aged 69.0±11.8years. Skeletal muscle index (skeletal muscle mass/height2), handgrip strength, 6-m gait speed and strength of hip flexion and knee extension were measured. Sarcopenia was defined according to the Asian Working Group for Sarcopenia 2019. Endothelial dysfunction and PAD were assessed using the vascular reactivity index (VRI) and ankle-brachial index (ABI), respectively. A VRI<1.0 was classified as poor endothelial function, and an ABI<0.9 was defined as PAD. Additionally, endothelial and inflammatory biomarkers, including intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), asymmetric dimethylarginine, endothelin-1 (ET-1) and interleukin-6, were measured in a subgroup of 262 patients. Among the participants, 103 (24.5%) were classified as having sarcopenia. Compared with patients without sarcopenia, those with sarcopenia had significantly lower ABI (1.04±0.16 vs. 1.08±0.15, P=0.028 for the right ABI; 1.01±0.16 vs. 1.06±0.16, P=0.002 for the left ABI) and VRI (0.83±0.57 vs. 1.08±0.56, P<0.001) and had higher serum levels of ICAM-1 (P<0.001), VCAM-1 (P=0.003) and ET-1 (P=0.037). Multivariate logistic regression revealed that, beyond age and body mass index, the average ABI (odds ratio [OR]: 0.81/0.1 increase; 95% confidence interval [CI]: 0.67-0.98; P=0.032) and VRI (OR: 0.93/0.1 increase; 95% CI: 0.88-0.98; P=0.010) were independently associated with sarcopenia. Among the endothelial biomarkers measured, ICAM-1 (OR: 2.47/1-SD increase; 95% CI: 1.62-3.75) and VCAM-1 (OR: 1.91/1-SD increase; 95% CI: 1.27-2.87) were independent predictors of sarcopenia. Group stratification based on the cut-offs of VRI and ABI showed that those with both poor VRI and ABI had the greatest risk for sarcopenia (OR: 4.22; 95% CI: 1.69-10.49), compared with those with normal VRI and ABI. Endothelial dysfunction and PAD are independently associated with sarcopenia in patients with stages 3-5 CKD, suggesting the dominant role of vascular dysfunction in sarcopenia.

Genetic associations of cardiovascular risk genes in European patients with coronary artery spasm.

Coronary artery spasm (CAS) is a frequent finding in patients presenting with angina pectoris. Although the pathogenesis of CAS is incompletely understood, previous studies suggested a genetic contribution. Our study aimed to elucidate genetic variants in a cohort of European patients with angina and unobstructed coronary arteries who underwent acetylcholine (ACh) provocation testing. A candidate association analysis of 208 genes previously associated with cardiovascular conditions was performed using genotyped and imputed variants in patients grouped in epicardial (focal, diffuse) CAS (n = 119) and microvascular CAS (n = 87). Patients with a negative ACh test result (n = 45) served as controls. We found no association below the genome-wide significance threshold of p < 5 × 10-8, thus not confirming variants in ALDH2, NOS3, and ROCK2 previously reported in CAS patients of Asian ancestry. However, the analysis identified suggestive associations (p < 10-05) for the groups of focal epicardial CAS (CDH13) and diffuse epicardial CAS (HDAC9, EDN1). Downstream analysis of the potential EDN1 risk locus showed that CAS patients have significantly increased plasma endothelin-1 levels (ET-1) compared to controls. An EDN1 haplotype comprising rs9349379 and rs2070698 was significantly associated to ET-1 levels (p = 0.01). In summary, we suggest EDN1 as potential genetic risk loci for patients with diffuse epicardial CAS, and European ancestry. Plasma ET-1 levels may serve as a potential cardiac marker.

Impact of Forced Swimming Stress on Serum Adiponectin and Endothelin-1 Levels in Wistar Rats: Comparative Analysis of Dietary Effects.

Aim This study aimed to assess the impact of forced repeated swimming stress on serum adiponectin and endothelin-1 levels in Wistar rats, comparing the effects between those fed a standard diet and those on a high-fat diet. Methods Twenty adult male Wistar rats were divided into two dietary groups: a standard food diet group (NFD, n=10) and a high-fat diet group (HFD, n=10). Both groups underwent daily forced swimming stress for six days, with durations increasing from 5 to 30 minutes. The protocol finished in an acute bout of swimming exercise on the seventh day with a duration of 40 minutes. Adherence to ethical guidelines was strictly maintained, and serum adiponectin and endothelin-1 levels were measured pre- and post-exercise using the ELISA method. Results Before swimming, the mean adiponectin levels were 4.30±1.50 ng/mL in the NFD group and 3.53±0.70 ng/mL in the HFD group. Post-exercise, these levels significantly decreased to 2.4±0.84 ng/mL (p=0.003) and 1.58±0.23 ng/mL (p=0.001), respectively. Endothelin-1 levels also showed significant decreases from 0.86 pg/mL (0.74-0.87) to 0.49 pg/mL (0.43-0.62) (p=0.003) in the NFD group, and from 0.89 pg/mL (0.86-0.93) to 0.69 pg/mL (0.60-0.75) (p=0.027) in the HFD group after swimming. Conclusion The study highlighted the significant effects of forced swimming stress on lowering serum adiponectin and endothelin-1 levels in Wistar rats, with more pronounced decreases observed in rats on a high-fat diet. The results of the study suggest the potential of exercise as a crucial component of strategies aimed at managing obesity and improving cardiovascular health, emphasizing the interaction between physical stress and dietary factors on metabolic and cardiovascular biomarkers.

Clinical and experimental parallels in the assessment of microcirculatory dysfunctions of the lungs in thoracic radiology

Introduction. Chronic obstructive disease (COPD) and thromboembolism of the branches of the pulmonary artery or pulmonary embolism (PE) remain the most common and socially significant lung diseases. Circulatory disorders in these pathologies play a critical role in the development of the disease. Radiation techniques, such as computed tomography/angiography and single-photon emission computed tomography (SPECT), are the leading ones in recognizing changes in the lung parenchyma and vessels. Many questions can be successfully solved experimentally. The main radiation techniques used to study the vascular bed when modeling lung diseases in experimental animals are microangiography and perfusion scintigraphy. Purpose. To show the capabilities of radiation diagnostic methods for identifying vascular disorders in COPD and PE in experiments and clinical manifestations. Material and methods. A clinical and experimental comparison of pulmonary circulation changes during COPD and PE was carried out based on the results of perfusion scintigraphy/SPECT in 55 rats with simulated COPD- like condition and 622 patients with COPD of varying severity, as well as 58 rats with simulated PE and 180 patients with PE. Results. When modelling lung diseases (PE and COPD), the results similar to blood circulation and structural changes in the lung parenchyma in patients with these nosologies were obtained. In COPD, changes in the lung parenchyma correlated with disturbances in functional indicators (microcirculation, PaO2, PaCO2, DSLzd, DLco/VA, SDPA) and increased levels of endothelin-1 (r=0.72) and endothelial growth factor (VEGF-A) (r=0.79). Analysis of experimental and clinical data on PE showed the importance of identifying minor forms of thromboembolism. Conclusion. 1. Persistent microcirculatory dysfunctions in the pulmonary circulation play an important role in the pathogenesis of chronic broncho-obstructive pathology. After just 7 days, initial signs of emphysema can be detected in the ischemic area of the lung tissue, and by the 60th day typical bullous cavities can be detected. 2. In the chronic course of minor forms of thromboembolism of small branches of the pulmonary artery, local emphysema is formed. 3. At the early stages of COPD formation, regional zones of emphysematous restructuring of the lung tissue with a predominance of ventilation dysfunctions and zones of «compressive ischemia» with prevailing vascular disorders are formed. As the pathological process progresses, the ischemia factor acquires independent significance in the pathogenesis of the disease.

Recombinant hirudin attenuates pulmonary hypertension and thrombosis in acute pulmonary embolism rat model.

Acute pulmonary embolism (APE) is classified as a subset of diseases that are characterized by lung obstruction due to various types of emboli. Current clinical APE treatment using anticoagulants is frequently accompanied by high risk of bleeding complications. Recombinant hirudin (R-hirudin) has been found to have antithrombotic properties. However, the specific impact of R-hirudin on APE remains unknown. Sprague-Dawley (SD) rats were randomly assigned to five groups, with thrombi injections to establish APE models. Control and APE group rats were subcutaneously injected with equal amounts of dimethyl sulfoxide (DMSO). The APE+R-hirudin low-dose, middle-dose, and high-dose groups received subcutaneous injections of hirudin at doses of 0.25 mg/kg, 0.5 mg/kg, and 1.0 mg/kg, respectively. Each group was subdivided into time points of 2 h, 6 h, 1 d, and 4 d, with five animals per point. Subsequently, all rats were euthanized, and serum and lung tissues were collected. Following the assessment of right ventricular pressure (RVP) and mean pulmonary artery pressure (mPAP), blood gas analysis, enzyme-linked immunosorbnent assay (ELISA), pulmonary artery vascular testing, hematoxylin-eosin (HE) staining, Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining, immunohistochemistry, and Western blot experiments were conducted. R-hirudin treatment caused a significant reduction of mPAP, RVP, and Malondialdehyde (MDA) content, as well as H2O2 and myeloperoxidase (MPO) activity, while increasing pressure of oxygen (PaO2) and Superoxide Dismutase (SOD) activity. R-hirudin also decreased wall area ratio and wall thickness to diameter ratio in APE rat pulmonary arteries. Serum levels of endothelin-1 (ET-1) and thromboxaneB2 (TXB2) decreased, while prostaglandin (6-K-PGF1α) and NO levels increased. Moreover, R-hirudin ameliorated histopathological injuries and reduced apoptotic cells and Matrix metalloproteinase-9 (MMP9), vascular cell adhesion molecule-1 (VCAM-1), p-Extracellular signal-regulated kinase (ERK)1/2/ERK1/2, and p-P65/P65 expression in lung tissues. R-hirudin attenuated pulmonary hypertension and thrombosis in APE rats, suggesting its potential as a novel treatment strategy for APE.

Hemp seeds attenuate loperamide-induced constipation in mice.

Constipation is a common gastrointestinal disease that seriously affects human physical and mental health. Studies have reported that hemp seeds can improve constipation, however the specific mechanism is still unclear. This study investigates that hemp seed (HS) and its water-ethanol extract (HSE) attenuates loperamide-induced constipation in mice. The research results show that: the fecal water content and small intestinal transit rate of mice in the hemp seed group and hemp seed hydroalcoholic extract group were significantly increased compared with MC group, and the first red feces defecation time was significantly shortened; HS and HSE significantly influence serum levels of Gastrin (Gas), motilin (MTL), substance P (SP), and endothelin (ET), potentially mediating their effects on gastrointestinal motility. HS and HSE can improve colon inflammation in constipated mice with H&E staining. Compared with the model of constipation group, the content of short-chain fatty acids in the HS group and HSE group increased significantly. Gut microbiome studies have shown that the structure and abundance of intestinal flora are altered. HS and HSE changed the abundance of Odoribacter, Bacteroide, Lactobacillus and Prevotella. Together, these results suggest that HS have the potential to stimulate the proliferation of beneficial gut microbes and promote intestinal motility, thereby improving gut health and relieving symptoms of constipation.

Endothelin Level in Menopausal Women with Comorbidity of the Acute Phase Middle Severity COVID-19 with Arterial Hypertension and Type 2 Diabetes Mellitus.

The levels of endothelins were assessed in menopausal women with arterial hypertension (AH) and type 2 diabetes mellitus (T2DM) in the acute phase of the moderate COVID-19. Women under observation (age 45-69 years) were divided into two groups. Control group consisted of women (n=16) who did not have COVID-19, were not vaccinated, and had no antibodies to SARS-CoV-2 (IgG). The main group included women (n=63) in the acute phase of the moderate COVID-19 accompanied by pneumonia. According to the clinical and anamnestic data analysis, the main group was divided into subgroups: without AH and T2DM (n=21); with AH and without T2DM (n=32); and with AH and T2DM (n=10). The parameters of clinical blood analysis, as well as endothelin-1, endothelin-2, and endothelin-3 levels were assessed. In women with a moderate COVID-19, the endothelin-1 and endothelin-2 levels were increased compared to the control regardless of AH and T2DM status. We found no statistically significant differences in the studied parameters of endothelial dysfunction between the subgroups of menopausal women in the acute phase of the moderate COVID-19.

546 Using Contingency Management to Understand the Cardiovascular, Immune and Psychosocial Benefits of Reduced Cocaine Use: A Protocol for a Randomized Controlled Trial

OBJECTIVES/GOALS: Contingency management (CM) procedures yield measurable reductions in cocaine use. This poster describes a trial aimed at using CM as a vehicle to show the biopsychosocial health benefits of reduced use, rather than total abstinence, the currently accepted metric for treatment efficacy. METHODS/STUDY POPULATION: In this 12-week, randomized controlled trial, CM was used to reduce cocaine use and evaluate associated improvements in cardiovascular, immune, and psychosocial well-being. Adults aged 18 and older who sought treatment for cocaine use (N=127) were randomized into three groups in a 1:1:1 ratio: High Value ($55) or Low Value ($13) CM incentives for cocaine-negative urine samples or a non-contingent control group. They completed outpatient sessions three days per week across the 12-week intervention period, totaling 36 clinic visits and four post-treatment follow-up visits. During each visit, participants provided observed urine samples and completed several assays of biopsychosocial health. RESULTS/ANTICIPATED RESULTS: Preliminary findings from generalized linear mixed effect modeling demonstrate the feasibility of the CM platform. Abstinence rates from cocaine use were significantly greater in the High Value group (47% negative; OR = 2.80; p = 0.01) relative to the Low Value (23% negative) and Control groups (24% negative;). In the planned primary analysis, the level of cocaine use reduction based on cocaine-negative urine samples will serve as the primary predictor of cardiovascular (e.g., endothelin-1 levels), immune (e.g., IL-10 levels) and psychosocial (e.g., Addiction Severity Index) outcomes using results from the fitted models. DISCUSSION/SIGNIFICANCE: This research will advance the field by prospectively and comprehensively demonstrating the beneficial effects of reduced cocaine use. These outcomes can, in turn, support the adoption of reduced cocaine use as a viable alternative endpoint in cocaine treatment trials.

Restoring brain health: Electroacupuncture at GB20 and LR3 for migraine mitigation through mitochondrial restoration.

Electroacupuncture (EA) is a promising alternative therapy for migraine, with mitochondrial dysfunction hypothesized as a pivotal mechanism in migraine pathophysiology. This research endeavors to investigate the therapeutic potential of EA in addressing migraines and shed light on the associated mechanisms linked to mitochondrial anomalies. Migraine in rats was induced by 10 mg/kg nitroglycerin, followed by 2/15 Hz EA treatment at GB20 and LR3. Nociceptive behavior was recorded via a camera and analyzed using EthoVision XT 12.0 software. The hind-paw withdrawal threshold was assessed using the von Frey test. We assessed the levels of calcitonin gene-related peptide (CGRP), nitric oxide (NO), and endothelin (ET) - key parameters in migraine pathophysiology using immunohistochemistry and enzyme-linked immunosorbent assay. Mitochondrial morphology in brain tissues was observed through transmission electron microscopy. Reactive oxygen species (ROS) level in mitochondria was measured by flow cytometry. The levels of PINK1 and Parkin were assessed using Western blot analysis. EA at GB20 and LR3 decreased nociceptive behaviors (resting and grooming) and increased exploratory and locomotor behaviors in migraine rats. The hind-paw withdrawal threshold in migraine rats was significantly elevated following EA treatment. Post-EA treatment, levels of CGRP and NO decreased, while ET level increased, suggesting an alteration in pain and vascular physiology. Notably, EA treatment mitigated the mitochondrial damage and reduced ROS level in the brain tissues of migraine rats. EA treatment upregulated the expression of PINK1 and Parkin in migraine rats. EA at GB20 and LR3 may treat migraine by alleviating PINK1/Parkin-mediated mitochondrial dysfunction.

CAPE ameliorates vascular damage caused by sepsis

parameters of vascular and oxidative damage caused by sepsis and to evaluated the effects of caffeic acid phenethyl ester (CAPE) on these damages.&#x0D; Materials and Methods: Wistar-Albino male rats were used for this study. Rats were divided into 4 groups (n = 10). Group 1 animals were intraperitoneally (i.p) injected with sterile saline (Control Group). Group 2 animals were i.p injected with lipopolysaccharide (LPS), 20 mg / kg-weight dose (Sepsis Group). Group 3 animals were i.p injected with lipopolysaccharide, 20 mg / kg-weight dose. Immediately after LPS injection, CAPE was i.p injected at single dose, 10 µmol / kg-body weight (Treatment Group). A single dose of CAPE, 10 µmol / kg-body weight / day, was injected i.p to Group 4 animals for 5 days. After 5th day CAPE injection, a single dose of LPS 20 mg / kg-weight was injected (Protective Group). At the 6th hour after the injections applied to all groups, blood sample were taken intracardiac and their serum were separated for the studies. Homocysteine (Hcy), asymmetric dimethyl arginine (ADMA), endothelin-1 (ET-1) and vascular cellular adhesion molecule-1 (VCAM-1) were measured by enzyme-linked immunosorbent assay (ELISA). In addition, the protective and therapeutic effects of CAPE on these parameters was investigated.&#x0D; Results: Control group Hcy, ADMA, ET-1 and VCAM-1 levels were found to be 4.987 ± 0.096 µmol/l, 0.803 ± 0.020 nmol/ml, 21.123 ± 2.575 ng/l, 3.155 ± 0.078 ng/ml, respectively. Sepsis group Hcy, ADMA, ET-1 and VCAM-1 levels were found to be 8.975 ± 0.160 µmol/l, 3.953 ± 0.678 nmol/ml, 52.446 ± 2.546 ng/l, 10.783 ± 1.068 ng/ml, respectively. Treatment group Hcy, ADMA, ET-1 and VCAM-1 levels were found to be 5.286 ± 0.037 µmol/l, 1.304 ± 0.040 nmol/ml, 27.995 ± 1.299 ng/l, 3.72 ± 0.073 ng/ml, respectively. Protective group Hcy, ADMA, ET-1 and VCAM-1 levels were found to be 5.401 ± 0.042 µmol/l, 1.431 ± 0.056 nmol/ml, 32.708 ± 1.326 ng/l, 4.058 ± 0.069 ng/ml, respectively. It was observed that the Hcy, ADMA, ET-1 and VCAM-1 levels of the sepsis group increased significantly compared to the control group (p0.05). It was observed that CAPE treatment significantly decreased these parameters levels. However, the use of CAPE as a protective was not as effective as its treatment effect.&#x0D; Conclusion: Our results demonstrated that sepsis resulted in increase Hcy, ADMA, ET-1, VCAM-1 levels. All these changes indicate that sepsis-mediated vascular damage is increased. Our results demonstrated that CAPE is more effective in preventing sepsis-mediated damages when given as a treatment.

Effect of Morning and Afternoon Exercise on the Improvement of Endothelial Dysfunction in Type-2 Diabetes Mellitus Patients

Introduction: Endothelial dysfunction, an early precursor to cardiovascular complications in Diabetes Mellitus (DM) patients, can be effectively managed through exercise, a non-pharmacological approach. However, it is significant to consider the timing of exercise as it can influence the benefits obtained in the management of vascular complications. This is because various systems in the body, including endothelial function, undergo fluctuations according to the circadian rhythm. This study aimed to explore the differences between morning and afternoon exercise in improving vascular endothelial dysfunction in the production of endothelial Nitric Oxide Synthase (eNOS) and Endothelin-1 (ET-1). Methods: Twenty two Type-2 Diabetes Mellitus (T2DM) patients were randomly divided into morning and afternoon exercise groups, and they were administered 10 weeks of Persadia diabetes gymnastics treatment. Before treatment, venous blood was taken as pre-exercise data, and after 10 weeks, it was obtained again for post-exercise examination. The examined data consisted of blood glucose, eNOS, and ET-1 levels. The data was processed statistically using the t-test. Results: eNOS level increased significantly in both samples at 6.66 ± 0.56 ng/ml and 5.46 ± 0.9 ng/ml (p&lt;0.001). However, the increment was higher in the morning group. Whereas, the level of ET-1 decreased significantly in both samples at 58.08 ± 4.01 pg/ml vs. 34.84 ± 4.75 pg/ml (p&lt;0.001), but the decrement was greater in the morning group. Conclusion: This study indicates that morning exercise is more influential in improving endothelial dysfunction by increasing the eNOS enzyme and significantly decreasing ET-1 in T2DM patients.

The Role of Microvascular Obstruction and Intra-Myocardial Hemorrhage in Reperfusion Cardiac Injury. Analysis of Clinical Data.

Microvascular obstruction (MVO) of coronary arteries promotes an increase in mortality and major adverse cardiac events in patients with acute myocardial infarction (AMI) and percutaneous coronary intervention (PCI). Intramyocardial hemorrhage (IMH) is observed in 41-50% of patients with ST-segment elevation myocardial infarction and PCI. The occurrence of IMH is accompanied by inflammation. There is evidence that microthrombi are not involved in the development of MVO. The appearance of MVO is associated with infarct size, the duration of ischemia of the heart, and myocardial edema. However, there is no conclusive evidence that myocardial edema plays an important role in the development of MVO. There is evidence that platelets, inflammation, overload, neuropeptide Y, and endothelin-1 could be involved in the pathogenesis of MVO. The role of endothelial cell damage in MVO formation remains unclear in patients with AMI and PCI. It is unclear whether nitric oxide production is reduced in patients with MVO. Only indirect evidence on the involvement of inflammation in the development of MVO has been obtained. The role of reactive oxygen species (ROS) in the pathogenesis of MVO is not studied. The role of necroptosis and pyroptosis in the pathogenesis of MVO in patients with AMI and PCI is also not studied. The significance of the balance of thromboxane A2, vasopressin, angiotensin II, and prostacyclin in the formation of MVO is currently unknown. Conclusive evidence regarding the role of coronary artery spasm in the development of MVhasn't been established. Correlation analysis of the neuropeptide Y, endothelin-1 levels and the MVO size in patients with AMI and PCI has not previously been performed. It is unclear whether epinephrine aggravates reperfusion necrosis of cardiomyocytes. Dual antiplatelet therapy improves the efficacy of PCI in prevention of MVO. It is unknown whether epinephrine or L-type channel blockers result in the long-term improvement of coronary blood flow in patients with MVO.

Effect and mechanism of acupuncture on airway smooth muscle relaxation during acute asthma attack in rats.

To explore the effect and mechanism of acupuncture at "Feishu" (BL 13) and "Dingchuan" (EX-B 1), and "Kongzui" (LU 6) and "Yuji" (LU 10) for relaxing the airway smooth muscle in the rats during acute asthma attack and compare the effect among the two pairs of acupoints and the acupoints combination. Forty SD male rats with SPF grade were randomly divided into a blank group, a model group, a pair-point A group (acupuncture at "Feishu" [BL 13] and "Dingchuan" [EX-B 1]), a pair-point B group (acupuncture at "Kongzui" [LU 6] and "Yuji" [LU 10]) and a point combination group (acupuncture at "Feishu" [BL 13] , "Dingchuan" [EX-B 1], "Kongzui" [LU 6] and "Yuji" [LU 10]), with 8 rats in each group. Except the rats in the blank group, the model of acute asthma attack was induced by ovalbumin (OVA) combined with aluminum hydroxide gel in the rest groups. Started on the 15th day of modeling, except in the blank group and the model group, acupuncture was delivered in the other groups, 30 min in each intervention, once daily, for 14 days. In each group, the latent period of asthma inducing was measured; the lung resistance (LR) and dynamic lung compliance (Cdyn) were determined using lung function detector; the levels of endothelin-1 (ET-1), tumor necrosis factor-α (TNF-α), cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) in serum and bronchoalveolar lavage fluid (BALF) were measured by ELISA; with Masson staining and electron microscopy adopted, the morphology and ultrastructure of airway smooth muscle of the rats were observed; the mRNA and protein expressions of ET-1 and beta-2 adrenergic receptor (β2-AR) were detected by quantitative real-time fluorescence and Western blot, respectively. Compared with the blank group, the latent period of asthma inducing was shortened (P<0.05), RL increased and Cdyn decreased (P<0.05) with the different concentrations of methacholine (0.025 mg/kg, 0.05 mg/kg, 0.1 mg/kg, 0.2 mg/kg) in the model group. In the pair-point A group, the pair-point B group and the point combination group, the latent period of asthma inducing was prolonged (P<0.05), RL decreased and Cdyn increased (P<0.05) with different concentrations of methacholine when compared with those in the model group; and the latent period of asthma inducing in the point combination group was longer than that in the pair-point A group (P<0.05). Compared with the blank group, the levels of ET-1, TNF-α and cGMP in the serum and BALF were elevated (P<0.05), and those of cAMP reduced (P<0.05) in the model group. The levels of ET-1, TNF-α and cGMP in the serum and BALF were reduced (P<0.05), and those of cAMP elevated (P<0.05) in the pair-point A group, the pair-point B group and the point combination group when compared with those in the model group. In the blank group, the lung tissue was normal structurally. In the model group, the collagen fibers were proliferated increasingly, the smooth muscle was thickened, the mitochondria were swollen, and their cristae disrupted and reduced massively. In the pair-point B group, the collagen fibers were proliferated, the smooth muscle was thicker compared with that in the blank group, the mitochondria were mildly swollen and their cristae disrupted partially. In the pair-point A group and the point combination group, the lung tissue changes were obviously alleviated in comparison with the model group, the mitochondria were slightly swollen and their cristae disrupted occasionally. Compared with the blank group, the mRNA and protein expression of ET-1 increased and that of β2-AR decreased in the lung tissue of the model group (P<0.05). In the pair-point A group, the pair-point B group and the point combination group, the mRNA and protein expression of ET-1 was reduced and that of β2-AR elevated in the lung tissue when compared with those in the model group (P<0.05). In comparison with the pair-point A group, the mRNA expression of β2-AR was elevated in the point combination group (P<0.05). When compared with the pair-point B group, the mRNA expression of β2-AR increased, the protein expression of ET-1 decreased (P<0.05) in the point combination group. Acupuncture at "Feishu" (BL 13) and "Dingchuan" (EX-B 1), "Kongzui" (LU 6) and "Yuji" (LU 10), two pairs of acupoints relieves the airway smooth muscle spasm in the rats during acute asthma attack, which may be related to inhibiting the mRNA and protein expression of ET-1 to reduce the excretion of ET-1 and TNF-α; while enhancing the mRNA and protein expression of β2-AR to balance the levels of cAMP and cGMP. The effect is optimal when acupuncture is delivered at two pairs of acupoints simultaneously.