Involvement of Bradykinin Receptor 2 in Nerve Growth Factor Neuroprotective Activity

Neurotrophin nerve growth factor (NGF) has been demonstrated to upregulate the gene expression of bradykinin receptor 2 (B2R) on sensory neurons, thus facilitating nociceptive signals. The aim of the present study is to investigate the involvement of B2R in the NGF mechanism of action in nonsensory neurons in vitro by using rat mixed cortical primary cultures (CNs) and mouse hippocampal slices, and in vivo in Alzheimer’s disease (AD) transgenic mice (5xFAD) chronically treated with NGF. A significant NGF-mediated upregulation of B2R was demonstrated by microarray, Western blot, and immunofluorescence analysis in CNs, indicating microglial cells as the target of this modulation. The B2R involvement in the NGF mechanism of action was also demonstrated by using a selective B2R antagonist which was able to reverse the neuroprotective effect of NGF in CNs, as revealed by viability assay, and the NGF-induced long-term potentiation (LTP) in hippocampal slices. To confirm in vitro observations, B2R upregulation was observed in 5xFAD mouse brain following chronic intranasal NGF treatment. This study demonstrates for the first time that B2R is a key element in the neuroprotective activity and synaptic plasticity mediated by NGF in brain cells.

THE COMBINATIONS OF GENETIC MARKERS OF BLOOD, THAT INFLUENCE ON THE DEVELOPMENT OF COMPLICATIONS OF ARTERIAL HYPERTENSION

В статье представлены исследования, в результате которых выявлены корреляции между сочетанием генетических маркеров крови (фенотипы гаптоглобина, группы крови, резус-фактор), полиморфизмом генов (альдостерон синтазы CYP11B2, рецептора брадикинина BDKRB2,) у больных с гипертонической болезнью. Исследование рассматривает современные аспекты диагностики, прогрессирование и развитие таких осложнений гипертонической болезни, как транзиторная ишемическая атака, острое нарушение мозгового кровообращения. Изучалось наличием отягощённого семейного анамнеза. В результате проведённого исследования установили, что пациенты, имеющие сочетание генетических характеристик (фенотип Нр 2-2, резус-фактор Rh(-), А(II) группа крови) и наличием полиморфизма CYP11B2 C(-344)T с генотипом Т/Т гена альдостерон синтазы, полиморфизма гена рецептора брадикинина BDKRB2 с генотипом (+9/+9)) имеет важное прогностическое значение высокого риска развития более тяжёлого течения артериальной гипертензии и развитием осложнений в виде острого нарушения мозгового кровообращения This article presents trials that have revealed correlations between the combination of genetic blood markers (phenotypes of haptoglobin, blood groups, Rh factor), gene polymorphism (aldosterone synthase CYP11B2, bradykinin receptor BDKRB2) in patients with arterial hypertension (AH). The article concerns up-to-date aspects of diagnosis, progression and development of such complications of AH as transient ischemic attack, acute cerebrovascular accident. It was investigated the family predisposition. As a result of the study, it was found that patients with a combination of genetic characteristics (phenotype Нр 2-2, Rh factor Rh (-), A (II) blood group) and the presence of CYP11B2 C (-344) T polymorphism with C / T genotypes and T / T of the aldosterone synthase gene, polymorphism of the bradykinin receptor BDKRB2 gene with genotype (+ 9 / + 9)) has an important prognostic value of a high risk of developing a more severe course of AH and the development of complications as a acute violation of cerebral circulation

Abstract 552: Bradykinin Receptor B1/ Matrix Metalloprotease 3 Pathway is a Novel Target in Preventing Cardiac Ischemia-reperfusion Injury

Introduction: Impaired endothelial function leads to the progression of heart failure after Ischemia-reperfusion (IR). Kinin activation of bradykinin receptor 1 (B1R), a G protein-coupled receptor that has been found to induce capillary leakage, may serve as a critical mediator in cardiac microvascular barrier dysfunction. However, the underlying mechanisms are not clear. We found that B1R inhibition abolished IR-induced endothelial matrix metalloprotease (MMP3) expression and improved endothelial barrier formation. Thus, we hypothesized that B1R antagonist protects against cardiac IR injury through an MMP3 pathway. Methods and Results: MMP3-/- mice and their littermate controls (WT) were subjected to either cardiac IR or sham control. The baseline characteristics of these mice showed minimal phenotypes. Cardiac function was determined at 3, 7 and 24 days post-IR by echocardiography. The MMP3-/- mice displayed improved cardiac function compared to the control mice, as determined by fractional shortening (26% ± 1.1 MMP3-/- vs. 21% ± 0.9 WT, p<0.05, n=5) and ejection fraction (48% ± 1.9 MMP3-/- vs. 42% ± 2.8.1 WT, p<0.05, n=5), and treating with B1R antagonist (300 μg/Kg) significantly reduced serum MMP3 levels (p<0.01). Compared to the control mice, MMP3-/- mice had significantly less infarction/area at risk 24 hours post-IR demonstrated through TTC staining. In vitro studies revealed that cellular hypoxia-reoxygenation (HR) injury significantly increased both B1R and MMP3 expression levels in primary isolated cardiac mice microvascular endothelial cells (mCMVEC). MMP3 levels were measured via ELISA. Moreover, B1R agonist treatment (1uM) increased MMP3 levels, while the use of the antagonist attenuated the increase of MMP3 in response to HR. Finally, the use of B1R antagonist improved MMP3 induced endothelial barrier dysfunction, which was measured by the electric cell-substrate impedance sensing (ECIS) system. Taken together, the results demonstrated that B1R antagonist abolished IR induced MMP3 induction and that the deletion of MMP3 is protective of cardiac function upon IR injury. Conclusions: MMP3 is a critical regulator of cardiac microvascular barrier function, and B1R/MMP3 could potentially serve as a novel therapeutic target for heart failure in response to IR injury.

Kinins and Cytokines in COVID-19: A Comprehensive Pathophysiological Approach

Most striking observations in COVID-19 patients are the hints on pulmonary edema (also seen on CT scans as ground glass opacities), dry cough, fluid restrictions to prevent more severe hypoxia, the huge PEEP that is needed while lungs are compliant, and the fact that anti-inflammatory therapies are not powerful enough to counter the severity of the disease. We propose that the severity of the disease and many deaths are due to a local vascular problem due to activation of B1 receptors on endothelial cells in the lungs. SARS-CoV-2 enters the cell via ACE2, a cell membrane bound molecule with enzymatic activity that next to its role in RAS is needed to inactivate des-Arg9 bradykinin, the potent ligand of the bradykinin receptor type 1 (B1). In contrast to bradykinin receptor 2 (B2), the B1 receptor on endothelial cells is upregulated by proinflammatory cytokines. Without ACE2 acting as a guardian to inactivate the ligands of B1, the lung environment is prone for local vascular leakage leading to angioedema. Angioedema is likely a feature already early in disease, and might explain the typical CT scans and the feeling of people that they drown. In some patients, this is followed by a clinical worsening of disease around day 9 due to the formation antibodies directed against the spike (S)-antigen of the corona-virus that binds to ACE2 that could contribute to disease by enhancement of local immune cell influx and proinflammatory cytokines leading to damage. In parallel, inflammation induces more B1 expression, and possibly via antibody-dependent enhancement of viral infection leading to continued ACE2 dysfunction in the lung because of persistence of the virus. In this viewpoint we propose that a bradykinin-dependent local lung angioedema via B1 and B2 receptors is an important feature of COVID-19, resulting in a very high number of ICU admissions. We propose that blocking the B1 and B2 receptors might have an ameliorating effect on disease caused by COVID-19. This kinin-dependent pulmonary edema is resistant to corticosteroids or adrenaline and should be targeted as long as the virus is present. In addition, this pathway might indirectly be responsive to anti-inflammatory agents or neutralizing strategies for the anti-S-antibody induced effects, but by itself is likely to be insufficient to reverse all the pulmonary edema. Moreover, we provide a suggestion of how to ventilate in the ICU in the context of this hypothesis.

Mitigation of Vascular Permeability Using Bradykinin Receptor Antagonist and Anti-Fibrinolytic Agent in Rats with Polytrauma and Hemorrhagic Shock

Introduction: Clinical studies suggest that one third of patients suffering major trauma develop acute traumatic coagulopathy (ATC), which is associated with increased morbidity and mortality. Recent evidence in both animal and clinical studies has shown that development of ATC is associated with an increase in fibrinolytic activity, initiated by an elevation in tissue plasminogen activator (tPA) immediately after trauma. Previous studies demonstrate that administration of the anti-fibrinolytic agent tranexamic acid (TXA) not only inhibits fibrinolytic activity in traumatized tissue but also attenuates lung edema after resuscitation in rats with polytrauma and hemorrhagic shock, suggesting a potential correlation between fibrinolysis and vascular permeability. One of the underlying mechanisms of acute elevation in vascular permeability after severe trauma is associated with an elevation of bradykinin activity due to activation of the tPA-plasmin-bradykinin pathway. This study will test the hypothesis that early intervention by administration of a bradykinin receptor antagonist with or without TXA prior to resuscitation attenuates edema in the tissues after resuscitation in rats with polytrauma/hemorrhagic shock. Method: Sprague-Dawley rats (350-450g) under anesthesia with isoflurane received polytrauma followed by hemorrhage (40% of blood volume). The rats either received vehicle (normal saline) or bradykinin receptor-2 antagonist (BR-2A (HOE140, 0.5mg/kg)), TXA (20mg/kg) or BR-2A/TXA via femoral vein at 45min after trauma (n=6-8/each group), followed by resuscitation with Lactated Ringer's solution (20% of blood volume) at 60min after trauma. Four normal rats, not undergoing surgical procedures were treated as normal control. At 2hr after trauma (1hr after resuscitation), the rats were euthanized and the tissues from internal organs were harvested and weighed for wet weight. The tissue samples were then transferred to a 600C oven for 10-14 days. The dry weight was determined when the weight of dried tissue was constant for three consecutive days. The wet-dry weight ratio was then calculated to represent the water content for each tissue. The wet-dry weight ratio was also measured in traumatized tissues collected from the injured sites of liver, intestine and skeletal muscle (at hindlimb). At 2hr after trauma, prothrombin time (PT) was measured for systemic hemostatic condition, and the resuscitation outcome was assessed by lactate levels. Results: The wet/dry weight ratio is different in organs due to underlying structure and water content. Trauma, hemorrhage and resuscitation led to a significant elevation of wet/dry weight ratio in a majority of the organs (except for heart, kidney, and stomach). The wet/dry weight ratio was significantly higher in traumatized tissues than non-traumatized tissues (at liver, intestine and skeletal muscle). BR-2A significantly decreased the wet/dry weight ratio of the brain (4.51±0.03 (BR-2A) vs 4.64±0.03 (Vehicle)), but had no effect on the other organs. TXA significantly reduces the wet/dry weight ratio of the lung and intestine (lung: 5.02±0.07 (BR-2A) vs 5.23±0.06 (Vehicle), p=0.003; Intestine: 4.16±0.10 (BR-2A) vs 4.45±0.07 (Vehicle), p=0.036) but not in the brain and other tissues. The combination of BR-2A with TXA significantly decreased elevation of wet/dry weight ratio of the brain, lung and intestine, but there was no synergistic effect on wet/dry weight ratio found in any organs. However, the rats treated with combination treatment of BR-2A and TXA had the lowest lactate level at 2hr after trauma as compared to rats treated with vehicle (1.85±0.15 vs 3.39±0.29 mmol/l, p=0.032) suggesting an overall improvement of resuscitation outcome. BR-2A, TXA or a combination of the two did not cause significant change in MAP and HR, and had no significant effect on systemic coagulopathy (elevation of prothrombin time) at 2hr after trauma and hemorrhage. Conclusions: This study suggests that BR-2A and TXA independently mitigate tissue-specific vascular permeability in trauma and hemorrhagic shock. Future studies are necessary to investigate the optimal dose and timing of administration of BR-2A or BR-2A with TXA in trauma and hemorrhagic shock to improve the outcome and efficacy of treatment. Disclosures No relevant conflicts of interest to declare.