Bradykinin Research Articles

KV7 channels are potential regulators of the exercise pressor reflex.

The exercise pressor reflex (EPR) originates in skeletal muscle and is activated by exercise-induced signals to increase arterial blood pressure and cardiac output. Muscle ischemia can elicit the EPR, which can be inappropriately activated in patients with peripheral vascular disease or heart failure to increase the incidence of myocardial infarction. We seek to better understand the receptor/channels that control excitability of group III and group IV muscle afferent fibers that give rise to the EPR. Bradykinin (BK) is released within contracting muscle and can evoke the EPR. However, the mechanism is incompletely understood. KV7 channels strongly regulate neuronal excitability and are inhibited by BK. We have identified KV7 currents in muscle afferent neurons by their characteristic activation/deactivation kinetics, enhancement by the KV7 activator retigabine, and block by KV7 specific inhibitor XE991. The blocking of KV7 current by different XE991 concentrations suggests that the KV7 current is generated by both KV7.2/7.3 (high affinity) and KV7.5 (low affinity) channels. The KV7 current was inhibited by 300 nM BK in neurons with diameters consistent with both group III and group IV afferents. The inhibition of KV7 by BK could be a mechanism by which this metabolic mediator generates the EPR. Furthermore, our results suggest that KV7 channel activators such as retigabine, could be used to reduce cardiac stress resulting from the exacerbated EPR in patients with cardiovascular disease.NEW & NOTEWORTHY KV7 channels control neuronal excitability. We show that these channels are expressed in muscle afferents and generate currents that are blocked by XE991 and bradykinin (BK). The XE991 block suggests that KV7 current is generated by KV7.2/3 and KV7.5 channels. The BK inhibition of KV7 channels may explain how BK activates the exercise pressor reflex (EPR). Retigabine can enhance KV7 current, which could help control the inappropriately activated EPR in patients with cardiovascular disease.

Investigation of the role of miR-221 in diabetic peripheral neuropathy and related molecular mechanisms.

Diabetic peripheral neuropathy (DPN) is one of the most common complications of diabetes, but the molecular mechanisms of DPN are still unclear. To investigate the role of miR-221 in DPN and the related molecular mechanisms. Streptozotocin (STZ) was used to establish an in vivo DPN model. An in vitro DPN model was established using high glucose-induced SH-SY5Y cells. The pain condition of rats was measured by evaluating the 50% paw withdrawal threshold (PWT) and paw withdrawal latency (PWL). Serum exosomes were extracted and identified. Expression of miR-221 in serum exosomes and serum SOCS3 expression were determined using reverse-transcription quantitative polymerase chain reaction (RT-qPCR). Western blotting was used to measure the protein levels of SOCS3, bradykinin (BK) and prostaglandin E2 (PEG2). The dual luciferase reporter assay was performed to confirm SOCS3 3'-UTR as a target of miR-221. The serum or cell supernatant levels of PEG2, BK, interleukin (IL)-6, IL-1β, and tumor necrosis factor alpha (TNF-α) were measured using enzyme-linked immunosorbent assay (ELISA). Induction of the lenti-miR-221 inhibitor significantly decreased the expression of miR-221 in DPN rats. Both 50% PWT and PWL values were markedly decreased in DPN rats. When miR-221 was inhibited, the 50% PWT and PWL values were both significantly increased. Knockdown of miR-221 significantly increased the expression of SOCS3 and decreased the expression of NF-κB. Furthermore, knockdown of miR-221 remarkably decreased the expression of PEG2, BK, IL-6, IL-1β, and TNF-α in both STZ-treated DPN rats and high glucose-induced SH-SY5Y cells, which was reversed by inhibition of SOCS3. The dual luciferase reporter assay showed that miR-221 directly targeted and negatively regulated SOCS3. Inhibition of miR-221 can reduce pain and decrease expression of inflammatory factors through targeting SOCS3 in DPN.

Angiotensin-(1\u20137) oral formulation improves physical performance in mountain bike athletes: a double\u2010blinded crossover study

BackgroundThe ECA2/Ang-(1–7)/Mas axis is shown to be involved in effects mediated by physical exercise, as it can induce the release of nitric oxide (ON) and bradykinin (BK), which are potent vasodilators. The vasodilating action the NO/BK can contribute to increased metabolic efficiency in muscle tissue and central nervous system. The formulation HPβ-CD-Ang-(1–7) through its mechanisms of action can be a promising supplement to aid in the maintenance and improvement of performance and may also favor recovery during competitions. The premise of this study was to investigate the effects of acute oral supplementation HPβ-CD-Ang-(1–7) on the performance of mountain bike (MTB) practitioners.MethodsFourteen recreational athletes, involved in training programs for at least one year, participated in this crossover design study. Subjects underwent two days of testing with a seven-day interval. HPβ-CD-Ang-(1–7) (1.75 mg) and HPβCD-Placebo were provided in capsules three hours prior to tests. To determine the safety of the HPβ-CD-Ang-(1–7) formulation associated with physical effort, cardiovascular parameters heart rate (HR) and blood pressure (BP) were analyzed. Physical performance was measured using maximal oxygen uptake (VO2), total exercise time (TET), mechanical work (MW), mechanical efficiency (ME), and rating of perceived exertion (RPE). Respiratory exchange coefficient (REC), lactate and non-esterified fatty acids (NEFAs) were measured. Maximal incremental tests were performed on a progressively loaded leg cycle ergometer.ResultsThere were no significant differences in terms of HR or BP at rest and maximum effort between the HPβ-CD-Ang-(1–7) and placebo groups. The VO2max showed significant differences (p = 0.04). It was higher in the Ang-(1–7)condition (66.15 mlO2.kg− 1.min− 1) compared to the placebo (60.72 mlO2.kg− 1.min− 1). This was also observed for TET (Ang-(1–7) 39.10 min vs. placebo 38.14 min; p = 0.04), MW (Ang-(1–7) 156.7 vs. placebo 148.2; p = 0.04), and at the lowest RPE (Ang-(1–7) vs. placebo; p = 0.009). No significant differences were observed for REC, NEFAs, or Lactate.ConclusionsThese results suggest that HPβ-CD-Ang-(1–7) improves the physical performance of MTB recreational athletes and could be a promising supplement.Trial registrationRBR-2 × 56pw8, registered January 15th, 2021. The study was prospectively registered.

Upregulating Aggregation-Induced-Emission Nanoparticles with Blood-Tumor-Barrier Permeability for Precise Photothermal Eradication of Brain Tumors and Induction of Local Immune Responses.

Compared to other tumors, glioblastoma (GBM) is extremely difficult to treat. Recently, photothermal therapy (PTT) has demonstrated advanced therapeutic efficacy; however, because of the relatively low tissue-penetration efficiency of laser light, its application in deep-seated tumors remains challenging. Herein, bradykinin (BK) aggregation-induced-emission nanoparticles (BK@AIE NPs) are synthesized; these offer selective penetration through the blood-tumor barrier (BTB) and strong absorbance in the near-infrared region (NIR). The BK ligand can prompt BTB adenosine receptor activation, which enhances transportation and accumulation inside tumors, as confirmed by T1 -weighted magnetic resonance and fluorescence imaging. The BK@AIE NPs exhibit high photothermal conversion efficiency under 980nm NIR laser irradiation, facilitating the treatment of deep-seated tumors. Tumor progression can be effectively inhibited to extend the survival span of mice after spatiotemporal PTT. NIR irradiation can eradicate tumor tissues and release tumor-associated antigens. It is observed that the PTT treatment of GBM-bearing mice activates natural killer cells, CD3+ T cells, CD8+ T cells, and M1 macrophages in the GBM area, increasing the therapeutic efficacy. This study demonstrates that NIR-assisted BK@AIE NPs represent a promising strategy for the improved systematic elimination of GBMs and the activation of local brain immune privilege.

Hypersensitivity Induced by Intrathecal Bradykinin Administration Is Enhanced by N-oleoyldopamine (OLDA) and Prevented by TRPV1 Antagonist.

Transient receptor potential vanilloid 1 (TRPV1) channels contribute to the development of several chronic pain states and represent a possible therapeutic target in many painful disease treatment. Proinflammatory mediator bradykinin (BK) sensitizes TRPV1, whereas noxious peripheral stimulation increases BK level in the spinal cord. Here, we investigated the involvement of spinal TRPV1 in thermal and mechanical hypersensitivity, evoked by intrathecal (i.t.) administration of BK and an endogenous agonist of TRPV1, N-oleoyldopamine (OLDA), using behavioral tests and i.t. catheter implantation, and administration of BK-induced transient thermal and mechanical hyperalgesia and mechanical allodynia. All these hypersensitive states were enhanced by co-administration of a low dose of OLDA (0.42 µg i.t.), which was ineffective only under the control conditions. Intrathecal pretreatment with TRPV1 selective antagonist SB366791 prevented hypersensitivity induced by i.t. co-administration of BK and OLDA. Our results demonstrate that both thermal and mechanical hypersensitivity evoked by co-administration of BK and OLDA is mediated by the activation of spinal TRPV1 channels.

AMP-activated protein kinase is a key regulator of acute neurovascular permeability.

ABSTRACTMany neuronal and retinal disorders are associated with pathological hyperpermeability of the microvasculature. We have used explants of rodent retinae to study acute neurovascular permeability, signal transduction and the role of AMP-activated protein kinase (AMPK). Following stimulation with either vascular endothelial growth factor (VEGF-A) or bradykinin (BK), AMPK was rapidly and strongly phosphorylated and acted as a key mediator of permeability downstream of Ca2+. Accordingly, AMPK agonists potently induced acute retinal vascular leakage. AMPK activation led to phosphorylation of endothelial nitric oxide synthase (eNOS, also known as NOS3), which in turn increased VE-cadherin (CDH5) phosphorylation on Y685. In parallel, AMPK also mediated phosphorylation of p38 MAP kinases (hereafter p38) and HSP27 (HSPB1), indicating that it regulated paracellular junctions and cellular contractility, both previously associated with endothelial permeability. Endothelial AMPK provided a missing link in neurovascular permeability, connecting Ca2+ transients to the activation of eNOS and p38, irrespective of the permeability-inducing factor used. Collectively, we find that, due to its compatibility with small molecule antagonists and agonists, as well as siRNA, the ex vivo retina model constitutes a reliable tool to identify and study regulators and mechanisms of acute neurovascular permeability.

High-dose ACEi might be harmful in COVID-19 patients with serious respiratory distress syndrome by leading to excessive bradykinin receptor activation.

We aimed to critically review the available information on the potential contribution of excessive kallikrein-kinin systems (KKSs) activation to severe respiratory inflammation in SARS-CoV-2 infection, and the likely consequence of ACE inhibition in seriously affected patients. The literature related to the above topic was reviewed including papers that analysed the connections, actions, interactions, consequences and occasionally suggestions for rational interventions. Severe broncho-alveolar inflammation seems to be caused, at least in part, by upregulation of the KKS that increases plasma and/or local tissue concentrations of bradykinin (BK) in patients with COVID-19 infection. Besides KKS activation, suppression of ACE activity results in decreased bradykinin degradation, and these changes in concert can lead to excessive BK B1 and B2 receptor (BKB1R/BKB2R) activation. Aminopeptidase P (APP), and carboxypeptidase N also degrade bradykinin, but their protein expression and activity are unclear in COVID-19 infection. On the other hand, ACE2 expression is upregulated in patients with COVID-19 infection, so ACE2 activity is unlikely to be decreased despite blockade of part of ACE2 by the virus for entry into the cells. ACE2 cleaves lys-des-arginine9BK and arg-des-arginine9BK, the active metabolites of bradykinin, which stimulate the BKB1R receptor. Stimulation of BKB1R/BKB2R can exacerbate the pulmonary inflammatory response by causing vascular leakage and edema, vasodilation, smooth muscle spasm and stimulation of pain afferent nerves. Despite all uncertainties, it seems rational to treat comorbid COVID patients with serious respiratory distress syndrome with ARBs instead of high-dose ACE inhibitor (ACEi) that will further decrease bradykinin degradation and enhance BKB1R/BKB2R activation, but ACEi may not be contraindicated in patients with mild pulmonary symptoms.

POTENTIAL SIGNIFICANCE OF BRADYKININ 2 RECEPTOR GENOTYPING IN KIDNEY TRANSPLANT RECIPIENTS

Bradykinin (BK), a vasoactive peptide, is considered to exert the renoprotective and cardioprotective effects. Furthermore, most of its effects are mediated by the activation of BK type 2 receptor (B2R), whose level of expression is influenced by insertion/deletion (+9/-9) gene polymorphism. The aim of this study was to investigate the potential influence of B2R +9/-9 gene polymorphism on kidney function and blood pressure in kidney transplant recipients (KTRs) within the first post-transplantation year. Secondary, the study analyzed the correlation between hypertensive status and graft function, with respect to the B2R +9/-9 gene polymorphism. The pharmacogenetic study included 95 KTRs with clinical and biochemical parameters evaluated in three time-points (3rd, 6th, 12th month). The graft function was assessed using the estimated glomerular filtration rate (eGFR), while systolic (SBP), diastolic (DBP), and mean blood pressure (MAP) were used as parameters for the evaluation of blood pressure control. The carriers of -9/-9 genotype had more stable graft function compared to both, heterozygotes and homozygous for +9 allele, not only in the early (up to 6 months) but as well in the late post-transplantation period (beyond 6 months). Also, the -9/-9 genotype carriers showed a tendency to significantly decrease MAP, SBP, and DBP in the observed period, whereby only in this patient group the significant association between hypertensive status and graft function was demonstrated. In conclusion, genotyping of B2R +9/-9 gene polymorphism alongside with biochemical and clinical parameters may identify patients with respect to the risk of more prompt decline of graft function.

The antihypertensive effect of MK on spontaneously hypertensive rats through the AMPK/Akt/eNOS/NO and ERK1/2/Cx43 signaling pathways

We investigated the antihypertensive effects of maximakinin (MK) on spontaneously hypertensive rats (SHRs). The effects of MK on arterial blood pressure in SHRs were observed, and flow cytometry and 4,5-diaminofluorescein-2 staining were used to examine MK-induced nitric oxide (NO) release in human umbilical vein endothelial cells (HUVECs). Western blotting was used to analyze the effects of MK on the expression of AMP-activated protein kinase (AMPK), Akt, Connexin 43, ERK1/2, p38, and p-eNOS in HUVECs. The results showed that MK induced a more significant antihypertensive effect on SHRs than bradykinin (BK). MK induced significant increases in endothelial nitric oxide synthase (eNOS) phosphorylation and NO release in HUVECs. MK also significantly increased the phosphorylation of Akt and AMPK in HUVECs. The AMPK inhibitor compound C blocked the effect of MK on the generation of NO. MK induced the phosphorylation of ERK1/2, p38, and Connexin 43. The expression of p-Connexin 43 was significantly decreased in the presence of the ERK1/2 inhibitor U0126 but not the p38 inhibitor SB203580. The effects of MK on the phosphorylation of AMPK and ERK1/2 were significantly decreased by the BK B2 receptor inhibitor HOE-140. In summary, MK can significantly reduce blood pressure in SHRs. The antihypertensive effect might be mediated through the activation of the BK B2 receptor, while the downstream AMPK/PI3K/Akt/eNOS/NO and ERK1/2/Connexin 43 signaling pathways play additional roles.

Malaria link of hypertension: a hidden syndicate of angiotensin II, bradykinin and sphingosine 1-phosphate.

In malaria-endemic countries, the burden of hypertension is on the rise. Although malaria and hypertension seem to have no direct link, several studies in recent years support their possible link. Three bioactive molecules such as angiotensin II (Ang II), bradykinin (BK) and sphingosine 1-phosphate (S1P) are crucial in regulating blood pressure. While the increased level of Ang II and S1P are responsible for inducing hypertension, BK is arthero-protective and anti-hypertensive. Therefore, in the present review, based on available literatures we highlight the present knowledge on the production and bioavailability of these molecules, the mechanism of their regulation of hypertension, and patho-physiological role in malaria. Further, a possible link between malaria and hypertension is hypothesized through various arguments based on experimental evidence. Understanding of their mechanisms of blood pressure regulation during malaria infection may open up avenues for drug therapeutics and management of malaria in co-morbidity with hypertension.

Abstract P808: The Main Peptide Ssubstrates of Neurolysin Enhance Brain Microvascular Permeability in a Human in vitro Model

Increased brain microvascular permeability and disruption of blood-brain barrier (BBB) function are among hallmarks of ischemic stroke. Recently, peptidase neurlysin (Nln) has been identified as a compensatory and cerebroprotective mechanism in the post-stroke brain that functions to process a diverse group of extracellular neuropeptides, including bradykinin (BK), neurotensin (NT) and substance P (SP). A number of studies suggest involvement of BK, NT and SP in BBB impairment and edema formation after stroke, however there is paucity of data in regards to the direct effects of these peptides on the brain microvascular endothelial cells (BMECs) and BBB. The purpose of this study was to evaluate the direct effects of BK, NT and SP on permeability of BBB in an in vitro model based on human, induced pluripotent stem cell (iPSC)-derived BMECs. Our data indicate that all three peptides increase BBB permeability in a concentration-dependent manner in an in vitro model formed from two different iPSC lines (CTR90F and CTR65M) and widely used hCMEC/D3 human BMECs. Combination of BK, NT and SP at a sub-effective concentration also resulted in increased BBB permeability in the iPSC-derived model. Furthermore, we observed abrogation of BK, NT and SP effects with pretreatment of pharmacological blockers targeting their specific receptors or in presence of recombinant neurolysin (Nln). This is the first experimental study to document increased permeability of BBB in response to direct action of NT in an in vitro model. In addition, our study confirms the expected but not well-documented, direct effect of SP on BBB permeability and adds to the well-recognized actions of BK on BBB. Lastly, we demonstrate that peptidase Nln can neutralize the effects of these peptides on BBB, suggesting potential therapeutic implications.

Is the Use of Surface-Enhanced Infrared Spectroscopy Justified in the Selection of Peptide Fragments That Play a Role in Substrate-Receptor Interactions? Adsorption of Amino Acids and Neurotransmitters on Colloidal Ag and Au Nanoparticles.

This paper describes an application of attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) and surface-enhanced infrared spectroscopy (SEIRA) to characterize the selective adsorption of four peptides present in body fluids such as neuromedin B (NMB), bombesin (BN), neurotensin (NT), and bradykinin (BK), which are known as markers for various human carcinomas. To perform a reliable analysis of the SERIA spectra of these peptides, curve fitting of these spectra in the spectral region above 1500 cm–1 and SEIRA measurements of sulfur-containing and aromatic amino acids were performed. On the basis of the analyses of the spectral profiles, specific conclusions were drawn regarding specific molecule–metal interactions and changes in the interaction during the substrate change from the surface of silver nanoparticles (AgNPs) to gold nanoparticles (AuNPs).

A Robust Bioassay of the Human Bradykinin B2 Receptor That Extends Molecular and Cellular Studies: The Isolated Umbilical Vein

Bradykinin (BK) has various physiological and pathological roles. Medicinal chemistry efforts targeted toward the widely expressed BK B2 receptor (B2R), a G-protein-coupled receptor, were primarily aimed at developing antagonists. The only B2R antagonist in clinical use is the peptide icatibant, approved to abort attacks of hereditary angioedema. However, the anti-inflammatory applications of B2R antagonists are potentially wider. Furthermore, the B2R antagonists notoriously exhibit species-specific pharmacological profiles. Classical smooth muscle contractility assays are exploited over a time scale of several hours and support determining potency, competitiveness, residual agonist activity, specificity, and reversibility of pharmacological agents. The contractility assay based on the isolated human umbilical vein, expressing B2R at physiological density, was introduced when investigating the first non-peptide B2R antagonist (WIN 64338). Small ligand molecules characterized using the assay include the exquisitely potent competitive antagonist, Pharvaris Compound 3 or the partial agonist Fujisawa Compound 47a. The umbilical vein assay is also useful to verify pharmacologic properties of special peptide B2R ligands, such as the carboxypeptidase-activated latent agonists and fluorescent probes. Furthermore, the proposed agonist effect of tissue kallikrein on the B2R has been disproved using the vein. This assay stands in between cellular and molecular pharmacology and in vivo studies.

Association of Ang-(1-7) and des-Arg9BK as new biomarkers of obesity and cardiometabolic risk factors in adolescents.

Children with obesity have a high risk of developing cardiovascular disease and hypertension, which is associated with the renin-angiotensin system (RAS) activation and kallikrein-kinin system (KKS) inactivation. Although recent studies have identified several peptide-based biomarkers for obesity, circulating peptides from the RAS and KKS in adolescents with obesity have not been described. The aim of this study was to examine circulating levels of RAS and KKS peptides in adolescents with obesity to investigate the turnover of these peptides and their relationship to metabolic disorders resulting from weight gain. The subjects (n = 104) were divided into normal weight (NW), overweight (OW), obese (OB), and morbidly obese (MO) groups. Anthropometric profiles were created by measuring height, weight, blood pressure, and skinfolds. Plasma levels of Ang I, II, (1-7), BK, and des-Arg9BK were quantified by high-performance liquid chromatography. The levels were as follows: Ang-(1-7)-MO 58.3 ± 50, OB 223.2 ± 150, OW 318.6 ± 190, NW 479.1 ± 160 pmol/mL, and Bradykinin (BK)-MO 367.6 ± 103, OB 253.8 ± 130, OW 484 ± 279, NW 874.9 ± 385 pmol/mL. Ang-(1-7) correlated inversely with weight, body mass index, leptin, diastolic blood pressure, and systolic blood pressure. BK and Ang-(1-7) levels correlated inversely with skinfolds, waist-hip ratio (WHR), leptin, and arm circumference. BK levels correlated with adiponectin and Ang-(1-7) levels. Plasma Ang I levelswere higher in the MO and OB groups than in the NW group, but plasma Ang II levels were similar in all groups. We suggest that Ang-(1-7) and des-Arg9BK metabolites are novel biomarkers of childhood obesity that are important for determining treatment strategies.

Effects of bradykinin on proliferation, apoptosis, and cycle of glomerular mesangial cells via the TGF-β1/Smad signaling pathway.

We aimed to assess the effects of bradykinin (BK) on the proliferation, apoptosis, and cycle of glomerular mesangial cells via the transforming growth factor-β 1 (TGF-β1)/Smad signaling pathway. Rat glomerular mesangial cells, HBZY-1, were divided into normal group (untreated), model group (5 ng/L TGF-β1), BK group (5 ng/L TGF-β1 + 1 ng/L BK), and inhibitor group [5 ng/L TGF-β1 + 1 ng/L LY2109761 (TGF-β1-specific inhibitor)]. The cell proliferation, cycle, apoptosis, expression of type I collagen (Col-1), and protein expressions of Col-1, TGF-β1, and phosphorylated Smad2 (p-Smad2) were detected by EdU labeling, flow cytometry, acridine orange/ethidium bromide (AO/EB) dual staining, immunofluorescence assay, and Western blotting, respectively. Compared with the normal group, the cell proliferation rate (P = 0.02) and protein expression levels of Col-1 (P = 0.02), TGF-β1 (P = 0.01), p-Smad2 (P = 0.02), and p-Smad7 (P = 0.00) in the model group significantly increased, and apoptosis rate (P = 0.01) significantly decreased. Compared with the model group, the BK and inhibitor groups significantly decreased in proliferation rate (P = 0.01) and protein expression levels of Col-1 (P = 0.01), TGF-β1 (P = 0.01), and p-Smad2 (P = 0.00). Also, they were significantly elevated in apoptosis rate (P = 0.02) and p-Smad7 protein expression (P = 0.02). BK regulates the proliferation, apoptosis, and the cycle of glomerular mesangial cells by inhibiting the TGF-β1/Smad signaling pathway.

Neurolysin substrates bradykinin, neurotensin and substance P enhance brain microvascular permeability in a human in vitro model.

Increased brain microvascular permeability and disruption of blood-brain barrier (BBB) function are among hallmarks of several acute neurodegenerative disorders, including stroke. Numerous studies suggest the involvement of bradykinin (BK), neurotensin (NT) and substance P (SP) in BBB impairment and oedema formation after stroke; however, there is paucity of data in regard to the direct effects of these peptides on the brain microvascular endothelial cells (BMECs) and BBB. The present study aimed to evaluate the direct effects of BK, NT and SP on the permeability of BBB in an in vitro model based on human induced pluripotent stem cell (iPSC)-derived BMECs. Our data indicate that all three peptides increase BBB permeability in a concentration-dependent manner in an in vitro model formed from two different iPSC lines (CTR90F and CTR65M) and widely used hCMEC/D3 human BMECs. The combination of BK, NT and SP at a sub-effective concentration also resulted in increased BBB permeability in the iPSC-derived model indicating potentiation of their action. Furthermore, we observed abrogation of BK, NT and SP effects with pretreatment of pharmacological blockers targeting their specific receptors. Additional mechanistic studies indicate that the short-term effects of these peptides are not mediated through alteration of tight-junction proteins claudin-5 and occludin, but likely involve redistribution of F-actin and secretion of vascular endothelial growth factor. This is the first experimental study to document the increased permeability of the BBB in response to direct action of NT in an in vitro model. In addition, our study confirms the expected but not well-documented, direct effect of SP on BBB permeability and adds to the well-recognised actions of BK on BBB. Lastly, we demonstrate that peptidase neurolysin can neutralise the effects of these peptides on BBB, suggesting potential therapeutic implications.

Intra-arterial injection of bradykinin produces reflex cardiorespiratory changes involving histamine receptors in anesthetized rats

Objectives: It is well known that intra-arterial injection of nociceptive agent produces vasosensory reflex responses altering cardiorespiratory parameters. The role of various inflammatory mediators is also implicated in the regulation of these reflex responses. However, the role of histamine in this regard is not clear. This study was performed to understand the role of H1 and H2 receptors in modulating the cardiorespiratory responses evoked after i.a. injection of bradykinin (BK). Materials and Methods: Male albino rats were anesthetized with an intra-peritoneal injection of urethane (1.5 g/kg). Tracheostomy was performed to keep the respiratory tract patent. The femoral artery was cannulated proximally by pediatric i.v. cannula (24 G, double ported). This cannulation was used for the blood pressure (BP) recording as well as for the drugs instillation as it contains double port with injection valve. The effect of BK (1 µM) on BP, electrocardiographic, and respiration was recorded for 30 min. The respiratory frequency, respiratory minute volume, mean arterial pressure, and heart rate were computed from the original tracings and the data were presented as mean ± SEM. Results: Intra-arterial injection of BK produced immediate hyperventilatory (50% from initial), hypotensive (40% from initial), and bradycardiac responses (17% from initial) of shorter latency (5–8 s) indicating the neural mechanisms in producing the responses. Pre-treatment with pheniramine maleate significantly attenuated the BK-induced hyperventilatory (11% from initial), hypotensive (8% from initial), and bradycardiac responses (2% from initial). Conclusion: Our data provide evidences for the involvement of H1 and H2 receptors in producing the BK-induced vasosensory reflex responses modulating the cardiovascular parameters in anesthetized rats.

Vascular Cells Proteome Associated with Bradykinin and Leptin Inflammation and Oxidative Stress Signals.

Among the primary contributors to cardiovascular diseases are inflammation and oxidative imbalance within the vessel walls as well as the fibrosis of rat aortic smooth muscle cell (RASMC). Bradykinin (BK) and leptin are inflammatory modulators that are linked to vascular injury. In this study, we employed tandem LC-MS/MS to identify protein signatures that encompass protein abundance in RASMC treated with BK or leptin followed by systems biology analyses to gain insight into the biological pathways and processes linked to vascular remodeling. In the study, 1837 proteins were identified in control untreated RASMC. BK altered the expression of 72 (4%) and 120 (6.5%) proteins, whereas leptin altered the expression of 189 (10.2%) and 127 (6.5%) proteins after 24 and 48 h, respectively, compared to control RASMC. BK increased the protein abundance of leptin receptor, transforming growth factor-β. On the other hand, leptin increased the protein abundance of plasminogen activator inhibitor 1 but decreased the protein abundance of cofilin. BK and leptin induced the expression of inflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-1β (IL-1β) and pathway analysis revealed the activation of mitogen-activated protein kinases (MAPKs) and AKT pathways. The proteome profile in response to BK and leptin revealed mechanistic interplay of multiple processes that modulate inflammation and oxidative stress signals in the vasculature.

The COX-2-derived PGE2 autocrine contributes to bradykinin-induced matrix metalloproteinase-9 expression and astrocytic migration via STAT3 signaling

BackgroundThe matrix metalloproteinase-9 (MMP-9) is up-regulated by several proinflammatory mediators in the central nervous system (CNS) diseases. Increasing reports show that MMP-9 expression is an inflammatory biomarker of several CNS disorders, including the CNS inflammation and neurodegeneration. Bradykinin (BK) is a common proinflammatory mediator and elevated in several brain injury and inflammatory disorders. The raised BK may be detrimental effects on the CNS that may aggravate brain inflammation through MMP-9 up-regulation or cyclooxygenase-2 (COX-2)-derived prostaglandin E2 (PGE2) production in brain astrocytes. However, the relationship between BK-induced MMP-9 expression and COX-2-derived PGE2 release in brain astrocytes remains unclear.MethodsHerein we used rat brain astrocytes (RBA) to investigate the role of the COX-2/PGE2 system in BK-induced MMP-9 expression. We used zymographic, RT-PCR, EIA, and Western blotting analyses to confirm that BK induces MMP-9 expression via a COX-2/PGE2-dependent pathway.ResultsOur results show activation of native COX-2 by BK led to PGE2 production and release. Subsequently, PGE2 induced MMP-9 expression via PGE2 receptor (EP)-mediated c-Src, Jak2, ERK1/2, and then activated signal transducer and activator of transcription 3 (STAT3) signaling pathway. Finally, up-regulation of MMP-9 by BK via the pathway may promote astrocytic migration.ConclusionThese results demonstrated that a novel autocrine pathway for BK-induced MMP-9 protein expression is mediated through activation of STAT3 by native COX-2/PGE2-mediated c-Src/Jak2/ERK cascades in brain astrocytes.9AsGp9CBXvHHi3gF-G6FRAVideo

GRK2 Dictates a Functional Switch of the Peripheral Mu-Opioid Receptor.

The peripheral mu-opioid receptor (MOR) has been recognized as a potential target to provide safer analgesia with reduced central side effects. Although analgesic incompetence of the peripheral MOR in the absence of inflammation was initially identified more than a decade ago, there has been very limited investigation into the underlying signaling mechanisms. Here we identify that G protein-coupled receptor kinase 2 (GRK2) constitutively interacts with the MOR in peripheral sensory neurons to suppress peripheral MOR activity. Brief exposure to bradykinin (BK) causes uncoupling of GRK2 from the MOR and subsequent restoration of MOR functionality in dorsal root ganglion (DRG) neurons. Interestingly, prolonged BK treatment induces constitutive activation of the MOR through a mechanism that involves protein kinase C (PKC) activation. After silencing Raf kinase inhibitory protein (RKIP) by RNA interference, BK-induced constitutive MOR activation is completely abrogated, which agrees with previous findings that BK activates PKC signaling to initiate GRK2 sequestration by RKIP. Furthermore, we demonstrate that constitutive, peripheral MOR activity requires GRK2 uncoupling and that the FDA-approved SSRI paroxetine promotes this state of uncoupling. Collectively, these results indicate that GRK2 tightly regulates MOR functional states and controls constitutive MOR activity in peripheral sensory neurons, supporting the potential for targeting the kinase to provide safer analgesia.