Sigma-1 receptor modulation fine-tunes KV1.5 channels and impacts pulmonary vascular function.

KV1.5 channels are key players in the regulation of vascular tone and atrial excitability and their impairment is associated with cardiovascular diseases including pulmonary arterial hypertension (PAH) and atrial fibrillation (AF). Unfortunately, pharmacological strategies to improve KV1.5 channel function are missing. Herein, we aimed to study whether the chaperone sigma-1 receptor (S1R) is able to regulate these channels and represent a new strategy to enhance their function. By using different electrophysiological and molecular techniques in X. laevis oocytes and HEK293 cells, we demonstrate that S1R physically interacts with KV1.5 channels and regulate their expression and function. S1R induced a bimodal regulation of KV1.5 channel expression/activity, increasing it at low concentrations and decreasing it at high concentrations. Of note, S1R agonists (PRE084 and SKF10047) increased, whereas the S1R antagonist BD1047 decreased, KV1.5 expression and activity. Moreover, PRE084 markedly increased KV1.5 currents in pulmonary artery smooth muscle cells and attenuated vasoconstriction and proliferation in pulmonary arteries. We also show that both KV1.5 channels and S1R, at mRNA and protein levels, are clearly downregulated in samples from PAH and AF patients. Moreover, the expression of both genes showed a positive correlation. Finally, the ability of PRE084 to increase KV1.5 function was preserved under sustained hypoxic conditions, as an in vitro PAH model. Our study provides insight into the key role of S1R in modulating the expression and activity of KV1.5 channels and highlights the potential role of this chaperone as a novel pharmacological target for pathological conditions associated with KV1.5 channel dysfunction.

Intranasal metformin treatment ameliorates cognitive functions via insulin signaling pathway in ICV-STZ-induced mice model of Alzheimer's disease.

AIMS: The relationship between type 2 diabetes and Alzheimer's disease (AD) provides evidence that insulin and insulin sensitizers may be beneficial for the treatment of AD. The present study investigated the effect and mechanism of action of intranasal metformin treatment on impaired cognitive functions in an experimental mice model of AD. MAIN METHODS: Intracerebroventricularly (ICV) streptozotocin (STZ)-injected mice were treated with intranasal or oral metformin for 4 weeks. Learning and memory functions were evaluated using Morris water maze. Metformin and Aß42 concentrations were determined by liquid chromatography tandem mass spectrometry and ELISA respectively. The expressions of insulin receptor, Akt and their phosphorylated forms were determined in the hippocampi and cerebral cortices of mice. KEY FINDINGS: ICV-STZ-induced AD mice displayed impaired learning and memory functions which were improved by metformin treatment. ICV-STZ injection or intranasal/oral metformin treatments had no effect on blood glucose concentrations. Intranasal treatment yielded higher concentration of metformin in the hippocampus and lower in the plasma compared to oral treatment. ICV-STZ injection and metformin treatments did not change amyloid ß-42 concentration in the hippocampus of mice. In hippocampal and cortical tissues of ICV-STZ-induced AD mice, insulin receptor (IR) and Akt expressions were unchanged, while phosphorylated insulin receptor (pIR) and pAkt expressions decreased compared to control. Metformin treatments did not change IR and Akt expressions but increased pIR and pAkt expressions. SIGNIFICANCE: The present study showed for the first time that intranasal metformin treatment improved the impaired cognitive functions through increasing insulin sensitivity in ICV-STZ-induced mice model of AD.

Scutellaria brevibracteata Stapf and active principles with anti-inflammatory effects through regulation of NF-κB/COX-2/iNOS pathways.

Inflammation is a critical defense mechanism for body which is crucial for wound healing. However, when it spirals out of control and becomes chronic, it possesses a risk for diseases such as metabolic disorders or cancer. Scutellaria L. species are known with their potent anti-inflammatory effects. We aim to investigate the potential anti-inflammatory effects of Scutellaria brevibracteata Stapf and its active principles with underlying mechanisms using both in vitro and in vivo methods. Aqueous extract of S. brevibracteata (SB) was chromatographed on several columns to get 6 main fractions and 2 pure compounds. The levels of IL-6, NO, and TNF-α were measured in LPS-induced RAW 264.7 cells. In vivo carrageenan-induced paw edema test and immunoblotting in these paws were performed. SB was significantly reduced the inflammation in LPS-induced RAW cells via IL-6 and TNF-α cytokines at 200 µg/mL (P < 0.001) and paw edema via COX-2 and iNOS (P < 0.001). 3-O-methyl kaempferol, isolated from SB, reduced the production of NO, IL-6 and TNF-α in LPS-stimulated macrophages at 50 µM (P < 0.001); it significantly diminished inflammation-induced edema and COX-2, iNOS, and NF-κB protein levels were reduced compared to control group (P < 0.001). It was noticed that the methoxylation of C3 position has no effect on the anti-inflammatory mechanism of action comparing with kaempferol. In vivo studies indicated 3-O-methyl kaempferol was as effective as indomethacin. Considering the side effects caused by indomethacin, this compound can be evaluated as a novel drug candidate in inflammatory diseases with fewer side effects.

Anti-inflammatory effects of oral and intraperitoneal administration of cerium oxide nanoparticles on experimental hepatic ischemia-reperfusion injury.

Objectives: Hepatic ischemia-reperfusion (IR) injury occurs in liver surgery, resection, and transplantation. Reactive oxygen species (ROS) produced following IR starts the cascade of cell damage, necrosis/apoptosis, and proinflammatory responses by activating intracellular signaling cascade to drive hepatocellular damage. Cerium oxide nanoparticles (CONPs) act as anti-inflammatory and antioxidant agents. Thus, we evaluated the protective effects of oral (o.g.) and intraperitoneal (i.p.) administration of CONPs on hepatic IR injury. Material and Methods: Mice were randomly divided into five groups: control, sham, IR protocol, CONP+IR (i.p.), and CONP+IR (o.g.). Mouse hepatic IR protocol was applied to the animals in the IR group. CONPs (300 µg/kg) were administered 24 hours before IR protocol. Blood and tissue samples were taken after the reperfusion period. Results: Hepatic IR injury markedly increased enzyme activities, tissue lipid peroxidation, myeloperoxidase (MPO), xanthine oxidase (XO), nitrite oxide (NO), and tissue nuclear factor kappa-B (NF-κB) p65 levels, plasma pro-inflammatory cytokines, chemokines, and adhesion molecules while decreasing antioxidant markers and caused pathological changes in hepatic tissue. The expression of tumor necrosis factor alpha (TNF-α), matrix metalloproteinase 2 (MMP-2), and 9 increased, and tissue inhibitor matrix metalloproteinase 1 (TIMP-1) expression decreased in the IR group. Pretreatment with CONPs o.g. and i.p. 24 hours before hepatic ischemia improved the biochemical parameters above and alleviated the histopathological findings. Conclusion: Results of the present study demonstrate a significant reduction in liver degeneration by administering CONPs via i.p. and o.g. route in an experimental liver IR model, suggesting that CONPs have the extensive potential to prevent hepatic IR injury.

Investigation of the relationship between chronic montelukast treatment, asthma and depression-like behavior in mice.

In 2008, the Food and Drug Administration of the US issued a warning about the neuropsychiatric side effects of montelukast. Previous clinical studies on montelukast have reported conflicting results and, to the best of our knowledge, no experimental studies concerning these side effects had been conducted. In the current study, the effect of montelukast on depression-like behavior in an ovalbumin (OVA)-induced mouse model was investigated. A total of 3 OVA challenges were applied at 2 week intervals for the persistence of asthma. Depression-like behavior was assessed using forced swim tests following each challenge and locomotor activities were evaluated using open field tests. At the end of the current study, plasma montelukast concentrations were measured and the development of asthma and effect of montelukast treatment were histopathologically examined. Inflammation scores that were increased in the OVA mice following all challenges were indicated to be reduced by montelukast treatment. The immobility time of mice increased beginning with the first challenge and this was also reduced by montelukast treatment. Montelukast administration to the control mice did not alter immobility times. Moreover, motor activity of the OVA and montelukast-treated mice were not altered. The results indicated there was no association between chronic montelukast treatment and depression. Furthermore, the chronic administration of montelukast to non-asthmatic mice did not increase immobility. However, depressive behavior increased at all time points in the OVA mice. These results indicated that chronic montelukast treatment is not associated with depression-like behavior and confirmed the association between asthma and depression. Further studies are required to provide an improved understanding of the neuropsychiatric side effects of montelukast.

New 2-Pyrazoline and Hydrazone Derivatives as Potent and Selective Monoamine Oxidase A Inhibitors.

Thirty compounds having 1-[2-(5-substituted-2-benzoxazolinone-3-yl) acetyl]-3,5-disubstitutedphenyl-2-pyrazoline structure and nine compounds having N'-(1,3-disubstitutedphenylallylidene)-2-(5-substituted-2-benzoxazolinone-3-yl)acetohydrazide skeleton were synthesized and evaluated as monoamine oxidase (MAO) inhibitors. All of the compounds exhibited selective MAO-A inhibitor activity in the nanomolar or low micromolar range. The results of the molecular docking for hydrazone derivatives supported the in vitro results. Five compounds, 6 (0.008 µM, Selectivity Index (SI): 9.70 × 10-4), 7 (0.009 µM, SI: 4.55 × 10-5), 14 (0.001 µM, SI: 8.00 × 10-4), 21 (0.009 µM, SI: 1.37 × 10-5), and 42 (0.010 µM, SI: 5.40 × 10-6), exhibiting the highest inhibition and selectivity toward hMAO-A and nontoxic to hepatocytes were assessed for antidepressant activity as acute and subchronic in mice. All of these five compounds showed significant antidepressant activity with subchronic administration consistent with the increase in the brain serotonin levels and the compounds crossed the blood-brain barrier according to parallel artificial membrane permeation assay. Compounds 14, 21, and 42 exhibited an ex vivo MAO-A profile, which is highly consistent with the in vitro data.

The Cardiopulmonary Effects of the Calcitonin Gene-related Peptide Family.

Cardiopulmonary diseases are very common among the population. They are high-cost diseases and there are still no definitive treatments. The roles of members of the calcitonin-gene related-peptide (CGRP) family in treating cardiopulmonary diseases have been studied for many years and promising results obtained. Especially in recent years, two important members of the family, adrenomedullin and adrenomedullin2/intermedin, have been considered new treatment targets in cardiopulmonary diseases. In this review, the roles of CGRP family members in cardiopulmonary diseases are investigated based on the studies performed to date.

Mechanism of adrenomedullin 2/intermedin mediated vasorelaxation in rat main pulmonary artery.

Adrenomedullin 2/intermedin (AM2/IMD) is a member of calcitonin related gene peptide family and an important nitric oxide mediated vasorelaxant in various vascular beds. However, the mechanism of post receptor-interaction is not clear and may differ depending on tissue type and species. In this study, we aimed to investigate the exact mechanism and the role of BKCa and calcium channels on the vasorelaxant effect of AM2/IMD in rat PA. Changes in the AM2/IMD-mediated vasorelaxation were evaluated in the presence of various inhibitors. CGRP(8-37) (10-6 M), L-NAME (10-4 M), ODQ (10-5 M), SQ22536 (10-4 M), H89 (10-6 M), TEA (10-2 M), iberiotoxin (3 × 10-7 M), and verapamil (10-5 M), all partly or completely inhibited the vasorelaxation. The relaxation was also abolished by removal of the endothelium, or in KCl precontracted PAs. AM2/IMD did not elicit vasorelaxation in the Ca2+-free conditions. However, the vasorelaxation was not inhibited with AM(22-52) (10-6 M), 4-AP (3 × 10-3 M), glibenclamide (10-5 M), apamin (3 × 10-7 M), TRAM-34 (10-5 M), and La+3 (10-4 M). AM2/IMD -induced changes in intracellular calcium levels and isometric force were monitored simultaneously in fura-2-loaded, endothelium-intact PAs. The AM2/IMD-induced increase in intracellular Ca2+ concentration was inhibited in the presence of iberiotoxin and verapamil, whereas no change was observed with La3+ incubation. Our data suggest that the cAMP/PKA pathway is one of the important pathways AM2/IMD-induced vasorelaxation. AM2/IMD acts through activation of endothelial BKCa and subsequently causes hyperpolarization of the endothelial cell membrane. The hyperpolarization induces Ca2+ influx, which leads to NO production and subsequent vasorelaxation.

Effect of intermedin/adrenomedullin2 on the pulmonary vascular bed in hypoxia-induced pulmonary hypertensive rats.

AIMS: This study aimed to investigate the effect and mechanism of action of intermedin/adrenomedullin2 (IMD/AM2) on the pulmonary vascular bed in pulmonary hypertensive rats. MATERIALS AND METHODS: Male Sprague-Dawley rats were exposed to hypobaric hypoxia for 3 weeks to induce pulmonary hypertension (PHT). The development of PHT was confirmed by histopathological analyses and measurement of hematocrit, basal perfusion pressure, and right ventricle hypertrophy. Subsequently, the effect of IMD/AM2 in pulmonary hypertensive rats was assessed with both, isolated organ bath and isolated lung perfusion studies. KEY FINDINGS: In the PHT group, the basal perfusion pressure and % hematocrit were increased, and right ventricle hypertrophy occurred after 3 weeks of hypoxia exposure. Increased medial wall thickness was also observed in the pulmonary artery with histopathological analysis. In the PHT, the nitric oxide-mediated vasodilation caused by IMD/AM2 in the pulmonary vascular bed and this was as potent as the control group. Acetylcholine responses were also protected in pulmonary hypertensive rats. SIGNIFICANCE: Our results showed for the first time in in vitro studies that IMD/AM2 administration causes potent, concentration-dependent vasodilation in the main and resistance pulmonary arteries of rats with PHT. Based on these results, IMD/AM2 might be considered as a future therapeutic target for PHT treatment.

The effects of 5-hydroxytryptophan on carrageenan-induced mouse paw oedemas / Efeito de 5-hidroxitriptofano no edema de pata de camundongo induzido por carragenina

ABSTRACT Objective 5-Hydroxytryptophan is the precursor compound of serotonin biosynthesis. The oral absorption of 5-Hydroxytryptophan is close to 100% and, unlike serotonin, it crosses the blood-brain barrier freely. 5-Hydroxytryptophan has been used as a food supplement for many years to treat anxiety and depression. Recent studies have shown that 5-Hydroxytryptophan suppresses the pro-inflammatory mediators and is effective in some inflammatory diseases, such as arthritis and allergic asthma. However, the role of 5-Hydroxytryptophan supplements on acute peripheral inflammation has not been investigated yet. In this study, the in vivo anti-inflammatory activity of 5-Hydroxytryptophan was evaluated with a carrageenan-induced paw oedema test in mice. Methods For the investigation of the acute antiinflammatory activity, single oral doses of 5-Hydroxytryptophan (1.5, 5 and 20mg/kg) were given to mice 1.5 hours prior to the carrageenan test. For chronic activity, the same oral doses were administered daily for two weeks prior to the carrageenan test on the 14th day. To induce inflammation, 0.01mL of 2% carrageenan was injected into the paws of mice. Results Supplementation with 5-Hydroxytryptophan significantly reduced inflammation in a dose-independent manner which was irrespective of the duration of exposure (per cent inhibition in acute experiments was 35.4%, 20.9%, 24.0%, and per cent inhibition in chronic experiments was 29.5%, 35.3%, 40.8% for the doses of 1.5, 5, and 20mg/kg, respectively). Conclusion Our findings demonstrate for the first time that 5-HTP supplements have the potential of suppressing the measures of acute peripheral inflammation. It is suggested that, apart from several diseases where serotonin is believed to play an important role, including depression, patients with inflammatory conditions may also benefit from 5-HTP.

RESUMO Objetivo O 5-hidroxitriptofano (5-HTP) é o composto precursor da biossíntese da serotonina. A absorção oral do 5-HTP é próxima a 100% e, ao contrário da serotonina, atravessa a barreira hematoencefálica livremente. O 5-HTP tem sido usado como suplemento alimentar por muitos anos na ansiedade e na depressão. Estudos recentes demonstraram que o 5-HTP suprime os mediadores pró-inflamatórios e é eficaz em algumas doenças inflamatórias, como artrite e asma alérgica. No entanto, o papel dos suplementos de 5-HTP na inflamação periférica aguda ainda não foi investigado. Neste estudo, a atividade anti-inflamatória in vivo do 5-HTP foi avaliada por meio do teste de edema de pata induzido por carragenina em ratos. Métodos Para a atividade aguda, doses orais únicas de 5 -HTP (1,5, 5 e 20 mg/kg) foram dados aos ratos 1,5 horas antes do teste da carragenina. Para a atividade crônica, as mesmas doses orais foram dadas cada dia durante duas semanas antes do teste da carragenina no 14º dia. 0,01ml da carragenina a 2% foi injetado nas patas dos ratos a fim de induzir a inflamação. Resultados A suplementação com 5-HTP reduziu significativamente a inflamação de uma maneira independente da dose, que foi independente da duração da exposição (por cento de inibição em experimentos agudos; 35,4%, 20,9%, 24,0% e por cento de inibição em experimentos crônicos; 29,5%, 35,3%, 40,8% para as doses de 1.5, 5 e 20 mg/kg respectivamente). Conclusão Nossas conclusões demonstram pela primeira vez que os suplementos de 5-HTP têm potencial para suprimir os sintomas de inflamação periférica aguda. É sugerido que, além de várias doenças em que se acredita que a serotonina tem uma função importante, incluindo a depressão, os pacientes com doenças inflamatórias também podem se beneficiar do 5-HTP.