High magnesium mitigates the vasoconstriction mediated by different types of calcium influx from monocrotaline-induced pulmonary hypertensive rats.

NEW FINDINGS: What is the central question of this study? What is the involvement of Mg2+ in mitigating the vasoconstriction in pulmonary arteries and smaller pulmonary arteries in the monocrotaline-induced pulmonary arterial hypertension (MCT-PAH) rat model? What are the main finding and its importance? Both store-operated Ca2+ entry- and receptor-operated Ca2+ entry-mediated vasoconstriction were enhanced in the MCT-PAH model. High magnesium inhibited vasoconstriction by directly antagonizing Ca2+ and increasing NO release, and this was more notable in smaller pulmonary arteries. ABSTRACT: Increased extracellular magnesium concentration has been shown to attenuate the endothelin-1-induced contractile response via the release of nitric oxide (NO) from the endothelium in proximal pulmonary arteries (PAs) of chronic hypoxic mice. Here, we further examined the involvement of Mg2+ in the inhibition of vasoconstriction in PAs and distal smaller pulmonary arteries (sPAs) in a monocrotaline-induced pulmonary arterial hypertension (MCT-PAH) rat model. The data showed that in control rats vasoconstriction in sPAs is more intense than that in PAs. In MCT-PAH rats, store-operated Ca2+ entry (SOCE)- and receptor-operated Ca2+ entry (ROCE)-mediated contraction were significantly strengthened. However, there was no upregulation of the vasoconstriction mediated by voltage-dependent calcium entry (VDCE). Furthermore, high magnesium greatly inhibited VDCE-mediated contraction in PAs rather than sPAs, which was the opposite of the ROCE-mediated contraction. Moreover, monocrotaline pretreatment partly eliminated the endothelium-dependent vasodilatation in PAs, which in sPAs, however, was still promoted by magnesium due to the increased NO release in pulmonary microvascular endothelial cells (PMVECs). In conclusion, the findings suggest that both SOCE- and ROCE-mediated vasoconstriction in the MCT-PAH model are enhanced, especially in sPAs. The inhibitory effect of high magnesium on vasoconstriction can be achieved partly by its direct role as a Ca2+ antagonist and partly by increasing NO release in PMVECs.

Analysis of oral microbiota in patients with obstructive sleep apnea-associated hypertension.

Obstructive sleep apnea-hypopnea syndrome (OSAHS) is an independent risk factor for hypertension (HTN). The oral microbiota plays a pathophysiological role in cardiovascular diseases; however, there are few reports directly investigating and identifying the organisms involved in OSAHS-related HTN. Therefore, this study aimed to identify those organisms. We obtained 139 oral samples and determined the microbiome composition using pyrosequencing and bioinformatic analyses of the 16S rRNA. We examined the fasting levels of cytokines and homocysteine in all participants and analyzed the correlations between the oral microbiota and homocysteine levels. We determined the molecular mechanism underlying HTN by investigating the genetic composition of the strains in the blood. We detected higher relative abundances of Porphyromonas and Aggregatibacter and elevated proinflammatory cytokines in patients with OSAHS of varying severity compared with individuals without OSAHS; however, the two organisms were not measured in the blood samples from all participants. High levels of specific Porphyromonas bacteria were detected in patients with OSAHS with and without HTN, whereas the relative abundance of Aggregatibacter was negatively correlated with the homocysteine level. The receiver operating characteristic curve analysis of controls and patients with OSAHS resulted in area under the curve values of 0.759 and 0.641 for patients with OSAHS with or without HTN, respectively. We found that the predictive function of oral microbiota was different in patients with OSAHS with and without HTN. However, there was no direct invasion by the two organisms causing endothelial cell injury, leading to speculation regarding the other mechanisms that may lead to HTN. Elucidating the differences in the oral microbiome will help us understand the pathogenesis of OSAHS-related HTN.

Disruption of sleep architecture in Prevotella enterotype of patients with obstructive sleep apnea-hypopnea syndrome.

INTRODUCTION: Intermittent hypoxia and sleep fragmentation are critical pathophysiological processes involved in obstructive sleep apnea-hypopnea syndrome (OSAHS). Those manifestations independently affect similar brain regions and contribute to OSAHS-related comorbidities that are known to be related to the host gut alteration microbiota. We hypothesized that gut microbiota disruption may cross talk the brain function via the microbiota-gut-brain axis. Thus, we aim to survey enterotypes and polysomnographic data of patients with OSAHS. METHODS: Subjects were diagnosed by polysomnography, from whom fecal samples were obtained and analyzed for the microbiome composition by variable regions 3-4 of 16S rRNA pyrosequencing and bioinformatic analyses. We examined the fasting levels of interleukin-6 and tumor necrosis factor-alpha of all subjects. RESULTS: Three enterotypes Bacteroides, Ruminococcus, and Prevotella were identified in patients with OSAHS. Arousal-related parameters or sleep stages are significantly disrupted in apnea-hypopnea index (AHI) ≥15 patients with Prevotella enterotype; further analysis this enterotype subjects, obstructive, central, and mixed apnea indices, and mean heart rate are also significantly elevated in AHI ≥15 patients. However, blood cytokines levels of all subjects were not significantly different. CONCLUSIONS: This study indicates the possibility of pathophysiological interplay between enterotypes and sleeps structure disruption in sleep apnea through a microbiota-gut-brain axis and offers some new insight toward the pathogenesis of OSAHS.

Increased caveolin-1 expression enhances the receptor-operated Ca2+ entry in the aorta of two-kidney, one-clip hypertensive rats.

NEW FINDINGS: What is the central question of this study? The aim was to examine and compare the contributions of caveolin-1 to the contractile responses mediated by L-type voltage-dependent calcium channels, store-operated Ca2+ channels and receptor-operated Ca2+ channels in two different types of arteries from two-kidney, one-clip hypertensive rats. What is the main finding and its importance? We demonstrated that the density of caveolae and caveolin-1 expression were significantly upregulated in the aorta of two-kidney, one-clip hypertensive rats, but not in the third-order branches of mesenteric arteries. We highlight that caveolin-1 plays an important role in aortic constriction by enhancing receptor-operated Ca2+ entry in the hypertensive rat model. ABSTRACT: Calcium and its multiple regulatory mechanisms are crucial for the development of hypertension. Among these regulatory mechanisms, store-operated Ca2+ entry (SOCE) and receptor-operated Ca2+ entry (ROCE) mediate agonist-induced calcium influx, contributing to vascular contraction. The SOCE and ROCE are regulated by a variety of mechanisms involving caveolin-1 (Cav1), which has been found to be strongly associated with hypertension in gene polymorphism. In the present study, we investigated the role of Cav1 during the enhanced activity of calcium channels in hypertensive arteries. We demonstrated that the expression level of Cav1 was significantly increased in the aorta of two-kidney, one-clip (2K1C) hypertensive rats. The disruption of caveolae by methyl-ß-cyclodextrin did not cause a marked difference in agonist-induced vasoconstriction in the third-order branches of the mesenteric arteries but strongly suppressed the aortic contractile response to endothelin-1 in the 2K1C group, which was not found in the control group. The increase in Cav1 by introduction of Cav1 scaffolding domain enhancing peptide promoted the 1-oleoyl-2-acetyl-glycerol-induced ROCE in hypertensive aortic smooth muscle cells but did not enhance the cyclopiazonic acid-induced SOCE. In the resistance arteries, similar changes were not observed, and no statistical changes of Cav1 expression were evident in the third-order branches of the mesenteric arteries. Our results indicate that increased Cav1 expression might promote the altered [Ca2+ ]i -induced aortic vasoreactivity by enhancing ROCE and be involved in the pathogenesis of hypertension.

Magnesium attenuates endothelin-1-induced vasoreactivity and enhances vasodilatation in mouse pulmonary arteries: Modulation by chronic hypoxic pulmonary hypertension.

NEW FINDINGS: What is the central question of this study? The central goal of this study was to elucidate the role of magnesium in the regulation of pulmonary vascular reactivity in relationship to hypoxic pulmonary hypertension. What is the main finding and its importance? We found that magnesium is essential for normal vasoreactivity of the pulmonary artery. Increasing the magnesium concentration attenuates vasoconstriction and improves vasodilatation via release of nitric oxide. Pulmonary hypertension is associated with endothelial dysfunction resulting in the suppression of magnesium modulation of vasodilatation. These results provide evidence that magnesium is important for the modulation of pulmonary vascular function. ABSTRACT: Pulmonary hypertension (PH) is characterized by enhanced vasoreactivity and sustained pulmonary vasoconstriction, arising from aberrant Ca2+ homeostasis in pulmonary arterial (PA) smooth muscle cells. In addition to Ca2+ , magnesium, the most abundant intracellular divalent cation, also plays crucial roles in many cellular processes that regulate cardiovascular function. Recent findings suggest that magnesium regulates vascular functions by altering the vascular responses to vasodilator and vasoactive agonists and affects endothelial function by modulating endothelium-dependent vasodilatation in hypertension. Administration of magnesium also decreased pulmonary arterial pressure and improved cardiac output in animal models of PH. However, the role of magnesium in the regulation of pulmonary vascular function related to PH has not been studied. In this study, we examined the effects of magnesium on endothelin-1 (ET-1)-induced vasoconstriction, ACh-induced vasodilatation and the generation of NO in PAs of normoxic mice and chronic hypoxia (CH)-treated mice. Our data showed that removal of extracellular magnesium suppressed vasoreactivity of PAs to both ET-1 and ACh. A high concentration of magnesium (4.8 mm) inhibited ET-1-induced vasoconstriction in endothelium-intact or endothelium-disrupted PAs of normoxic and CH-treated mice, and enhanced the ACh-induced production of NO in PAs of normoxic mice. Moreover, magnesium enhanced ACh-induced vasodilatation in PAs of normoxic mice, and the enhancement was completely abolished after exposure to CH. Hence, in this study we demonstrated that increasing the magnesium concentration can attenuate the ET-1-induced contractile response and improve vasodilatation via release of NO from the endothelium. We also demonstrated that chronic exposure to hypoxia can cause endothelial dysfunction resulting in suppression of the magnesium-dependent modulation of vasodilatation.

[Comparison of the synchronous changes of vascular tension and intracellular Ca2+ signal in third-order branches of mesenteric arteries under the different objective amplification of confocal microscope].

This study was aimed to establish an optimized method to observe the synchronous changes of vascular tension and intracellular Ca2+ signal in the third-order branches of mesenteric arteries (sMA, diameter: 100-300 µm). The vascular tension and intracellular Ca2+ signal changes in response to potassium chloride (KCl), endothelin-1 (ET-1) and Gd3+ were detected using confocal wire myograph system and confocal laser scanning microscopy imaging technique, respectively. The experimental results were analyzed to explore the optimal experimental conditions. The results showed that KCl caused contraction in sMA significantly, and the intracellular Ca2+ level of vascular smooth muscle cells (VSMCs) was also increased under 20× and 40× objective lens. Compared with those under the 40× objective lens, the Ca2+ signal change was larger and the fluorescence value was more stable under the 20× objective lens, whereas the Ca2+ signal change was not obvious under the 10× objective lens. ET-1 (1-10 nmol/L) caused concentration dependent contraction in sMA significantly, and the intracellular Ca2+ signal of VSMCs was also enhanced in a concentration dependent manner. Additionally, Gd3+ significantly reduced the contraction of sMA and the intracellular Ca2+ signal of VSMCs caused by ET-1. The results suggest that the intracellular Ca2+ signal of VSMCs changes with vascular contraction or relaxation caused by the agonists or antagonists of Ca2+ channels. We successfully recorded both changes synchronously using confocal wire myograph system and confocal laser scanning microscopy imaging technique at the same time. Based on the analysis of the experimental results, we concluded that 20× objective lens provides the best experimental condition. Compared to combination of vascular tone detection method and real-time cellular fluorescence imaging technique, the present synchronous method is convenient and helpful to reduce experimental error.

[Comparison of agonists-induced contraction between main and the third-order branches of pulmonary arteries in rats].

This study was designed to observe the differences between main pulmonary arteries and the third-order branches of pulmonary arteries in the contractile response to phenylephrine (Phen), endothelin-1 (ET-1) and potassium chloride (KCl). The vascular tension changes of main and the third-order branches of pulmonary arteries induced by KCl, ET-1 and Phen were recorded by traditional vascular tone detection methods and microvascular ring technique, respectively. The results showed that Phen could cause a significant contraction in main pulmonary arteries, but did not induce apparent contraction in the third-order branches of pulmonary arteries. Compared with main pulmonary arteries, ET-1 contracted the third-order branches of pulmonary arteries with reduced maximal response value and PD2 value. In comparison with the main pulmonary arteries, contraction caused by KCl was enhanced in the third-order branches of pulmonary arteries. The results suggest that the vascular reactivity of main and the third-order branches of pulmonary arteries is different and it is important to study the vascular function of small branches of pulmonary arteries. This study could provide an important experimental basis for the further study on vascular function of small branches of pulmonary arteries and the functional changes in pulmonary hypertension.

Alterations in Caveolin-1 Expression and Receptor-Operated Ca2+ Entry in the Aortas of Rats with Pulmonary Hypertension.

BACKGROUND/AIMS: Alterations in intracellular Ca2+ concentration ([Ca2+]i) underlie the pathogenesis of various cardiovascular diseases. Caveolin-1 (Cav-1) is the primary functional protein associated with caveolae, which are invaginations in the plasma membrane, and is a regulator of [Ca2+]i signaling. Caveolae and Cav-1 increase the activity of store-operated Ca2+ channels (SOCC) in rat pulmonary arterial smooth muscle cells (PASMCs), and these enhancing effects were more pronounced in rats with pulmonary hypertension (PH). Classical transient receptor potential (TRPC) proteins are highly expressed in vascular smooth muscle cells, and these proteins form functional receptor-operated Ca2+ channels (ROCC) and SOCC in PASMCs. Previous studies suggested that functional and structural changes in aortas might occur during the pathological process of PH. Our data demonstrated that Cav-1 and TRPC were also abundant in the aorta smooth muscle cells (AoSMCs) of PH rats. However, previous PH research primarily focused on Ca2+ channels in pulmonary arteries, but not functional changes in Ca2+ channels in aortas. The contribution of Cav-1 of AoSMCs to alterations of Ca2+ signaling in aortic functions during the pathological process of PH has not been fully characterized. Therefore, this study investigated alterations in Cav-1 expression and the relationship of these changes to Ca2+ channels in AoSMCs of PH rats. METHODS: The present study examined physiological caveolae and Cav-1 expression and characterized the function of altered Cav-1 expression in rat aortas with PH. RESULTS: The appearance of caveolae with Cav-1 expression increased significantly in the aortas of rats with PH, but TRPC1 and TRPC6 expression was not altered. In vitro experiments demonstrated that caveolae contributed to phenylephrine, endothelin-1, and 1-oleoyl-2-acetyl-sn-glycerol (OAG)-induced aortic vasoreactivity, but KCl and cyclopiazonic acid had no effect, which suggests the vital ability of Cav-1 to regulate ROCC activity. The introduction of Cav-1 scaffolding domain peptide enhanced OAG-induced ROCC function in primary AoSMCs. CONCLUSION: Cav-1 is specifically associated with ROCC in aortas and plays a vital role in altering vasoreactivity, which affects cardiovascular diseases pathology. Caveolae and Cav-1 up-regulation may affect the function of ROCC in rat models of PH.

Increase in caveolae and caveolin-1 expression modulates agonist-induced contraction and store- and receptor-operated Ca(2+) entry in pulmonary arteries of pulmonary hypertensive rats.

Caveolin-1 (Cav-1) is a major component protein associated with caveolae in the plasma membrane and has been identified as a regulator of store-operated Ca(2+) entry (SOCE) and receptor-operated Ca(2+) entry (ROCE). However, the contributions of caveolae/Cav-1 of pulmonary arterial smooth muscle cells (PASMCs) to the altered Ca(2+) signaling pathways in pulmonary arteries (PAs) during pulmonary hypertension (PH) have not been fully characterized. The present study quantified caveolae number and Cav-1 expression, and determined the effects of caveolae disruption on ET-1, cyclopiazonic acid (CPA) and 1-Oleoyl-2-acetyl-glycerol (OAG)-induced contraction in PAs and Ca(2+) influx in PASMCs of chronic hypoxia (CH)- and monocrotaline (MCT)-induced PH rats. We found that the number of caveolae, and the Cav-1 mRNA and protein levels were increased significantly in PASMCs in both PH models. Disruption of caveolae by cholesterol depletion with methyl-ß-cyclodextrin (MßCD) significantly inhibited the contractile response to ET-1, CPA and OAG in PAs of control rats. ET-1, SOCE and ROCE-mediated contractile responses were enhanced, and their susceptibility to MßCD suppression was potentiated in the two PH models. MßCD-induced inhibition was reversed by cholesterol repletion. Introduction of Cav-1 scaffolding domain peptide to mimic Cav-1 upregulation caused significant increase in CPA- and OAG-induced Ca(2+) entry in PASMCs of control, CH and MCT-treated groups. Our results suggest that the increase in caveolae and Cav-1 expression in PH contributes to the enhanced agonist-induced contraction of PA via modulation of SOCE and ROCE; and targeting caveolae/Cav-1 in PASMCs may provide a novel therapeutic strategy for the treatment of PH.