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Chronic hypertension is associated with an impaired vascular relaxation caused by an increased vascular tone; however, the underlying mechanisms are not fully understood in human patients. The present study was to investigate whether large-conductance Ca 2+ - and voltage-activated K + (BK Ca ) channels are involved in dysfunctional relaxation of artery in Han Chinese patients with hypertension using the perforated patch clamp, inside-out single-channel, and macromembrane patch recording techniques to determine whole-cell current, spontaneous transient outward current, open probability, and Ca 2+ sensitivity and the reverse transcription polymerase chain reaction and Western blot analysis to examine the gene and protein expression of α-subunit (KCa1.1) and β1-subunit (KCNMB1) of BK Ca channels in isolated human vascular smooth muscle cells and mesenteric arteries from normotensive and hypertensive patients. It was found that whole-cell current density, spontaneous transient outward current, and Ca 2+ sensitivity, but not single-channel open probability and slope conductance, were significantly decreased in vascular smooth muscle cells from patients with hypertension. Interestingly, mRNA and protein levels of KCNMB1, but not KCa1.1, were reduced in the arterial tissue from patients with hypertension. These results demonstrate for the first time that whole-cell current, spontaneous transient outward current, and Ca 2+ sensitivity of BK Ca channels are reduced in human vascular smooth muscle cells, which resulted from downregulation of β 1-subunit of the channel. This may account, at least in part, for the dysfunction of artery relaxation in Han Chinese patients with primary hypertension.

Background Large‐conductance calcium‐ and voltage‐activated potassium channels ( BK C a channels) play important roles in the maintenance of vascular tone, and their dysregulation is associated with abnormal vascular relaxation and contraction. We tested the changes in BK C a channel properties in patients at different ages to assess the effects of hypertension and aging on the functional changes of BK C a channels. Methods and Results Patch clamp was performed to detect the activities of BK C a channels in freshly isolated human mesenteric artery smooth muscle cells from younger patients (aged ≤45 years) without hypertension, older patients (aged ≥65 years) without hypertension, and older patients with hypertension. The expression of mRNA and protein from BK C a channels was evaluated by reverse transcription polymerase chain reaction and Western blot analysis, respectively. Results showed that the whole‐cell current density, spontaneous transient outward current, and Ca 2+ sensitivity of the artery smooth muscle cells were significantly decreased in the older patients with hypertension; the decreases were insignificant in the older patients without hypertension, although a clear tendency to have spontaneous transient outward current was detected in these patients. The expression of both mRNA and protein of BK C a subunits α and β1 was significantly decreased in the older patients with hypertension but not in the older patients without hypertension compared with the younger patients without hypertension. Conclusions Our findings demonstrate for the first time that hypertension is an important factor for the pathological alteration of the properties of BK C a channels in human mesenteric artery smooth muscle cells, and aging itself may also be a factor in these changes in the cells.

Introduction: Up4A, a novel endothelium-derived vasoactive agent, is proposed to play a role in cardiovascular disorder via activation of P2 receptors (P2R). Here, we evaluated the vasoactive effect of Up4A on coronary flow (CF) in ApoE knockout mice (KO) with and without high fat diet (HFD). Methods: Functional studies were conducted in isolated mouse hearts using Langendorff and isolated coronary arteries (diameter≈100 μm) using wire Myograph. Immunofluorescence was performed for P2X1R expression in isolated coronary arteries. Results: There was no significant difference in baseline CF (ml/min/g) among wild type (WT, 18.4±0.5), ApoE KO (17.6±1.1) and ApoE KO+HFD (15.9±1.0). Infusion of Up4A (10-9-10-5 M) resulted in a dose-dependent reduction in CF in WT (by ~38%), which was similar to ApoE KO (by ~40%). In isolated coronary arteries of WT, Up4A produced a mild contraction up to ~20%, indicating a vasoconstrictor effect of Up4A. Notably, Up4A further decreased CF in ApoE+HFD as compared to ApoE. Moreover, selective P2X1R antagonist MRS2159 (10 μM) restored Up4A-mediated decrease in CF more in ApoE+HFD than ApoE (Figure). Finally, the percentage expression of vasoconstrictor P2X1R on smooth muscle cells (SMC) of coronary arteries was higher in ApoE+HFD than ApoE, which was due to a markedly reduced vasodilator P2X1R expression on EC in ApoE+HFD (8.7±0.6 fluorescence intensity in A.U.) than ApoE (22.1±0.5), while P2X1R expression on SMC was similar between ApoE+HFD (6.3±0.4) and ApoE (8.4±0.3). Conclusion: Up4A exerts a vasoconstrictor influence in mouse coronary microcirculation. Up4A decreases CF in WT to a similar extent in ApoE, whereas HFD exacerbates Up4A-decreased CF in ApoE possibly through downregulation of vasodilator P2X1R on EC. These findings indicate that Up4A alters CF in atherosclerosis and P2X1R may represent a potential therapeutic target.

Previous data have showed that application of ivabradine, a selective blocker of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, leaded to divergent trends regarding proarrhythmic and antiarrhythmic effects in atrial fibrillation (AF). The aim of this study was to investigate the

Myricetin (Myr) is a flavonoid that exerts anti-arrhythmic effects. However, its potential effects on ion channels have remained elusive. The aim of the present study was to investigate the effects of Myr on Kv1.5 channels in HEK293 cells. The current of Kv1.5 channels (Ikur) in HEK293 cells was recorded using the whole-cell patch-clamp technique and the expression of the Kv1.5 protein was measured using western blot analysis 24 h after treatment with Myr. The results showed that 5 µM Myr significantly reduced Ikur from 215.04 ± 40.59 to 77.72 ± 17.94 pA/pF (P

Functional cardiomyocytes generated from human induced stem (h-iPS) cells can potentially offer an unlimited cell source for therapies in cardiac disease, cardiovascular regeneration medicine, and the study of cardiac development. In this study, we aimed to establish a method for differentiating h

Small conductance calcium activated potassium channels (SK) were found closely related to Atrial Fibrillation (AF). Dronedarone is recently used for the patients with paroxysmal AF or after conversion of persistent AF. The purpose of this study was to confirm the inhibitory effects of dronedarone on

Objectives: Atrial fibrillation (AF) is associated with a decrease in atrial ERP and ERP adaptation to rate as well as changes in atrial conduction velocity. The cellular changes in repolarization and the underlying ionic mechanisms in human AF are only poorly understood. Methods: Action potentials (AP) and ionic currents were studied with the patch clamp technique in single atrial myocytes from patients in chronic AF and compared to those from patients in stable sinus rhythm (SR). Results: The presence of AF was associated with a marked shortening of the AP duration and a decreased rate response of atrial repolarization. L-type calcium current ( I Ca,L) and the transient outward current ( I to) were both reduced about 70% in AF, whereas an increased steady-state outward current was detectable at test potentials between –30 and 0 mV. The inward rectifier potassium current ( I K1) and the acetylcholine-activated potassium current ( I KACh) were increased in AF at hyperpolarizing potentials. Voltage-dependent inactivation of the fast sodium current ( I Na) was shifted to more positive voltages in AF. Conclusions: AF in humans leads to important changes in atrial potassium and calcium currents that likely contribute to the decrease in APD and APD rate adaptation. These changes contribute to electrical remodeling in AF and are therefore important factors for the perpetuation of the arrhythmia.

Introduction: Traditional vectors for gene therapy are associated with various side effects including low delivery and/or transduction efficiency, immunogenicity, carcinogenicity, inability to pass...

According to the requirement that the higher education of China in the new term needs new educational strategy “to enhance the teaching quality”, the concept and the formation and operation of the effective mechanism of the refined teaching management of college are introduced in this article, to seek an optimal solution to enhance the teaching quality.

Induced pluriptent stem (IPS) cells have great potential for regenerative cardiac medicine, since it could serve as a source of regenerative cardiomyocytes due to their high potentials for self-renewal, proliferation, differentiation. Here we aimed to establish a method for reprogramming the human

Oncogenesis, development, invasion and metastasis of lung cancer are modulated by relative genes of lung cancer, and the expression, deletion or mutation of these genes are regulated by cell membrane signal transduction system and cell membrane ionic channels. The aim of this study is to explore the electrophysiological properties of human lung adenocarcinoma cell line A549 and cell proliferation affected by tetraethylammonium chloride (TEA), one potassium channel blocker, so as to know whether the voltage-gated potassium channels are required for the proliferation of A549 cell line.Ionic currents were recorded by whole-cell patch-clamp recording technique. The proliferation activity of A549 cells was measured by MTT assay.Membrane current was observed when cells were held at -70mV and test potentials ranged from -30 to +110mV. The current exhibited properties of voltage-dependent, outward rectification and no or little inactivation over the 500ms voltage pulse. Exposure of tumor cells to 10mmol/L TEA reduced the peak outward potassium current (evoked by depolarization to +110mV) from (1057.52±59.17)pA to (212.26±11.96)pA, the ratio of suppression was 79.92% (P0.01). Obvious inhibitive effect of TEA with different concentrations ranged from 20 to 60mmol/L on proliferation activity of the cells was found.The voltage-gated potassium channels exist in human lung adenocarcinoma cell line A549 and play a great role in proliferation of A549 cells. TEA can inhibit proliferation of A549 cell through blocking the potassium channels and the inhibition is dose-dependent.

To investigate the degeneration mechanics of outer hair cells in guinea pig.The mechanics of outer hair cells isolated by enzyme were observed under inverted microscope for 6-8 h continuously.Over half of living outer hair cells could keep good conditions in 6 hours. During the degeneration there was always a longitudinal fold line from tip to base. Presence or absence of calcium, as well as lossing of stereociliary bundle, couldn't change the conditions of out hair cells.Neither calcium nor stereociliary bundle is the decisive cause in keeping outer hair cells alive, and its degeneration may be basically related with something surrounding the cell.

The instructional pattern of Problem-based Learning, which requires teachers to be “organizers, guides and cooperators” of their students’ learning, is now becoming a trend in the development of medical education around the world. In order to be competent in all the above mentioned roles, teachers need to be equipped with corresponding qualities. In this article, basic qualities under the ideal PBL conditions are summarized from relevant literatures and teachers’ practical ability was also understood through analysis of requirements. Then the difference between the ideals and reality was discovered, also a question data base was set up accordingly as the starting point to train teachers and seek a solution to intrigue them to implement PBL. Thus, a teachers’ training system of “Five in One”, the integration of requirement analysis, objectives setting, methods selection, content design and quality assurance, was constructed.

PBL is a studying process that emphasizes how to enquire, understand and solve problems. Problem Base is a form of resource for PBL’s implementation and is indispensable to the overall practice of the latter. This article first sets about the two types of problems: disciplinary and conditional to collect problems from 6 aspects-- teacher’s reflection, students, colleagues, teaching supervision and reference literature; then manages the problems from 3 levels – junior, middle and senior–according to the above mentioned two types, and finally constructs a problem Base of “two types, six aspects and three levels”.

In order to meet the needs of electrophysiology outer hair cells, we investigate a method to separate outer hair cells simply and effectively.The basal membrane was dissected combined with spiral ligament and axis of cochlea, then incubated with enzyme, and then mechanically triturated by micropipette.A large number of living outer hair cells were obtained. Of 80% cells could keep good condition in 6 hours.If increasing the enzyme concentration slightly and the large part of cochlea without outer bone was enzymes, we can get living outer hair cells easily.

The purpose of the present study was to further study the characteristics and regulation of spontaneous transient outward currents (STOCs) in freshly isolated porcine coronary artery smooth muscle cells (ASMCs). STOCs were recorded using the perforated whole-cell patch-clamp configuration. STOCs were voltage-dependent and superimposed stochastically onto whole-cell Ca(2+)-activated-K(+) (BK(Ca)) currents. Charybdotoxin (ChTX, 200 nmol/L), a selective blocker of BK(Ca) channels, completely inhibited STOCs within 10 min. STOCs activity was greatly suppressed when extracellular Ca(2+) concentration decreased from 1.8 mmol/L to 200 nmol/L, further removal of Ca(2+) abolished STOCs activity. Ca(2+) ionophore A23187 (10 micromol/L) increased STOCs activity significantly. Verapamil (20 micromol/L) and CdCl(2) (200 micromol/L), two kinds of organic L-type voltage-dependent Ca(2+) channels (L-VDCCs) antagonists, had little effect on STOCs. In addition, the ryanodine receptors (RyRs) agonist caffeine (5 mmol/L) significantly activated STOCs. Application of ryanodine (50 micromol/L) to block RyRs abolished STOCs, subsequent washout of ryanodine or application of caffeine failed to reproduce STOCs activity. Inhibition of inositol 1,4,5-trisphosphate receptors (IP(3)Rs) by 2APB (40 micromol/L) greatly suppressed the activity of STOCs, application of caffeine (5 mmol/L) in the presence of 2APB caused a burst of outward currents followed by inhibition of STOCs. These results suggest that STOCs in porcine coronary ASMCs are mediated by BK(Ca) channels. Extracellular Ca(2+) is essential for STOCs activity, while Ca(2+) entry through L-VDCCs has little effect on STOCs. Intracellular Ca(2+) release induced by RyRs is responsible for the regulation of STOCs, whereas IP3Rs might also be involved.

Recent evidence has demonstrated that neural stem cells (NSC) can be expanded from a variety of sources, including embryos, fetuses, and adult bone marrow and brain tissue. We have previously reported the generation of adult rat bone marrow-derived cellular spheres that are morphologically and phenotypically similar to neurospheres derived from brain NSC. Here we show that adult human bone marrow-derived neural stem cells (HBM-NSC) are capable of generating spheres that are similar to brain neural-derived neurospheres. Additionally, we sought to promote proliferation and differentiation of HBM-NSC through transduction with nonreplicative recombinant adenovirus encoding the cDNA sequence for Gli, rADV-Gli-1; sonic hedgehog, rADV-Shh; or Nurr1, rADV-Nurr1. Immunocytochemistry and RT-PCR analysis showed that HBM-NSC could be efficiently expanded and differentiated in vitro and that HBM-NSC transduced with rADV-Gli-1 or rADV-Shh dramatically increased NSC time-related proliferation; however, Nurr1 had no effect on proliferation. We also transplanted HBM-NSC into chicken embryos to examine their potential function in vivo. We found that transduction of HBM-NSC with rADV-Gli-1 or rADV-Shh and subsequent transplantation into chicken embryos increased HBM-NSC proliferation, whereas rADV-Nurr1 promoted migration and differentiation in vivo. Our findings suggest that HBM-NSC can be efficiently expanded and differentiated in vitro and in vivo by overexpressing Gli-1, Shh or Nurr1.

Scalp acupuncture (SA) therapy on strokes has been empirically established and widely used in clinics in China. SA is particularly effective at ameliorating paralyses and speech disturbances, and the recovery rate is twice that for those treated with medication alone. To investigate the effects of SA on a scientific basis, we have developed a new experimental system that provides reliable controls and excludes psychological effects by using a genetic strain of rats, spontaneous hypertensive rats-stroke prone. Here we report that SA indeed has rapid and powerful effects to remove limb paralyses caused either by cerebral infarct or by cerebral haemorrhage. This model is well suited to study the mechanism of the effects of SA in parallel with clinical studies, and to describe the whole recovery process after the stroke onset.

Molecular and electrical remodeling of ion channels determining action potential duration has been proposed as a major mechanism in chronic atrial fibrillation. We investigated the mRNA expression of the cardiac L-type Ca2+-channel subunits alpha1c, alpha2/delta1, beta1a, and beta1b/c in atrial tissue of patients with chronic atrial fibrillation compared to patients in sinus rhythm. In addition, the mRNA expression of the 5-hydroxytryptamine type 4-, beta1-, and beta2-adrenergic receptors, which are known to stimulate the L-type Ca2+-current in human atrium, was analyzed and the effect of chronic beta-blocker treatment on the mRNA expression of these receptors and of the L-type Ca2+-channel subunits was assessed. Total RNA was isolated from right atrial appendages of patients in sinus rhythm and of patients with chronic atrial fibrillation. Then, semiquantitative RT-PCR using 18S RNA as the "housekeeping gene" was performed. In patients with chronic atrial fibrillation, there were only mild reductions in mRNA expression of the alpha1c-subunit (-15.5 %, p = 0.13), and of the beta1-subunit isoforms a and c (-13.3 %, p = 0.14 and -16.6%, p = 0.18, respectively). However, mRNA expression of the alpha2/delta1-subunit (-31.5 %, p < 0.01) and of the beta1-subunit isoform b (-39.9 %, p < 0.0005) was significantly reduced in patients with chronic AF. Taken together, the mRNA expression of the beta1-subunit isoforms b and c, which are splice variants, was significantly down-regulated by 26.5 % (p < 0.05) in these patients. The analysis of the beta1c/beta1b ratio resulted in a significant shift by 39.2 % (p < 0.0001) in favor of beta1c in patients with chronic atrial fibrillation. In the AF patients, the abundance of the 5-HT4-receptor transcript was significantly reduced by 36 % (p < 0.05). The beta-adrenoreceptor transcription was unchanged. In both SR and AF patients, chronic beta-blocker treatment did neither significantly effect the mRNA expression of the L-type Ca2+-channel subunits, the beta-adrenoreceptor subtypes 1 and 2, nor that of the 5-HT4-receptor. Our data show that chronic AF is associated with a decrease in the atrial mRNA amount of auxiliary subunits of the L-type Ca2+-channel and of the 5-HT4-receptor. This supports the hypothesis that the observed alterations in mRNA transcription in AF patients may lead to a decrease in the availability of functional L-type Ca2+-channels and 5-HT4-receptors and/or reduce L-type Ca2+-current amplitude and density, thus, promoting and stabilizing the arrhythmia.

Objectives The small conductance calcium-activated potassium (SK Ca ) channel is a key factor in the regulation of vessel tone and blood pressure. It has been proposed that activation of endothelial SK3 channel plays an important role in the arteriolar dilation through EDHF signal pathway. Nowdays, the expression of SK3 channel has been researched in a variety of hypertensive rat models. However, the results are not completely identical and the expression of SK3 channel in human with hypertension is undefined. To clarify the alteration of SK3 channel in human with hypertension, we quantified the expression of SK3 mRNA and protein in mesenteric artery of hypertension patients. Methods Mesenteric arterial tissues were collected from the removed tissues by abdominal operations in Han Chinese normotensive and primary hypertensive patients. The human tissue collection protocol was approved by the Ethics Committee of Luzhou medical College. Informed consent was obtained from the patients for the use of vascular tissue (which is usually discarded). The expression of mRNA was detected using quantitative Real-time PCR and analyzed with 2 -ΔΔCT method. The artery protein extractions were detected using western blot with SK3 antibody (alomone labs, APC-025). The statistical analysis was done using independent-samples T test by SPSS 19.0. Results Average age of hypertension group (HP) used in this research was 55.0000 ± 10.2425 (n = 12), while the age of normotensive group (NP) was 58.5000 ± 12.2808 (n = 12). The value of ΔCt in HP was 10.6808 ± 1.4035 (n = 12), while it was 12.1015 ± 1.3541 (n = 12) in NP. The expression of SK3 mRNA in HP was 2.36 times higher than in NP. The relative expression of SK3 protein in HP was 1.0748 ± 0.4089 (n = 12), while it was 0.7671 ± 0.1643 (n = 12) in NP, P Conclusions The alteration of SK3 channel in human with hypertension is obvious enhancement but not decrease. This is contrary to results which were reported in spontaneously hypertensive rats. The nosogenesis of hypertension is quite complex, it is a result of synergism involving a series of factors and pathways. The up-regulation of SK3 channel maybe compensate deficient activity of other factors in the vasculature of hypertensive patients.

MicroRNAs have been investigated to involve in electrical remodeling in atrial fibrillation (AF). MicroRNA-101 (miR-101) has a high expression level in heart, but the role of miR-101 in AF remains elusive. The goal of this study is to evaluate the changes of miR-101 in patients with persistent AF

Small-conductance Ca2+-activated K+ (SK) channels are recognized as a new ion channel which associated with atrial fibrillation (AF). This study aims to investigate regulatory functions and the mechanism of protein kinase C (PKC) and verapamil to SK2 in human patients with atrial fibrillation. 30