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Background: KCNH2 gene mutations disrupting rapid component of I(K) (I(Kr)) underlie type 2 congenital long QT syndrome (LQT2). Startled auditory stimuli are specific symptomatic triggers in LQT2, thus suggesting fast arrhythmogenic mechanism.Objective: We investigated acute alpha(1A)- and cyclic adenosine monophosphate (cAMP)-related beta-adrenergic modulation of I(Kr) in HL-1 cardiomyocytes, wild type (WT)- and 2 LQT2-associated mutant Kv11.1 channels (Y43D- and K595E-Kv11.1) reconstituted in Chinese hamster ovary (CHO) cells.Methods: I(Kr) and Kv11.1 currents were recorded using the whole-cell patch-clamp technique and confocal microscopy of HL-1 cardiomyocytes transfected with green fluorescent protein (GFP)-tagged pleckstrin homology domain of phospholipase C-delta(1) visualized fluctuations of membrane phosphatidylinositol 4,5-bisphosphate (PIP(2)) content.Results: In HL-1 cardiomyocytes expressing human alpha(1A)-adrenoceptor, superfusion with phenylephrine significantly reduced I(Kr) amplitude, shifted current activation to more positive potentials, and accelerated kinetics of deactivation. Confocal images showed a decline of membrane PIP(2) content during phenylephrine exposure. Simultaneous application of adenylyl cyclase activator forskolin and phosphodiesterase inhibitor 3-isobutyl-1-methylxantine (IBMX) shifted I(Kr) activation to more negative potentials and decreased tail current amplitudes after depolarizations between +10 and +50 mV. In CHO cells, alpha(1A)-adrenoceptor activation downregulated WT-Kv11.1 channels and forskolin/IBMX produced a dual effect. Expressed alone, the Y43D-Kv11.1 or K595E-Kv11.1 channel had no measurable function. However, co-expression of WT-Kv11.1 and each mutant protein evoked currents with loss-of-function alterations but identical to WT-Kv11.1 alpha(1A)- and forskolin/IBMX-induced regulation.Conclusion: Acute adrenergic regulation of at least 2 Kv11.1 mutant channels is preserved as in WT-Kv11.1 and native I(Kr). Suppression of alpha(1A)-adrenoceptor-related transduction might have therapeutic implications in some cases of LQT2. [ABSTRACT FROM AUTHOR]

Pregnancy causes changes in the uterus, such as increased cell volume and altered water content. However, the mechanisms that protect the structure and maintain the function of uterine smooth muscle cells against these changes during pregnancy have not been clarified. This study focused on the volume-regulated anion channel (VRAC), which opens with cell swelling under low osmotic pressure and releases Cl - ions and various organic osmolytes to resist cell swelling and regulates a wide range of biological processes such as cell death. In this study, myometrial smooth muscle (MSM) tissues and cells (MSMCs) were collected from non-pregnant and pregnant mice. Using western blotting and immunocytochemistry, leucine-rich repeat containing protein 8A (LRRC8A), an essential membrane protein that constitutes part of the VRAC, was determined to be diffused throughout MSMCs including in the cell membrane. Patch-clamp experiments were performed to investigate the electrophysiology of swelling-induced Cl - currents (I Cl, swell ) mediated by the VRAC. No significant changes between non-pregnancy and pregnancy groups were observed in either the expression density of LRRC8A or the current density of I Cl, swell , however the presence of LRRC8A on the cell membrane was significantly increased in the third trimester of pregnancy compared to the non-pregnancy. This study suggests that the VRAC may play a role, such as maintaining cellular homeostasis in the pregnant MSM.

Propofol is a broadly used intravenous anesthetic agent that can cause cardiovascular effects, including bradycardia and asystole. A possible mechanism for these effects is slowing cardiac pacemaker activity due to inhibition of the hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels. However, it remains unclear how propofol affects the allosteric nature of the voltage- and cAMP-dependent gating mechanism in HCN channels. To address this aim, we investigated the effect of propofol on HCN channels (HCN4 and HCN2) in heterologous expression systems using a whole-cell patch clamp technique. The extracellular application of propofol substantially suppressed the maximum current at clinical concentrations. This was accompanied by a hyperpolarizing shift in the voltage dependence of channel opening. These effects were significantly attenuated by intracellular loading of cAMP, even after considering the current modification by cAMP in opposite directions. The differential degree of propofol effects in the presence and absence of cAMP was rationalized by an allosteric gating model for HCN channels, where we assumed that propofol affects allosteric couplings between the pore, voltage-sensor, and cyclic nucleotide-binding domain (CNBD). The model predicted that propofol enhanced autoinhibition of pore opening by unliganded CNBD, which was relieved by the activation of CNBD by cAMP. Taken together, these findings reveal that propofol acts as an allosteric modulator of cAMP-dependent gating in HCN channels, which may help us to better understand the clinical action of this anesthetic drug.

The atrioventricular (AV) node is the only conduction pathway where electrical impulse can pass from atria to ventricles and exhibits spontaneous automaticity. This study examined the function of the rapid- and slow-activating delayed rectifier K + currents (I Kr and I Ks ) in the regulation of AV node automaticity. Isolated AV node cells from guinea pigs were current- and voltage-clamped to record the action potentials and the I Kr and I Ks current. The expression of I Kr or I Ks was confirmed in the AV node cells by immunocytochemistry, and the positive signals of both channels were localized mainly on the cell membrane. The basal spontaneous automaticity was equally reduced by E4031 and HMR-1556, selective blockers of I Kr and I Ks , respectively. The nonselective β-adrenoceptor agonist isoproterenol markedly increased the firing rate of action potentials. In the presence of isoproterenol, the firing rate of action potentials was more effectively reduced by the I Ks inhibitor HMR-1556 than by the I Kr inhibitor E4031. Both E4031 and HMR-1556 prolonged the action potential duration and depolarized the maximum diastolic potential under basal and β-adrenoceptor-stimulated conditions. I Kr was not significantly influenced by β-adrenoceptor stimulation, but I Ks was concentration-dependently enhanced by isoproterenol (EC 50 : 15 nM), with a significant negative voltage shift in the channel activation. These findings suggest that both the I Kr and I Ks channels might exert similar effects on regulating the repolarization process of AV node action potentials under basal conditions; however, when the β-adrenoceptor is activated, I Ks modulation may become more important., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Abstract: Dexmedetomidine (DEX), an α2-adrenoreceptor (α2-AR) and imidazoline receptor agonist, is most often used for the sedation of patients in the intensive care unit. Its administration is associated with an increased incidence of bradycardia; however, the precise mechanism of DEX-induced bradycardia has yet to be fully elucidated. This study was undertaken to examine whether DEX modifies pacemaker activity and the underlying ionic channel function through α2-AR and imidazoline receptors. The whole-cell patch-clamp techniques were used to record action potentials and related ionic currents of sinoatrial node cells in guinea pigs. DEX (≥10 nM) reduced sinoatrial node automaticity and the diastolic depolarization rate. DEX reduced the amplitude of hyperpolarization-activated cation current (If or Ih) the pacemaker current, even within the physiological pacemaker potential range. DEX slowed the If current activation kinetics and caused a significant shift in the voltage dependence of channel activation to negative potentials. In addition, efaroxan, an α2-AR and imidazoline I1 receptor antagonist, attenuated the inhibitory effects of DEX on sinoatrial node automaticity and If current activity, whereas yohimbine, an α2-AR-selective antagonist, did not. DEX did not affect the current activities of other channels, including rapidly and slowly activating delayed rectifier K+ currents (IKr and IKs), L-type Ca2+ current (ICa,L), Na+/Ca2+ exchange current (INCX), and muscarinic K+ current (IK,ACh). Our results indicate that DEX, at clinically relevant concentrations, induced a negative chronotropic effect on the sinoatrial node function through the downregulation of If current through an imidazoline I1 receptor other than the α2-AR in the clinical setting., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Delayed rectifier K + current (I Ks ) is a key contributor to repolarization of action potentials. This study investigated the mechanisms underlying the adrenoceptor-induced potentiation of I Ks in pulmonary vein cardiomyocytes (PVC). PVC were isolated from guinea pig pulmonary vein. The action potentials and I Ks current were recorded using perforated and conventional whole-cell patch-clamp techniques. The expression of I Ks was examined using immunocytochemistry and Western blotting. KCNQ1, a I Ks pore-forming protein was detected as a signal band approximately 100 kDa in size, and its immunofluorescence signal was found to be mainly localized on the cell membrane. The I Ks current in PVC was markedly enhanced by both β 1 - and β 2 -adrenoceptor stimulation with a negative voltage shift in the current activation, although the potentiation was more effectively induced by β 2 -adrenoceptor stimulation than β 1 -adrenoceptor stimulation. Both β-adrenoceptor-mediated increases in I Ks were attenuated by treatment with the adenylyl cyclase (AC) inhibitor or protein kinase A (PKA) inhibitor. Furthermore, the I Ks current was increased by α 1 -adrenoceptor agonist but attenuated by the protein kinase C (PKC) inhibitor. PVC exhibited action potentials in normal Tyrode solution which was slightly reduced by HMR-1556 a selective I Ks blocker. However, HMR-1556 markedly reduced the β-adrenoceptor-potentiated firing rate. The stimulatory effects of β- and α 1 -adrenoceptor on I Ks in PVC are mediated via the PKA and PKC signal pathways. HMR-1556 effectively reduced the firing rate under β-adrenoceptor activation, suggesting that the functional role of I Ks might increase during sympathetic excitation under in vivo conditions., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Background and Purpose: The slowly activating delayed rectifier K + channel (I Ks ), composed of pore-forming KCNQ1 α-subunits and ancillary KCNE1 β-subunits, regulates ventricular repolarization in human heart. Propofol, at clinically used concentrations, modestly inhibits the intact (wild-type) I Ks channels and is therefore unlikely to appreciably prolong QT interval in ECG during anaesthesia. However, little information is available concerning the inhibitory effect of propofol on I Ks channel associated with its gene variants implicated in QT prolongation. The KCNE1 single nucleotide polymorphism leading to D85N is associated with drug-induced QT prolongation and therefore regarded as a clinically important genetic variant. This study examined whether KCNE1-D85N affects the sensitivity of I Ks to inhibition by propofol., Experimental Approach: Whole-cell patch-clamp and immunostaining experiments were conducted in HEK293 cells and/or mouse cardiomyocyte-derived HL-1 cells, transfected with wild-type KCNQ1, wild-type or variant KCNE1 cDNAs., Key Results: Propofol inhibited KCNQ1/KCNE1-D85N current more potently than KCNQ1/KCNE1 current in HEK293 cells and HL-1 cells. Immunostaining experiments in HEK293 cells revealed that pretreatment with propofol (10 μM) did not appreciably affect cell membrane expression of KCNQ1 and KCNE1 proteins in KCNQ1/KCNE1 and KCNQ1/KCNE1-D85N channels., Conclusion and Implications: The KCNE1 polymorphism D85N significantly elevates the sensitivity of I Ks to inhibition by propofol. This study detects a functionally important role of KCNE1-D85N polymorphism in conferring genetic susceptibility to propofol-induced QT prolongation and further suggests the possibility that the inhibitory action of anaesthetics on ionic currents becomes exaggerated in patients carrying variants in genes encoding ion channels., (© 2021 The British Pharmacological Society.)

Background: The anterior pedicle screw (APS) technique for L5 and S1 is crucial for proper anterior lumbar interbody fusion (ALIF). This study aimed to determine the projection, screw trajectory angle, and bone screw passageway length (BSPL), as well as the screw insertion regularity and the operating area within which it is safe to perform insertion., Methods: Forty patients with low back pain, all of whom had lumbar computed tomography scans available, was included in this retrospective analysis. Radiographic parameters were measured, including: the distances from the projection to the upper endplate, lower endplate, and midline; the transverse and sagittal screw angles; and the BSPL. In addition, 10 fresh adult cadaveric lumbosacral spine segments were selected to determine the safe anatomic area in which to operate. Finally, APSs were inserted in L5 and S1 to determine the regularity of APS insertion., Results: We measured the anterior projection parameters, including: the distances to the upper endplate (L5: 12.5±1.3 mm; S1: 4.54±0.87 mm), lower endplate (L5: 17.3±1.6 mm), and midline (L5: 6.6±0.7 mm; S1: 6.6±0.6 mm); the screw trajectory angle, including the transverse screw angle (L5: 25.3±2.8°; S1: 25.7±2.6°), sagittal screw angle (L5: 17.1±1.7°; S1: 22.4±1.1°); and the BSPL (L5: 48.6±3.5 mm; S1: 48.0±3.5 mm). The regularity of APS insertion in L5 and S1 was determined. Upon the needle reaching a point in the lateral view, it reached the corresponding point in the anteroposterior (AP) view. The anatomic parameters of the safe operating area were as follows: the distance from the abdominal aortic bifurcation to the L5 lower edge (40.50±9.40 mm); the distance from the common iliac vein confluence to the L5 lower edge (27.80±8.60 mm); and the horizontal distance from the inner edge of the common iliac vein to the L5 lower edge (37.50±1.30 mm). We also determined the distance between S1 holes (29.30±1.30 mm), the L5/S1 intervertebral height (17.20±1.50 mm), and the safe operating area (2,058.20±84.30 mm 2 )., Conclusions: This study has determined the projection, screw trajectory angle, and BSPL of APSs in L5 and S1, their insertion regularity, and the area in which the operation can be safely performed., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/atm-21-2143). The authors report that the study was supported by Zhejiang Province Public Welfare Technology Application Research Project (CN) (LGF19H060006). The authors have no other conflicts of interest to declare., (2021 Annals of Translational Medicine. All rights reserved.)

Background: We studied the characteristics and regularity of appropriate insertion points for percutaneous pedicle screw placement in the lumbar spine using C-arm X-ray fluoroscopy. The purpose of this study was to improve the accuracy of percutaneous pedicle screw placement and reduce the incidence of superior-level facet joint violation., Methods: Six normal spinal specimens were included. Three different methods for placing percutaneous pedicle screws in the lumbar spine were applied, including the Roy-Camille method, Magerl method and Weinstein method. The relationships among the insertion point, pedicle projection and proximal facet joint on C-arm X-ray films were studied. The projection morphology of the vertebral pedicle in different segments of the lumbar spine was observed. The relationship between the outer edge of the pedicle projection and the outer edge of the cranial articular process was also studied. The distance between the insertion point and the facet joint (M1), the distance between the insertion point and outer edge of the cranial articular process (M2), and the distance between the insertion point and the projection center of the pedicle (M) were measured., Results: In this study, we found that the projection shape of the vertebral pedicle differed across segments of the lumbar spine: the shape for L1-L3 was oval, and that for L4-L5 was round. The radiographic study showed that the outer edge of the cranial articular process was located on the lateral side of the outer edge of the pedicle projection and did not overlap with the pedicle projection. M for the Weinstein group was larger than that for the Roy-Camille group (P <  0.05). M1 for the Weinstein group was larger than that for the Roy-Camille and Magerl groups (P <  0.05). M2 for the Roy-Camille group was negative, M2 for the Magerl group was 0, and M2 for the Weinstein group was positive., Conclusion: Under C-arm X-ray fluoroscopy, we were able to accurately identify the characteristics and regularity of the appropriate insertion point for percutaneous pedicle screw placement in the lumbar spine, which was important for improving the accuracy of percutaneous pedicle screw placement and reducing the incidence of superior-level facet joint violation.

Background and Purpose: Volatile anaesthetics have been shown to differentially modulate mammalian Shaker-related voltage-gated potassium (K v 1.x) channels. This study was designed to investigate molecular and cellular mechanisms underlying the modulatory effects of desflurane or sevoflurane on human K v 1.5 (hK v 1.5) channels., Experimental Approach: Thirteen single-point mutations were constructed within pore domain of hK v 1.5 channel using site-directed mutagenesis. The effects of desflurane or sevoflurane on heterologously expressed wild-type and mutant hK v 1.5 channels were examined by whole-cell patch-clamp technique. A computer simulation was conducted to predict the docking pose of desflurane or sevoflurane within hK v 1.5 channel., Key Results: Both desflurane and sevoflurane increased hK v 1.5 current at mild depolarizations but decreased it at strong depolarizations, indicating that these anaesthetics produce both stimulatory and inhibitory actions on hK v 1.5 channels. The inhibitory effect of desflurane or sevoflurane on hK v 1.5 channels arose primarily from its open-channel blocking action. The inhibitory action of desflurane or sevoflurane on hK v 1.5 channels was significantly attenuated in T480A, V505A, and I508A mutant channels, compared with wild-type channel. Computational docking simulation predicted that desflurane or sevoflurane resides within the inner cavity of channel pore and has contact with Thr479, Thr480, Val505, and Ile508., Conclusion and Implications: Desflurane and sevoflurane exert an open-channel blocking action on hK v 1.5 channels by functionally interacting with specific amino acids located within the channel pore. This study thus identifies a novel molecular basis mediating inhibitory modulation of hK v 1.5 channels by desflurane and sevoflurane., (© 2020 The British Pharmacological Society.)

Background: Early repolarization syndrome (ERS) is characterized by J-point elevation on electrocardiograms and ventricular fibrillation (VF). Early repolarization arises from augmentation of the transmural electrical gradient in the cardiac action potential; therefore, the transient outward potassium current (I to ) has been regarded as a key candidate current for elucidating the mechanism of ERS. KCND3 encoding Kv4.3, an α-subunit of the I to channel, is considered as one of target genes., Objective: The purpose of this study was to search for novel KCND3 mutations associated with ERS and to clarify the pathogenesis., Methods: We performed genetic screening for 11 unrelated probands with ERS and analyzed the electrophysiological properties of detected mutations by patch-clamp methods., Results: A novel de novo KCND3 heterozygous mutation, Gly306Ala (c.917g>c), was found in 1 proband. The proband was a 12-year-old boy, who suffered VF storm and showed significant J-point elevation in multiple leads. Intravenous isoproterenol and subsequent administration of quinidine were effective in preventing VF recurrence and restored the J-point elevation. In electrophysiological analysis, cultured cells expressing mutant Kv4.3 showed significantly increased current densities, slow inactivation, and slow recovery from inactivation compared to wild type. Extracellular application of quinidine significantly restored the inactivation time course in mutant Kv4.3. A simulation study confirmed the relationship between the novel KCND3 mutation and early repolarization on electrocardiograms., Conclusion: A novel KCND3 heterozygous mutation was found to be associated with ERS. The pathogenesis can be explained by the increased I to . Genetic screening for KCND3 could be useful for understanding the pathogenesis and selecting effective treatment., (Copyright © 2019. Published by Elsevier Inc.)

Study Design: A prospective randomized controlled trial., Objective: The purpose of this study was to evaluate the clinical effect and safety of a new multimodal nutritional management (MNM) protocol for patients receiving primary lumbar spine surgery., Summary of Background Data: Poor nutritional status is common in the perioperative period in primary lumbar spine surgery, and may impede recovery after surgery., Methods: A total of 187 patients were included in this prospective randomized controlled trial. They were randomly assigned to the MNM group or the control group. Albumin (ALB) infusion, postoperative ALB level, electrolyte disorders, postoperative electrolyte levels, transfusion rate, postoperative hemoglobin level, length of stay (LOS), and complications were compared between the groups., Results: Compared with the control group, the rate and the total amount of ALB infusion were lower in the MNM group, and the postoperative level of ALB in the MNM group was higher on the first postoperative day, and the third postoperative day. The incidence of hypokalemia, hyponatremia, and hypocalcemia were lower in the MNM group. In the MNM group, the postoperative levels of sodium, potassium, and calcium were higher than the control group. The transfusion rate was similar between the two groups. The hemoglobin level was similar between the two groups on first postoperative day, but was higher in the MNM group on third postoperative day. LOS in the MNM group was shorter than in the control group. The incidence of wound drainage was lower in the MNM group. No statistical differences were observed regarding surgical complications between the two groups., Conclusion: The MNM protocol effectively reduced ALB infusion, the incidence of electrolyte disorders, and wound drainage, increased the postoperative levels of ALB, sodium, potassium, and calcium, and reduced the LOS without increasing the rate of postoperative complications., Level of Evidence: 2.

The human Kv1.5 channel (hKv1.5) produces the ultrarapid delayed rectifier potassium current (I Kur ), which is important for determining the repolarization of action potential in the cardiac atrium. However, the expression of I Kur is reduced in patients with chronic atrial fibrillation. 4-Aminopyridine (4-AP) can specifically suppress I Kur , suggesting that it modifies hKv1.5 as a chaperone molecule. Herein, the effects of long-term 4-AP treatment on hKv1.5 protein expression and function were investigated in HEK cells. 4-AP treatment (24 h) improved hKv1.5 protein levels, promoted hKv1.5 glycosylation, and facilitated the hKv1.5 current in a time-dependent manner. Long-term 4-AP treatment also markedly enhanced hKv1.5 localization in the cell membrane, endoplasmic reticulum, and Golgi. Importantly, the Ile508 residue located in the hKv1.5 channel pore was found to be important for 4-AP inhibitory activity. These results provide insight into developing hKv1.5 channel blocker that can functionally rescue I Kur in patients with chronic atrial fibrillation., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Objective: Endothelial progenitor cells (EPCs) can improve endothelial integrity. This study aimed to examine the effects and the mechanism of EPCs on lung ischemia-reperfusion injury (LIRI)., Methods: Wistar rats were randomized into the sham or the left lung transplantation group. The recipients were randomized and treated with vehicle as the LIRI group, with EPC as the EPC group, or with N5-(1-iminoethyl)-l-ornithine-pretreated EPC as the EPC/L group (n = 8 per group). The ratios of arterial oxygen partial pressure to fractional inspiratory oxygen were measured. The lung wet-to-dry weight ratios, protein levels, and injury, as well as the levels of plasma cytokines, were examined. The levels of endothelin (ET)-1, endothelial nitric oxide synthase (eNOS), phosphorylated eNOS, inducible NOS, phosphorylated myosin light chain, nuclear factor-κBp65, Bax, Bcl-2, cleaved caspase-3, and myeloperoxidase in the graft lungs were detected., Results: Compared with the LIRI group, EPC treatment significantly increased the ratios of arterial oxygen partial pressure to fractional inspiratory oxygen and decreased the lung wet-to-dry weight ratios and protein levels in the grafts, accompanied by increasing eNOS expression and phosphorylation, but decreasing endothelin-1, inducible NOS, phosphorylated nuclear factor-kBp65, phosphorylated myosin light chain expression, and myeloperoxidase activity. EPCs reduced lung tissue damage and apoptosis associated with decreased levels of Bax and cleaved caspase-3 expression, but increased Bcl-2 expression. EPC treatment significantly reduced the levels of serum proinflammatory factors, but elevated levels of interleukin-10. In contrast, the protective effect of EPCs were mitigated and abrogated by N5-(1-iminoethyl)-l-ornithine pretreatment., Conclusions: Data indicated that EPC ameliorated LIRI by increasing eNOS expression., (Copyright © 2018 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.)

Background/aims: The phenylalkylamine class of L-type Ca 2+ channel antagonist verapamil prolongs the effective refractory period (ERP) of human atrium, which appears to contribute to the efficacy of verapamil in preventing reentrant-based atrial arrhythmias including atrial fibrillation. This study was designed to investigate the molecular and electrophysiological mechanism underlying the action of verapamil on human Kv1.5 (hKv1.5) channel that determines action potential duration and ERP in human atrium., Methods: Site-directed mutagenesis created 10 single-point mutations within pore region of hKv1.5 channel. Wholecell patch-clamp method investigated the effect of verapamil on wild-type and mutant hKv1.5 channels heterologously expressed in Chinese hamster ovary cells. Docking simulation was conducted using open-state homology model of hKv1.5 channel pore., Results: Verapamil preferentially blocked hKv1.5 channel in its open state with IC 50 of 2.4±0.6 μM (n = 6). The blocking effect of verapamil was significantly attenuated in T479A, T480A, I502A, V505A, I508A, L510A, V512A and V516A mutants, compared with wild-type hKv1.5 channel. Computer docking simulation predicted that verapamil is positioned within central cavity of channel pore and has contact with Thr479, Thr480, Val505, Ile508, Ala509, Val512, Pro513 and Val516., Conclusion: Verapamil acts as an open-channel blocker of hKv1.5 channel, presumably due to direct binding to specific amino acids within pore region of hKv1.5 channel, such as Thr479, Thr480, Val505, Ile508, Val512 and Val516. This blocking effect of verapamil on hKv1.5 channel appears to contribute at least partly to prolongation of atrial ERP and resultant antiarrhythmic action on atrial fibrillation in humans., Competing Interests: The authors have no conflicts of interest to declare., (© Copyright by the Author(s). Published by Cell Physiol Biochem Press.)

Objective: To observe the open angle (OA), cervical curvature angle (CA), preoperative spinal cord compression rate(PSCR), postoperative spinal cord shift (PSCS) in patients with chronic compressive cervical myelopathy undergoing C3-7 single open laminoplasty, and to explore the possible mechanism and influencing factors of postoperative average spinal cord drift, so as to provide objective basis for predicting PSCS., Methods: From May 2012 to July 2016, 32 patients with multi-segmental chronic compressive cervical myelopathy who underwent single-door laminoplasty in our department were analyzed retrospectively, including 14 cases of cervical spondylotic myelopathy, 8 cases of developmental cervical spinal stenosis with cervical myelopathy, and 10 cases of ossification of posterior longitudinal ligament. The OA of cervical spine was measured on CT, the CA was measured on X-ray, the PSCR and PSCS were measured on MRI. The patients were divided into two groups according to PSCS(group A>=2.5 mm, group B<2.5 mm). In the group A, there were 11 males and 6 females, with an average age of (56.58±9.80) years old, a mean course of the disease of (23.52±7.86) months; while in group B, there were 6 males and 9 females, with an average age of (58.46±12.53) years old, a mean course of disease of (21.13±7.75) months. The correlation analysis between PSCS and OA, CA and PSCR was performed, and multiple linear analysis of correlated parameters was carried out., Results: The OA in group A was (40.47±9.45) °, in group B was (27.84±5.67) °. The OA in group A was higher than that in group B ( P <0.01), and OA was moderately correlated with PSCS ( r =0.794, P =0.000). The CA was(11.56± 4.99) ° in group A and(6.64±3.28) ° in group B . The CA in group A was higher than that in group B ( P <0.01), and CA was moderately correlated with PSCS ( r =0.632, P =0.000). The PSCR was (27.70±2.92) % in group A , was (24.59±2.80) % in group B . The PSCR in group A was higher than that in group B( P <0.01), PSCR was moderately correlated with PSCS( r =0.667, P =0.000). The CA dependent variable was kicked out( P >0.1), and the partial regression coefficients of OA and PSCR were 0.113 and 0.059 respectively., Conclusions: PSCS is the result of OA, CA and PSCR, among which PSCR has the most important influence, OA is the second, CA is the least. PSCS can be predicted by 0.059×OA+0.113×PSCR-2.266 equation, which provides a theoretical basis for preoperative evaluation of spinal cord decompression after surgery., Competing Interests: The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose., (Copyright© 2018 by the China Journal of Orthopaedics and Traumatology Press.)

Objective: To explore the influencing factors and possible mechanism of axial symptoms(AS) after C₃-C₇ single open-door laminoplasty in patients with chronic compression cervical myelopathy., Methods: The clinical data of 32 patients with multi-segment chronic compression cervical cord disease treated by C₃-C₇ single open-door laminectomy from May 2012 to July 2016 were retrospectively analyzed. Including cervical spondylotic myelopathy of 14 cases, developmental cervical stenosis complicated with cervical myelopathy of 8 cases, ossification of posterior longitudinal ligament(OPLL) of 10 cases. There were 17 males and 15 females, aged from 47 to 82 years old with an average of 57.46 year, the course of disease was 5 to 35 months with an average of 22.4 months. The opening angle(OA), cervical curvature angle(CA), preoperative spinal cord compression rate(PSCR) and postoperative spinal cord shift (PSCS) were recorded. After 2 weeks of surgery, determining whether occurred an AS condition according to the AS assessment criteria, the patients were divided into a axial symptom group and a non-axial symptom group, the general data and imaging parameters of the two groups were compared and the factors that may be postoperative AS were analyzed by binary Logistic regression analysis., Results: At 2 weeks after operation, 13 patients occurred AS. There was no significant difference in gender, age and course of disease between axial symptom group and a non-axial symptom group ( P >0.05). In axial symptom group, OA was(36.76±9.35)°, CA was(11.53±4.36)°, PSCR was(27.83±1.72)%, PSCS was (3.17±0.81) mm, while in non-axial symptom group, above items were (33.03±10.52)°, (7.71±4.73)°, (25.16±3.59)%, (2.43±0.95) mm, respectively, there was significant difference in CA, PSCR, PSCS between two groups( P <0.05), and there was no significant difference in OA between two groups( P >0.05). The results of the binary Logistic regression analysis of 3 parameters(OA, PSCR, PSCS) and AS showed OA and PSCR were eliminated in dependent variables, and the partial regression coefficient of PSCR was 0.311, and P =0.031., Conclusions: CA, PSCR, and PSCS are related influencing factors of AS, and PSCS is a high risk factor for AS. C₄,₅ nerve traction caused by posterior spinal movement, postoperative dural self-expansion causes greater traction of the spinal cord, excessive deformation of the cervical spinal cord causes autonomic nerve damage or necrosis that dominates blood vessels may be the pathogenesis of AS, but this is only a theoretical inference, and further improved experiment is necessary to verify it in the future., Competing Interests: The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose., (Copyright© 2018 by the China Journal of Orthopaedics and Traumatology Press.)

BackgroundIn the clinical setting, verapamil is contraindicated in neonates and infants, because of the perceived risk of hypotension or bradyarrhythmia. However, it remains unclear whether there is an age-dependent difference in the sensitivity of cardiac L-type Ca 2+ channel current (I Ca,L ) to inhibition by verapamil.MethodsVentricular myocytes were enzymatically dissociated from the hearts of six different age groups (0, 7, 14, 21, 28 days, and 10-15 weeks) of mice, using a similar Langendorff-perfusion method. Whole-cell patch-clamp technique was applied to examine the sensitivity of I Ca,L to inhibition, by three classes of structurally different L-type Ca 2+ channel antagonists.ResultsVerapamil, nifedipine, and diltiazem concentration-dependently blocked the ventricular I Ca,L in all six age groups. However, although nifedipine and diltiazem blocked ventricular I Ca,L with a similar potency in all age groups, verapamil more potently blocked ventricular I Ca,L in day 0, day 7, day 14, and day 21 mice, than in day 28, and 10-15-week mice.ConclusionIn a mouse heart model, ventricular I Ca,L before the weaning age (~21 days of age) exhibited a higher sensitivity to inhibition by verapamil than that after the weaning age, which may explain one possible mechanism associated with the development of verapamil-induced hypotension in human neonates and infants.

Dr. Wei-Guang Ding's given name and family name were inadvertently interchanged initially. The correct names are as shown above.

Reperfusion of ischemic myocardium is accompanied by intracellular Ca overload, leading to cardiac dysfunction. However, the mechanisms underlying intracellular Ca overload have yet to be fully elucidated. The mechanism may involve the activation of store-operated Ca entry, which is primarily mediated through the transient receptor potential canonical (TRPC) channels. This study was undertaken to examine the possible involvement of TRPC channels in the development of contractile dysfunction associated with reperfusion of ischemic myocardium using a mouse heart model. The functional expression of TRPC channels was confirmed in mouse ventricular myocytes using immunocytochemistry, Western blotting, and patch-clamp experiments. The left ventricular functions were assessed by measuring left ventricular end-diastolic pressure, left ventricular developed pressure, and its first derivatives in a Langendorff-perfused mouse heart subjected to 30 minutes of normothermic (37°C) global ischemia followed by 60 minutes of reperfusion. Under control conditions, left ventricular functions were deteriorated during reperfusion, which was significantly ameliorated by administration of the TRPC channel blockers 2-aminoethoxydiphenyl borate and La during initial 5 minutes of reperfusion. Our findings suggest that TRPC channels are involved in mediating contractile dysfunction during reperfusion of ischemic myocardium and detect TRPC channels as a potential therapeutic target for preventing myocardial ischemia/reperfusion injury.

Background: Missense mutations in KCNH2, a gene encoding the Kv11.1 channel, cause long QT syndrome (LQTS) type 2 primarily by disrupting the intracellular transport of Kv11.1 to the plasma membrane. The present study aimed to clarify the functional changes by two novel KCNH2 missense mutations., Methods: We performed genetic screening of three unrelated symptomatic LQTS probands with family histories of cardiac symptoms. Chinese hamster ovary cells were transfected with wild-type (WT) and/or mutant KCNH2 plasmid and examined by patch-clamp technique. Immunostaining and confocal microscopy were performed to evaluate the intracellular localization of WT and homozygous mutant Kv11.1 in human embryonic kidney cells. For the study of trafficking rescue, we used low-temperature incubation (30°C). We also examined pharmacological rescue of homozygous mutant Kv11.1 current in cells treated with E-4031 or dofetilide., Results: We identified two novel KCNH2 missense mutations, G785D and T826I. Electrophysiological study showed that both mutant channels were nonfunctional in homozygous condition and reduced current densities by half in heterozygous condition compared with WT Kv11.1. Heterozygous Kv11.1-G785D produced a significant positive shift in activation and a significant negative shift in inactivation, whereas heterozygous Kv11.1-T826I caused no kinetic changes. Immunostaining revealed that both were transport-refractory mutations. Incubation at 30°C rescued plasma membrane expression of Kv11.1-T826I but not G785D. We confirmed low-temperature-induced restoration of homozygous Kv11.1-T826I transport by functional current measurements. In contrast, incubation with E-4031 or dofetilide failed to produce measurable currents in both homozygous mutant channels., Conclusions: Two novel KCNH2 mutations disrupted the intracellular transport of Kv11.1. Low-temperature incubation rescued plasma membrane expression of Kv11.1-T826I but not G785D. Both mutations exerted loss-of-function effects on Kv11.1 and explained the phenotypes of the mutation carriers., (Copyright © 2017 Japanese College of Cardiology. Published by Elsevier Ltd. All rights reserved.)

The sustained inward Na + current (I st ) identified in the sinoatrial node (SAN) cell has been suggested to play a pivotal role in cardiac pacemaking. However, the composition of cells in the SAN is heterogeneous and cell-to-cell variability in the magnitude of I st remains to be fully characterized. The present study investigated the current density of I st in morphologically different types of pacemaker cells dissociated from guinea pig SAN. I st was preferentially detected in spontaneously active spindle or spider-shaped cells, but was less well expressed in larger-sized elongated spindle-type cells and practically absent in clearly striated atrial-like cells, despite clear expression of the funny current (I f ). The current density of I st in spindle and spider cells varied from 0.7 to 1.6 pA pF -1 and was significantly reduced in non-beating cells with similar morphologies. By linear regression analysis, we identified a positive correlation between the current densities of I st and the L-type Ca 2+ current (I Ca,L ), which was specifically observed in spindle and spider cells. These cells exhibited a more negative voltage for half maximal I Ca,L activation than atrial-like cells, suggesting a variable ratio between Ca V 1.2- and Ca V 1.3-mediated I Ca,L in SAN cells. Consistent single-cell transcript measurements confirmed a higher relative expression of Ca V 1.3, which activates at more negative potentials, in spindle cells than in atrial-like cells. Taken together, these results can be interpreted as indicating that I st plays a specific role in primary pacemaker cells and that its presence is closely correlated with functional levels of Ca V 1.3-mediated I Ca,L .

Congenital long QT syndrome (LQTS) caused by compound mutations is usually associated with more severe clinical phenotypes. We identified a LQTS family harboring three compound mutations in different genes (KCNQ1-R174C, hERG-E1039X and SCN5A-E428K). KCNQ1-R174C, hERG-E1039X and SCN5A-E428K mutations and/or relevant wild-type (WT) cDNAs were respectively expressed in mammalian cells. I Ks -like, I Kr -like, I Na -like currents and the functional interaction between KCNQ1-R174C and hERG-E1039X channels were studied using patch-clamp and immunocytochemistry techniques. (1) Expression of KCNQ1-R174C alone showed no I Ks . Co-expression of KCNQ1-WT + KCNQ1-R174C caused a loss-of-function in I Ks and blunted the activation of I Ks in response to isoproterenol. (2) Expression of hERG-E1039X alone and co-expression of hERG-WT + hERG-E1039X negatively shifted inactivation curves and decelerated the recovery time from inactivation. (3) Expression of SCN5A-E428K increased peak I Na , but had no effect on late I Na . (4) I Ks and I Kr interact, and hERG-E1039X caused a loss-of-function in I Ks . (5) Immunocytochemical studies indicated that KCNQ1-R174C is trafficking defective and hERG-E1039X is defective in biosynthesis/degradation, but the abnormities were rescued by co-expression with WT. Thus, KCNQ1-R174C and hERG-E1039X disrupted I Ks and I Kr functions, respectively. The synergistic lesion, caused by KCNQ1-R174C and hERG-E1039X in I Ks , is very likely why patients showed more severe phenotypes in the compound mutation case.

Propofol blocks the voltage-gated human Kv1.5 (hKv1.5) channel by preferentially affecting in its open state. A previous mutational study suggested that several amino acids within the pore region of the hKv1.5 channel are involved in mediating the blocking action of propofol. The present investigation was undertaken to elucidate the predicted binding modes of propofol within the pore cavity of the open-state hKv1.5 channel, using computational docking and mutagenesis approaches. The docking simulation using a homology model of the hKv1.5 channel, constructed based on the crystal structure of the Kv1.2 channel, predicted that propofol was positioned at the base of the pore cavity of hKv1.5 channel, adjacent to 4 amino acids Thr479, Thr480, Val505, and Ile508, and formed arene-H interactions with Val505. The patch-clamp experiments on wild-type and mutant hKv1.5 channels constructed by site-directed mutagenesis revealed that the blocking potency of propofol was significantly reduced in T480A, V505A, and I508A but not in T479A mutants compared with wild-type hKv1.5 channel. These computational docking and experimental mutational analyses suggest that propofol is positioned at the base of the pore cavity and forms functional contact with Thr480, Val505, and Ile508 to directly block the hKv1.5 channel.

Background: One complication of total knee arthroplasty (TKA) is patella baja (PB). Patellar tendon shortening and joint line elevation are two main causes of PB. The purpose of this study was to determine the incidence of PB before and after TKA by measuring the patellar height and provide evidence for choosing a suitable index., Methods: In total, 256 consecutive patients who underwent primary TKA were included in this study. Radiographic measurements were performed; the Insall-Salvati (IS) index, modified IS (MIS) index, Blackburne-Peel (BP) index, and Caton-Deschamps (CD) index were computed; and the incidence of PB was calculated before and after the operation. The consistency between the IS and MIS indices and between the BP and CD indices was analyzed., Results: The preoperative incidence of true PB (TPB) and pseudo-PB (PPB) was 9.4 and 0.8%, respectively. The postoperative incidence of TPB and PPB was 10.2 and 9.0%, respectively. The consistency between the IS and MIS indices was moderate preoperatively (pre-kappa = 0.602) and postoperatively (post-kappa = 0.742). The consistency between the BP and CD indices was moderate preoperatively (pre-kappa = 0.742) and good postoperatively (post-kappa = 0.797)., Conclusion: The incidence of PB, especially PPB, increased after TKA. The CD and BP indices are of greater importance for the diagnosis of PB after TKA. The MIS index is a better choice than the IS index to measure the length of the patellar tendon. To measure the height of the joint line, the BP index is better postoperatively and the CD index is better preoperatively.

Objective: The purpose of this systematic review and meta-analysis of randomized controlled trials (RCTs) and non-RCTs was to evaluate the effect of using a bipolar sealer to prevent surgical bleeding in spine surgery., Methods: In June 2017, the PubMed, Embase, Cochrane controlled trials register, Web of Science, Google, and Chinese Wanfang databases were used to identify RCTs and non-RCTs comparing the effects of intraoperative placement of a bipolar sealer versus standard electrocautery with regard to blood loss and blood transfusion. Stata 12.0 software was used to perform the meta-analysis. Weighted mean differences with 95% confidential intervals (CIs) were used for continuous outcomes, and relative risks with 95% CIs were used for discontinuous outcomes. After testing for publication bias and heterogeneity across studies, the data were aggregated and assessed with a random effects model when necessary., Results: In total, 6 clinical trials with 560 patients were included in this meta-analysis. The pooled results indicated that the use of a bipolar sealer decreased the estimated blood loss (MD = -165.06, 95% CI -236.73 to -93.40, P < 0.001), the need for a blood transfusion (RR = 0.46, 95% CI 0.31 to 0.68, P < 0.001), the transfusion units used (MD = -0.41, 95% CI -0.60 to -0.21, P < 0.001), the operative time (MD = -12.98, 95% CI -21.82 to -4.15, P = 0.004) and the length of hospital stay (MD = -2.77, 95% CI -5.45 to -0.10, P = 0.042). No significant difference was observed for the occurrence of infection (RR = 0.52, 95% CI 0.19 to 1.39, P = 0.192)., Conclusion: Based on the current meta-analysis, the use of a bipolar sealer is superior to standard electrocautery for reducing intraoperative blood loss and is therefore recommend for use during spine surgery., (Copyright © 2017. Published by Elsevier Ltd.)

The spontaneous activity of sinoatrial node (SAN) pacemaker cells is generated by a functional interplay between the activity of ionic currents of the plasma membrane and intracellular Ca 2+ dynamics. The molecular correlate of a dihydropyridine (DHP)-sensitive sustained inward Na + current (I st ), a key player in SAN automaticity, is still unknown. Here we show that I st and the L-type Ca 2+ current (I Ca,L ) share Ca V 1.3 as a common molecular determinant. Patch-clamp recordings of mouse SAN cells showed that I st is activated in the diastolic depolarization range, and displays Na + permeability and minimal inactivation and sensitivity to I Ca,L activators and blockers. Both Ca V 1.3-mediated I Ca,L and I st were abolished in Ca V 1.3-deficient (Ca V 1.3 -/- ) SAN cells but the Ca V 1.2-mediated I Ca,L current component was preserved. In SAN cells isolated from mice expressing DHP-insensitive Ca V 1.2 channels (Ca V 1.2 DHP-/- ), I st and Ca V 1.3-mediated I Ca,L displayed overlapping sensitivity and concentration-response relationships to the DHP blocker nifedipine. Consistent with the hypothesis that Ca V 1.3 rather than Ca V 1.2 underlies I st , a considerable fraction of I Ca,L was resistant to nifedipine inhibition in Ca V 1.2 DHP-/- SAN cells. These findings identify Ca V 1.3 channels as essential molecular components of the voltage-dependent, DHP-sensitive I st Na + current in the SAN.

Background: Long QT syndrome (LQTS) presents two clinical phenotypes, congenital and acquired forms. This study aims to evaluate the genetic contribution of a KCNH2 variant for the two LQTS phenotypes., Methods: From 1996 to 2014, genetic screening for LQTS probands was performed for five major genes: KCNQ1, KCNH2, SCN5A, KCNE1, and KCNE2 and 389 probands were found to be mutation carriers. We analyzed the clinical phenotypes of p.His492Tyr carriers in KCNH2., Results: Heterozygous p.His492Tyr variant was identified in 10 LQTS families. Six probands (mean age, 26±23 years) carried another mutation, and two of six had syncope associated with emotional stress or telephone ringing. The remaining four probands were significantly older at diagnosis (mean age, 42±33 years) and carried no other compound mutations. All the four probands had fatal arrhythmic events in the presence of additional precipitating factors such as culprit drugs in 2, hypokalemia in 1, and bradycardia in 1. The QTc interval of carriers with p.His492Tyr alone was 445±10ms and significantly shorter than that in double mutation carriers (481±40ms, p=0.041)., Conclusions: KCNH2 p.His492Tyr variant presented Romano-Ward syndrome in the presence of another mutation and heterozygous carriers had mild phenotypes while even heterozygous carriers should be cared for not to encounter secondary factors because incidental factors could manifest "latent" form of p.His492Tyr heterozygous carriers., (Copyright © 2016 Japanese College of Cardiology. Published by Elsevier Ltd. All rights reserved.)

Human Kv1.5 channels (hKv1.5) conduct the ultra-rapid delayed rectifier potassium current (I Kur ), which plays an important role in action potential repolarization of atrial myocytes. The present study was undertaken to examine the effects of acidic pH on hKv1.5 wild-type (WT) and its pore mutant channels heterologously expressed in Chinese hamster ovary (CHO) cells using site-directed mutagenesis combined with whole-cell patch-clamp technique. Both extracellular and intracellular acidifications equally and reversely reduced the amplitude of hKv1.5 currents. The extracellular acidification significantly shifted the voltage dependence of current activation to more depolarized potentials and accelerated deactivation kinetics of the current. The ancillary β subunits Kvβ1.3 and Kvβ1.2, known to modify the pharmacological sensitivities of hKv1.5, enhanced the extracellular proton-induced inhibitory effect on hKv1.5 current. In addition, several mutants (T462C, T479A, T480A, and I508A) exhibited significantly higher sensitivity to acidic pH-induced inhibition compared with WT channel, whereas the inhibitory effect of acidic pH was markedly reduced in H463G mutant. These observations indicate that (1) extracellular acidification modifies hKv1.5 gating and activity, (2) β subunits and several residues (T462, T479, T480, and I508) play critical roles in determining the sensitivity of the channel to acidic exposure, and (3) H463 may be a critical sensor for the channel inhibition by extracellular protons.

Long QT syndrome type 1 (LQT1) is a subtype of a congenital cardiac syndrome caused by mutation in the KCNQ1 gene, which encodes the α- subunit of the slow component of delayed rectifier K + current ( I Ks ) channel. Arrhythmias in LQT1 are characterized by prolongation of the QT interval on ECG, as well as the occurrence of life-threatening cardiac events, frequently triggered by adrenergic stimuli (e.g., physical or emotional stress). During the past two decades, much advancement has been made in understanding the molecular pathogenesis underlying LQT1. Uncovering the genotype-phenotype correlations in LQT1 is of clinical importance to better understand the gene-specific differences that may influence the propensity for developing life-threatening arrhythmias under specific conditions. Elucidation of these mechanisms will also help to improve the diagnosis and management of this cardiac disorder based on gene-specific considerations. This review describes the current medical consensus and recent developments regarding the molecular pathogenesis of LQT1 and provides a novel insight into the adrenergic regulation of this disease.

The intravenous anesthetic propofol modulates various ion channel functions. It is generally accepted that approximately 98% of propofol binds to blood constituents and that the free (unbound) drug preferentially affects target proteins including ion channels. However, modulatory effects of propofol on ion channels have not been previously explored in the presence of serum albumin. This study was designed to investigate the effects of serum albumin on the blocking action of propofol on the human Kv1.5 (hKv1.5) current. Whole-cell patch-clamp method was used to record the hKv1.5 channel current, heterologously expressed in Chinese hamster ovary cells, in the absence and presence of bovine serum albumin (BSA). Propofol induced a time-dependent decline of the hKv1.5 current during depolarizing steps and slowed the time course of tail current decay upon repolarization, supporting that propofol acts as an open-channel blocker. This blocking effect was reversible and concentration-dependent with an IC50 of 62.9±3.1μM (n = 6). Bath application of 1% BSA markedly reduced the blocking potency of propofol on hKv1.5 current (IC50 of 1116.0±491.4μM; n = 6). However, in the presence of BSA, the propofol-induced inhibition of hKv1.5 current was also accompanied by a gradual decline of activated current during depolarization and deceleration of deactivating tail current upon repolarization. The presence of BSA greatly attenuated the blocking potency of propofol on hKv1.5 channel without affecting the mode of action of propofol on the channel. Serum albumin thus appears to bind to propofol and thereby reducing effective concentrations of the drug for inhibition of hKv1.5 channel., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Silicon nitride (SiN(x)) films containing nanocrystalline silicon (nc-Si) were deposited on crystalline silicon substrate by facing-target sputtering technique. Thermal annealing process was performed at 450 degrees C for 50 min in a conventional furnace under FG(10% H2, 90% N2) ambient. The photoluminescece (PL) properties of the SiN(x) films with nc-Si were investigated by steady/transient PL spectra measurements by Fluorescence spectrometer with different temperatures. The PL processes could be attributed to the quantum confinement effect of nc-Si and the defects in the film. The PL peak position exhibits a small blue shift with the increasing of the excitation energy, which indicates that the PL portion of the nc-Si increased with smaller size. In addition, the PL lifetime increases and the PL intensity exhibits exponential increase as a result of the decreased temperature which supressed the nonradiative recombination process and then improved the radiative recombination. The PL lifetime of the film significantly reduces with the decreasing of the detection wavelength, which indicates that the PL process related to the the quantum confinement effect strongly depends on temperature.

Background and Purpose: In human heart, the Kv 1.5 channel contributes to repolarization of atrial action potentials. This study examined the electrophysiological and molecular mechanisms underlying arachidonic acid (AA)-induced inhibition of the human Kv 1.5 (hKv 1.5) channel., Experimental Approach: Site-directed mutagenesis was conducted to mutate amino acids that reside within the pore domain of the hKv 1.5 channel. Whole-cell patch-clamp method was used to record membrane currents through wild type and mutant hKv 1.5 channels heterologously expressed in CHO cells. Computer docking simulation was conducted to predict the putative binding site(s) of AA in an open-state model of the Kv 1.5 channel., Key Results: The hKv 1.5 current was minimally affected at the onset of depolarization but was progressively reduced during depolarization by the presence of AA, suggesting that AA acts as an open-channel blocker. AA itself affected the channel at extracellular sites independently of its metabolites and signalling pathways. The blocking effect of AA was attenuated at pH 8.0 but not at pH 6.4. The blocking action of AA developed rather rapidly by co-expression of Kv β1.3. The AA-induced block was significantly attenuated in H463C, T480A, R487V, I502A, I508A, V512A and V516A, but not in T462C, A501V and L510A mutants of the hKv 1.5 channel. Docking simulation predicted that H463, T480, R487, I508, V512 and V516 are potentially accessible for interaction with AA., Conclusions and Implications: AA itself interacts with multiple amino acids located in the pore domain of the hKv 1.5 channel. These findings may provide useful information for future development of selective blockers of hKv 1.5 channels., (© 2015 The British Pharmacological Society.)

The intravenous anesthetic propofol affects the function of a diversity of ligand-gated and voltage-gated ion channels. However, there is little information as to whether propofol directly interacts with voltage-gated ion channel proteins to modulate their functions. The Kv1.5 channel is activated by membrane depolarization during action potentials and contributes to atrial repolarization in the human heart. This study was undertaken to examine the effect of propofol on voltage-gated human Kv1.5 (hKv1.5) channel and to elucidate the underlying molecular determinants. Site-directed mutagenesis was carried out through six amino acids that reside within the pore domain of hKv1.5 channel. Whole-cell patch-clamp technique was used to record membrane currents through the wild type and mutant hKv1.5 channels heterologously expressed in Chinese hamster ovary cells. Propofol (≥5 μM) reversibly and concentration-dependently (IC50 of 49.3±9.4 μM; n=6) blocked hKv1.5 current. Propofol-induced block of hKv1.5 current gradually progressed during depolarizing voltage-clamp steps and was enhanced by higher frequency of activation, consistent with a preferential block of the channels in their open state. The degree of current block by propofol was significantly attenuated in T480A, I502A, I508A and V516A, but not in H463C and L510A mutants of hKv1.5 channel. Thus, several amino acids near the selectivity filter (Thr480) or within S6 (Ile502, Ile508 and Val516) are found to be critically involved in the blocking action of propofol. This study provides the first evidence suggesting that direct interaction with specific amino acids underlies the blocking action of propofol on voltage-gated hKv1.5 channel., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The authors prepared nc-SiOx: H thin films using plasma enhanced chemical vapor deposition methods (PECVD) and investigated the influence of oxygen incorporation on the microstructure and band gap properties of the films. The results indicated that with the increase in oxygen mixing ratio (CO2/SiH4), the grain size of the nanocrystal-silicon grain as well as the crystallinity of the film reduced, and the surface tensile stress of the nanocrystal-silicon grain first increased and then decreased. Fourier infrared absorption spectra analysis indicated that, with the increase in oxygen mixing ratio, the intensity of the oxygen rich Si--O bond increased while that of the silicon rich Si--O bond decreased and the structure factor reduced in the meantime accompanied by the improved order degree of thin films. The structure factor increased when the oxygen mixing ratio exceeded 0.08, which shows that the order degree of thin films dropped. In addition, the optical gap increased and the band tail width first increased and then decreased as a result of the incorporation of the oxygen. As a result, the microstructure and band gap properties of the films can be controlled by incorporating oxygen. And the crystallinity and optical gap of the material was high, and the microstructure of the films was improved at the same time when the oxygen mixing ratio was 0. 08, so it can be used as intrinsic layer of the thin-film solar cells.

The slow component of the delayed rectifier K(+) current (I Ks) plays an important role in the repolarization of action potentials in cardiac pacemaker cells and ventricular myocytes, and is regulated by various signaling pathways. Recent evidence has shown that calmodulin (CaM) is involved in modulation of diverse ion channels in cardiac myocytes under physiological and pathophysiological conditions. In the present study, we examined regulation of I Ks by Ca(2+)/CaM in guinea pig sinoatrial (SA) node cells using the whole-cell patch-clamp method. The density of I Ks was larger during intracellular dialysis with a higher Ca(2+) concentration (pCa 7, Ca (+)) compared to that with a low Ca(2+) concentration (pCa 10, Ca (-)). Intracellular application of CaM (400 nM) markedly potentiated I Ks with a Ca (+) pipette solution but not with a Ca (-) solution, thus showing that CaM potentiates I Ks in an intracellular Ca(2+)-dependent manner. Intracellular application of a specific Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) inhibitor, autocamtide-2 inhibitory peptide (AIP, 500 nM), markedly reduced I Ks activity in the presence of higher intracellular Ca(2+). Similarly, bath application of another inhibitor, KN-93 (1 μM) also significantly suppressed I Ks. Finally, the stimulatory action on I Ks of Ca(2+)/CaM was abolished by pretreatment with KN-93. Taken together, these observations suggest that Ca(2+)/CaM stimulates I Ks in guinea pig SA node cells through activation of CaMKII. This enhancement of I Ks by CaMKII may be involved in modulation of SA node automaticity under physiological or pathophysiological condition.

The human ether-a-go-go-related gene (HERG) potassium current (IHERG) has been shown to decrease in amplitude following stimulation with Gq protein-coupled receptors (GqRs), such as α1-adrenergic and M1-muscarinic receptors (α1R and M1R, respectively), at least partly via the reduction of membrane phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). The present study was designed to investigate the modulation of HERG channels by PI(4,5)P2 and phosphatidylinositol4-phosphate 5-kinase (PI(4)P5-K), a synthetic enzyme of PI(4,5)P2. Whole-cell patch-clamp recordings were used to examine the activity of HERG channels expressed heterologously in Chinese Hamster Ovary cells. The stimulation of α1R with phenylephrine or M1R with acetylcholine decreased the amplitude of IHERG accompanied by a significant acceleration of deactivation kinetics and the effects on IHERG were significantly attenuated in cells expressing PI(4)P5-K. The density of IHERG in cells expressing GqRs alone was significantly increased by the coexpression of PI(4)P5-K without significant differences in the voltage dependence of activation and deactivation kinetics. The kinase-deficient substitution mutant, PI(4)P5-K-K138A did not have these counteracting effects on the change in IHERG by M1R stimulation. These results suggest that the current density of IHERG is closely dependent on the membrane PI(4,5)P2 level, which is regulated by PI(4)P5-K and GqRs and that replenishing PI(4,5)P2 by PI(4)P5-K recovers IHERG., (Copyright © 2014 Japanese Pharmacological Society. Production and hosting by Elsevier B.V. All rights reserved.)

Aims: CACNA1C mutations have been reported to cause LQTS type 8 (LQT8; Timothy syndrome), which exhibits severe phenotypes, although the frequency of patients with LQT8 exhibiting only QT prolongation is unknown. This study aimed to elucidate the frequency of CACNA1C mutations in patients with long QT syndrome (LQTS), except those with Timothy syndrome and investigate phenotypic variants., Methods and Results: CACNA1C gene screening was performed in 278 probands negative for LQTS-related gene mutations. Functional analysis of mutant channels using a whole-cell patch-clamp technique was also performed. Using genetic screening, we identified five novel CACNA1C mutations: P381S, M456I, A582D, R858H, and G1783C in seven (2.5%) unrelated probands. Seven mutation carriers showed alternative clinical phenotypes. Biophysical assay of CACNA1C mutations revealed that the peak calcium currents were significantly larger in R858H mutant channels than those of wild-type (WT). In contrast, A582D mutant channels displayed significantly slower inactivation compared with WT. The two mutant channels exerted different gain-of-function effects on calcium currents., Conclusion: In patients with LQTS, the frequency of CACNA1C mutations was higher than reported. Even without typical phenotypes of Timothy syndrome, CACNA1C mutations may cause QT prolongation and/or fatal arrhythmia attacks., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2014. For permissions please email: journals.permissions@oup.com.)

Introduction: Stretch of the atrial membrane upregulates the slow component of delayed rectifier K(+) current (I(Ks)). Blockade of angiotensin II subtype 1 receptors (AT(1)R) attenuates this increase in I(Ks). The present study aimed to examine the effects of irbesartan, a selective AT(1)R blocker (ABR), on both the enhancement of I(Ks) and the shortening of action potential duration (APD) induced by stretching atrial myocytes for exploring the mechanisms underlying the prevention of atrial fibrillation (AF) by ABR., Methods: Hyposmotic solution (Hypo-S) was used to stretch guinea pig atrial myocytes. I(Ks) and APD were recorded using the whole-cell patch-clamp technique., Results: Irbesartan (1-50 μM) attenuated the Hypo-S-induced increase in I(Ks) and shortening of APD90. Hypo-S increased the I(Ks) by 113.4%, whereas Hypo-S + 1 μM irbesartan and Hypo-S + 50 μM irbesartan increased the I(Ks) by only 74.5% and 70.3%, respectively. In addition, Hypo-S shortened the APD(90) by 19.0%, whereas Hypo-S + 1 μM irbesartan and Hypo-S + 50 μM irbesartan shortened the APD90 by 12.1% and 12.0%, respectively., Conclusion: The actions of irbesartan on electrical changes induced by stretching atrial myocytes are associated with blocking AT(1)R. These actions may be beneficial for treating AF., (© The Author(s) 2013.)

Background: To compare the effectiveness of tenocutaneous suture and conventional Kessler suture techniques in treating acute closed Achilles tendon rupture., Methods: A total of 33 patients with acute closed Achilles tendon rupture who were admitted to our hospital from February 1998 to December 2008 underwent repair with either a tenocutaneous suture or Kessler suture technique. All patients were followed up for 1-5 years (mean, 3 years)., Results: According to the American Orthopaedic Foot and Ankle Society ankle-hindfoot scale, the excellence rate was 91% in the Kessler suture group and 98% in the tenocutaneous suture group, with a significant difference between groups., Conclusion: Our tenocutaneous suture technique is an effective method for treating Achilles tendon rupture. It has certain advantages compared with the conventional incision method and is worthy of wide clinical application., (Copyright © 2014 European Foot and Ankle Society. Published by Elsevier Ltd. All rights reserved.)

Background: Brugada syndrome (BrS) is an inherited disease characterized by right precordial ST segment elevation on electrocardiograms (ECGs) that predisposes patients to sudden cardiac death as a result of polymorphic ventricular tachyarrhythmia or ventricular fibrillation (VF). In BrS patients, except for SCN5A, mutations in other responsible genes are poorly elucidated., Methods and Results: We identified 4 KCNH2 mutations, T152I, R164C, W927G, and R1135H, in 236 consecutive probands with BrS or Brugada-like ECG. Three of these mutation carriers showed QTc intervals shorter than 360 milliseconds and 1 experienced VF. We performed patch-clamp analyses on I(Kr) reconstituted with the KCNH2 mutations in Chinese hamster ovary cells and compared the phenotypes of the patients with different genotypes. Three mutations, R164C, W927G, and R1135H, increased I(Kr) densities. Three mutations, T152I, R164C, and W927G, caused a negative shift in voltage-dependent activation curves. Only the R1135H mutant channel prolonged the deactivation time constants. We also identified 20 SCN5A and 5 CACNA1C mutation carriers in our cohort. Comparison of probands' phenotypes with 3 different genotypes revealed that KCNH2 mutation carriers showed shorter QTc intervals and SCN5A mutation carriers had longer QRS durations., Conclusions: All KCNH2 mutations that we identified in probands with BrS exerted gain-of-function effects on I(Kr) channels, which may partially explain the ECG findings in our patients., (© 2014 Wiley Periodicals, Inc.)

We herein investigated the effect of low temperature exposure on the expression, degradation, localization and activity of human Kv1.5 (hKv1.5). In hKv1.5-expressing CHO cells, the currents were significantly increased when cultured at a reduced temperature (28°C) compared to those observed at 37°C. Western blot analysis indicated that the protein levels (both immature and mature proteins) of hKv1.5 were significantly elevated under the hypothermic condition. Treatment with a proteasome inhibitor, MG132, significantly increased the immature, but not the mature, hKv1.5 protein at 37°C, however, there were no changes in either the immature or mature hKv1.5 proteins at low temperature following MG132 exposure. These observations suggest that the enhancement of the mature hKv1.5 protein at reduced temperature may not result from the inhibition of proteolysis. Moreover, the hKv1.5 fluorescence signal in the cells increased significantly on the cell surface at 28°C versus those cultured at 37°C. Importantly, the low temperature treatment markedly shifted the subcellular distribution of the mature hKv1.5, which showed considerable overlap with the trans-Golgi component. Experiments using tunicamycin, an inhibitor of N-glycosylation, indicated that the N-glycosylation of hKv1.5 is more effective at 28°C than at 37°C. Finally, the hypothermic treatment also rescued the protein expression and currents of trafficking-defective hKv1.5 mutants. These results indicate that low temperature exposure stabilizes the protein in the cellular organelles or on the plasma membrane, and modulates its maturation and trafficking, thus enhancing the currents of hKv1.5 and its trafficking defect mutants.

Objectives: This study sought to explore molecular mechanisms underlying the adrenergic-induced QT prolongation associated with KCNQ1 mutations., Background: The most frequent type of congenital long QT syndrome is LQT1, which is caused by mutations in the gene (KCNQ1) that encodes the alpha subunit of the slow component of delayed rectifier K(+) current (IKs) channel. We identified 11 patients from 4 unrelated families that are heterozygous for KCNQ1-G269S. Most patients remained asymptomatic, and their resting corrected QT intervals ranged from normal to borderline but were prolonged significantly during exercise., Methods: Wild-type (WT) KCNQ1 and/or KCNQ1-G269S (G269S) were expressed in mammalian cells with KCNE1. IKs-like currents were measured in control conditions or after isoproterenol or protein kinase A (PKA) stimulation using the patch-clamp technique. Additionally, experiments that incorporated the phosphomimetic KCNQ1 substitution, S27D, in WT or KCNQ1-G269S were also performed., Results: The coexpression of WT-KCNQ1 with varying amounts of G269S decreased IKs, shifted the current-voltage I-V relation of IKs to more positive potentials, and accelerated the IKs deactivation rates in a concentration-dependent manner. In addition, the coexpression of G269S and WT blunted the activation of IKs in response to isoproterenol or PKA stimulation. Lastly, a phosphomimetic substitution in G269S did not show an increased IKs., Conclusions: G269S modestly affected IKs in control conditions, but it almost completely blunted IKs responsiveness in conditions that simulate or mimic PKA phosphorylation of KCNQ1. This insensitivity to PKA stimulation may explain why patients with G269S mutation showed an excessive prolongation of QT intervals on exercise., (Copyright © 2014 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Background: Atrial fibrillation (AF) is one of the most common cardiac arrhythmias. In some patients, the disease is inheritable; however, hereditary aspects of AF remain not fully elucidated., Objective: The purpose of this study was to identify genetic backgrounds that contribute to juvenile-onset AF and to define the mechanism., Methods: In 30 consecutive juvenile-onset AF patients (onset age <50 years), we screened AF-related genes (KCNQ1, KCNH2, KCNE1-3, KCNE5, KCNJ2, SCN5A). We analyzed the function of mutant channels using whole-cell patch-clamp techniques and computer simulations., Results: Among the juvenile-onset AF patients, we identified three mutations (10%): SCN5A-M1875T, KCNJ2-M301K, and KCNQ1-G229D. Because KCNQ1 variant (G229D) identified in a 16-year-old boy was novel, we focused on the proband. The G229D-IKs was found to induce a large instantaneous activating component without deactivation after repolarization to -50 mV. In addition, wild-type (WT)/G229D-IKs (WT and mutant coexpression) displayed both instantaneous and time-dependent activating currents. Compared to WT-IKs, the tail current densities in WT/G229D-IKs were larger at test potentials between -130 and -40 mV but smaller at test potentials between 20 and 50 mV. Moreover, WT/G229D-IKs resulted in a negative voltage shift for current activation (-35.2 mV) and slower deactivation. WT/G229D-IKs conducted a large outward current induced by an atrial action potential waveform, and computer simulation incorporating the WT/G229D-IKs results revealed that the mutation shortened atrial but not ventricular action potential., Conclusion: A novel KCNQ1-G229D mutation identified in a juvenile-onset AF patient altered the IKs activity and kinetics, thereby increasing the arrhythmogenicity to AF., (© 2013 Heart Rhythm Society Published by Heart Rhythm Society All rights reserved.)

Background:  Loss-of-function mutations in the HCN4 gene have been shown to be associated with sinus dysfunction, but there are no reports on HCN4-mediated atrioventricular (AV) block. A novel missense HCN4 mutation G1097W was identified in a 69 year-old Japanese male with AV block, and we characterized the functional consequences of If-like channels reconstituted with the heterozygous HCN4 mutation., Methods and Results:  Wild-type (WT) HCN4 or/and HCN4-G1097W were expressed in a heterologous cell expression system. A functional assay using a whole-cell patch-clamp demonstrated that the mutant If-like currents were activated at more negative voltages compared to WT currents, while they retained the sensitivity to changes in intracellular cyclic adenosine monophosphate (cAMP) levels. Co-expression of G1097W with WT channels showed dominant-negative effects, including a reduction in peak currents and a negative voltage shifting on reconstituted currents., Conclusions:  The HCN4-G1097W mutant channels displayed a loss-of-function type modulation on cardiac If channels and thus could predispose them to AV nodal dysfunction. These data provide a novel insight into the genetic basis for the AV block.

Background: Fatal ventricular arrhythmias in the early period of life have been associated with cardiac channelopathies for decades, and postmortem analyses in SIDS victims have provided evidence of this association. However, the prevalence and functional properties of cardiac ion channel mutations in infantile fatal arrhythmia cases are not clear., Methods and Results: Seven infants with potentially lethal arrhythmias at age < 1 year (5 males, age of onset 44.1 ± 72.1 days) were genetically analyzed for KCNQ1, KCNH2, KCNE1-5, KCNJ2, SCN5A, GJA5, and CALM1 by using denaturing high-performance liquid chromatography and direct sequencing. Whole-cell currents of wildtype and mutant channels were recorded and analyzed in Chinese hamster ovary cells transfected with SCN5A and KCNH2 cDNA. In 5 of 7 patients, we identified 4 mutations (p.N1774D, p.T290fsX53, p.F1486del and p.N406K) in SCN5A, and 1 mutation (p.G628D) in KCNH2. N1774D, F1486del, and N406K in SCN5A displayed tetrodotoxin-sensitive persistent late Na(+) currents. By contrast, SCN5A-T290fsX53 was nonfunctional. KCNH2-G628D exhibited loss of channel function., Conclusion: Genetic screening of 7 patients was used to demonstrate the high prevalence of cardiac channelopathies. Functional assays revealed both gain and loss of channel function in SCN5A mutations, as well as loss of function associated with the KCNH2 mutation., (© 2014 Wiley Periodicals, Inc.)

Achilles tendon rupture is a common injury, and its complications can impair function. Numerous operations have been described for reconstructing the ruptured tendon, but these methods can compromise the microcirculation in the tendon and can seriously damage healing of the tendon. Suturing with a minimally invasive tenocutaneous technique soon after the rupture and systematic functional exercise can greatly reduce the possibility of complications. From June 1996 to February 2007, we treated 20 patients (14 males), who ranged in age from 21 to 66 years old, with this method. After follow-up period of 1 to 7 years, the mean American Orthopedic Foot and Ankle Society Ankle Hindfoot score was 95 (range 90 to 98), and the maximum length of postoperative scarring was 3 cm. One patient again ruptured his Achilles tendon 1 year after surgery in an accident; however, after 10 months, the repaired tendon was still intact. In another patient, the nervus suralis was damaged during surgery by piercing the tension suture at the near end, causing postoperative numbness and swelling. The tension suture was quickly removed, and the patient recovered well with conservative treatment. No large irregular scars, such as those sustained during immobilization, were present over the Achilles tendon. Minimally invasive percutaneous suturing can restore the original length and continuity of the Achilles tendon, is minimally invasive, and results in fewer postoperative complications than other methods., (Copyright © 2013 American College of Foot and Ankle Surgeons. Published by Elsevier Inc. All rights reserved.)

Objective: To study the clinical efficacy of needle-knife to cut off the medial branch of the lumbar posterior ramus under C-arm guiding to treat low back pain caused by lumbar facet osteoarthritis., Methods: From July 2009 to June 2011, 60 patients with low back pain caused by lumbar facet osteoarthritis were reviewed,including 34 males and 26 females, ranging in age from 39 to 73 years old,averaged 61.9 years old; the duration of the disease ranged from 6 to 120 months, with a mean of 18.9 months. All the patients were divided into two groups, 30 patients (18 males and 12 females, ranging in age from 39 to 71 years old, needle-knife group) were treated with needle-knife to cut off medial branch of the lumbar posterior ramus under C -arm guiding and the other 30 patients(16 males and 14 females, ranging in age from 41 to 73 years old, hormone injection group) were treated with hormone injection in lumbar facet joint under C-arm guiding. The preoperative JOA scores and the scores at the 1st, 12th and 26th weeks after treatment were analyzed., Results: Before treatment,the JOA scores between the two groups had no significant difference (P= 0.479); after 1 week of treatment, the JOA scores between the two groups had significant difference (P= 0.040), the improvement rate of hormone injection group was superior than that of the needle-knife group,which were (58.73+/-18.20)% in needle-knife group and (71.10+/-22.19)% in hormone injection group; after 12 weeks of treatment, the JOA scores between the two groups had no significant difference(P=0.569), and the improvement rate between the two groups had no significant difference,which were (50.09+/-19.33)% in the needle-knife group and (48.70+/-18.36)%) in the hormone injection group; after 26 weeks of treatment,the JOA scores between the two groups had significant difference (P=0.000), the improvement rate of hormone injection group was superior than that of the needle-knife group,which were (48.56+/-28.24)% in needle-knife group and (15.62+/-11.23 )% in hormone injection group., Conclusion: Using needle-knife to cut off the medial branch of the lumbar posterior ramus could get longer efficacy than hormone injection in the treatment of lumbar facet osteoarthritis.