Beta-adrenergic stimulation reverses the I Kr–I Ks dominant pattern during cardiac action potential

β-Adrenergic stimulation differentially modulates different K⁺ channels and thus fine-tunes cardiac action potential (AP) repolarization. However, it remains unclear how the proportion of I _Ks, I _Kr, and I _K1 currents in the same cell would be altered by β-adrenergic stimulation, which would change the relative contribution of individual K⁺ current to the total repolarization reserve. In this study, we used an innovative AP-clamp sequential dissection technique to directly record the dynamic I _Ks, I _Kr, and I _K1 currents during the AP in guinea pig ventricular myocytes under physiologically relevant conditions. Our data provide quantitative measures of the magnitude and time course of I _Ks, I _Kr, and I _K1 currents in the same cell under its own steady-state AP, in a physiological milieu, and with preserved Ca²⁺ homeostasis. We found that isoproterenol treatment significantly enhanced I _Ks, moderately increased I _K1, but slightly decreased I _Kr in a dose-dependent manner. The dominance pattern of the K⁺ currents was I _Kr?>?I _K1?>?I _Ks at the control condition, but reversed to I _Kr?I _K1?I _Ks following β-adrenergic stimulation. We systematically determined the changes in the relative contribution of I _Ks, I _Kr, and I _K1 to cardiac repolarization during AP at different adrenergic states. In conclusion, the β-adrenergic stimulation fine-tunes the cardiac AP morphology by shifting the power of different K⁺ currents in a dose-dependent manner. This knowledge is important for designing antiarrhythmic drug strategies to treat hearts exposed to various sympathetic tones.     

Probing the bradycardic drug binding receptor of HCN-encoded pacemaker channels

I_f (or I_h), encoded by the hyperpolarization-activated, cyclic nucleotide-gated (HCN1–4) channel gene family, contributes significantly to cardiac pacing. Bradycardic agents such as ZD7288 that target HCN channels have been developed, but the molecular configuration of their receptor is poorly defined. Here, we probed the drug receptor by systematically introducing alanine scanning substitutions into the selectivity filter (C347A, I348A, G349A, Y350A, G351A in the P-loop), outer (P355A, V356A, S357A, M358A in the P-S6 linker), and inner (M377A, F378A, V379A in S6) pore vestibules of HCN1 channels. When heterologously expressed in human embryonic kidney 293 cells for patch-clamp recordings, I348A, G349A, Y350A, G351A, P355A, and V356A did not produce measurable currents. The half-blocking concentration (IC₅₀) of wild type (WT) for ZD7288 was 25.8?±?9.7 μM. While the IC₅₀ of M358A was identical to WT, those of C347A, S357A, F378A, and V379A markedly increased to 137.6?±?56.4, 113.3?±?34.1, 587.1?±?167.5, and 1726.3?±?673.4 μM, respectively (p?<?0.05). Despite the proximity of the S6 residues studied, M377A was hypersensitive (IC₅₀?=?5.1?±?0.7 μM; p?<?0.05) implicating site specificity. To explore the energetic interactions among the S6 residues, double and triple substitutions (M377A/F378A, M377A/V379A, F378A/V379A, and M377A/F378A/V379A) were generated for thermodynamic cycle analysis. Specific interactions with coupling energies (ΔΔG) >1 kT for M377–F378 and F378–V379 but not M377–V379 were identified. Based on these new data and others, we proposed a refined drug-blocking model that may lead to improved antiarrhythmics and bioartificial pacemaker designs.     

Angiotensin II type 1 receptor mediates partially hyposmotic-induced increase of I Ks current in guinea pig atrium

A repolarizing conduction in the heart augmented by hyposmotic or mechanically induced membrane stretch is the slow component of delayed rectifier K⁺ current (I _Ks). I _Ks upregulation is recognized as a factor promoting appearance of atrial fibrillation (AF) since gain-of-function mutations of the channel genes have been detected in congenital AF. Mechanical stretch activates angiotensin II type 1 (AT₁) receptor in the absence of its physiological ligand angiotensin II. We investigated the functional role of AT₁ receptor in I _Ks enhancement in hyposmotically challenged guinea pig atrial myocytes using the whole-cell patch-clamp method. In atrial myocytes exposed to hyposmotic solution with osmolality decreased to 70% of the physiological level, I _Ks was enhanced by 84.1%, the duration of action potential at 90% repolarization (APD₉₀) was decreased by 16.8%, and resting membrane potential was depolarized (+4.9 mV). The hyposmotic-induced effects on I _Ks and APD₉₀ were significantly attenuated by specific AT₁ receptor antagonist candesartan (1 and 5 μM). Pretreatment of atrial myocytes with protein tyrosine kinase inhibitors tyrphostin A23 and A25 suppressed but the presence of tyrosine phosphatase inhibitor orthovanadate augmented hyposmotic stimulation of I _Ks. The above results implicate AT₁ receptor and tyrosine kinases in the hyposmotic modulation of atrial I _Ks and suggest acute antiarrhythmic properties of AT₁ antagonists in the settings of stretch-related atrial tachyarrhythmias.     

Insulin suppresses IKs (KCNQ1/KCNE1) currents,which require β-subunit KCNE1

Abnormal QT prolongation in diabetic patients has become a clinical problem because it increases the risk of lethal ventricular arrhythmia. In an animal model of type 1 diabetes mellitus, several ion currents, including the slowly activating delayed rectifier potassium current (I_Ks), are altered. The I_Ks channel is composed of KCNQ1 and KCNE1 subunits, whose genetic mutations are well known to cause long QT syndrome. Although insulin is known to affect many physiological and pathophysiological events in the heart, acute effects of insulin on cardiac ion channels are poorly understood at present. This study was designed to investigate direct electrophysiological effects of insulin on I_Ks (KCNQ1/KCNE1) currents. KCNQ1 and KCNE1 were co-expressed in Xenopus oocytes, and whole cell currents were measured by a two-microelectrode voltage-clamp method. Acute application of insulin suppressed the KCNQ1/KCNE1 currents and phosphorylated Akt and extracellular signal-regulated kinase (ERK), the two major downstream effectors, in a concentration-dependent manner. Wortmannin (10^?6 M), a phosphoinositide 3-kinase (PI3K) inhibitor, attenuated the suppression of the currents and phosphorylation of Akt by insulin, whereas U0126 (10^?5 M), a mitogen-activated protein kinase kinase (MEK) inhibitor, had no effect on insulin-induced suppression of the currents. In addition, insulin had little effect on KCNQ1 currents without KCNE1, which indicated an essential role of KCNE1 in the acute suppressive effects of insulin. Mutagenesis studies revealed amino acid residues 111–118 within the distal third C-terminus of KCNE1 as an important region. Insulin has direct electrophysiological effects on I_Ks currents, which may affect cardiac excitability.     

Haematological and acute-phase response of dogs with experimental skin <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> infection to treatment with antibiotic and parthenolide

Changes in white blood cells, leukogram patterns, the positive acute-phase protein (APP) fibrinogen and negative APPs (albumin and arylesterase) were monitored to evaluate their potential as sensitive indicators throughout the course of therapy in canine skin Pseudomonas aeruginosa infection. The study was performed on 15 male mixed-breed dogs, divided in three groups of 5 dogs each. Dogs from group A were injected subcutaneously with P. aeruginosa bacterial culture (1?×?10⁸ CFU/mL) at a dose of 0.3 mL/kg and treated with enrofloxacin (5 mg/kg, s.c.) on post infection hour 48 for 10 consecutive days. Dogs from group B were infected and treated with a combination of enrofloxacin (at above-mentioned dose and intervals) and parthenolide (feverfew extract 90 mg, 0.7 % parthenolide). The schedule consisted of daily oral intake of two capsules of feverfew beginning on post infection hour 4 and continued for 6 days. The control group C included healthy dogs, injected s.c. with 0.3 mL/kg physiological saline. The haematological indices and APPs were assayed before infection and on 4th, 24th, 48th and 72nd hours and on 7th, 10th and 14th days after infection. Infected and antibiotic-treated dogs responded with significant leukocytosis, left shift, eosinopaenia and lymphopaenia between hours 24 and 72. In this group, fibrinogen increased substantially by post infection hours 24 (p?<?0.01 vs 0 h; p?<?0.05 vs group C), 48 (p?<?0.001 vs 0 h; p?<?0.05 vs group C) and 72 (p?<?0.001 vs 0 h; p?<?0.01 vs group C) while albumin reduction was marked by hours 48 (p?<?0.05 vs 0 h) and 72 (p?<?0.05 vs 0 h; p?<?0.001 vs group C) and day 7 (p?<?0.01 vs 0 h; p?<?0.001 vs group C). The combination of enrofloxacin and parthenolide modified, at a significant extent, the deviations in studied parameters except for eosinophil percentage, which persisted low.     

The inhibitory effect of β-stimulation on the Na/K pump current in guinea pig ventricular myocytes is mediated by a cAMP-dependent PKA pathway

 The β-agonist isoproterenol (ISO) reduces the Na/K pump current (I _p) via β-adrenergic receptors when the intracellular calcium concentration ([Ca²⁺]_i) is below 150 nM [8]. In the present study, the intracellular signaling pathway was investigated with whole-cell patch-clamp of isolated guinea pig ventricular myocytes. The inhibitory effect of ISO could be mimicked by external application of the membrane-permeant cAMP analog chlorophenylthio-cAMP (0.5 mM), the phosphodiesterase inhibitor isobutyl-1-methylxanthine (IBMX, 100 μM), or the adenylyl cyclase activator forskolin (50 μM). Intracellular application of the synthetic peptide inhibitor of protein kinase A (PKA), PKI (5 μM), prevented the effect of ISO. These results suggest that the inhibitory effect of ISO on I _p is mediated via a phosphorylation step induced by a cAMP-dependent PKA pathway. Neither the non-specific protein kinase inhibitor H₇ (100 μM) nor the protein phosphatase inhibitor calyculin A (0.5 μM) had any effect on I _p in the absence of ISO. However, H₇ could increase I _p and calyculin A could reduce it in the presence of ISO (1 μM and 12 nM respectively). These results indicate that there is a low basal level of phosphorylation which makes the effects of H₇ and calyculin A difficult to detect in the absence of an ISO-induced increase in phosphorylation level. Received: 21 July1997 / Received after revision: 14 October 1997 / Accepted: 15 October 1997     

Larger transient outward K+ current and shorter action potential duration in Gα11 mutant mice

The α₁-adrenoceptor as well as the AT₁- and the ET_A-receptor couple to G-proteins of the Gα_q/11 family and contribute to the regulation of the transient outward K⁺ current (I_to,f) under pathological conditions such as cardiac hypertrophy or failure. This suggests an important role of Gα_q/11-signalling in the physiological regulation of I_to,f. Here, we investigate mice deficient of the Gα₁₁ protein (gna11^?/?) to clarify the physiological role of Gα₁₁ signalling in cardiac ion channel regulation. Myocytes from endocardial and epicardial layers were isolated from the left ventricular free wall and investigated using the ruptured-patch whole-cell patch-clamp technique. At +40 mV, epicardial myocytes from gna11^?/? mice displayed a 23% larger I_to,f than controls (52.6?±?4.1 pApF^?1, n?=?20 vs 42.7?±?2.8 pApF^?1, n?=?26, p?<?0.05). Endocardial I_to,f was similar in gna11^?/? mice and controls. With the except of minor changes in endocardial myocytes, I_to,f kinetics were similar in both groups. In the epicardial layer, western blot analysis revealed a 19% higher expression of the K⁺-channel α-subunit Kv4.2 in gna11^?/? mice than in wild type (wt; p?<?0.05). The β-subunit KChIP2b was upregulated by 102% in epicardial myocytes of gna11^?/? mice (p?<?0.01, n?=?4). Consistent with the difference in I_to,f, action potential duration was shorter in epicardial cells of gna11^?/? mice than in wt (p?<?0.05), while no difference was found in endocardial myocytes. These results suggest that Gα₁₁-coupled signalling is a central pathway in the regulation of I_to,f. It physiologically exerts a tonic inhibitory influence on the expression of I_to,f and thereby contributes to the regulation of cardiac repolarisation.     

Isoflurane depolarizes bronchopulmonary C neurons by inhibiting transient A-type and delayed rectifier potassium channels

Inhalation of isoflurane (ISO), a widely used volatile anesthetic, can produce clinical tachypnea. In dogs, this response is reportedly mediated by bronchopulmonary C-fibers (PCFs), but the relevant mechanisms remain unclear. Activation of transient A-type potassium current (I_A) channels and delayed rectifier potassium current (I_K) channels hyperpolarizes neurons, and inhibition of both channels by ISO increases neural firing. Due to the presence of these channels in the cell bodies of rat PCFs, we determined whether ISO could stimulate PCFs to produce tachypnea in anesthetized rats, and, if so, whether this response resulted from ISO-induced depolarization of the pulmonary C neurons via the inhibition of I_A and I_K. We recorded ventilatory responses to 5% ISO exposure in anesthetized rats before and after blocking PCF conduction and the responses of pulmonary C neurons (extracellularly recorded) to ISO exposure. ISO-induced (1 mM) changes in pulmonary C neuron membrane potential and I_A/I_K were tested using the perforated patch clamp technique. We found that: (1) ISO inhalation evoked a brief tachypnea (∼7 s) and that this response disappeared after blocking PCF conduction; (2) the ISO significantly elevated (by 138%) the firing rate of most pulmonary C neurons (17 out of 21) in the nodose ganglion; and (3) ISO perfusion depolarized the pulmonary C neurons in the vitro and inhibited both I_A and I_K, and this evoked-depolarization was largely diminished after blocking both I_A and I_K. Our results suggest that ISO is able to stimulate PCFs to elicit tachypnea in rats, at least partly, via inhibiting I_A and I_K, thereby depolarizing the pulmonary C neurons.     

Role of slow delayed rectifying potassium current in dynamics of repolarization and electrical memory in swine ventricles

Dynamics of repolarization, quantified as restitution and electrical memory, impact conduction stability. Relatively less is known about role of slow delayed rectifying potassium current, I _Ks, in dynamics of repolarization and memory compared to the rapidly activating current I _Kr. Trans-membrane potentials were recorded from right ventricular tissues from pigs during reduction (chromanol 293B) and increases in I _Ks (mefenamic acid). A novel pacing protocol was used to explicitly control diastolic intervals to quantify memory. Restitution hysteresis, a consequence of memory, increased after chromanol 293B (loop thickness and area increased 27 and 38 %) and decreased after mefenamic acid (52 and 53 %). Standard and dynamic restitutions showed an increase in average slope after chromanol 293B and a decrease after mefenamic acid. Increase in slope and memory are hypothesized to have opposite effects on electrical stability; therefore, these results suggest that reduction and enhancement of I _Ks likely also have offsetting components that affect stability.     

Time-dependent changes in glucose and insulin regulation during intermittent hypoxia and continuous hypoxia

Hypoxia manifests in many forms including the short repetitive intermittent hypoxia (IH) of sleep apnoea and the continuous hypoxia (CH) of altitude, both of which may impact metabolic function. Based on our own previous studies and the available literature, we hypothesized that whereas acute exposure to IH and CH would lead to comparable metabolic dysfunction, with longer-term exposure, metabolism would normalize to a greater extent with CH than IH. Studies were conducted in lean C57BL/6J mice exposed to either IH or CH for 1 day or 4 weeks and compared to either intermittent air (IA) or unhandled (UN) controls, respectively. We utilized the frequently sampled intravenous glucose tolerance test and minimal model analyses to determine insulin-dependent (insulin sensitivity; S _I) and insulin-independent (glucose effectiveness; S _g) glucose disposal, as well as the insulin response to glucose (acute insulin response to glucose; AIR_g). Our data show that 1-day exposure impaired the glucose tolerance and caused reductions in S _g and AIR_g in both the IH and CH groups, but only IH caused a significant decrease in S _I (7.5 ± 2.7 vs. 17.0 ± 5.3 μU ml^?1 min^?1; p < 0.05). After 4-week exposure, there was evidence of metabolic adaptation in both hypoxic groups, however, in the CH group, there was a supranormal increase in S _I relative to both UN and IH groups. We conclude that in lean mice, the marked metabolic dysfunction that occurs with acute exposure to hypoxia is reversed to a greater extent with chronic CH exposure than chronic IH exposure.     

兔左室肥厚心肌跨室壁复极不均一性及其膜离子流变化的实验研究

目的:探讨兔左心室肥厚心肌心外膜下、中层、心内膜下3层心肌细胞动作电位及膜离子流变化的不均一性。方法: 心肌肥厚组以腹主动脉缩窄术复制兔压力超负荷心肌肥厚模型,并设正常对照组以作比较。胶原酶两步消化法分离获取兔单个心室肌细胞,其中用植皮刀分离左室游离壁内膜下、中层、外膜下3层心肌。以全细胞膜片钳记录单细胞跨膜动作电位和离子电流。结果: 肥厚组3层心肌细胞动作电位时程(APD₉₀)较对照组3层心肌细胞APD₉₀均分别有明显延长,而以中层心肌细胞APD₉₀延长最为明显(延长比例:中层26.0%±2.7%,外膜14.0%±1.6%,内膜10.0%±1.1%),使肥厚心肌跨室壁复极不均一性明显大于对照组。肥厚组各层心肌细胞瞬时外向钾电流(I_to)和缓慢激活的延迟整流钾电流(I_Ks)密度均低于对照组,且均以中层细胞下降的幅度最大。肥厚组各层心肌细胞L型钙电流(I_Ca,L)与快速激活的延迟整流钾电流(I_Kr) 密度与对照组均无明显差异。肥厚组各层心肌细胞内向整流钾电流(I_K1) 均明显低于对照组,但各层变低的幅度无明显差异。 结论:兔肥厚心肌跨室壁复极不均一性明显增大,I_to及I_Ks的跨室壁不均一性下降可能是其主要原因。     

Low total haemoglobin mass, blood volume and aerobic capacity in men with type 1 diabetes

Blood O₂ carrying capacity affects aerobic capacity (VO_2max). Patients with type 1 diabetes have a risk for anaemia along with renal impairment, and they often have low VO_2max. We investigated whether total haemoglobin mass (tHb-mass) and blood volume (BV) differ in men with type 1 diabetes (T1D, n = 12) presently without complications and in healthy men (CON, n = 23) (age-, anthropometry-, physical activity-matched), to seek an explanation for low VO_2max. We determined tHb-mass, BV, haemoglobin concentration ([Hb]), and VO_2max in T1D and CON. With similar (mean ± SD) [Hb] (144 vs. 145 g l^?1), T1D had lower tHb-mass (10.1 ± 1.4 vs. 11.0 ± 1.1 g kg^?1, P < 0.05), BV (76.8 ± 9.5 vs. 83.5 ± 8.3 ml kg^?1, P < 0.05) and VO_2max (35.4 ± 4.8 vs. 44.9 ± 7.5 ml kg^?1 min^?1, P < 0.001) than CON. VO_2max correlated with tHb-mass and BV both in T1D (r = 0.71, P < 0.01 and 0.67, P < 0.05, respectively) and CON (r = 0.54, P < 0.01 and 0.66, P < 0.001, respectively), but not with [Hb]. Linear regression slopes were shallower in T1D than CON both between VO_2max and tHb-mass (2.4 and 3.6 ml kg^?1 min^?1 vs. g kg^?1, respectively) and VO_2max and BV (0.3 and 0.6 ml kg^?1 min^?1 vs. g kg^?1, respectively), indicating that T1D were unable to reach similar VO_2max than CON at a given tHb-mass and BV. In conclusion, low tHb-mass and BV partly explained low VO_2max in T1D and may provide early and more sensitive markers of blood O₂ carrying capacity than [Hb] alone.     

The effect of self-licking behavior on pharmacokinetics of eprinomectin and clinical efficacy against Psoroptes cuniculi in topically administered rabbits

Ear mange mite Psoroptes cuniculi, one of the predominant parasites in rabbits, can cause considerable weight loss, low favorable fee conversion rates, and meningitis. The present experiment was to investigate the difference of plasma disposition and the variation of clinical efficacy under the effect of animal self-licking behavior in topically administered rabbits. Ten rabbits for pharmacokinetic study in two groups (the self-licking and the non-licking) were topically administered with 1 mg kg^?1 of eprinomectin. In the self-licking group, rabbits were allowed to self-lick freely, while, to prevent self-licking, each animal in the non-licking group was fitted with a pet collar. Compared to the non-licking group, self-licking behavior contributed to an extremely significant shorter half-life of absorption (14.85?±?2.79 h in licking group vs. 29.44?±?7.81 h in non-licking group, p?<?0.01) and an extremely significant higher C _max value for eprinomectin (21.95?±?5.36 h in licking group vs. 6.98?±?0.72 ng ml^?1 in non-licking group, p?<?0.01) in plasma disposition. An extremely significantly shorter mean residence time (50.72?±?3.45 h) in self-licking group was also determined compared with the value obtained in non-licking group (106.66?±?7.39 h; p?<?0.01). Clinical efficacy study of eprinomectin was examined in rabbits naturally infested with P. cuniculi which were randomly allocated in three groups: the self-licking, the non-licking, and control groups. All rabbits in the self-licking and the non-licking groups were treated with topical eprinomectin at a single dose of 2 mg kg^?1 (day 0). Topical eprinomection led to a complete parasitological recovery in both treated groups on day 14 and remained free of live mites and clinical lesions from day 21 to the end of the study period (day 35).     

Efficacy evaluation of iclaprim in a neutropenic rat lung infection model with methicillin-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> entrapped in alginate microspheres

The objective of this study was to demonstrate the efficacy of iclaprim in a neutropenic rat lung infection model with methicillin-resistant Staphylococcus aureus (MRSA) entrapped in alginate beads. An inoculum of 5.25?×?10⁵ colony-forming units (CFU)/mL of S. aureus strain AH1252 was administered intratracheally to rats with prepared alginate bacteria suspensions. Beginning 2 h post-infection, rats received: (1) iclaprim 80 mg/kg (n?=?16); (2) iclaprim 60 mg/kg (n?=?16), or (3) vancomycin 50 mg/kg (n?=?24), for 3 days via subcutaneous (SC) injection every 12 h. Twelve hours after the last treatment, rats were euthanized and lungs collected for CFU determination. Iclaprim administered at 80 mg/kg or 60 mg/kg or vancomycin 50 mg/kg SC twice a day for 3 days resulted in a 6.05 log₁₀ CFU reduction (iclaprim 80 mg/kg compared with control, p?<?0.0001), 5.11 log₁₀ CFU reduction (iclaprim 60 mg/kg compared with control, p?<?0.0001), and 3.42 log₁₀ CFU reduction, respectively, from the controls (p?<?0.0001). Iclaprim 80 mg/kg and 60 mg/kg resulted in 2.59 and 1.69 log₁₀ CFU reductions, respectively, from vancomycin-treated animals (80 mg/kg iclaprim vs. vancomycin, p?=?0.0005; 60 mg/kg iclaprim vs. vancomycin, p?=?0.07). Animals receiving iclaprim, vancomycin, and controls demonstrated 100%, 91.7%, and 48.3% survival, respectively. In this neutropenic rat S. aureus lung infection model, rats receiving iclaprim demonstrated a greater CFU reduction than the controls or those receiving vancomycin.     

Expression of miRNAs and ZEB1 and ZEB2 correlates with histopathological grade in papillary urothelial tumors of the urinary bladder

Histopathological grading of papillary urothelial tumors (PUTs) of the urinary bladder is subjective and poorly reproducible. We investigated the relationship between the expression of frequently deregulated microRNAs (miRNAs) as well as their target genes (ZEB1/ZEB2) and bladder cancer histopathological grade in an attempt to find a miRNA that might allow more reliable grading of PUTs. We measured the expression levels of four miRNAs (miR-145, miR-205, miR-125b, and miR-200c) in 120 formalin-fixed, paraffin-embedded bladder tumor tissue samples using real-time PCR assays. ZEB1 and ZEB2 expression was assessed in the same bladder tissues by immunohistochemistry. MiR-205 distinguished low-grade papillary urothelial carcinoma (LG) from high-grade papillary urothelial carcinoma (HG), and miR-145 distinguished HG from infiltrating carcinoma (CA) with an area under the receiver operator characteristic curve (AUC) of 0.992 and 0.997, respectively (sensitivity/specificity of 95.8/96.7 % and 100/91.7 %, respectively; p?<?0.05). The expression level of miR-125b was significantly lower in LG than in PUNLMP, with an AUC value of 0.870 (93.3 % sensitivity and 84.2 % specificity; p?<?0.05). ZEB1 immunoreactivity was more frequently detected in HG than in LG (57 % vs 13 %, p?<?0.01) and in HG than in CA (57 % vs 17 %, p?<?0.01). ZEB2 immunoreactivity was more frequent in CA than in HG (83 % vs 54 %, p?<?0.05). ZEB1/ZEB2 and miRNAs expression seems to reliably distinguish between different grades of PUTs of the urinary bladder. They might well serve as useful complementary diagnostic biomarkers for grading of papillary urothelial tumors.     

A KCNQ1 V205M missense mutation causes a high rate of long QT syndrome in a First Nations community of northern British Columbia: a community-based approach to understanding the impact

PurposeHereditary long QT syndrome is named for a prolonged QT interval reflecting predisposition to ventricular arrhythmias and sudden death. A high rate in a remote, northern Canadian First Nations community was brought to attention.MethodsTwo severely affected index cases and 122 relatives were ascertained using community-based participatory research principles. Genetic sequencing of five known genes responsible for long QT syndrome was carried out on the index cases, leading to the identification of a novel missense mutation. Functional properties of the identified mutation were studied in transfected mouse ltk- cells using whole cell patch clamp techniques. Corrected QT interval measurements were obtained from participants and subsequent genotyping of relatives was carried out.ResultsIn the two index cases, a novel missense mutation (V205M) was identified in the S3 transmembrane helix of KvLQT1, the pore forming domain of the I_Ks channel complex. In transfected mouse ltk-cells the V205M mutation suppressed I_Ks by causing a dramatic depolarizing shift in activation voltage coupled with acceleration of channel deactivation. Twenty-two mutation carriers had a significantly higher mean corrected QT interval than noncarriers (465 ± 28 milliseconds vs. 434 ± 26 milliseconds, P < 0.0001); however, 30% of carriers had a corrected QT interval below 440 milliseconds.ConclusionA novel KCNQ1 mutation in this founder population likely confers increased susceptibility to arrhythmias because of decreased I_Ks current. Even with a common mutation within a relatively homogenous population, clinical expression remains variable, exemplifying the multifactorial nature of long QT syndrome, and supporting the difficulty of definitive diagnosis without genetic testing. A community participatory approach enabled a comprehensive evaluation of the impact.     

Activity of ethanol and daptomycin lock on biofilm generated by an in vitro dynamic model using real subcutaneous injection ports

Vancomycin lock solution (LS) is recommended for the conservative treatment of subcutaneous injection port (SIP)-related infections, but may be associated with failure. We used an in vitro dynamic model of biofilm formation in an SIP, based on a continuous flow circulating via a real SIP, to assess the effectiveness of vancomycin (5 mg/ml), daptomycin (5 mg/ml) and ethanol 40 % LS in eradicating a pre-established Staphylococcus epidermidis biofilm. Heparin, Ringer’s lactate and enoxaparin sodium LS were used as controls. The logarithmic reductions of colony-forming units (CFU) were compared by Student’s t-test. After 24 h of exposure, the vancomycin LS did not exert a greater bactericidal effect than the heparin LS control (mean logarithmic reduction: 2.27?±?0.58 vs. 1.34?±?0.22, respectively, p?=?0.3). The mean logarithmic reduction was greater with daptomycin LS (5.45?±?0.14 vs. 0.39?±?0.12, p?<?0.01) and ethanol LS (6.79?±?1.03 vs. 1.43?±?0.54, p?=?0.02). Bacterial revival after exposure to 24 h of LS was assessed. The mean viable bacteria count was significantly higher for vancomycin LS (9.36?±?0.10 log₁₀CFU) and daptomycin LS (9.16?±?0.02 log₁₀CFU) than for ethanol LS (2.95?±?1.65 log₁₀CFU). Ethanol appeared to be the most attractive option to treat SIP-related infection, but its poor ability to entirely disrupt the biofilm structure may require its use in association with a dispersal agent to avoid renewal of the biofilm.     

Can muscle shortening alone, explain the energy cost of muscle contraction in vivo?

Purpose

Decreased whole-body energy cost of running has been associated with an increased Achilles tendon stiffness. It is usually assumed that this lower energy cost can be attributed to less muscle fascicle shortening with a stiffer tendon. Increased fiber shortening is an important determinant of muscle energetics in vitro. However, other factors, like increased muscle activation may be important when considering whole muscle energetics in vivo.

Methods

To determine the effects of a small additional muscle shortening on skeletal muscle energy requirement, 19 subjects performed 30 plantarflexions on two separate occasions: isometric (ISO) and isokinetic (KIN, 6.98 rad s^–1), each with a target of 50 % of maximum isometric torque. Medial gastrocnemius muscle fascicle length (FL) was measured by ultrasound and rate of oxyhemoglobin (HbO₂) desaturation was measured during blood flow occlusion using near-infrared spectroscopy.

Results

KIN resulted in significantly greater muscle shortening (23.8 ± 1.3 mm) than ISO (18.3 ± 1.0 mm, p < 0.001, mean ± SEM), and greater shortening velocity (KIN = 2.5 ± 0.3 FL s^–1, ISO = 1.1 ± 0.1 FL s^–1, p < 0.001). Rate of HbO₂ desaturation was 19 ± 7 %, greater in KIN than ISO (p < 0.01), despite 19 ± 2 % lower mean torque (p < 0.001) and 9.8 ± 1.6 Nm s lower mean impulse per contraction (p < 0.001) in KIN compared to ISO. Root mean square for EMG was significantly greater (p < 0.05) during KIN (73 ± 3 %) than during ISO (63 ± 2 %).

Conclusion

These results illustrate that muscle energy requirement is greater when muscle fascicle shortening and/or velocity of shortening is increased, and suggest that greater activation contributes to that increased energy requirement.