Activation of CK1ɛ by PP2A/PR61ɛ is required for the initiation of Wnt signaling.

This corrects the article DOI: 10.1038/onc.2016.209.

Activation of CK1ɛ by PP2A/PR61ɛ is required for the initiation of Wnt signaling.

Canonical Wnt signaling induces the stabilization of ß-catenin, its translocation to the nucleus and the activation of target promoters. This pathway is initiated by the binding of Wnt ligands to the Frizzled receptor, the association of the LRP5/6 co-receptor and the formation of a complex comprising Dvl-2, Axin and protein kinases CK1α, ɛ, γ and GSK3. Among these, activation of CK1ɛ, constitutively bound to LRP5/6 through p120-catenin, is required for the association of the rest of the components. We describe here that CK1ɛ is activated by the PP2A/PR61ɛ phosphatase. Binding of Wnt ligands promotes the interaction of LRP5/6-associated CK1ɛ with Frizzled-bound PR61ɛ regulatory subunit, facilitating the access of PP2A catalytic subunit to CK1ɛ and its activation, what enables the recruitment of Dvl-2 to the receptor complex and the initiation of the Wnt pathway. Our results uncover the mechanism of activation of the canonical Wnt pathway by its ligands.

A survey of the risk of zoonoses for veterinarians.

The objectives of this study were to identify factors associated with zoonotic infections in veterinarians, the incidence of physician consultation and treatment and the incidence of diagnostic and treatment errors. Veterinarians in any area of practice were solicited to participate in an online survey through an invitation letter sent to the Oregon Veterinary Medical Association. Proportions of respondents to various factors were analyzed for differences among gender, age, time since graduation and type of practice in which they worked. In all, 216 complete responses were received. In all, 13.9% of respondents had never been vaccinated against rabies, and 20.8% had been exposed to suspect rabid animals, mostly (64.4%) a single time. Other zoonoses were reported by 47.2% of respondents: mostly diseases transmitted via contact (57.4%) especially ringworm, followed by those with oral transmission (21.7%). Most zoonotic infections were reportedly acquired by young veterinarians working in primary care veterinary practice. Cats were the species most commonly reported as the animal source of a zoonotic infection. Veterinarians likely self-diagnosed zoonotic diseases, especially those transmitted by contact. Medical care providers were consulted for diagnosis of more serious diseases. Diagnosis and treatment errors were uncommon. Results of this study emphasize the need to educate future veterinarians during their early years in veterinary school about the risks associated with their future jobs.

Golgi apparatus and endoplasmic reticulum dynamic during meiotic development in canine oocytes.

The Golgi apparatus (GA) and endoplasmic reticulum (ER) play a central role in the events related to intracellular trafficking distribution. This work evaluated the dynamics and localization of the GA and ER in canine oocytes during meiotic development in vitro. Cumulus-oocytes complexes (COCs) from ovaries of adult bitches were incubated for IVM for 0, 48, 72 and 96 h. At each time, the nuclear status was determined using DAPI staining, and the GA was evaluated by immunofluorescence using two antibodies against Golgi proteins: GM130 and Giantin. ER was analysed with fluorescent lipid probes (ER-Tracker), for living cells. Golgi structures were homogeneous in the cytoplasm in non-matured oocytes, mainly in those GV-arrested oocytes. In contrast, at 48 h and from GVBD stage, the immunolocalization began to be subcortical, increasing at 72 h and 96 h. Meiotic development increased with time and the majority of oocytes at MI-MII stages showed cortical distribution of Golgi structure. Living ZP intact non-matured oocytes showed a reticular pattern of ER that covered oocyte cortex. Confocal microscopy showed that, in all levels cuts the fluorescence marks were located in the cortical region, irrespective of culture time. The changes and localization in these organelles during IVM might be related to meiotic development, but in a non-synchronous manner.

Nefrectomía laparoscópica en niños / Laparoscopic nephrectomy in children

Introducción: la nefrectomía laparoscópica en niños, a diferencia de la población adulta, ha tenido una evolución más lenta debido al erróneo concepto de que los niños requieren incisiones pequeñas y se recuperan con mayor rapidez. El objetivo de este artículo es presentar nuestra experiencia en cirugía renal laparoscópica en una población pediátrica. Material y métodos: se revisaron las historias clínicas de los niños que fueron sometidos a cirugía renal laparoscópica en nuestra institución entre mayo de 1993 y septiembre de 2010. Los datos fueron registrados prospectivamente y analizados de forma retrospectiva. Las variables analizadas fueron la indicación quirúrgica, edad, sexo, procedimiento y abordaje, tiempo de quirófano, estancia hospitalaria, sangrado intraoperatorio, complicaciones perioperatorias, conversión a cirugía abierta, reintervención y tasas de transfusión de sangre. Resultados: entre mayo de 1993 y septiembre de 2010 un total de 72 pacientes fueron operados; se practicaron 45 nefrectomías (62,5%), 13 nefroureterectomías (18%) y 14 heminefrectomías(19,4%). La edad media fue de 68 meses (rango: 3-168). El tiempo medio operatorio (OR) fue de 80,4 minutos (rango: 25-270 min) con una pérdida estimada sanguínea promedio de 37,5 ml (rango: 0 -1.000 ml). Hubo tres (4,1%) conversiones a cirugía abierta. Una fue debido a un sangrado intraoperatorio y las otras dos secundarias a una difícil disección debido a presentar cirugías renales previas. La estancia media hospitalaria fue de 3,17 días (rango: 1-30 días) y hubo 7 (9,72%) complicaciones postoperatorias. No hubo mortalidad. Conclusiones: consideramos que el abordaje laparoscópico para la cirugía renal en pacientes pediátricos es un procedimiento factible, seguro y eficaz que debe ser considerado como la primera opción para el tratamiento de la patología benigna en este grupo de población (AU)

Introduction: Laparoscopic nephrectomy in children, not as in the adult population, has evolved more slowly due to the misleading concept that children require small incisions and recover faster. The aim of this article is to present our experience in laparoscopic renal surgery in a paediatric population. Material and methods: We checked the clinical records of children that were subjected to laparoscopic renal surgery at our institution between May 1993 and September 2010. We recorded the data prospectively and analysed it retrospectively. The variables we studied were surgical indication, age, sex, procedure and approach, surgery time, hospital stay, intraoperative bleeding, perioperative complications, conversion to open surgery, reintervention and blood transfusion rates. Results: Between May 1993 and September 2010, we operated a total of 72 patients, 45 nephrectomies (62.5%), 13 nephroureterectomies (18%) and 14 heminephrectomies (19.4%). The mean age was 68 months (range 3-168). The mean surgery time was 80.4 min. (range: 25-270 min.) with a estimated mean blood loss of 37.5 ml (range: 0 -1.000 ml). There were 3 (4.1%) conversions to open surgery. One was due to intraoperative bleeding and the other two were secondary to a difficult dissection due to a history of prior renal surgical interventions. The mean hospital stay was 3.17 days (range: 1-30 days) and there were 7 (9.72%) postoperative complications. There were no deaths. Conclusions: We consider the laparoscopic approach for renal surgery in paediatric patients to be a feasible, safe and effective procedure that must be considered as the first option for the treatment of benign pathology in this population group (AU)

[Laparoscopic nephrectomy in children]. / Nefrectomía laparoscópica en niños.

INTRODUCTION: Laparoscopic nephrectomy in children, not as in the adult population, has evolved more slowly due to the misleading concept that children require small incisions and recover faster. The aim of this article is to present our experience in laparoscopic renal surgery in a paediatric population. MATERIAL AND METHODS: We checked the clinical records of children that were subjected to laparoscopic renal surgery at our institution between May 1993 and September 2010. We recorded the data prospectively and analysed it retrospectively. The variables we studied were surgical indication, age, sex, procedure and approach, surgery time, hospital stay, intraoperative bleeding, perioperative complications, conversion to open surgery, reintervention and blood transfusion rates. RESULTS: Between May 1993 and September 2010, we operated a total of 72 patients, 45 nephrectomies (62.5%), 13 nephroureterectomies (18%) and 14 heminephrectomies (19.4%). The mean age was 68 months (range 3-168). The mean surgery time was 80.4 min. (range: 25-270 min.) with a estimated mean blood loss of 37.5 ml (range: 0 -1.000 ml). There were 3 (4.1%) conversions to open surgery. One was due to intraoperative bleeding and the other two were secondary to a difficult dissection due to a history of prior renal surgical interventions. The mean hospital stay was 3.17 days (range: 1-30 days) and there were 7 (9.72%) postoperative complications. There were no deaths. CONCLUSIONS: We consider the laparoscopic approach for renal surgery in paediatric patients to be a feasible, safe and effective procedure that must be considered as the first option for the treatment of benign pathology in this population group.

Duração da gestação e do parto em éguas Puro Sangue Arabe / Length of gestation and parturition in Arabian Thoroughbred mares

The length of gestation and parturition in 147 Arabian Thoroughbred mares were studied. Length of gestation averaged 330 days and was influenced by the age of mare. They were 335, 330 and 328 days for mares aged up to 4 years, 5 to 9 years and over 10 years, respectively. Parturition lasted 56 minutes on average and was not influenced by the reproductive condition of the mare. Most births occurred at night.

Effect of age of donor on embryo production in Morada Nova (white variety) ewes participating in a conservation programme in Brazil.

In order to evaluate embryo production in Morada Nova (white variety) ewes superovulated with porcine follicle-stimulating hormone, 20 cycling ewes were used as embryo donors and allocated into two groups according to age: group 1 (ewes aged 1-2 years; n = 9) or group 2 (ewes aged 3-4 years; n = 11). Embryo recovery was performed by laparotomy 5-6 days after oestrus. The evaluation of embryos was made under stereomicroscope according to International Embryo Transfer Society rules. The overall recovery rate was 64.6% (5.0 +/- 0.7 structures per ewe) and 86.3% of the recovered structures were fertilized. Group 1 was superior (p < 0.05) to group 2 according to recovered (6.6 +/- 0.9 vs 3.6 +/- 0.8) and fertilized structures (5.6 +/- 1.1 vs. 3.5 +/- 0.7) per ewe. In conclusion, the ovarian response and the embryo production in Morada Nova (white variety) sheep subjected to a standard superovulation treatment were considered satisfactory. In addition, the use of multiple ovulation and embryo transfer in younger ewes ( < or = 2 years old) of this sheep breed appears to be an efficient tool to accelerate the preservation of the Morada Nova (white variety) breed, since younger ewes are as responsive as older ones.

Intravenous and subcutaneous pharmacokinetics of florfenicol in sheep.

Pharmacokinetic parameters of florfenicol were determined in 10 adult sheep (five wethers and five ewes) after a single 40 mg/kg intravenous (i.v.) dose, and three daily subcutaneous (s.c.) doses of 40 mg/kg of a commercial preparation (Nuflor((R))). The concentration of florfenicol in serum samples was assayed using a proprietary HPLC assay method, and pharmacokinetic parameters derived for individual animal data by each route using compartmental and noncompartmental approaches. Two animals (one male and one female) were excluded due to observed i.v. dosing problems, and a biexponential model was found to fit the i.v. data well for six of the other eight animals. Data from two males showed prolonged low concentrations of florfenicol in serum and were better fit by a three-compartment model. The mean +/- SD for the half-lives of the distribution and elimination phases for the six sheep best fit with a two-compartment model were 0.069 +/- 0.018 and 1.01 +/- 0.09 h respectively, and for the V(d(ss)) and clearances were 0.503 +/- 0.035 L/kg and 366 +/- 53 mL/h/kg respectively. The data collected during the s.c. multiple dose study were analyzed using noncompartmental methods only. The bioavailability (F%) after s.c. dosing was calculated in three ways to compare estimation methods as steady-state had not been reached and single dose s.c. data were not obtained past 24 h. Using the AUC(0--24) and AUC(0--> infinity ) from the first dose, the F% values averaged 27 and 40% respectively. Using the AUC(0--> infinity ) for all doses, the F% was 65%. Calculations of the mean time during which the serum concentration exceeded 0.5 and 1.0 microg/mL were 105 +/- 3.9 and 74.7 +/- 12.2 h respectively.

Validación del Cuestionario de la enfermedad respiratoria crónica en pacientes chilenos con limitación crónica del flujo aéreo / Validation of the chronic respiratory questionnaire in Chilean patients with chronic airflow limitation

Chronic respiratory diseases are an important problem in Chile, causing disability and worsening the quality of life. Aim: To validate the Spanish version of the Chronic Respiratory Questionnaire (CRQ) to be used in Chilean population. Material and methods: The CRQ was applied to 38 patients with chronic respiratory diseases. At the same time the functional and physiologic status of patients (the 6 Minute Walking Test and spirometry) was assessed in an independent and blinded fashion. Results: CRQ scores were associated with 6 Minute Walking Test and forced expiratory volume results (p <0.01 adjusted by age, sex and smoking). The Internal Consistency analysis showed a Cronbach a between 0.60 and 0.89. Conclusions: Considering the construct validation as an on-going process, the present results give additional evidence supporting the use of the CRQ in Chilean population to evaluate the health-related quality of life of patients with chronic respiratory diseases (Rev Méd Chile 2003; 131: 1243-50).

Localization of KCNQ5 in the normal and epileptic human temporal neocortex and hippocampal formation.

The KCNQ family of voltage-dependent non-inactivating K+ channels is composed of five members, four of which (KCNQ2-5) are expressed in the CNS and are responsible for the M-current. Mutations in either KCNQ2 or KCNQ3 lead to a hereditary form of dominant generalized epilepsy. Using specific antisera to the KCNQ2, KCNQ3 and KCNQ5 subunits, we found that KCNQ3 co-immunoprecipitated with KCNQ2 and KCNQ5 subunits, but no association was detected between KCNQ2 and KCNQ5. Intense KCNQ5 immunoreactivity was found to be widely distributed throughout the temporal neocortex and the hippocampal formation. In these structures, both pyramidal and non-pyramidal neurons and a population of glial cells in the white matter expressed the KCNQ5 subunit. In the sclerotic areas of the CA fields of epileptic patients, a marked loss of KCNQ5 immunoreactive pyramidal neurons was found in relation with the loss of neurons in these regions. However, in the regions adjacent to the sclerotic areas, the distribution and intensity of KCNQ5 immunostaining was apparently normal. The widespread distribution of KCNQ5 subunits, its persistence in pharmacoresistant epilepsy, along with the significant role of the M-current in the control of neuronal excitability, makes this protein a possible target for the development of anticonvulsant drugs.

Avaliaçäo da transferência passiva da imunidade através da proteína total sangüínea, em potros da raça Árabe / Measurement of blood total protein for assessment of passive immunity transference in the newborn Arabian foals

To study the passive immunity transference to the new born foal via colostrum, the total serum protein of 27 foals, being 13 females and 14 males, born from multiparae and primiparae mares was estimated. The blood samples were colected at 6, 12, 18, 24 and 30 hours after the first suckling. The total protein values increase significantly between 6 and 12 hours after the first suckling, being not influenced by the sex of the foal. The total protein values of new borns from multiparae mares were significantly higher than primiparae mares at 12 hours after the first suckling and was mantained in the same levels for 30 hours. It was concluded that the newborn Arabian foals had the highest immunoglobulin absortion between 6 and 12 hours after the first suckling and it is higher in foals born from multiparae than from primiparae mares

Control del dolor y tenesmo rectal con Fentanilo-TTS en pacientes tratados con radioterapia pélvica / Pain and rectal tenesmus control with fentanyl-TTS in patients under pelvic radiotherapy

La mayoría de los pacientes que reciben tratamiento de radioterapia pélvica presentan en las últimas semanas de irradiación, entre otros síntomas rectitis, que se manifiesta con dolor y tenesmo rectal, que les produce una gran incomodidad, por la constante sensación de ganas de defecar. Ante el mal control de estos síntomas con analgésicos no opioides, u opioides débiles, utilizamos Fentanilo-TTS para intentar controlar el dolor producido en la rectitis, evaluando la disminución del dolor rectal, con Escala Analógica Visual (EVA) y el tenesmo rectal por el número de intentos de defecación. Se estudiaron 22 pacientes en tratamiento radioterápico pélvico, evidenciándose una disminución del dolor rectal de EVA 7,2 a 2,7 (p< 0.001) y del tenesmo rectal de una media de12,4 a 2,7 episodios / día (p<0.001). La tolerancia al tratamiento fue muy buena, presentando únicamente y de forma leve: somnolencia el 36 por ciento, estreñimiento el 18 por ciento y sensación de nausea el 4,5 por ciento de los pacientes. El Fentanilo TTS produce un control eficaz del dolor rectal, consiguiendo una disminución de la graduación de la EVA y del número de episodios de tenesmo rectal, en pacientes tratados con radioterapia pélvica, logrando que el tratamiento radioterápico se realice con muchas menos molestias para el paciente, mejorando la calidad de vida durante el mismo (AU)

Sex differences in the acetylcholine receptor kinetics of postnatal and denervated rat muscle.

Single-channel recording from visualised endplates in freshly dissociated muscles from postnatal and denervated rat muscle revealed the presence of a low conductance, fetal type of acetylcholine receptor. Kinetic analysis showed a main component in the burst durations with a mean of 10.8 +/- 2.7 ms (n = 29). Receptors from female rats had an additional 27.3 +/- 5.5 ms (n = 5) kinetic component which was found in one-third of the 15 female endplates. Recordings from male and female denervated muscles gave more homogeneous kinetics with single time constants of 7.2 +/- 1.3 and 7.4 +/- 1.3 ms, respectively. It is concluded that the acetylcholine receptor channels present during early development are different from those of denervated muscle.

Intersubunit cooperativity in the NMDA receptor.

Opening of the NMDA receptor channel requires simultaneous binding of glutamate and glycine. Although the binding sites for each agonist are in different subunits, the presence of one agonist influences the binding of the other. We have localized regions in the S1 binding domain of both subunits required for the transmission of allosteric signals from the glutamate binding NR2A subunit to the glycine binding NR1 subunit. Three-dimensional modeling indicates that these segments are not directly involved in ligand binding, but likely form solvent-accessible loops protruding out of the binding pocket, making them suitable to relay interactions between adjacent subunits. Thus, these segments mediate negative allosteric coupling between the two subunit types that form the NMDA receptor.

Four residues of the extracellular N-terminal domain of the NR2A subunit control high-affinity Zn2+ binding to NMDA receptors.

NMDA receptors are allosterically inhibited by Zn2+ ions in a voltage-independent manner. The apparent affinity for Zn2+ of the heteromeric NMDA receptors is determined by the subtype of NR2 subunit expressed, with NR2A-containing receptors being the most sensitive (IC50, approximately 20 nM) and NR2C-containing receptors being the least sensitive (IC50, approximately 30 microM). Using chimeras constructed from these two NR2 subtypes, we show that the N-terminal LIVBP-like domain of the NR2A subunit controls the high-affinity Zn2+ inhibition. Mutations at four residues in this domain markedly reduce Zn2+ affinity (by up to >500-fold) without affecting either receptor activation by glutamate and glycine or inhibition by extracellular protons and Ni2+ ions, indicating that these residues most likely participate in high-affinity Zn2+ binding.

Protection by imidazol(ine) drugs and agmatine of glutamate-induced neurotoxicity in cultured cerebellar granule cells through blockade of NMDA receptor.

This study was designed to assess the potential neuroprotective effect of several imidazol(ine) drugs and agmatine on glutamate-induced necrosis and on apoptosis induced by low extracellular K+ in cultured cerebellar granule cells. Exposure (30 min) of energy deprived cells to L-glutamate (1-100 microM) caused a concentration-dependent neurotoxicity, as determined 24 h later by a decrease in the ability of the cells to metabolize 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) into a reduced formazan product. L-glutamate-induced neurotoxicity (EC50=5 microM) was blocked by the specific NMDA receptor antagonist MK-801 (dizocilpine). Imidazol(ine) drugs and agmatine fully prevented neurotoxicity induced by 20 microM (EC100) L-glutamate with the rank order (EC50 in microM): antazoline (13)>cirazoline (44)>LSL 61122 [2-styryl-2-imidazoline] (54)>LSL 60101 [2-(2-benzofuranyl) imidazole] (75)>idazoxan (90)>LSL 60129 [2-(1,4-benzodioxan-6-yl)-4,5-dihydroimidazole](101)>RX82 1002 (2-methoxy idazoxan) (106)>agmatine (196). No neuroprotective effect of these drugs was observed in a model of apoptotic neuronal cell death (reduction of extracellular K+) which does not involve stimulation of NMDA receptors. Imidazol(ine) drugs and agmatine fully inhibited [3H]-(+)-MK-801 binding to the phencyclidine site of NMDA receptors in rat brain. The profile of drug potency protecting against L-glutamate neurotoxicity correlated well (r=0.90) with the potency of the same compounds competing against [3H]-(+)-MK-801 binding. In HEK-293 cells transfected to express the NR1-1a and NR2C subunits of the NMDA receptor, antazoline and agmatine produced a voltage- and concentration-dependent block of glutamate-induced currents. Analysis of the voltage dependence of the block was consistent with the presence of a binding site for antazoline located within the NMDA channel pore with an IC50 of 10-12 microM at 0 mV. It is concluded that imidazol(ine) drugs and agmatine are neuroprotective against glutamate-induced necrotic neuronal cell death in vitro and that this effect is mediated through NMDA receptor blockade by interacting with a site located within the NMDA channel pore.