On near-redundancy and identifiability of parametric hazard regression models under censoring.

We study parametric inference on a rich class of hazard regression models in the presence of right-censoring. Previous literature has reported some inferential challenges, such as multimodal or flat likelihood surfaces, in this class of models for some particular data sets. We formalize the study of these inferential problems by linking them to the concepts of near-redundancy and practical nonidentifiability of parameters. We show that the maximum likelihood estimators of the parameters in this class of models are consistent and asymptotically normal. Thus, the inferential problems in this class of models are related to the finite-sample scenario, where it is difficult to distinguish between the fitted model and a nested nonidentifiable (i.e., parameter-redundant) model. We propose a method for detecting near-redundancy, based on distances between probability distributions. We also employ methods used in other areas for detecting practical nonidentifiability and near-redundancy, including the inspection of the profile likelihood function and the Hessian method. For cases where inferential problems are detected, we discuss alternatives such as using model selection tools to identify simpler models that do not exhibit these inferential problems, increasing the sample size, or extending the follow-up time. We illustrate the performance of the proposed methods through a simulation study. Our simulation study reveals a link between the presence of near-redundancy and practical nonidentifiability. Two illustrative applications using real data, with and without inferential problems, are presented.

Functional forms of socio-territorial inequities in breast cancer screening - A French cross-sectional study using hierarchical generalised additive models.

To reduce the breast cancer burden, the French National Organised Breast Cancer Screening Programme (FNOBCSP) was implemented in 2004. The recommended participation rate has never been achieved and socio-territorial inequities in participation have been reported on several occasions. We investigated the functional forms and consistency of the relationships between neighbourhood deprivation, travel time to the nearest accredited radiology centre and screening uptake. We used two-level hierarchical generalised additive models in 8 types of territories classified by socio-demographic and economic factors. The first level was 368,201 women aged 50-72 invited to the 2013-2014 screening campaign in metropolitan France. They were nested in 41 départements, the level of organisation of the FNOBCSP. The effect of travel time showed two main patterns: it was either linear (with participation decreasing as travel time increased) or participation first increased with increasing travel time to a peak around 5-15 min and decreased afterward. In nearly all types and départements, the probability of participation decreased linearly with increasing deprivation. Territorial inequities in participation were more context-dependent and complex than social inequities. Inequities in participation represent a loss of opportunity for individuals who already have the worst cancer outcomes. Evidence-based public health policies are needed to increase the effectiveness and equity of breast cancer screening.

Individual frailty excess hazard models in cancer epidemiology.

Unobserved individual heterogeneity is a common challenge in population cancer survival studies. This heterogeneity is usually associated with the combination of model misspecification and the failure to record truly relevant variables. We investigate the effects of unobserved individual heterogeneity in the context of excess hazard models, one of the main tools in cancer epidemiology. We propose an individual excess hazard frailty model to account for individual heterogeneity. This represents an extension of frailty modeling to the relative survival framework. In order to facilitate the inference on the parameters of the proposed model, we select frailty distributions which produce closed-form expressions of the marginal hazard and survival functions. The resulting model allows for an intuitive interpretation, in which the frailties induce a selection of the healthier individuals among survivors. We model the excess hazard using a flexible parametric model with a general hazard structure which facilitates the inclusion of time-dependent effects. We illustrate the performance of the proposed methodology through a simulation study. We present a real-data example using data from lung cancer patients diagnosed in England, and discuss the impact of not accounting for unobserved heterogeneity on the estimation of net survival. The methodology is implemented in the R package IFNS.

A tractable Bayesian joint model for longitudinal and survival data.

We introduce a numerically tractable formulation of Bayesian joint models for longitudinal and survival data. The longitudinal process is modeled using generalized linear mixed models, while the survival process is modeled using a parametric general hazard structure. The two processes are linked by sharing fixed and random effects, separating the effects that play a role at the time scale from those that affect the hazard scale. This strategy allows for the inclusion of nonlinear and time-dependent effects while avoiding the need for numerical integration, which facilitates the implementation of the proposed joint model. We explore the use of flexible parametric distributions for modeling the baseline hazard function which can capture the basic shapes of interest in practice. We discuss prior elicitation based on the interpretation of the parameters. We present an extensive simulation study, where we analyze the inferential properties of the proposed models, and illustrate the trade-off between flexibility, sample size, and censoring. We also apply our proposal to two real data applications in order to demonstrate the adaptability of our formulation both in univariate time-to-event data and in a competing risks framework. The methodology is implemented in rstan.

On models for the estimation of the excess mortality hazard in case of insufficiently stratified life tables.

In cancer epidemiology using population-based data, regression models for the excess mortality hazard is a useful method to estimate cancer survival and to describe the association between prognosis factors and excess mortality. This method requires expected mortality rates from general population life tables: each cancer patient is assigned an expected (background) mortality rate obtained from the life tables, typically at least according to their age and sex, from the population they belong to. However, those life tables may be insufficiently stratified, as some characteristics such as deprivation, ethnicity, and comorbidities, are not available in the life tables for a number of countries. This may affect the background mortality rate allocated to each patient, and it has been shown that not including relevant information for assigning an expected mortality rate to each patient induces a bias in the estimation of the regression parameters of the excess hazard model. We propose two parametric corrections in excess hazard regression models, including a single-parameter or a random effect (frailty), to account for possible mismatches in the life table and thus misspecification of the background mortality rate. In an extensive simulation study, the good statistical performance of the proposed approach is demonstrated, and we illustrate their use on real population-based data of lung cancer patients. We present conditions and limitations of these methods and provide some recommendations for their use in practice.

Internal Hemipelvectomy and Reconstruction Assisted by 3D Printing Technology Using Premade Intraoperative Cutting and Placement Guides in a Patient With Pelvic Sarcoma: A Case Report.

CASE: We describe the case of a 75-year-old patient diagnosed with high-grade epithelioid hemangioendothelioma in the left hemipelvis. She underwent an internal hemipelvectomy, followed by reconstruction with a tumor prosthesis with iliac anchorage using 3D-printed cutting and placement guides. Eighteen months postoperatively, she is pain-free and walks without appliances. CONCLUSIONS: Using 3D-printed guides could be an appropriate alternative for patients with aggressive bone tumors in the pelvic area that require hemipelvectomy and reconstruction using a prosthesis with iliac anchorage. 3D-printed cutting guides allow precise resection with appropriate margins, could reduce the risk of injuring critical structures, and facilitate proper prosthetic component positioning.

On a general structure for hazard-based regression models: An application to population-based cancer research.

The proportional hazards model represents the most commonly assumed hazard structure when analysing time to event data using regression models. We study a general hazard structure which contains, as particular cases, proportional hazards, accelerated hazards, and accelerated failure time structures, as well as combinations of these. We propose an approach to apply these different hazard structures, based on a flexible parametric distribution (exponentiated Weibull) for the baseline hazard. This distribution allows us to cover the basic hazard shapes of interest in practice: constant, bathtub, increasing, decreasing, and unimodal. In an extensive simulation study, we evaluate our approach in the context of excess hazard modelling, which is the main quantity of interest in descriptive cancer epidemiology. This study exhibits good inferential properties of the proposed model, as well as good performance when using the Akaike Information Criterion for selecting the hazard structure. An application on lung cancer data illustrates the usefulness of the proposed model.

Prelimbic cortex is a common brain area activated during cue-induced reinstatement of cocaine and heroin seeking in a polydrug self-administration rat model.

Many preclinical studies examined cue-induced relapse to heroin and cocaine seeking in animal models, but most of these studies examined only one drug at a time. In human addicts, however, polydrug use of cocaine and heroin is common. We used a polydrug self-administration relapse model in rats to determine similarities and differences in brain areas activated during cue-induced reinstatement of heroin and cocaine seeking. We trained rats to lever press for cocaine (1.0 mg/kg per infusion, 3-hr/day, 18 day) or heroin (0.03 mg/kg per infusion) on alternating days (9 day for each drug); drug infusions were paired with either intermittent or continuous light cue. Next, the rats underwent extinction training followed by tests for cue-induced reinstatement where they were exposed to either heroin- or cocaine-associated cues. We observed cue-selective reinstatement of drug seeking: the heroin cue selectively reinstated heroin seeking and the cocaine cue selectively reinstated cocaine seeking. We used Fos immunohistochemistry to assess cue-induced neuronal activation in different subregions of the medial prefrontal cortex, dorsal striatum, nucleus accumbens, and amygdala. Fos expression results indicated that only the prelimbic cortex (PL) was activated by both heroin and cocaine cues; in contrast, no significant cue-induced neuronal activation was observed in other brain areas. RNA in situ hybridization indicated that the proportion of glutamatergic and GABAergic markers in PL Fos-expressing cells was similar for the heroin and cocaine cue-activated neurons. Overall, the results indicate that PL may be a common brain area involved in both heroin and cocaine seeking during polydrug use.

Tractable Bayesian variable selection: beyond normality.

Bayesian variable selection often assumes normality, but the effects of model misspecification are not sufficiently understood. There are sound reasons behind this assumption, particularly for large p: ease of interpretation, analytical and computational convenience. More flexible frameworks exist, including semi- or non-parametric models, often at the cost of some tractability. We propose a simple extension that allows for skewness and thicker-than-normal tails but preserves tractability. It leads to easy interpretation and a log-concave likelihood that facilitates optimization and integration. We characterize asymptotically parameter estimation and Bayes factor rates, under certain model misspecification. Under suitable conditions misspecified Bayes factors induce sparsity at the same rates than under the correct model. However, the rates to detect signal change by an exponential factor, often reducing sensitivity. These deficiencies can be ameliorated by inferring the error distribution, a simple strategy that can improve inference substantially. Our work focuses on the likelihood and can be combined with any likelihood penalty or prior, but here we focus on non-local priors to induce extra sparsity and ameliorate finite-sample effects caused by misspecification. We show the importance of considering the likelihood rather than solely the prior, for Bayesian variable selection. The methodology is in R package 'mombf'.

Prevención de la muerte súbita por miocardiopatía arritmogénica del ventrículo derecho en deportistas / Arrhythmogenic right ventricular cardiomyopathy. Prevention of sudden death in athletes

Introducción: La miocardiopatía arritmogénica del ventrículo derecho (ARVC) es una enfermedad hereditaria caracterizada por la sustitución progresiva y parcheada del tejido miocárdico por tejido fibroadiposo, lo cual puede originar arritmias ventriculares y muerte súbita (SCD), incluso como primera manifestación. Las manifestaciones de la enfermedad se ven favorecidas por el ejercicio físico, siendo una de las principales causas de SCD en deportistas menores de 35 años. Material y método: Se ha realizado una revisión sistemática en las diferentes bases de datos científicas relacionada con la ARVC. La búsqueda inicial de 938 artículos se redujo finalmente a 36, tras aplicar los diferentes criterios de inclusión y exclusión. Resultados: En nuestro medio, la historia clínica, la exploración física y el electrocardiograma (ECG) son las principales herramientas usadas en la prevención de la SCD a partir de los 12 años de edad (evaluación cardiovascular básica). En deportistas profesionales o con alto riesgo, se añade ecocardiografía y prueba de esfuerzo máxima (evaluación cardiovascular avanzada). Aun así, la prueba de elección para el diagnóstico de ARVC es la resonancia magnética cardiaca (RMC). El test genético juega un papel importante tanto en el estudio de pacientes sospechosos como en la evaluación de los familiares de pacientes ya diagnosticados. El tratamiento de la ARVC consiste en el uso de fármacos antiarrítmicos, la implantación de un desfibrilador automático implantable (DAI) en función del riesgo de SCD y la restricción de la actividad física. Discusión: La falta de estudios estandarizados de grandes poblaciones de deportistas y la ausencia de registros de muerte súbita dificultan la obtención de una evidencia sólida en la interpretación de los resultados de los artículos revisados. Conclusiones: El screening preparticipativo a todos los deportistas debería incluir: historia clínica, exploración física completa y ECG de 12 derivaciones; considerándose altamente recomendable la realización de una ecocardiografía

Introduction: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited disease characterized by progressive replacement of myocardial tissue and patched by fibrofatty tissue, which can cause ventricular arrhythmias and sudden cardiac death (SCD), even as a first manifestation. The manifestations of the disease are favoured by physical exercise, so it is one of the main causes of SCD in athletes under 35 years old. Material and method: A systematic review in different scientific databases related to ARVC has been made. The initial research on 938 publications was eventually reduced to 36, after applying the different criteria of inclusion and exclusion. Results: In our environment, medical history, physical examination and electrocardiogram (ECG) are the main tools used in the screening of SCD from 12 years of age (basic cardiovascular evaluation). However, in professional or high risk athletes, echocardiography and maximal exercise test are added to the initial screening (advanced cardiovascular assessment). Still, the gold standard test for the diagnosis of ARVC is cardiac magnetic resonance (RMC). The genetic test plays an important role in the study of suspected patients as well as in the evaluation of the relatives of patients who have already been diagnosed. ARVC treatment involves the use of antiarrhythmic drugs, implantation of an implantable cardioverter defibrillator (DAI) based on the risk of SCD and restriction of physical activity. Discussion: The lack of standardized studies on large populations of athletes and the absence of sudden death registries difficult to obtain solid evidence in the interpretation of the results of the reviewed articles. Conclusions: The preparticipative screening of all athletes should include medical history, complete physical examination and 12-lead ECG; considering running an echocardiography being highly recommended

Knee Viscosupplementation: Cost-Effectiveness Analysis between Stabilized Hyaluronic Acid in a Single Injection versus Five Injections of Standard Hyaluronic Acid.

Given the wide difference in price per vial between various presentations of hyaluronic acid, this study seeks to compare the effectiveness and treatment cost of stabilized hyaluronic acid (NASHA) in a single injection with standard preparations of hyaluronic acid (HA) in five injections in osteoarthritis (OA) of the knee. Fifty-four patients with knee osteoarthritis (Kellgren-Lawrence Grade II and III) and the Western Ontario and McMaster Universities Arthritis Index (WOMAC) pain score greater than 7, with a homogeneous distribution of age, sex, BMI, and duration of disease, were included in this study. Patients were randomized into two groups: Group I was treated with NASHA (Durolane®) and Group II with HA (Go-ON®). Patient's evolution was followed up at the 1st, 2nd, 4th, 8th, 12th, and 26th week after treatment. A statistically significant improvement in WOMAC score was observed for patients treated with NASHA versus those who received HA at Week 26. In addition, the need for analgesia was significantly reduced at Week 26 in the NASHA-treated group. Finally, the economic analysis showed an increased cost of overall treatment with HA injections. Our data support the use of the NASHA class of products in the treatment of knee OA.

Efectividad de un programa de ejercicio físico individualizado no supervisado, de cuatro meses de duración, sobre la tolerancia al esfuerzo, percepción de fatiga y variables antropométricas en pacientes sedentarios con factores de riesgo cardiovascular / Effectiveness of a customised, unsupervised 4 month exercise programme, on exercise tolerance, perception of fatigue and anthropometric variables in sedentary patients with cardiovascular risk factors

Introducción: La mejora de la condición física se relaciona con beneficios para la salud. El objetivo de este estudio es valorar la efectividad de un programa de ejercicio físico individualizado no supervisado sobre variables antropométricas, la percepción de fatiga y la tolerancia al esfuerzo (test de los 6 minutos) en pacientes sedentarios con factores de riesgo cardiovascular. Material y métodos: Se estudiaron 119 pacientes, de los cuales terminaron el estudio 75 (45 mujeres y 30 hombres), con edades comprendidas entre los 21 y 77 años, sedentarios con factores de riesgo cardiovascular. Previo al inicio del programa de ejercicio físico se sometieron a un examen médico-deportivo que incluyó: anamnesis, exploración por aparatos, toma de tensión arterial (TA), electrocardiograma de reposo (ECG), estudio antropométrico (peso, talla, IMC e impedanciometría). Al principio y final del estudio se realizó el test de lo 6 minutos que mide la distancia recorrida y se valoró al inicio y final del test: TA, la frecuencia cardiaca, la saturación de oxígeno y solo al terminar, la percepción de esfuerzo. El programa de ejercicio físico, de 4 meses de duración, incluyó: caminar 30-60 minutos /día, bicicleta estática: 3 días/semana, 30 minutos/sesión, con una intensidad del 40-60% de la capacidad funcional individual máxima, abdominales isométricos y estiramientos estáticos. Se realizó estudio estadístico descriptivo y comparación de medias para datos apareados. Resultados: Los datos al comienzo versus el final del programa de ejercicio físico fueron los siguientes: Peso: 100,63 (24,29) vs. 99,6 (23,32) (p <0,05) kg; IMC: 36,62 (8,47) vs. 36,23 (8,12) (p <0,05) kg/m2; Percepción de fatiga. Escala de Borg: 4,15 (2,40) vs. 2,93 (1,81). (p <0,001). Las distancias recorridas fueron: 474 (61) vs. 514,6 (69,2) metros. (p <0,001). Discusión y conclusiones: Los datos del estudio confirman que el modelo de programa de ejercicio físico individualizado no supervisado aplicado en nuestro centro en pacientes sedentarios con factores de riesgo cardiovascular mejora, de forma estadísticamente significativa: la tolerancia al esfuerzo, la sensación de fatiga y, aunque discretamente, el peso y el IMC

Introduction: Improving the physical condition is related to health benefits. The objective of this study is to assess the effectiveness of an individualized unsupervised exercise program on anthropometric variables, the perception of fatigue and physical effort tolerance (6 minutes walking test) in sedentary patients with cardiovascular risk factors. Material and Methods: We studied 119 sedentary patients with cardiovascular risk factors, aged between 21 and 77 years old. Only 75 patients completed the study (45 women and 30 men). Before beginning the exercise program a medical examination was conducted, including: medical history, physical exam, blood pressure measurement (BP), rest-electrocardiogram (rest-ECG), anthropometrical measurements (weight, height, body mass index (BMI) and impedanciometry. The six minutes walk test was performed at the beginning and end of the study. The distance, BP, heart rate, oxygen saturation and perceived effort were measured. The 4 month exercise program included: walking for about 30-60 minutes/day, cycloergometer: 3 days/week, 30 minutes each session, intensity of 40-60% of individual maximum functional capacity, isometric abdominal and static stretching. A descriptive statistical study and a comparison of means for paired data were realized. Results: The data at the beginning versus the end of the exercise program were: Weight: 100,63 (24,29) vs. 99,6 (23,32) (p<0,05) kg; BMI: 36,62 (8,47) vs. 36,23 (8,12) (p <0,05) kg/m2. Fatigue perception, Borg Scale: 4,15 (2,37) vs. 2,93 (1,81) (p <0,001). The distances covered were: 474 (61) vs. 514,6 (69,2) (p <0,001) meters. Discussion and Conclusions: The results of the study confirm that the exercise program implemented in our center improves exercise tolerance, reduces the perception of fatigue and even slightly decreases the weight and the BMI, in sedentary patients with cardiovascular risk factors

Bayesian linear regression with skew-symmetric error distributions with applications to survival analysis.

We study Bayesian linear regression models with skew-symmetric scale mixtures of normal error distributions. These kinds of models can be used to capture departures from the usual assumption of normality of the errors in terms of heavy tails and asymmetry. We propose a general noninformative prior structure for these regression models and show that the corresponding posterior distribution is proper under mild conditions. We extend these propriety results to cases where the response variables are censored. The latter scenario is of interest in the context of accelerated failure time models, which are relevant in survival analysis. We present a simulation study that demonstrates good frequentist properties of the posterior credible intervals associated with the proposed priors. This study also sheds some light on the trade-off between increased model flexibility and the risk of over-fitting. We illustrate the performance of the proposed models with real data. Although we focus on models with univariate response variables, we also present some extensions to the multivariate case in the Supporting Information. Copyright © 2016 John Wiley & Sons, Ltd.

Fútbol femenino categorías inferiores. Características antropométricas y fisiológicas. Evolución a lo largo de una temporada / Young women soccer players. Anthropometric and physiological characteristics. Evolution in a Sports season

Introducción: El fútbol femenino ha experimentado un importante aumento de practicantes en las últimas décadas. Se aportan datos antropométricos y de condición física de jugadoras de fútbol en formación valorando su evolución a lo largo de una temporada y comparándolos con los existentes en la literatura. Material y métodos: Se estudiaron 21 jugadoras de edades comprendidas entre 12 y 15 años, que entrenaban 2 días/semana, 90 minutos/sesión, más el partido del fin de semana. No se diferenció por posiciones en el terreno deportivo. Todas ellas realizaron un examen médico-deportivo al inicio y al final del estudio, que incluyó: anamnesis, exploración física, tensión arterial, ECG de reposo, antropometría (peso, talla, 6 pliegues) y Test de Banco de Astrand. Se realizó estudio estadístico descriptivo y comparación de medias para datos apareados. Resultados: A lo largo del año se observa un aumento del peso: media de 48,83 (8,17) a 52,82 (7,69) Kg, de la talla: media de 158,5 (6,19) a 160,7 (5,33) cm , del % de grasa: media de 14,7 (3,84) % a 16,9 (3,98) % y un aumento del VO2 max: media de 42,95 (6,13) a 44,58 (9,37) ml/Kg/min. Los valores del % de grasa son algo inferiores a los descritos en jugadoras de categoría senior de equipos de elite (rango de 17,5-28,3%), mientras que el VO2 max se sitúa por debajo del rango de referencia para jugadoras europeas de elite (47-57 ml/kg/min). Discusión y conclusiones: Las diferencias halladas entre los dos controles son estadísticamente significativas en el peso (p<0.0001), talla (p<0.0001), % graso (p=0.002) y VO2 max en valores absolutos (p=0,009) y no en valores referidos al peso. En las edades objeto de estudio es difícil atribuir en qué proporción estas variaciones se deben al crecimiento y desarrollo y que parte al entrenamiento físico

Background: Female’s football has had a great improvement and in the number of players over the last decades. Our goal is to analyse both anthropometrical characteristics and physical capacity of young women football players, comparing our results with current literature and assess the evolution during a season. Methods: 21 women football players were examined. All between 12-15 years old and used to train twice a week during 90 minutes each session; playing a match at the weekend as well. Players positions were not discriminated. They all passed a sports physical exam at the beginning and at the end of the study. This check-up included a thorough medical history, a physical exam, blood pressure, rest-electrocardiogram, anthropometry (weight, height, 6 skin-fold thickness) and the Astrand step test. Descriptive statistical analysis and paired means comparison were performed. Results: We observed a weight gain, a growth in height and a rise in body fat percentage throughout the season. The average weight increased from 48.83 (8.17) to 52.82 (7.69) kg. Height augmentation was from 158.5 (6.19) to 160.7 (5.33) cm, and body fat percentage moved up from 14.7 (3.84) to 16.9 (3.98) %. Maximal oxygen uptake incremented from 42.95 (6.13) to 44.58 (9.37) ml/kg/min. The body fat percentage results are slightly lower than reference values in senior elite women football players (17.5-28.3%) while maximal oxygen uptake is lower than reference range for European women elite football players (47-57 ml/kg/min). Discussion and conclusions: The results concerning weight (p<0.0001), height (p<0.0001), body fat percentage (p=0.002) and absolute values of maximal oxygen uptake (p=0.009) are statistically significant. Given the age of the players, it is difficult to attribute which part of these results is due to growth itself and which one is due to training

Músculo sóleo accesorio de una deportista. A propósito de un caso y revisión de la bibliografía / Accessory soleus muscle in an athlete. Presentation of a case and a literature review

Según los estudios necrópsicos, la incidencia del músculo sóleo accesorio fluctúa entre el 0,5 y el 6,0% de la población. La presentación típica es una tumoración blanda del tercio distal posteromedial de la pierna que aumenta de tamaño con la actividad física, especialmente con la flexión plantar. Se acompaña de dolor con el ejercicio en el 67% de los casos publicados. El tratamiento de elección es conservador, pero cuando provoca un síndrome compartimental se debe realizar la fasciotomía y, en el caso de tener síntomas de claudicación o compresión nerviosa, se necesitará una escisión completa del músculo. Presentamos el caso clínico de una deportista que presenta un músculo sóleo accesorio sintomático que fue estudiado mediante radiología, ecografía y resonancia magnética nuclear (RMN)

The incidence of an accessory soleus muscle, according to autopsy studies, ranged from 0.5 to 6.0% of the population. The typical presentation is a soft mass in the posteromedial distal third of the leg, which increases in size with physical activity, especially plantar flexion. It is accompanied by pain with exercise in 67% of reported cases. The treatment of choice is conservative, but when it causes compartment syndrome, fasciotomy should be performed. If a patient has symptoms of claudication or nerve compression, a complete excision of the muscle is required. We report the case of an athlete who had a symptomatic accessory soleus muscle, which was studied by standard X-ray, ultrasound and magnetic resonance imaging (MRI)

Detection of molecular alterations in methamphetamine-activated Fos-expressing neurons from a single rat dorsal striatum using fluorescence-activated cell sorting (FACS).

Methamphetamine and other drugs activate a small proportion of all neurons in the brain. We previously developed a fluorescence-activated cell sorting (FACS)-based method to characterize molecular alterations induced selectively in activated neurons that express the neural activity marker Fos. However, this method requires pooling samples from many rats. We now describe a modified FACS-based method to characterize molecular alterations in Fos-expressing dorsal striatal neurons from a single rat using a multiplex pre-amplification strategy. Fos and NeuN (a neuronal marker) immunohistochemistry indicate that 5-6% of dorsal striatum neurons were activated 90 min after acute methamphetamine injections (5 mg/kg, i.p.) while less than 0.5% of neurons were activated by saline injections. We used FACS to separate NeuN-labeled neurons into Fos-positive and Fos-negative neurons and assessed mRNA expression using RT-qPCR from as little as five Fos-positive neurons. Methamphetamine induced 3-20-fold increases of immediate early genes arc, homer-2, c-fos, fosB, and its isoforms (ΔfosB and a novel isoform ΔfosB-2) in Fos-positive but not Fos-negative neurons. Immediate early gene mRNA induction was 10-fold lower or absent when assessed in unsorted samples from single dorsal striatum homogenates. Our modified method makes it feasible to study unique molecular alterations in neurons activated by drugs or drug-associated cues in complex addiction models. Methamphetamine and other drugs activate a small proportion of all neurons in the brain. We here report an improved method to characterize molecular alterations induced selectively in activated neurons that express the neural activity marker Fos. We used FACS along with targeted PCR pre-amplification to assess acute methamphetamine-induced gene expression from as few as 5 Fos-expressing neurons from a single rat dorsal striatum. Methamphetamine induced 3-20-fold increases of immediate early genes (IEGs) in Fos-positive but not Fos-negative neurons. Targeted PCR pre-amplification makes it feasible to study unique molecular alterations in neurons activated by drugs or drug-associated cues in complex addiction models.

The glycolytic enzyme aldolase C is up-regulated in rat forebrain microsomes and in the cerebrospinal fluid after repetitive fluoxetine treatment.

The antidepressant drug fluoxetine is widely used for the treatment of a broad range of psychiatric disorders. Its mechanism of action is thought to involve cellular adaptations that are induced with a slow time course after initiation of treatment. To gain insight into the signaling pathways underlying such changes, the expression levels of proteins in a microsomal sub-fraction enriched in intracellular membranes from the rat forebrain was analyzed after two weeks of treatment with fluoxetine. Proteins were separated by two-dimensional gel electrophoresis, and the differentially regulated protein spots were identified by mass spectrometry. Protein network analysis suggested that most of the identified proteins could potentially be regulated by the insulin family of proteins. Among them, Fructose-bisphosphate aldolase C (AldoC), a glycolytic/gluconeogenic enzyme primarily expressed in forebrain astrocytes, was up-regulated 7.6-fold. An immunohistochemical analysis of the dorsal hippocampus revealed a robust decrease (43±2%) in the co-localization of AldoC and the astrocyte marker GFAP and a diffuse staining pattern, compatible with AldoC secretion into the extracellular space. Consistently, AldoC, contained in an exosome-like fraction in astrocyte conditioned medium, increased significantly in the cerebrospinal fluid. Our findings strongly favor a non-canonic signaling role for AldoC in cellular adaptations induced by repetitive fluoxetine treatment.

Long-term fluoxetine treatment induces input-specific LTP and LTD impairment and structural plasticity in the CA1 hippocampal subfield.

Antidepressant drugs are usually administered for several weeks for the treatment of major depressive disorder. However, they are also prescribed in several additional psychiatric conditions as well as during long-term maintenance treatments. Antidepressants induce adaptive changes in several forebrain structures which include modifications at glutamatergic synapses. We recently found that repetitive administration of the selective serotonin reuptake inhibitor (SSRI) fluoxetine to naïve adult male rats induced an increase of mature, mushroom-type dendritic spines in several forebrain regions. This was associated with an increase of GluA2-containing α-amino-3-hydroxy-5-methylisoxazole-4-propionate receptors (AMPA-Rs) in telencephalic postsynaptic densities. To unravel the functional significance of such a synaptic re-arrangement, we focused on glutamate neurotransmission in the hippocampus. We evaluated the effect of four weeks of 0.7 mg/kg fluoxetine on long-term potentiation (LTP) and long-term depression (LTD) in the CA1 hippocampal subfield. Recordings in hippocampal slices revealed profound deficits in LTP and LTD at Schaffer collateral-CA1 synapses associated to increased spine density and enhanced presence of mushroom-type spines, as revealed by Golgi staining. However, the same treatment had neither an effect on spine morphology, nor on LTP and LTD at perforant path-CA1 synapses. Cobalt staining and immunohistochemical experiments revealed decreased AMPA-R Ca(2+) permeability in the stratum radiatum (s.r.) together with increased GluA2-containing Ca(2+) impermeable AMPA-Rs. Therefore, 4 weeks of fluoxetine treatment promoted structural and functional adaptations in CA1 neurons in a pathway-specific manner that were selectively associated with impairment of activity-dependent plasticity at Schaffer collateral-CA1 synapses.

Cell therapy using induced pluripotent stem cells or somatic stem cells: this is the question.

A lot of effort has been developed to bypass the use of embryonic stem cells (ES) in human therapies, because of several concerns and ethical issues. Some unsolved problems of using stem cells for human therapies, excluding the human embryonic origin, are: how to regulate cell plasticity and proliferation, immunological compatibility, potential adverse side-effects when stem cells are systemically administrated, and the in vivo signals to rule out a specific cell fate after transplantation. Currently, it is known that almost all tissues of an adult organism have somatic stem cells (SSC). Whereas ES are primary involved in the genesis of new tissues and organs, SSC are involved in regeneration processes, immuno-regulatory and homeostasis mechanisms. Although the differentiating potential of ES is higher than SSC, several studies suggest that some types of SSC, such as mesenchymal stem cells (MSC), can be induced epigenetically to differentiate into tissue-specific cells of different lineages. This unexpected pluripotency and the variety of sources that they come from, can make MSC-like cells suitable for the treatment of diverse pathologies and injuries. New hopes for cell therapy came from somatic/mature cells and the discovery that could be reprogrammed to a pluripotent stage similar to ES, thus generating induced pluripotent stem cells (iPS). For this, it is necessary to overexpress four main reprogramming factors, Sox2, Oct4, Klf4 and c-Myc. The aim of this review is to analyze the potential and requirements of cellular based tools in human therapy strategies, focusing on the advantage of using MSC over iPS.