Highly efficient neural differentiation of human somatic stem cells, isolated by minimally invasive periodontal surgery

Neural stem cells (NSCs) are potential sources for cell therapy of neurodegenerative diseases and for drug screening. Despite their potential benefits, ethical and practical considerations limit the application of NSCs derived from human embryonic stem cells (ES) or adult brain tissue. Thus, alternative sources are required to satisfy the criteria of ready accessibility, rapid expansion in chemically defined media and reliable induction to a neuronal fate. We isolated somatic stem cells from the human periodontium that were collected during minimally invasive periodontal access flap surgery as part of guided tissue regeneration therapy. These cells could be propagated as neurospheres in serum-free medium, which underscores their cranial neural crest cell origin. Culture in the presence of epidermal growth factor (EGF) and fibroblast growth factor-2 (FGF-2) under serum-free conditions resulted in large numbers of nestin-positive/Sox-2-positive NSCs. These periodontium-derived (pd) NSCs are highly proliferative and migrate in response to chemokines that have been described as inducing NSC migration. We used immunocytochemical techniques and RT-PCR analysis to assess neural differentiation after treatment of the expanded cells with a novel induction medium. Adherence to substrate, growth factor deprivation, and retinoic acid treatment led to the acquisition of neuronal morphology and stable expression of markers of neuronal differentiation by more than 90% of the cells. Thus, our novel method might provide nearly limitless numbers of neuronal precursors from a readily accessible autologous adult human source, which could be used as a platform for further experimental studies and has potential therapeutic implications.

Towards pain-free diagnosis of skin diseases through multiplexed microneedles: biomarker extraction and detection using a highly sensitive blotting method

Immunodiagnostic microneedles provide a novel way to extract protein biomarkers from the skin in a minimally invasive manner for analysis in vitro. The technology could overcome challenges in biomarker analysis specifically in solid tissue, which currently often involves invasive biopsies. This study describes the development of a multiplex immunodiagnostic device incorporating mechanisms to detect multiple antigens simultaneously, as well as internal assay controls for result validation. A novel detection method is also proposed. It enables signal detection specifically at microneedle tips and therefore may aid the construction of depth profiles of skin biomarkers. The detection method can be coupled with computerised densitometry for signal quantitation. The antigen specificity, sensitivity and functional stability of the device were assessed against a number of model biomarkers. Detection and analysis of endogenous antigens (interleukins 1α and 6) from the skin using the device was demonstrated. The results were verified using conventional enzyme-linked immunosorbent assays. The detection limit of the microneedle device, at ≤10 pg/mL, was at least comparable to conventional plate-based solid-phase enzyme immunoassays.

Prior history of sexually transmitted diseases in women living with AIDS in Sao Paulo, Brazil

Objectives: To describe the epidemiological profile, risk behaviors, and the prior history of sexually transmitted diseases (STDs) in women living with acquired immunodeficiency syndrome (AIDS). Methods: Cross-sectional study, performed at the Centro de Referencia e Treinamento em DST/AIDS of Sao Paulo. The social, demographic, behavioral, and clinical data such as age, schooling, marital status, age at first sexual intercourse, number of sexual partners, parity, use of drugs, time of HIV diagnosis, CD4 count, and viral load determination were abstracted from the medical records of women living with AIDS who had gynecological consultation scheduled in the period from June 2008 to May 2009. Results: Out of 710 women who were scheduled to a gynecological consultation during the period of the study, 598 were included. Previous STD was documented for 364 (60.9%; 95% CI: 56.9%-64.8%) women. The associated factors with previous STDs and their respective risks were: human development index (HDI) <0.50 (ORaj = 5.5; 95% CI: 2.8-11.0); non-white race (ORaj = 5.2; 95% CI: 2.5-11.0); first sexual intercourse at or before 15 years of age (ORaj = 4.4; 95% CI: 2.3-8.3); HIV infection follow-up time of nine years or more (ORaj = 4.2; 95% CI: 2.3-7.8)]; number of sexual partners during the entire life between three and five partners (ORaj = 2.2; 95% CI: 1.1-4.6), and six or more sexual partners (ORaj = 3.9; 95% CI: 1.9-8.0%); being a sex worker (ORaj = 1.9; 95% CI: 1.1-3.1). Conclusions: A high prevalence of a prior history of STDs in the studied population was found. It is essential to find better ways to access HIV infection prevention, so that effective interventions can be more widely implemented. (C) 2012 Elsevier Editora Ltda. All rights reserved.

New molecular targets for PET and SPECT imaging in neurodegenerative diseases

The pathophysiology of neurodegenerative diseases (ND) such as Alzheimer's disease (AD) and Parkinson's disease (PD) has not yet been completely elucidated. However, in the past few years, there have been great knowledge advances about intra-and extracellular proteins that may display impaired function or expression in AD, PD and other ND, such as amyloid beta (AB), alpha-synuclein, tau protein and neuroinfiammatory markers. Recent developments in the imaging techniques of positron emission tomography (PET) and single photon emission computed tomography (SPECT) now allow the non-invasive tracking of such molecular targets of known relevance to ND in vivo. This article summarizes recent findings of PET and SPECT studies using these novel methods, and discusses their potential role in the field of drug development for ND as well as future clinical applications in regard to differential diagnosis of ND and monitoring of disease progression.

Invasive candidiasis and oral manifestations in premature newborns

Objective: To investigate prevalence of invasive candidiasis in a Neonatal Intensive Care Unit and to evaluate oral diseases and Candida spp. colonization in low birth weight preterm newborns. Methods: A descriptive epidemiological study performed in two stages. First, prevalence of candidiasis was analyzed in a database of 295 preterm patients admitted to hospital for over 10 days and birth weight less than 2,000g. In the second stage, oral changes and Candida spp. colonization were assessed in 65 patients weighing less than 2,000g, up to 4 week-old, hospitalized for over 10 days and presenting oral abnormalities compatible with fungal lesions. Swab samples were collected in the mouth to identify fungi. Results: Prevalence of candidiasis was 5.4% in the database analyzed. It correlated with prolonged hospital length of stay (p<0.001), in average, 31 days, and 85% risk of developing infection in the first 25 days. It correlated with low birth weight (p<0.001), with mean of 1,140g. The most frequent alterations were white soft plaques, detachable, in oral mucosa and tongue. Intense oral colonization by Candida spp was observed (80%). Conclusions: The frequency of invasive candidiasis was low and correlated with low birth weight and prolonged hospital stay. The most common oral changes were white plaques compatible with pseudomembranous candidiasis and colonization by Candida spp. was above average.

Biomarkers for Hepatocellular Apoptosis in the Management of Liver Diseases

Apoptosis is a rare event in normal hepatocytes. However, multiple signals can trigger apoptosis in hepatocytes and it plays a role in the pathogenesis of many liver diseases. This review summarizes the mechanisms of hepatocellular apoptosis and the importance of apoptosis in the pathological processes of liver disease. The potential for non-invasive biomarkers of apoptosis to gauge the extent and follow the evolution of clinical disease is emphasized.

Vascular Diseases of the Spinal Cord: A Review

OPINION STATEMENT: • In acute spinal cord ischemia syndrome (ASCIS), treatment recommendations are derived from data of cerebral ischemic stroke, atherosclerotic vascular disease and acute spinal cord injury. Besides acute management, secondary prevention is of major importance. Pathologies affecting the aorta as well as underlying cerebrovascular conditions should be treated whenever possible.• ASCIS may occur after aortic surgery, less often after thoracic endovascular aortic repair (TEVAR). Protocols are proposed.• Acute spinal cord hemorrhage can be treated surgically and/or pharmacologically.• Symptomatic treatment in patients with a spinal cord lesion is of major importance. Depending on level and extension of the lesion, there is a risk for systemic and neurological complications, which may be life-threatening.• Each spinal vascular malformation is a unique lesion that needs an individualized treatment algorithm. In case of a symptomatic vascular malformation, endovascular intervention is the primary treatment option.• Spinal dural Arteriovenous fistula (AVF) may be treated endovascularly or surgically. If preoperative localization of the fistula is possible, surgery is feasible with a low complication rate. In comparison, endovascular approaches are less invasive.• Spinal AVM are rather treated endovascularly than surgically or in a stepwise multidisciplinary approach.• Symptomatic and exophytic spinal cavernous angiomas should be treated surgically. Deep spinal cavernous angiomas that are asymptomatic or only show mild symptoms can be observed.

A randomized, blinded, multicenter trial of lipid-associated amphotericin B alone versus in combination with an antibody-based inhibitor of heat shock protein 90 in patients with invasive candidiasis

BACKGROUND: Mycograb (NeuTec Pharma) is a human recombinant monoclonal antibody against heat shock protein 90 that, in laboratory studies, was revealed to have synergy with amphotericin B against a broad spectrum of Candida species. METHODS: A double-blind, randomized study was conducted to determine whether lipid-associated amphotericin B plus Mycograb was superior to amphotericin B plus placebo in patients with culture-confirmed invasive candidiasis. Patients received a lipid-associated formulation of amphotericin B plus a 5-day course of Mycograb or placebo, having been stratified on the basis of Candida species (Candida albicans vs. non-albicans species of Candida). Inclusion criteria included clinical evidence of active infection at trial entry plus growth of Candida species on culture of a specimen from a clinically significant site within 3 days after initiation of study treatment. The primary efficacy variable was overall response to treatment (clinical and mycological resolution) by day 10. RESULTS: Of the 139 patients enrolled from Europe and the United States, 117 were included in the modified intention-to-treat population. A complete overall response by day 10 was obtained for 29 (48%) of 61 patients in the amphotericin B group, compared with 47 (84%) of 56 patients in the Mycograb combination therapy group (odds ratio [OR], 5.8; 95% confidence interval [CI], 2.41-13.79; P<.001). The following efficacy criteria were also met: clinical response (52% vs. 86%; OR, 5.4; 95% CI, 2.21-13.39; P<.001), mycological response (54% vs. 89%; OR, 7.1; 95% CI, 2.64-18.94; P<.001), Candida-attributable mortality (18% vs. 4%; OR, 0.2; 95% CI, 0.04-0.80; P = .025), and rate of culture-confirmed clearance of the infection (hazard ratio, 2.3; 95% CI, 1.4-3.8; P = .001). Mycograb was well tolerated. CONCLUSIONS: Mycograb plus lipid-associated amphotericin B produced significant clinical and culture-confirmed improvement in outcome for patients with invasive candidiasis.

EcPV2 DNA in equine papillomas and in situ and invasive squamous cell carcinomas supports papillomavirus etiology

Equine penile papillomas, in situ carcinomas, and invasive carcinomas are hypothesized to belong to a continuum of papillomavirus-induced diseases. The former ones clinically present as small grey papules, while the latter 2 lesions are more hyperplasic or alternatively ulcerated. To test the hypothesis that these lesions are papillomavirus-induced, samples of 24 horses with characteristic clinical and histologic findings of penile papillomas or in situ or invasive squamous cell carcinomas were collected. As controls, 11 horses with various lesions--namely, Balanoposthitis (6 cases), melanoma (3 cases), follicular cyst (1 case), and amyloidosis (1 case)--were included. DNA was extracted and polymerase chain reaction applied to amplify papillomavirus DNA. The respective primers were designed to amplify DNA of the recently discovered equine papillomavirus EcPV2. All tested papilloma and squamous cell carcinoma samples were found to contain DNA of either of 2 previously published EcPV2 variants. Among the other samples 6 of 11 were found to contain EcPV2 DNA. To further support the findings and to determine where the papillomavirus DNA was located within the lesions, an in situ hybridization for the detection of EcPV2 DNA was established. The samples tested by this technique were found to clearly contain papillomavirus nucleic acid concentrated in the nucleus of the koilocytes. The findings of this study support previous data and the hypothesis that papillomaviruses induce the described penile lesions in horses.

IL1B and DEFB1 Polymorphisms Increase Susceptibility to Invasive Mold Infection After Solid Organ Transplantation.

BACKGROUND  Single nucleotide polymorphisms (SNPs) in immune genes have been associated with susceptibility to invasive mold infection (IMI) among hematopoietic stem cell (HSCT) but not solid organ transplant (SOT) recipients. METHODS  24 SNPs from systematically selected genes were genotyped among 1101 SOT recipients (715 kidneys, 190 liver, 102 lungs, 79 hearts, 15 other) from the Swiss Transplant Cohort Study. Association between SNPs and the endpoint were assessed by log-rank test and Cox regression models. Cytokine production upon Aspergillus stimulation was measured by ELISA in PBMCs from healthy volunteers and correlated with relevant genotypes. RESULTS  Mold colonization (N=45) and proven/probable IMI (N=26) were associated with polymorphisms in interleukin-1 beta (IL1B, rs16944; log-rank test, recessive mode, colonization P=0.001 and IMI P=0.00005), interleukin-1 receptor antagonist (IL1RN, rs419598; P=0.01 and P=0.02) and β-defensin-1 (DEFB1, rs1800972; P=0.001 and P=0.0002, respectively). The associations with IL1B and DEFB1 remained significant in a multivariate regression model (IL1B rs16944 P=0.002; DEFB1 rs1800972 P=0.01). Presence of two copies of the rare allele of rs16944 or rs419598 was associated with reduced Aspergillus-induced IL-1β and TNFα secretion by PBMCs. CONCLUSIONS  Functional polymorphisms in IL1B and DEFB1 influence susceptibility to mold infection in SOT recipients. This observation may contribute to individual risk stratification.

PTX3 Polymorphisms and Invasive Mold Infections after Solid Organ Transplantation

Donor PTX3 polymorphisms were shown to influence the risk of invasive aspergillosis among hematopoietic stem cell transplant recipients. Here, we show that PTX3 polymorphisms are independent risk factors for invasive mold infection among 1101 solid organ transplant recipients, thereby strengthening their role in mold infection pathogenesis and patient's risk stratification.

DIMETER: a haptic master device for tremor diagnosis in neurodegenerative diseases

In this study, a device based on patient motion capture is developed for the reliable and non-invasive diagnosis of neurodegenerative diseases. The primary objective of this study is the classification of differential diagnosis between Parkinson's disease (PD) and essential tremor (ET). The DIMETER system has been used in the diagnoses of a significant number of patients at two medical centers in Spain. Research studies on classification have primarily focused on the use of well-known and reliable diagnosis criteria developed by qualified personnel. Here, we first present a literature review of the methods used to detect and evaluate tremor; then, we describe the DIMETER device in terms of the software and hardware used and the battery of tests developed to obtain the best diagnoses. All of the tests are classified and described in terms of the characteristics of the data obtained. A list of parameters obtained from the tests is provided, and the results obtained using multilayer perceptron (MLP) neural networks are presented and analyzed.

Non-invasive user monitoring in daily tasks for cognitive rehabilitation

El uso de técnicas para la monitorización del movimiento humano generalmente permite a los investigadores analizar la cinemática y especialmente las capacidades motoras en aquellas actividades de la vida cotidiana que persiguen un objetivo concreto como pueden ser la preparación de bebidas y comida, e incluso en tareas de aseo. Adicionalmente, la evaluación del movimiento y el comportamiento humanos en el campo de la rehabilitación cognitiva es esencial para profundizar en las dificultades que algunas personas encuentran en la ejecución de actividades diarias después de accidentes cerebro-vasculares. Estas dificultades están principalmente asociadas a la realización de pasos secuenciales y al reconocimiento del uso de herramientas y objetos. La interpretación de los datos sobre la actitud de este tipo de pacientes para reconocer y determinar el nivel de éxito en la ejecución de las acciones, y para ampliar el conocimiento en las enfermedades cerebrales, sus consecuencias y severidad, depende totalmente de los dispositivos usados para la captura de esos datos y de la calidad de los mismos. Más aún, existe una necesidad real de mejorar las técnicas actuales de rehabilitación cognitiva contribuyendo al diseño de sistemas automáticos para crear una especie de terapeuta virtual que asegure una vida más independiente de estos pacientes y reduzca la carga de trabajo de los terapeutas. Con este objetivo, el uso de sensores y dispositivos para obtener datos en tiempo real de la ejecución y estado de la tarea de rehabilitación es esencial para también contribuir al diseño y entrenamiento de futuros algoritmos que pudieran reconocer errores automáticamente para informar al paciente acerca de ellos mediante distintos tipos de pistas como pueden ser imágenes, mensajes auditivos o incluso videos. La tecnología y soluciones existentes en este campo no ofrecen una manera totalmente robusta y efectiva para obtener datos en tiempo real, por un lado, porque pueden influir en el movimiento del propio paciente en caso de las plataformas basadas en el uso de marcadores que necesitan sensores pegados en la piel; y por otro lado, debido a la complejidad o alto coste de implantación lo que hace difícil pensar en la idea de instalar un sistema en el hospital o incluso en la casa del paciente. Esta tesis presenta la investigación realizada en el campo de la monitorización del movimiento de pacientes para proporcionar un paso adelante en términos de detección, seguimiento y reconocimiento del comportamiento de manos, gestos y cara mediante una manera no invasiva la cual puede mejorar la técnicas actuales de rehabilitación cognitiva para la adquisición en tiempo real de datos sobre el comportamiento del paciente y la ejecución de la tarea. Para entender la importancia del marco de esta tesis, inicialmente se presenta un resumen de las principales enfermedades cognitivas y se introducen las consecuencias que tienen en la ejecución de tareas de la vida diaria. Más aún, se investiga sobre las metodologías actuales de rehabilitación cognitiva. Teniendo en cuenta que las manos son la principal parte del cuerpo para la ejecución de tareas manuales de la vida cotidiana, también se resumen las tecnologías existentes para la captura de movimiento de manos. Una de las principales contribuciones de esta tesis está relacionada con el diseño y evaluación de una solución no invasiva para detectar y seguir las manos durante la ejecución de tareas manuales de la vida cotidiana que a su vez involucran la manipulación de objetos. Esta solución la cual no necesita marcadores adicionales y está basada en una cámara de profundidad de bajo coste, es robusta, precisa y fácil de instalar. Otra contribución presentada se centra en el reconocimiento de gestos para detectar el agarre de objetos basado en un sensor infrarrojo de última generación, y también complementado con una cámara de profundidad. Esta nueva técnica, y también no invasiva, sincroniza ambos sensores para seguir objetos específicos además de reconocer eventos concretos relacionados con tareas de aseo. Más aún, se realiza una evaluación preliminar del reconocimiento de expresiones faciales para analizar si es adecuado para el reconocimiento del estado de ánimo durante la tarea. Por su parte, todos los componentes y algoritmos desarrollados son integrados en un prototipo simple para ser usado como plataforma de monitorización. Se realiza una evaluación técnica del funcionamiento de cada dispositivo para analizar si es adecuada para adquirir datos en tiempo real durante la ejecución de tareas cotidianas reales. Finalmente, se estudia la interacción con pacientes reales para obtener información del nivel de usabilidad del prototipo. Dicha información es esencial y útil para considerar una rehabilitación cognitiva basada en la idea de instalación del sistema en la propia casa del paciente al igual que en el hospital correspondiente. ABSTRACT The use of human motion monitoring techniques usually let researchers to analyse kinematics, especially in motor strategies for goal-oriented activities of daily living, such as the preparation of drinks and food, and even grooming tasks. Additionally, the evaluation of human movements and behaviour in the field of cognitive rehabilitation is essential to deep into the difficulties some people find in common activities after stroke. This difficulties are mainly associated with sequence actions and the recognition of tools usage. The interpretation of attitude data of this kind of patients in order to recognize and determine the level of success of the execution of actions, and to broaden the knowledge in brain diseases, consequences and severity, depends totally on the devices used for the capture of that data and the quality of it. Moreover, there is a real need of improving the current cognitive rehabilitation techniques by contributing to the design of automatic systems to create a kind of virtual therapist for the improvement of the independent life of these stroke patients and to reduce the workload of the occupational therapists currently in charge of them. For this purpose, the use of sensors and devices to obtain real time data of the execution and state of the rehabilitation task is essential to also contribute to the design and training of future smart algorithms which may recognise errors to automatically provide multimodal feedback through different types of cues such as still images, auditory messages or even videos. The technology and solutions currently adopted in the field don't offer a totally robust and effective way for obtaining real time data, on the one hand, because they may influence the patient's movement in case of marker-based platforms which need sensors attached to the skin; and on the other hand, because of the complexity or high cost of implementation, which make difficult the idea of installing a system at the hospital or even patient's home. This thesis presents the research done in the field of user monitoring to provide a step forward in terms of detection, tracking and recognition of hand movements, gestures and face via a non-invasive way which could improve current techniques for cognitive rehabilitation for real time data acquisition of patient's behaviour and execution of the task. In order to understand the importance of the scope of the thesis, initially, a summary of the main cognitive diseases that require for rehabilitation and an introduction of the consequences on the execution of daily tasks are presented. Moreover, research is done about the actual methodology to provide cognitive rehabilitation. Considering that the main body members involved in the completion of a handmade daily task are the hands, the current technologies for human hands movements capture are also highlighted. One of the main contributions of this thesis is related to the design and evaluation of a non-invasive approach to detect and track user's hands during the execution of handmade activities of daily living which involve the manipulation of objects. This approach does not need the inclusion of any additional markers. In addition, it is only based on a low-cost depth camera, it is robust, accurate and easy to install. Another contribution presented is focused on the hand gesture recognition for detecting object grasping based on a brand new infrared sensor, and also complemented with a depth camera. This new, and also non-invasive, solution which synchronizes both sensors to track specific tools as well as recognize specific events related to grooming is evaluated. Moreover, a preliminary assessment of the recognition of facial expressions is carried out to analyse if it is adequate for recognizing mood during the execution of task. Meanwhile, all the corresponding hardware and software developed are integrated in a simple prototype with the purpose of being used as a platform for monitoring the execution of the rehabilitation task. Technical evaluation of the performance of each device is carried out in order to analyze its suitability to acquire real time data during the execution of real daily tasks. Finally, a kind of healthcare evaluation is also presented to obtain feedback about the usability of the system proposed paying special attention to the interaction with real users and stroke patients. This feedback is quite useful to consider the idea of a home-based cognitive rehabilitation as well as a possible hospital installation of the prototype.

Uncoupling signal transducers from oncogenic MET mutants abrogates cell transformation and inhibits invasive growth

The assumption that genes encoding tyrosine kinase receptors could play a role in human cancers has been confirmed by the identification of oncogenic mutations in the kinase domain of RET and KIT. Recently, homologous residues were found mutated in MET, in papillary renal carcinomas (PRCs). The link coupling these genetic lesions to cellular transformation is still unclear. METPRC mutations result in increased kinase activity and—in some instances, i.e., M1250T substitution—in changes in substrate specificity. A direct correlation occurs between the transforming potential of METPRC mutants and their ability to constitutively associate with signal transducers through two phosphorylated tyrosines (Y1349VHVNATY1356VNV) located in the receptor tail. Substitution of these “docking tyrosines” with phenylalanines leaves unaffected the altered properties of the kinase but abrogates transformation and invasiveness in vitro. Uncoupling the receptor from signal transducers with a tyrosine-phosphorylated peptide derivative (YpVNV) inhibits invasive growth induced by METPRC mutants. These data indicate that constitutive receptor coupling to downstream signal transducers is a key mechanism in neoplastic transformation driven by mutated MET and suggest a therapeutic strategy to target neoplastic diseases associated with this oncogene.