Parte 1: Impostazione e preparare la pelle <ol><li> Impostare i seguenti sensori respirazione cintura, EMG, ECG, conduttanza cutanea, temperatura, terreno; </li><li> Preparare la pelle strofinando delicatamente con un batuffolo imbevuto d&#39;alcol nelle seguenti località: la fronte sopra gli occhi per EMG, all&#39;interno di entrambi i polsi per ECG, dorso della mano sinistra per terra (posizione del dell&#39;elettrodo di massa può variare a seconda macchina di occasione); </li><li> Aggiungere sensori per la pelle nelle seguenti località: respirazione cintura intorno alla vita, EMG sulla fronte sopra ciascun occhio, ECG all&#39;interno di ogni polso, conduttanza cutanea sul dito medio della mano sinistra, termometro sul puntatore della mano sinistra, a terra sul dorso della mano sinistra; </li><li> Seleziona sessione per respirazione HRV +; </li><li> Assicurarsi che impedenza è accettabile per tutti i sensori e che i sensori sono sensibili al movimento del cliente e la produzione di letture ragionevole. </li></ol> Parte 2: Registrazione <ol><li> Spiegare la procedura per il cliente. </li><li> (Min 0-2) Istruire il cliente a convivere tranquillamente con gli occhi aperti. Record di due minuti basale. </li><li> (Min 2-4) Al termine dei due minuti, iniziano a due minuti a stress cognitivo - problemi aritmetici che aumenta gradualmente in difficoltà da risolvere senza l&#39;uso della penna e carta o calcolatrice (se l&#39;aritmetica è particolarmente difficile per il cliente, chiedere loro per sottrarre ripetutamente sette a partire da 100, vale a dire, &quot;sette seriale&quot;). Dare le seguenti istruzioni al cliente: &quot;Nei prossimi 2 minuti, sarò richiesto di risolvere alcuni problemi di matematica. Si prega di lavoro il più velocemente possibile e mi dica la risposta &quot;. Non fornire un feedback se rispondere al cliente è corretto o non corretti ed offrire incoraggiamento minima (es., &quot;basta fare il meglio che puoi&quot;) in modo da non eliminare le lievi proprietà dello stress inducendo di questo esercizio. </li><li> (Min 4-6) Lasciate che il cliente sa che questa parte della valutazione è finita e chiedere a lui / lei a convivere tranquillamente con gli occhi aperti per 2 minuti. </li><li> (Min 6-8) Iniziare fattore di stress fisico: accendere il forte rumore nocivi (bambini che piangono, incidenti stradali, ecc) e chiedere al cliente di ascoltare i suoni senza messa fuori. </li><li> (Min 8-10) Spegnere il rumore, e chiedere al cliente di convivere tranquillamente con gli occhi aperti per 2 minuti. </li><li> (Min 10-12) Iniziare fattore di stress emotivo: chiedere al cliente di parlare di un incidente stressante che ricordo bene, sono i dettagli della manifestazione, i loro pensieri e sentimenti. Fornire minima risposta empatica al fine di non ridurre la risposta allo stress. </li><li> (Min 12-14) Interrompere la storia del cliente, anche se lui / lei non è finito, e istruire il client a convivere tranquillamente con gli occhi aperti. </li><li> Interrompere la registrazione, salvare i dati, e lasciare che il cliente sa che la valutazione è fatta. </li></ol> Parte 3: Debriefing e pianificazione del trattamento <ol><li> Chiedere al cliente circa la loro esperienza durante la valutazione. Quali parti sono state particolarmente stressanti e perché? Forse lui / lei si sentono le alterazioni funzionali succedendo? </li><li> Revisione dati secondo i criteri descritti nell&#39;appendice I. </li><li> Ha scelto i parametri fisiologici che devono essere affrontate sulla base di analisi di cui sopra. Se la lettura del sensore al basale è superiore o inferiore alla norma, l&#39;obiettivo terapeutico è quello di aiutare il cliente a portare quel parametro fisiologico alla normalità in un ambiente non stressante. Se la lettura del sensore durante l&#39;evento stressante è superiore o inferiore alla norma, l&#39;obiettivo terapeutico è quello di aiutare il cliente ad identificare i fattori di stress ed essere in grado di mantenere il normale livello di funzionamento fisiologico sotto stress. Se la lettura del sensore nel periodo di inattività indica la mancanza di recupero, l&#39;obiettivo terapeutico è quello di aiutare il cliente a portare quel parametro fisiologico a livello normale rapidamente a seguito di un fattore di stress. Qualsiasi o di tutti questi obiettivi possono essere inclusi nel piano di trattamento. </li></ol> Appendice I Valutazione dello stress interpretazione linee guida (Tutti &quot;bold&quot; risposte indicano le zone da trattare) <table border="0"><tbody><tr><td colspan="3"> EMG (normale lettura &lt;3 microvolt) </td></tr><tr><td> 1. È EMG elevati al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. EMG è elevato oltre la linea di base in uno dei tre fattori di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero di EMG a livello di base durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td colspan="3"> Frequenza cardiaca (range normale 60-80 battiti al minuto) </td></tr><tr><td> 1. È la frequenza cardiaca elevata al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. Frequenza cardiaca elevata è passata la linea di base in uno dei tre fattori di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero della frequenza cardiaca a livello di base durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td colspan="3"> Frequenza respiratoria (normale tasso &lt;12 respiri al minuto) </td></tr><tr><td> 1. È il tasso di respirazione elevati al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. Tasso di respirazione è elevato oltre la linea di base per ogni fattore di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero della respirazione tasso di riferimento durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td colspan="3"> Conduttanza cutanea </td></tr><tr><td> 1. È conduttanza cutanea elevati al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. È la conduttanza cutanea elevata passato la linea di base per qualsiasi fattore di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero di conduttanza cutanea al basale durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td colspan="3"> Dito temperatura (normale lettura&gt; 90F) </td></tr><tr><td> 1. È la temperatura delle dita bassa al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. La temperatura diminuisce dito passato la linea di base per qualsiasi fattore di stress? </td><td> y es _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero della temperatura dito al basale durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td colspan="3"> Variabilità della frequenza cardiaca (norme variano in base all&#39;età) </td></tr><tr><td> 1. È basso tasso di variabilità cardiaca al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. La variabilità della frequenza cardiaca diminuisce passato la linea di base per qualsiasi fattore di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero della variabilità della frequenza cardiaca durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr></tbody></table> Appendice I Valutazione dello stress interpretazione: <table border="0"><tbody><tr><td colspan="3"> EMG (normale lettura &lt;3 microvolt) </td></tr><tr><td> 1. È EMG elevati al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. EMG è elevato oltre la linea di base in uno dei tre fattori di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero di EMG a livello di base durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td> Frequenza cardiaca (range normale 60-80 battiti al minuto) </td></tr><tr><td> 1. È la frequenza cardiaca elevata al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. Frequenza cardiaca elevata è passata la linea di base in uno dei tre fattori di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero della frequenza cardiaca a livello di base durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td> Frequenza respiratoria (normale tasso &lt;12 respiri al minuto) </td></tr><tr><td> 1. È il tasso di respirazione elevati al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. Tasso di respirazione è elevato oltre la linea di base per ogni fattore di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero della respirazione tasso di riferimento durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td> Conduttanza cutanea (normale lettura &lt;5 ohm micro) </td></tr><tr><td> 1. È conduttanza cutanea elevati al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. È la conduttanza cutanea elevata passato la linea di base per qualsiasi fattore di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero di conduttanza cutanea al basale durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td> Dito temperatura (normale lettura&gt; 90F) </td></tr><tr><td> 1. È la temperatura delle dita bassa al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. La temperatura diminuisce dito passato la linea di base per qualsiasi fattore di stress? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 3. C&#39;è il recupero della temperatura dito al basale durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr><tr><td> Variabilità della frequenza cardiaca (norme variano in base all&#39;età) </td></tr><tr><td> 1. È basso tasso di variabilità cardiaca al basale? </td><td> sì _ </td><td> no _ </td></tr><tr><td> 2. La variabilità della frequenza cardiaca diminuisce passato la linea di base per qualsiasi fattore di stress? </td><td> sì _ </td><td> no _ </td></tr><tr> &lt;td&gt; 3. C&#39;è il recupero della variabilità della frequenza cardiaca durante i periodi di riposo? </td><td> sì _ </td><td> no _ </td></tr></tbody></table> Pianificazione del trattamento: Se la lettura è superiore o inferiore rispetto al normale al basale, obiettivo del trattamento è quello di aiutare clienti che portare parametro fisiologico alla normalità in tutte le circostanze Se la lettura è superiore o inferiore del normale durante l&#39;evento stressante, obiettivo del trattamento è quello di aiutare clienti di identificare i fattori di stress ed essere in grado di mantenere il normale livello di funzionamento fisiologico sotto stress Se una lettura indica mancanza di recupero nel periodo di inattività, obiettivo del trattamento è quello di aiutare clienti che portare parametro fisiologico a livello normale rapidamente a seguito di un fattore di stress.

<ol>
	<li>Arena, J., Schwartz, M. S., Schwartz, M. S., Andrasik, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Psychophysiological+assessment+and+biofeedback+baselines.">Psychophysiological assessment and biofeedback baselines.</a> A Practitioners Guide. , 128-158 (2005).</li><li>Schwartz, N. M., Schwartz, M. S., Schwartz, M. S., Andrasik, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Definitons+of+biofeedback+and+applied+psychophysiology.">Definitons of biofeedback and applied psychophysiology.</a> A Practitioners Guide. , 27-42 (2005).</li></ol>

The authors have nothing to disclose.

Psicofisiologiche completato nella valutazione dello stress appendici fornisce un professionista biofeedback con le informazioni necessarie per formulare un piano di trattamento. Questa valutazione fornisce informazioni su entrambi stereotipie risposta individuale, o uniformità di alcune risposte fisiologiche a tutti i fattori di stress, e di stimolo-risposta specificità, o individuo risposte uniche fisiologiche a diversi tipi di stress. Con queste informazioni, il cliente è poi in grado di imparare a prevedere che t...

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psychophysiological stress assessment using biofeedback 

In mezzo secolo, la ricerca in biofeedback ha dimostrato fino a che punto la mente umana può influenzare il funzionamento del sistema nervoso autonomo, si pensava di essere al di fuori del controllo cosciente. Consentendo agli utenti di osservare i segnali dal proprio corpo, il biofeedback permette loro di sviluppare una maggiore consapevolezza delle proprie reazioni fisiologiche e psicologiche, come lo stress, e imparare a modificare tali reazioni. Professionisti biofeedback può facilitare questo processo, valutando le reazioni della gente s ad eventi moderatamente stressante e la formulazione di un biofeedback basato piano di trattamento. Durante la valutazione dello stress il praticante primo record una base per letture fisiologici, e quindi presenta il client con diversi fattori di stress lieve, come un cognitivi, stress fisico ed emotivo. Varietà di fattori di stress è presentato al fine di determinare una persona stimolo-risposta specificità, o differenze di reazione di ogni persona a stimoli qualitativamente diverso. Questo video illustra il processo di valutazione dello stress psicofisico utilizzando le linee guida generali biofeedback e presente per la pianificazione del trattamento.

Watch this Scientific Journal Video about Psychophysiological Stress Assessment Using Biofeedback at JoVE.com

Psychophysiological Stress Assessment Using Biofeedback

In questo video si descrive un metodo per valutare la reazione psicofisiologica allo stress della persona utilizzando il biofeedback. Saremo anche presenti linee guida generali per la pianificazione del trattamento.

Lo stress di valutazione psicofisiologica Utilizzando Biofeedback

In the last half century, research in biofeedback has shown the extent to which the human mind can influence the functioning of the autonomic nervous ...

psychophysiological-stress-assessment-using-biofeedback

Research

JoVE Journal

Biology

Psychophysiological Stress Assessment Using Biofeedback 

13.2K Views.  Cambridge Health Alliance. In questo video si descrive un metodo per valutare la reazione psicofisiologica allo stress della persona utilizzando il biofeedback. Saremo anche presenti linee guida generali per la pianificazione del trattamento.

Video: Psychophysiological Stress Assessment Using Biofeedback

Monitoring Plant Hormones During Stress Responses

Plant hormones and related signaling compounds play an important role in the regulation of plant responses to various environmental stimuli and stresses. Among the most severe stresses are insect herbivory, pathogen infection, and drought stress. For each of these stresses a specific set of hormones and/or combinations thereof are known to fine-tune the responses, thereby ensuring the plant's survival. The major hormones involved in the regulation of these responses are jasmonic acid (JA), salicylic acid (SA), and abscisic acid (ABA). To better understand the role of individual hormones as well as their potential interaction during these responses it is necessary to monitor changes in their abundance in a temporal as well as in a spatial fashion. For the easy, sensitive, and reproducible quantification of these and other signaling compounds we developed a method based on vapor phase extraction and gas chromatography/mass spectrometry (GC/MS) analysis (1, 2, 3, 4). After extracting these compounds from the plant tissue by acidic aqueous 1-propanol mixed with dichloromethane the carboxylic acid-containing compounds are methylated, volatilized under heat, and collected on a polymeric absorbent. After elution into a sample vial the analytes are separated by gas chromatography and detected by chemical ionization mass spectrometry. The use of appropriate internal standards then allows for the simple quantification by relating the peak areas of analyte and internal standard.

A simple method is provided that allows for the rapid extraction and analysis of multiple plant hormones from small tissue samples. The procedure uses vapor phase extraction as the solemn purification step. Samples are analyzed by GC/MS with chemical ionization that produces mainly (M+1)+ ions.

Monitoraggio ormoni vegetali Durante risposte allo stress

Plant hormones and related signaling compounds play an important role in the regulation of plant responses to various environmental stimuli and stresses. ...

Ricerca

Biologia

Microsurgical Clip Obliteration of Middle Cerebral Aneurysm Using Intraoperative Flow Assessment

Cerebral aneurysms are abnormal widening or ballooning of a localized segment of an intracranial blood vessel.  Surgical clipping is an important treatment for aneurysms which attempts to exclude blood from flowing into the aneurysmal segment of the vessel while preserving blood flow in a normal fashion.  Improper clip placement may result in residual aneurysm with the potential for subsequent aneurysm rupture or partial or full occlusion of distal arteries resulting in cerebral infarction.  Here we describe the use of an ultrasonic flow probe to provide quantitative evaluation of arterial flow before and after microsurgical clip placement at the base of a middle cerebral artery aneurysm.  This information helps ensure adequate aneurysm reconstruction with preservation of normal distal blood flow.

Description of the surgical obliteration of a cerebral aneurysm utilizing an ultrasonic flow probe to assess arterial flow prior to and after aneurysm clip placement.

Microchirurgico Obliteration clip del Medio aneurisma cerebrale mediante valutazione del flusso intraoperatoria

Cerebral aneurysms are abnormal widening or ballooning of a localized segment of an intracranial blood vessel.  Surgical clipping is an important ...

A Quantitative Assessment of The Yeast Lipidome using Electrospray Ionization Mass Spectrometry

Lipids are one of the major classes of biomolecules and play important roles membrane dynamics, energy storage, and signalling1-4. The budding yeast Saccharomyces cerevisiae, a genetically and biochemically manipulable unicellular eukaryote with annotated genome and very simple lipidome, is a valuable model for studying biological functions of various lipid species in multicellular eukaryotes2,3,5. S. cerevisiae has 10 major classes of lipids with chain lengths mainly of 16 or 18 carbon atoms and either zero or one degree of unsaturation6,7. Existing methods for lipid identification and quantification - such as high performance liquid chromatography, thin-layer chromatography, fluorescence microscopy, and gas chromatography followed by MS - are well established but have low sensitivity, insufficiently separate various molecular forms of lipids, require lipid derivitization prior to analysis, or can be quite time consuming. Here we present a detailed description of our experimental approach to solve these inherent limitations by using survey-scan ESI/MS for the identification and quantification of the entire complement of lipids in yeast cells. The described method does not require chromatographic separation of complex lipid mixtures recovered from yeast cells, thereby greatly accelerating the process of data acquisition. This method enables lipid identification and quantification at the concentrations as low as g/ml and has been successfully applied to assessing lipidomes of whole yeast cells and their purified organelles. Lipids extraction from whole yeast cells for using this method of lipid analysis takes two to three hours. It takes only five to ten minutes to run each sample of extracted and dried lipids on a Q-TOF mass spectrometer equipped with a nano-electrospray source.

We describe a new quantitative lipidomics method for identifying numerous lipid species in yeast using survey-scan electrospray ionization mass spectrometry (ESI/MS). This method exceeds currently available methods for lipid identification and quantification in the ability to resolve various molecular forms of lipids, sensitivity, and speed.

Una valutazione quantitativa del Lipidome lievito con ionizzazione Electrospray Spettrometria di massa

Lipids are one of the major classes of biomolecules and play important roles membrane dynamics, energy storage, and signalling1-4. The budding yeast ...

The use of Biofeedback in Clinical Virtual Reality: The INTREPID Project

Generalized anxiety disorder (GAD) is a psychiatric disorder characterized by a constant and unspecific anxiety that interferes with daily-life activities. Its high prevalence in general population and the severe limitations it causes, point out the necessity to find new efficient strategies to treat it. Together with the cognitive-behavioral treatments, relaxation represents a useful approach for the treatment of GAD, but it has the limitation that it is hard to be learned. The INTREPID project is aimed to implement a new instrument to treat anxiety-related disorders and to test its clinical efficacy in reducing anxiety-related symptoms. The innovation of this approach is the combination of virtual reality and biofeedback, so that the first one is directly modified by the output of the second one. In this way, the patient is made aware of his or her reactions through the modification of some features of the VR environment in real time. Using mental exercises the patient learns to control these physiological parameters and using the feedback provided by the virtual environment is able to gauge his or her success. The supplemental use of portable devices, such as PDA or smart-phones, allows the patient to perform at home, individually and autonomously, the same exercises experienced in therapist's office. The goal is to anchor the learned protocol in a real life context, so enhancing the patients' ability to deal with their symptoms. The expected result is a better and faster learning of relaxation techniques, and thus an increased effectiveness of the treatment if compared with traditional clinical protocols.

The Project on Virtual Reality Intelligent Multi-sensor System for the Treatment of Anxiety-related Disorders (INTREPID) is aimed at developing a multi-sensor context-aware virtual reality system for the treatment of anxiety-related disorders.

L&#39;uso del Biofeedback in Clinica Realtà Virtuale: Il Progetto INTREPID

Generalized anxiety disorder (GAD) is a psychiatric disorder characterized by a constant and unspecific anxiety that interferes with daily-life ...

Assessment of GFP Expression and Viability Using the Tali Image-Based Cytometer

Single-cell and population information are commonly obtained either by flow cytometry or fluorescence microscopy. However, these two methods provide different information. Flow cytometry gives quantitative multi-parametric information about physical characteristics and staining or expression, but doesn't allow for visualization. Stand-alone fluorescence microscopy provides visual data, but doesn't allow for straightforward quantitative measurements1.
Image-based cytometry bridges the gap between these two methods, enabling the quick visualization and simultaneous quantitative analysis of thousands of cells in heterogeneous populations2. Here, we present a method for performing cell viability and green fluorescent protein (GFP) expression assays using the Tali Image-Based Cytometer3. The Tali instrument is a 3-channel (bright field, green fluorescence, red fluorescence) benchtop assay platform that offers several advantages over flow cytometry and fluorescence microscopy. The Tali cytometer is less expensive, takes up less bench space, requires less maintenance, and the work flow has been simplified so that the operation and analysis is much simpler and quicker. The Tali cytometer is capable of performing a range of suspension cell-based assays, including GFP and red fluorescent protein (RFP) expression, apoptosis4-6 and cell viability analysis with propidium iodide (PI)7-11.
Here, we demonstrate the use of the Tali instrument in performing a cell viability assay in cells expressing GFP. GFP-transduced cells are stained using the Tali Viability Kit - Dead Cell Red. The cells are then pipetted into a Tali Cellular Analysis Slide and loaded into the cytometer. Bright field, red fluorescence and green fluorescence images are captured and analyzed using assay specific algorithms. Histograms are then generated to display cell size, PI fluorescence intensity, and GFP fluorescence intensity. These parameters can then be thresholded to home in on a specific cell population.
A side-by side comparison of the Tali Image-Based Cytometer and traditional flow cytometry demonstrates that the two methods provide comparable data regarding cell viability and protein expression. However, the Tali instrument provides additional visual information about the cell population that cannot be obtained using a flow cytometer.

This protocol describes how to perform cell viability and fluorescence expression assays using the Tali Image-Based Cytometer.

Valutazione di espressione GFP e sostenibilità Utilizzando la Image-Based Tali Citometro

Single-cell and population information are commonly obtained either by flow cytometry or fluorescence microscopy. However, these two methods provide ...

Analysis of Oxidative Stress in Zebrafish Embryos

High levels of reactive oxygen species (ROS) may cause a change of cellular redox state towards oxidative stress condition. This situation causes oxidation of molecules (lipid, DNA, protein) and leads to cell death. Oxidative stress also impacts the progression of several pathological conditions such as diabetes, retinopathies, neurodegeneration, and cancer. Thus, it is important to define tools to investigate oxidative stress conditions not only at the level of single cells but also in the context of whole organisms. Here, we consider the zebrafish embryo as a useful in vivo system to perform such studies and present a protocol to measure in vivo oxidative stress. Taking advantage of fluorescent ROS probes and zebrafish transgenic fluorescent lines, we develop two different methods to measure oxidative stress in vivo: i) a &#8220;whole embryo ROS-detection method&#8221; for qualitative measurement of oxidative stress and ii) a &#8220;single-cell&#160;ROS detection method&#8221; for quantitative measurements of oxidative stress. Herein, we demonstrate the efficacy of these procedures by increasing oxidative stress in tissues by oxidant agents and physiological or genetic methods. This protocol is amenable for forward genetic screens and it will help address cause-effect relationships of ROS in animal models of oxidative stress-related pathologies such as neurological disorders and cancer.

Here we report a protocol to measure oxidative stress in living zebrafish embryos. This procedure allows reactive oxygen species (ROS) detection in both whole embryo tissues and single-cell populations. This protocol will accomplish both qualitative and quantitative analyses.

Analisi dello Stress Ossidativo in embrioni di zebrafish

High levels of reactive oxygen species (ROS) may cause a change of cellular redox state towards oxidative stress condition. This situation causes ...

Method for the Assessment of Effects of a Range of Wavelengths and Intensities of Red/near-infrared Light Therapy on Oxidative Stress In Vitro

Red/near-infrared light therapy (R/NIR-LT), delivered by laser or light emitting diode (LED), improves functional and morphological outcomes in a range of central nervous system injuries in vivo, possibly by reducing oxidative stress. However, effects of R/NIR-LT on oxidative stress have been shown to vary depending on wavelength or intensity of irradiation. Studies comparing treatment parameters are lacking, due to absence of commercially available devices that deliver multiple wavelengths or intensities, suitable for high through-put in vitro optimization studies. This protocol describes a technique for delivery of light at a range of wavelengths and intensities to optimize therapeutic doses required for a given injury model. We hypothesized that a method of delivering light, in which wavelength and intensity parameters could easily be altered, could facilitate determination of an optimal dose of R/NIR-LT for reducing reactive oxygen species (ROS) in vitro.
Non-coherent Xenon light was filtered through narrow-band interference filters to deliver varying wavelengths (center wavelengths of 440, 550, 670 and 810nm) and fluences (8.5 x 10-3 to 3.8 x 10-1 J/cm2) of light to cultured cells. Light output from the apparatus was calibrated to emit therapeutically relevant, equal quantal doses of light at each wavelength. Reactive species were detected in glutamate stressed cells treated with the light, using DCFH-DA and H2O2 sensitive fluorescent dyes.&#160;
We successfully delivered light at a range of physiologically and therapeutically relevant wavelengths and intensities, to cultured cells exposed to glutamate as a model of CNS injury. While the fluences of R/NIR-LT used in the current study did not exert an effect on ROS generated by the cultured cells, the method of light delivery is applicable to other systems including isolated mitochondria or more physiologically relevant organotypic slice culture models, and could be used to assess effects on a range of outcome measures of oxidative metabolism.

Method for the Assessment of Effects of a Range of Wavelengths and Intensities of Red/near-infrared Light Therapy on Oxidative Stress In Vitro

Non-coherent Xenon light was passed through narrow-band interference and neutral density filters to deliver light of varying wavelength and intensity to cultured cells. This protocol was used to assess the effects of red/near-infrared light therapy on production of reactive species in vitro: no effects were observed using the tested parameters.

Metodo per la valutazione degli effetti di un intervallo di lunghezze d&#39;onda e intensità di / vicino-infrarosso di terapia della luce rossa sullo stress ossidativo<em&gt; In Vitro</em

Red/near-infrared light therapy (R/NIR-LT), delivered by laser or light emitting diode (LED), improves functional and morphological outcomes in a range of ...

A Protocol for Using Gene Set Enrichment Analysis to Identify the Appropriate Animal Model for Translational Research

Recent studies that compared transcriptomic datasets of human diseases with datasets from mouse models using traditional gene-to-gene comparison techniques resulted in contradictory conclusions regarding the relevance of animal models for translational research. A major reason for the discrepancies between different gene expression analyses is the arbitrary filtering of differentially expressed genes. Furthermore, the comparison of single genes between different species and platforms often is limited by technical variance, leading to misinterpretation of the con/discordance between data from human and animal models. Thus, standardized approaches for systematic data analysis are needed. To overcome subjective gene filtering and ineffective gene-to-gene comparisons, we recently demonstrated that gene set enrichment analysis (GSEA) has the potential to avoid these problems. Therefore, we developed a standardized protocol for the use of GSEA to distinguish between appropriate and inappropriate animal models for translational research. This protocol is not suitable to predict how to design new model systems a-priori, as it requires existing experimental omics data. However, the protocol describes how to interpret existing data in a standardized manner in order to select the most suitable animal model, thus avoiding unnecessary animal experiments and misleading translational studies.

We provide a standardized protocol for the use of gene set enrichment analysis of transcriptomic data to identify an ideal mouse model for translational research. 
This protocol can be used with DNA microarray and RNA sequencing data and can further be extended to other omics data if data are available.

Un protocollo per l'utilizzo di Gene imposta analisi di arricchimento per identificare il modello animale adeguato per la ricerca traslazionale

Recent studies that compared transcriptomic datasets of human diseases with datasets from mouse models using traditional gene-to-gene comparison ...

Fish Sperm Assessment Using Software and Cooling Devices

For gamete quality evaluation, there are innovative, rapid, and quantitative techniques that can provide useful data for aquaculture. Computerized systems for sperm analysis were developed to measure several parameters and one of the most commonly measured is the sperm motility.
Initially, this computer technology was designed for mammalian species, although it can also be used for fish sperm analysis. Fish have specific features that can affect sperm assessment such as a short motility time after activation and, in some cases, adaptation to lower temperatures. Thus, it is necessary to modify both software and hardware components to make motility analysis more efficient for fish sperm analysis. For mammalian sperm, the heating plate is used to maintain optimal temperatures of spermatozoa. However, for some fish species, it is advantageous to use a lower temperature to prolong the duration of motility, since the sperm remain active for less than 2 min. Therefore, cooling devices are necessary to refrigerate samples at constant temperature over the time of analysis, including on the optical microscope. This protocol describes the analysis of fish sperm motility using software for sperm analysis and new cooling devices to optimize the results.

The present protocol describes a procedure of fish sperm assessment using computer-assisted sperm analysis and cooling devices. The software gives a rapid, accurate and quantitative analysis of fish sperm quality based on spermatozoa motility, which can be a useful tool in aquaculture to improve reproduction success.

Valutazione di sperma di pesce utilizzando Software e dispositivi di raffreddamento

For gamete quality evaluation, there are innovative, rapid, and quantitative techniques that can provide useful data for aquaculture. Computerized systems ...

An Air-liquid Interface Bronchial Epithelial Model for Realistic, Repeated Inhalation Exposure to Airborne Particles for Toxicity Testing

For toxicity testing of airborne particles, air-liquid interface (ALI) exposure systems have been developed for in vitro tests in order to mimic realistic exposure conditions. This puts specific demands on the cell culture models. Many cell types are negatively affected by exposure to air (e.g., drying out) and only remain viable for a few days. This limits the exposure conditions that can be used in these models: usually relatively high concentrations are applied as a cloud (i.e., droplets containing particles, which settle down rapidly) within a short period of time. Such experimental conditions do not reflect realistic long-term exposure to low concentrations of particles. To overcome these limitations the use of a human bronchial epithelial cell line, Calu-3 was investigated. These cells can be cultured at ALI conditions for several weeks while retaining a healthy morphology and a stable monolayer with tight junctions. In addition, this bronchial model is suitable for testing the effects of repeated exposures to low, realistic concentrations of airborne particles using an ALI exposure system. This system uses a continuous airflow in contrast to other ALI exposure systems that use a single nebulization producing a cloud. Therefore, the continuous flow system is suitable for repeated and prolonged exposure to airborne particles while continuously monitoring the particle characteristics, exposure concentration, and delivered dose. Taken together, this bronchial model, in combination with the continuous flow exposure system, is able to mimic realistic, repeated inhalation exposure conditions that can be used for toxicity testing.

Described is the cell culture and exposure method of an in vitro bronchial model for realistic, repeated inhalation exposure to particles for toxicity testing.

Un modello epiteliale bronchiale interfaccia aria-liquido per un'esposizione realistica e ripetuta all'inalazione alle particelle trasportate dall'aria per i test di tossicità

For toxicity testing of airborne particles, air-liquid interface (ALI) exposure systems have been developed for in vitro tests in order to mimic realistic ...