In recent years, there has been the difficulty in finding more effective therapies against cancer with less systemic side effects. Therefore Photodynamic Therapy is a novel approach for a more tumor selective treatment.
Photodynamic Therapy (PDT) that makes use of a nontoxic photosensitizer (PS), which, upon activation with light of a specific wavelength in the presence of oxygen, generates oxygen radicals that elicit a cytotoxic response1. Despite its approval almost twenty years ago by the FDA, PDT is nowadays only used to treat a limited number of cancer types (skin, bladder) and nononcological diseases (psoriasis, actinic keratosis)2.
The major advantage of the use of PDT is the ability to perform a local treatment, which prevents systemic side effects. Moreover, it allows the treatment of tumors at delicate sites (e.g. around nerves or blood vessels). Here, an intraoperative application of PDT is considered in osteosarcoma (OS), a tumor of the bone, to target primary tumor satellites left behind in tumor surrounding tissue after surgical tumor resection. The treatment aims at decreasing the number of recurrences and at reducing the risk for (postoperative) metastasis.
In the present study, we present in vitro PDT procedures to establish the optimal PDT settings for effective treatment of widely used OS cell lines that are used to reproduce the human disease in well established intratibial OS mouse models. The uptake of the PS mTHPC was examined with a spectrophotometer and phototoxicity was provoked with laser light excitation of mTHPC at 652 nm to induce cell death assessed with a WST-1 assay and by the counting of surviving cells. The established techniques enable us to define the optimal PDT settings for future studies in animal models. They are an easy and quick tool for the evaluation of the efficacy of PDT in vitro before an application in vivo.
22 Related JoVE Articles!
Antifouling Self-assembled Monolayers on Microelectrodes for Patterning Biomolecules
Institutions: Texas A&M University (TAMU), Texas A&M University (TAMU).
We present a procedure for forming a poly(ethylene glycol) (PEG) trimethoxysilane self-assembled monolayer (SAM) on a silicon substrate with gold microelectrodes. The PEG-SAM is formed in a single assembly step and prevents biofouling on silicon and gold surfaces. The SAM is used to coat microelectrodes patterned with standard, positive-tone lithography. Using the microtubule as an example, we apply a DC voltage to induce electrophoretic migration to the SAM-coated electrode in a reversible manner. A flow chamber is used for imaging the electrophoretic migration and microtubule patterning in situ
using epifluorescence microscopy. This method is generally applicable to biomolecule patterning, as it employs electrophoresis to immobilize target molecules and thus does not require specific molecular interactions. Further, it avoids problems encountered when attempting to pattern the SAM molecules directly using lithographic techniques. The compatibility with electron beam lithography allows this method to be used to pattern biomolecules at the nanoscale.
Biomedical Engineering, Issue 30, protein patterning, self-assembly, tubulin, kinesin, biofouling, bioNEMS, biosensor
The NeuroStar TMS Device: Conducting the FDA Approved Protocol for Treatment of Depression
Institutions: Beth Israel Deaconess Medical Center, Inc..
The Neuronetics NeuroStar Transcranial Magnetic Stimulation (TMS) System is a class II medical device that produces brief duration, pulsed magnetic fields. These rapidly alternating fields induce electrical currents within localized, targeted regions of the cortex which are associated with various physiological and functional brain changes.1,2,3
In 2007, O'Reardon et al.
, utilizing the NeuroStar device, published the results of an industry-sponsored, multisite, randomized, sham-stimulation controlled clinical trial in which 301 patients with major depression, who had previously failed to respond to at least one adequate antidepressant treatment trial, underwent either active or sham TMS over the left dorsolateral prefrontal cortex (DLPFC). The patients, who were medication-free at the time of the study, received TMS five times per week over 4-6 weeks.4
The results demonstrated that a sub-population of patients (those who were relatively less resistant to medication, having failed not more than two good pharmacologic trials) showed a statistically significant improvement on the Montgomery-Asberg Depression Scale (MADRS), the Hamilton Depression Rating Scale (HAMD), and various other outcome measures. In October 2008, supported by these and other similar results5,6,7
, Neuronetics obtained the first and only Food and Drug Administration (FDA) approval for the clinical treatment of a specific form of medication-refractory depression using a TMS Therapy device (FDA approval K061053).
In this paper, we will explore the specified FDA approved NeuroStar depression treatment protocol (to be administered only under prescription and by a licensed medical profession in either an in- or outpatient setting).
Neuroscience, Issue 45, Transcranial Magnetic Stimulation, Depression, Neuronetics, NeuroStar, FDA Approved
The Spared Nerve Injury (SNI) Model of Induced Mechanical Allodynia in Mice
Institutions: Aarhus University, University of Copenhagen.
Peripheral neuropathic pain is a severe chronic pain condition which may result from trauma to sensory nerves in the peripheral nervous system. The spared nerve injury (SNI) model induces symptoms of neuropathic pain such as mechanical allodynia i.e. pain due to tactile stimuli that do not normally provoke a painful response .
The SNI mouse model involves ligation of two of the three branches of the sciatic nerve (the tibial nerve and the common peroneal nerve), while the sural nerve is left intact . The lesion results in marked hypersensitivity in the lateral area of the paw, which is innervated by the spared sural nerve. The non-operated side of the mouse can be used as a control. The advantages of the SNI model are the robustness of the response and that it doesn’t require expert microsurgical skills.
The threshold for mechanical pain response is determined by testing with von Frey filaments of increasing bending force, which are repetitively pressed against the lateral area of the paw , . A positive pain reaction is defined as sudden paw withdrawal, flinching and/or paw licking induced by the filament. A positive response in three out of five repetitive stimuli is defined as the pain threshold.
As demonstrated in the video protocol, C57BL/6 mice experience profound allodynia as early as the day following surgery and maintain this for several weeks.
Neuroscience, Issue 54, Sciatic, Injury, PNS, Mechanical allodynia, Neuropathic pain, von Frey
A Novel Rescue Technique for Difficult Intubation and Difficult Ventilation
Institutions: Children’s Hospital of Michigan, St. Jude Children’s Research Hospital.
We describe a novel non surgical technique to maintain oxygenation and ventilation in a case of difficult intubation and difficult ventilation, which works especially well with poor mask fit.
Can not intubate, can not ventilate" (CICV) is a potentially life threatening situation. In this video we present a simulation of the technique we used in a case of CICV where oxygenation and ventilation were maintained by inserting an endotracheal tube (ETT) nasally down to the level of the naso-pharynx while sealing the mouth and nares for successful positive pressure ventilation.
A 13 year old patient was taken to the operating room for incision and drainage of a neck abcess and direct laryngobronchoscopy. After preoxygenation, anesthesia was induced intravenously. Mask ventilation was found to be extremely difficult because of the swelling of the soft tissue. The face mask could not fit properly on the face due to significant facial swelling as well. A direct laryngoscopy was attempted with no visualization of the larynx. Oxygen saturation was difficult to maintain, with saturations falling to 80%. In order to oxygenate and ventilate the patient, an endotracheal tube was then inserted nasally after nasal spray with nasal decongestant and lubricant. The tube was pushed gently and blindly into the hypopharynx. The mouth and nose of the patient were sealed by hand and positive pressure ventilation was possible with 100% O2
with good oxygen saturation during that period of time. Once the patient was stable and well sedated, a rigid bronchoscope was introduced by the otolaryngologist showing extensive subglottic and epiglottic edema, and a mass effect from the abscess, contributing to the airway compromise. The airway was secured with an ETT tube by the otolaryngologist.This video will show a simulation of the technique on a patient undergoing general anesthesia for dental restorations.
Medicine, Issue 47, difficult ventilation, difficult intubation, nasal, saturation
Cortical Source Analysis of High-Density EEG Recordings in Children
Institutions: UCL Institute of Child Health, University College London.
EEG is traditionally described as a neuroimaging technique with high temporal and low spatial resolution. Recent advances in biophysical modelling and signal processing make it possible to exploit information from other imaging modalities like structural MRI that provide high spatial resolution to overcome this constraint1
. This is especially useful for investigations that require high resolution in the temporal as well as spatial domain. In addition, due to the easy application and low cost of EEG recordings, EEG is often the method of choice when working with populations, such as young children, that do not tolerate functional MRI scans well. However, in order to investigate which neural substrates are involved, anatomical information from structural MRI is still needed. Most EEG analysis packages work with standard head models that are based on adult anatomy. The accuracy of these models when used for children is limited2
, because the composition and spatial configuration of head tissues changes dramatically over development3
In the present paper, we provide an overview of our recent work in utilizing head models based on individual structural MRI scans or age specific head models to reconstruct the cortical generators of high density EEG. This article describes how EEG recordings are acquired, processed, and analyzed with pediatric populations at the London Baby Lab, including laboratory setup, task design, EEG preprocessing, MRI processing, and EEG channel level and source analysis.
Behavior, Issue 88, EEG, electroencephalogram, development, source analysis, pediatric, minimum-norm estimation, cognitive neuroscience, event-related potentials
Portable Intermodal Preferential Looking (IPL): Investigating Language Comprehension in Typically Developing Toddlers and Young Children with Autism
Institutions: University of Connecticut.
One of the defining characteristics of autism spectrum disorder (ASD) is difficulty with language and communication.1
Children with ASD's onset of speaking is usually delayed, and many children with ASD consistently produce language less frequently and of lower lexical and grammatical complexity than their typically developing (TD) peers.6,8,12,23
However, children with ASD also exhibit a significant social deficit, and researchers and clinicians continue to debate the extent to which the deficits in social interaction account for or contribute to the deficits in language production.5,14,19,25
Standardized assessments of language in children with ASD usually do include a comprehension component; however, many such comprehension tasks assess just one aspect of language (e.g.
or include a significant motor component (e.g.
, pointing, act-out), and/or require children to deliberately choose between a number of alternatives. These last two behaviors are known to also be challenging to children with ASD.7,12,13,16
We present a method which can assess the language comprehension of young typically developing children (9-36 months) and children with autism.2,4,9,11,22
This method, Portable Intermodal Preferential Looking (P-IPL), projects side-by-side video images from a laptop onto a portable screen. The video images are paired first with a 'baseline' (nondirecting) audio, and then presented again paired with a 'test' linguistic audio that matches only one of the video images. Children's eye movements while watching the video are filmed and later coded. Children who understand the linguistic audio will look more quickly to, and longer at, the video that matches the linguistic audio.2,4,11,18,22,26
This paradigm includes a number of components that have recently been miniaturized (projector, camcorder, digitizer) to enable portability and easy setup in children's homes. This is a crucial point for assessing young children with ASD, who are frequently uncomfortable in new (e.g.
, laboratory) settings. Videos can be created to assess a wide range of specific components of linguistic knowledge, such as Subject-Verb-Object word order, wh-questions, and tense/aspect suffixes on verbs; videos can also assess principles of word learning such as a noun bias, a shape bias, and syntactic bootstrapping.10,14,17,21,24
Videos include characters and speech that are visually and acoustically salient and well tolerated by children with ASD.
Medicine, Issue 70, Neuroscience, Psychology, Behavior, Intermodal preferential looking, language comprehension, children with autism, child development, autism
An Investigation of the Effects of Sports-related Concussion in Youth Using Functional Magnetic Resonance Imaging and the Head Impact Telemetry System
Institutions: University of Toronto, University of Toronto, University of Toronto, Bloorview Kids Rehab, Toronto Rehab, Sunnybrook Health Sciences Centre, University of Toronto.
One of the most commonly reported injuries in children who participate in sports is concussion or mild traumatic brain injury (mTBI)1
. Children and youth involved in organized sports such as competitive hockey are nearly six times more likely to suffer a severe concussion compared to children involved in other leisure physical activities2
. While the most common cognitive sequelae of mTBI appear similar for children and adults, the recovery profile and breadth of consequences in children remains largely unknown2
, as does the influence of pre-injury characteristics (e.g. gender) and injury details (e.g. magnitude and direction of impact) on long-term outcomes. Competitive sports, such as hockey, allow the rare opportunity to utilize a pre-post design to obtain pre-injury data before concussion occurs on youth characteristics and functioning and to relate this to outcome following injury. Our primary goals are to refine pediatric concussion diagnosis and management based on research evidence that is specific to children and youth. To do this we use new, multi-modal and integrative approaches that will:
1.Evaluate the immediate effects of head trauma in youth
2.Monitor the resolution of post-concussion symptoms (PCS) and cognitive performance during recovery
3.Utilize new methods to verify brain injury and recovery
To achieve our goals, we have implemented the Head Impact Telemetry (HIT) System. (Simbex; Lebanon, NH, USA). This system equips commercially available Easton S9 hockey helmets (Easton-Bell Sports; Van Nuys, CA, USA) with single-axis accelerometers designed to measure real-time head accelerations during contact sport participation 3 - 5
. By using telemetric technology, the magnitude of acceleration and location of all head impacts during sport participation can be objectively detected and recorded. We also use functional magnetic resonance imaging (fMRI) to localize and assess changes in neural activity specifically in the medial temporal and frontal lobes during the performance of cognitive tasks, since those are the cerebral regions most sensitive to concussive head injury 6
. Finally, we are acquiring structural imaging data sensitive to damage in brain white matter.
Medicine, Issue 47, Mild traumatic brain injury, concussion, fMRI, youth, Head Impact Telemetry System
Development of an IFN-γ ELISpot Assay to Assess Varicella-Zoster Virus-specific Cell-mediated Immunity Following Umbilical Cord Blood Transplantation
Institutions: Université de Montréal, Université de Montréal, Université de Montréal.
Varicella zoster virus (VZV) is a significant cause of morbidity and mortality following umbilical cord blood transplantation (UCBT). For this reason, antiherpetic prophylaxis is administrated systematically to pediatric UCBT recipients to prevent complications associated with VZV infection, but there is no strong, evidence based consensus that defines its optimal duration. Because T cell mediated immunity is responsible for the control of VZV infection, assessing the reconstitution of VZV specific T cell responses following UCBT could provide indications as to whether prophylaxis should be maintained or can be discontinued. To this end, a VZV specific gamma interferon (IFN-γ) enzyme-linked immunospot (ELISpot) assay was developed to characterize IFN-γ production by T lymphocytes in response to in vitro
stimulation with irradiated live attenuated VZV vaccine. This assay provides a rapid, reproducible and sensitive measurement of VZV specific cell mediated immunity suitable for monitoring the reconstitution of VZV specific immunity in a clinical setting and assessing immune responsiveness to VZV antigens.
Immunology, Issue 89, Varicella zoster virus, cell-mediated immunity, T cells, interferon gamma, ELISpot, umbilical cord blood transplantation
A Practical Guide to Phylogenetics for Nonexperts
Institutions: The George Washington University.
Many researchers, across incredibly diverse foci, are applying phylogenetics to their research question(s). However, many researchers are new to this topic and so it presents inherent problems. Here we compile a practical introduction to phylogenetics for nonexperts. We outline in a step-by-step manner, a pipeline for generating reliable phylogenies from gene sequence datasets. We begin with a user-guide for similarity search tools via online interfaces as well as local executables. Next, we explore programs for generating multiple sequence alignments followed by protocols for using software to determine best-fit models of evolution. We then outline protocols for reconstructing phylogenetic relationships via maximum likelihood and Bayesian criteria and finally describe tools for visualizing phylogenetic trees. While this is not by any means an exhaustive description of phylogenetic approaches, it does provide the reader with practical starting information on key software applications commonly utilized by phylogeneticists. The vision for this article would be that it could serve as a practical training tool for researchers embarking on phylogenetic studies and also serve as an educational resource that could be incorporated into a classroom or teaching-lab.
Basic Protocol, Issue 84, phylogenetics, multiple sequence alignments, phylogenetic tree, BLAST executables, basic local alignment search tool, Bayesian models
Surgical Management of Meatal Stenosis with Meatoplasty
Institutions: Johns Hopkins School of Medicine.
Meatal stenosis is a common urologic complication after circumcision. Children present to their primary care physicians with complaints of deviated urinary stream, difficult-to-aim, painful urination, and urinary frequency. Clinical exam reveals a pinpoint meatus and if the child is asked to urinate, he will usually have an upward, thin, occasionally forceful urinary stream with incomplete bladder emptying. The mainstay of management is meatoplasty (reconstruction of the distal urethra /meatus). This educational video will demonstrate how this is performed.
Medicine, Issue 45, Urinary obstruction, pediatric urology, deviated urinary stream, meatal stenosis, operative repair, meatotomy, meatoplasty
Simultaneous Quantification of T-Cell Receptor Excision Circles (TRECs) and K-Deleting Recombination Excision Circles (KRECs) by Real-time PCR
Institutions: Spedali Civili di Brescia.
T-cell receptor excision circles (TRECs) and K-deleting recombination excision circles (KRECs) are circularized DNA elements formed during recombination process that creates T- and B-cell receptors. Because TRECs and KRECs are unable to replicate, they are diluted after each cell division, and therefore persist in the cell. Their quantity in peripheral blood can be considered as an estimation of thymic and bone marrow output. By combining well established and commonly used TREC assay with a modified version of KREC assay, we have developed a duplex quantitative real-time PCR that allows quantification of both newly-produced T and B lymphocytes in a single assay. The number of TRECs and KRECs are obtained using a standard curve prepared by serially diluting TREC and KREC signal joints cloned in a bacterial plasmid, together with a fragment of T-cell receptor alpha constant gene that serves as reference gene. Results are reported as number of TRECs and KRECs/106
cells or per ml of blood. The quantification of these DNA fragments have been proven useful for monitoring immune reconstitution following bone marrow transplantation in both children and adults, for improved characterization of immune deficiencies, or for better understanding of certain immunomodulating drug activity.
Immunology, Issue 94, B lymphocytes, primary immunodeficiency, real-time PCR, immune recovery, T-cell homeostasis, T lymphocytes, thymic output, bone marrow output
Intraoperative Detection of Subtle Endometriosis: A Novel Paradigm for Detection and Treatment of Pelvic Pain Associated with the Loss of Peritoneal Integrity
Institutions: Greenville Hospital System, Duke University Health System, Duke University .
Endometriosis is a common disease affecting 40 to 70% of reproductive-aged women with chronic pelvic pain (CPP) and/or infertility. The purpose of this study was to demonstrate the use of a blue dye (methylene blue) to stain peritoneal surfaces during laparoscopy (L/S) to detect the loss of peritoneal integrity in patients with pelvic pain and suspected endometriosis. Forty women with CPP and 5 women without pain were evaluated in this pilot study. During L/S, concentrated dye was sprayed onto peritoneal surfaces, then aspirated and rinsed with Lactated Ringers solution. Areas of localized dye uptake were evaluated for the presence of visible endometriotic lesions. Areas of intense peritoneal staining were resected and some fixed in 2.5% buffered gluteraldehyde and examined by scanning (SEM) electron microscopy. Blue dye uptake was more common in women with endometriosis and chronic pelvic pain than controls (85% vs. 40%). Resection of the blue stained areas revealed endometriosis by SEM and loss of peritoneal cell-cell contact compared to normal, non-staining peritoneum. Affected peritoneum was associated with visible endometriotic implants in most but not all patients. Subjective pain relief was reported in 80% of subjects. Based on scanning electron microscopy, we conclude that endometrial cells extend well beyond visible implants of endometriosis and appear to disrupt the underlying mesothelium. Subtle lesions of endometriosis could therefore cause pelvic pain by disruption of peritoneal integrity, allowing menstrual or ovulatory blood and associated pain factors access to underlying sensory nerves. Complete resection of affected peritoneum may provide a better long-term treatment for endometriosis and CPP. This simple technique appears to improve detection of subtle or near invisible endometriosis in women with CPP and minimal visual findings at L/S and may serve to elevate diagnostic accuracy for endometriosis at laparoscopy.
Medicine, Issue 70, Anatomy, Physiology, Endocrinology, Obstetrics, Gynecology, Surgery, endometriosis, pelvic pain, dysmenorrhea, diagnostics, laparoscopy, peritoneum, scanning electron microscopy, SEM
Computerized Dynamic Posturography for Postural Control Assessment in Patients with Intermittent Claudication
Institutions: University of Sydney, University of Hull, Hull and East Yorkshire Hospitals, Addenbrookes Hospital.
Computerized dynamic posturography with the EquiTest is an objective technique for measuring postural strategies under challenging static and dynamic conditions. As part of a diagnostic assessment, the early detection of postural deficits is important so that appropriate and targeted interventions can be prescribed. The Sensory Organization Test (SOT) on the EquiTest determines an individual's use of the sensory systems (somatosensory, visual, and vestibular) that are responsible for postural control. Somatosensory and visual input are altered by the calibrated sway-referenced support surface and visual surround, which move in the anterior-posterior direction in response to the individual's postural sway. This creates a conflicting sensory experience. The Motor Control Test (MCT) challenges postural control by creating unexpected postural disturbances in the form of backwards and forwards translations. The translations are graded in magnitude and the time to recover from the perturbation is computed.
Intermittent claudication, the most common symptom of peripheral arterial disease, is characterized by a cramping pain in the lower limbs and caused by muscle ischemia secondary to reduced blood flow to working muscles during physical exertion. Claudicants often display poor balance, making them susceptible to falls and activity avoidance. The Ankle Brachial Pressure Index (ABPI) is a noninvasive method for indicating the presence of peripheral arterial disease and intermittent claudication, a common symptom in the lower extremities. ABPI is measured as the highest systolic pressure from either the dorsalis pedis or posterior tibial artery divided by the highest brachial artery systolic pressure from either arm. This paper will focus on the use of computerized dynamic posturography in the assessment of balance in claudicants.
Medicine, Issue 82, Posture, Computerized dynamic posturography, Ankle brachial pressure index, Peripheral arterial disease, Intermittent claudication, Balance, Posture, EquiTest, Sensory Organization Test, Motor Control Test
Collection, Isolation, and Flow Cytometric Analysis of Human Endocervical Samples
Institutions: University of Manitoba, University of Manitoba.
Despite the public health importance of mucosal pathogens (including HIV), relatively little is known about mucosal immunity, particularly at the female genital tract (FGT). Because heterosexual transmission now represents the dominant mechanism of HIV transmission, and given the continual spread of sexually transmitted infections (STIs), it is critical to understand the interplay between host and pathogen at the genital mucosa. The substantial gaps in knowledge around FGT immunity are partially due to the difficulty in successfully collecting and processing mucosal samples. In order to facilitate studies with sufficient sample size, collection techniques must be minimally invasive and efficient. To this end, a protocol for the collection of cervical cytobrush samples and subsequent isolation of cervical mononuclear cells (CMC) has been optimized. Using ex vivo
flow cytometry-based immunophenotyping, it is possible to accurately and reliably quantify CMC lymphocyte/monocyte population frequencies and phenotypes. This technique can be coupled with the collection of cervical-vaginal lavage (CVL), which contains soluble immune mediators including cytokines, chemokines and anti-proteases, all of which can be used to determine the anti- or pro-inflammatory environment in the vagina.
Medicine, Issue 89, mucosal, immunology, FGT, lavage, cervical, CMC
Measuring Attentional Biases for Threat in Children and Adults
Institutions: Rutgers University.
Investigators have long been interested in the human propensity for the rapid detection of threatening stimuli. However, until recently, research in this domain has focused almost exclusively on adult participants, completely ignoring the topic of threat detection over the course of development. One of the biggest reasons for the lack of developmental work in this area is likely the absence of a reliable paradigm that can measure perceptual biases for threat in children. To address this issue, we recently designed a modified visual search paradigm similar to the standard adult paradigm that is appropriate for studying threat detection in preschool-aged participants. Here we describe this new procedure. In the general paradigm, we present participants with matrices of color photographs, and ask them to find and touch a target on the screen. Latency to touch the target is recorded. Using a touch-screen monitor makes the procedure simple and easy, allowing us to collect data in participants ranging from 3 years of age to adults. Thus far, the paradigm has consistently shown that both adults and children detect threatening stimuli (e.g.,
snakes, spiders, angry/fearful faces) more quickly than neutral stimuli (e.g.,
flowers, mushrooms, happy/neutral faces). Altogether, this procedure provides an important new tool for researchers interested in studying the development of attentional biases for threat.
Behavior, Issue 92, Detection, threat, attention, attentional bias, anxiety, visual search
The Multiple Sclerosis Performance Test (MSPT): An iPad-Based Disability Assessment Tool
Institutions: Cleveland Clinic Foundation, Cleveland Clinic Foundation, Cleveland Clinic Foundation, Cleveland Clinic Foundation.
Precise measurement of neurological and neuropsychological impairment and disability in multiple sclerosis is challenging. We report a new test, the Multiple Sclerosis Performance Test (MSPT), which represents a new approach to quantifying MS related disability. The MSPT takes advantage of advances in computer technology, information technology, biomechanics, and clinical measurement science. The resulting MSPT represents a computer-based platform for precise, valid measurement of MS severity. Based on, but extending the Multiple Sclerosis Functional Composite (MSFC), the MSPT provides precise, quantitative data on walking speed, balance, manual dexterity, visual function, and cognitive processing speed. The MSPT was tested by 51 MS patients and 49 healthy controls (HC). MSPT scores were highly reproducible, correlated strongly with technician-administered test scores, discriminated MS from HC and severe from mild MS, and correlated with patient reported outcomes. Measures of reliability, sensitivity, and clinical meaning for MSPT scores were favorable compared with technician-based testing. The MSPT is a potentially transformative approach for collecting MS disability outcome data for patient care and research. Because the testing is computer-based, test performance can be analyzed in traditional or novel ways and data can be directly entered into research or clinical databases. The MSPT could be widely disseminated to clinicians in practice settings who are not connected to clinical trial performance sites or who are practicing in rural settings, drastically improving access to clinical trials for clinicians and patients. The MSPT could be adapted to out of clinic settings, like the patient’s home, thereby providing more meaningful real world data. The MSPT represents a new paradigm for neuroperformance testing. This method could have the same transformative effect on clinical care and research in MS as standardized computer-adapted testing has had in the education field, with clear potential to accelerate progress in clinical care and research.
Medicine, Issue 88, Multiple Sclerosis, Multiple Sclerosis Functional Composite, computer-based testing, 25-foot walk test, 9-hole peg test, Symbol Digit Modalities Test, Low Contrast Visual Acuity, Clinical Outcome Measure
Community-based Adapted Tango Dancing for Individuals with Parkinson's Disease and Older Adults
Institutions: Emory University School of Medicine, Brigham and Woman‘s Hospital and Massachusetts General Hospital.
Adapted tango dancing improves mobility and balance in older adults and additional populations with balance impairments. It is composed of very simple step elements. Adapted tango involves movement initiation and cessation, multi-directional perturbations, varied speeds and rhythms. Focus on foot placement, whole body coordination, and attention to partner, path of movement, and aesthetics likely underlie adapted tango’s demonstrated efficacy for improving mobility and balance. In this paper, we describe the methodology to disseminate the adapted tango teaching methods to dance instructor trainees and to implement the adapted tango by the trainees in the community for older adults and individuals with Parkinson’s Disease (PD). Efficacy in improving mobility (measured with the Timed Up and Go, Tandem stance, Berg Balance Scale, Gait Speed and 30 sec chair stand), safety and fidelity of the program is maximized through targeted instructor and volunteer training and a structured detailed syllabus outlining class practices and progression.
Behavior, Issue 94, Dance, tango, balance, pedagogy, dissemination, exercise, older adults, Parkinson's Disease, mobility impairments, falls
Analysis of Nephron Composition and Function in the Adult Zebrafish Kidney
Institutions: University of Notre Dame.
The zebrafish model has emerged as a relevant system to study kidney development, regeneration and disease. Both the embryonic and adult zebrafish kidneys are composed of functional units known as nephrons, which are highly conserved with other vertebrates, including mammals. Research in zebrafish has recently demonstrated that two distinctive phenomena transpire after adult nephrons incur damage: first, there is robust regeneration within existing nephrons that replaces the destroyed tubule epithelial cells; second, entirely new nephrons are produced from renal progenitors in a process known as neonephrogenesis. In contrast, humans and other mammals seem to have only a limited ability for nephron epithelial regeneration. To date, the mechanisms responsible for these kidney regeneration phenomena remain poorly understood. Since adult zebrafish kidneys undergo both nephron epithelial regeneration and neonephrogenesis, they provide an outstanding experimental paradigm to study these events. Further, there is a wide range of genetic and pharmacological tools available in the zebrafish model that can be used to delineate the cellular and molecular mechanisms that regulate renal regeneration. One essential aspect of such research is the evaluation of nephron structure and function. This protocol describes a set of labeling techniques that can be used to gauge renal composition and test nephron functionality in the adult zebrafish kidney. Thus, these methods are widely applicable to the future phenotypic characterization of adult zebrafish kidney injury paradigms, which include but are not limited to, nephrotoxicant exposure regimes or genetic methods of targeted cell death such as the nitroreductase mediated cell ablation technique. Further, these methods could be used to study genetic perturbations in adult kidney formation and could also be applied to assess renal status during chronic disease modeling.
Cellular Biology, Issue 90,
zebrafish; kidney; nephron; nephrology; renal; regeneration; proximal tubule; distal tubule; segment; mesonephros; physiology; acute kidney injury (AKI)
An Affordable HIV-1 Drug Resistance Monitoring Method for Resource Limited Settings
Institutions: University of KwaZulu-Natal, Durban, South Africa, Jembi Health Systems, University of Amsterdam, Stanford Medical School.
HIV-1 drug resistance has the potential to seriously compromise the effectiveness and impact of antiretroviral therapy (ART). As ART programs in sub-Saharan Africa continue to expand, individuals on ART should be closely monitored for the emergence of drug resistance. Surveillance of transmitted drug resistance to track transmission of viral strains already resistant to ART is also critical. Unfortunately, drug resistance testing is still not readily accessible in resource limited settings, because genotyping is expensive and requires sophisticated laboratory and data management infrastructure. An open access genotypic drug resistance monitoring method to manage individuals and assess transmitted drug resistance is described. The method uses free open source software for the interpretation of drug resistance patterns and the generation of individual patient reports. The genotyping protocol has an amplification rate of greater than 95% for plasma samples with a viral load >1,000 HIV-1 RNA copies/ml. The sensitivity decreases significantly for viral loads <1,000 HIV-1 RNA copies/ml. The method described here was validated against a method of HIV-1 drug resistance testing approved by the United States Food and Drug Administration (FDA), the Viroseq genotyping method. Limitations of the method described here include the fact that it is not automated and that it also failed to amplify the circulating recombinant form CRF02_AG from a validation panel of samples, although it amplified subtypes A and B from the same panel.
Medicine, Issue 85, Biomedical Technology, HIV-1, HIV Infections, Viremia, Nucleic Acids, genetics, antiretroviral therapy, drug resistance, genotyping, affordable
Functional Near Infrared Spectroscopy of the Sensory and Motor Brain Regions with Simultaneous Kinematic and EMG Monitoring During Motor Tasks
Institutions: National Institutes of Health.
There are several advantages that functional near-infrared spectroscopy (fNIRS) presents in the study of the neural control of human movement. It is relatively flexible with respect to participant positioning and allows for some head movements during tasks. Additionally, it is inexpensive, light weight, and portable, with very few contraindications to its use. This presents a unique opportunity to study functional brain activity during motor tasks in individuals who are typically developing, as well as those with movement disorders, such as cerebral palsy. An additional consideration when studying movement disorders, however, is the quality of actual movements performed and the potential for additional, unintended movements. Therefore, concurrent monitoring of both blood flow changes in the brain and actual movements of the body during testing is required for appropriate interpretation of fNIRS results. Here, we show a protocol for the combination of fNIRS with muscle and kinematic monitoring during motor tasks. We explore gait, a unilateral multi-joint movement (cycling), and two unilateral single-joint movements (isolated ankle dorsiflexion, and isolated hand squeezing). The techniques presented can be useful in studying both typical and atypical motor control, and can be modified to investigate a broad range of tasks and scientific questions.
Behavior, Issue 94, functional near infrared spectroscopy, fNIRS, brain activity, gait, motor tasks, cerebral palsy, coordination
Yeast Colony Embedding Method
Institutions: University of Missouri - Kansas City.
Patterning of different cell types in embryos is a key mechanism in metazoan development. Communities of microorganisms, such as colonies and biofilms also display patterns of cell types. For example, in the yeast S. cerevisiae
, sporulated cells and pseudohyphal cells are not uniformly distributed in colonies. The functional importance of patterning and the molecular mechanisms that underlie these patterns are still poorly understood.
One challenge with respect to investigating patterns of cell types in fungal colonies is that unlike metazoan tissue, cells in colonies are relatively weakly attached to one another. In particular, fungal colonies do not contain the same extensive level of extracellular matrix found in most tissues . Here we report on a method for embedding and sectioning yeast colonies that reveals the interior patterns of cell types in these colonies. The method can be used to prepare thick sections (0.5 μ) useful for light microscopy and thin sections (0.1 μ) suitable for transmission electron microscopy. Asci and pseudohyphal cells can easily be distinguished from ovoid yeast cells by light microscopy , while the interior structure of these cells can be visualized by EM.
The method is based on surrounding colonies with agar, infiltrating them with Spurr's medium, and then sectioning. Colonies with a diameter in the range of 1-2 mm are suitable for this protocol. In addition to visualizing the interior of colonies, the method allows visualization of the region of the colony that invades the underlying agar.
Cellular Biology, Issue 49, yeast, Saccharomyces cerevisiae, colony, embedding, sporulation, pattern formation, organization
Demonstration of Cutaneous Allodynia in Association with Chronic Pelvic Pain
Institutions: University of Calgary.
Pelvic pain is a common condition that is associated with dysmenorrhea and endometriosis. In some women the severe episodes of cyclic pain change and the resultant pain becomes continuous and this condition becomes known as Chronic Pelvic Pain. This state can be present even after the appropriate medical or surgical therapy has been instituted. It can be associated with pain and tenderness in the muscles of the abdomen wall and intra-pelvic muscles leading to severe dyspareunia. Additional symptoms of irritable bowel and interstitial cystitis are common. A common sign of the development of this state is the emergence of cutaneous allodynia
which emerges from the so-called viscero-somatic reflex. A simple bedside test for the presence of cutaneous allodynia
is presented that does not require excessive time or special equipment. This test builds on previous work associated with changes in sensation related to gall bladder function and the viscera-somatic reflex(1;2).
The test is undertaken with the subject s permission after an explanation of how the test will be performed. Allodynia
refers to a condition in which a stimulus that is not normally painful is interpreted by the subject as painful. In this instance the light touch associated with a cotton-tipped applicator would not be expected to be painful. A positive test is however noted by the woman as suddenly painful or suddenly sharp. The patterns of this sensation are usually in a discrete pattern of a dermatome of the nerves that innervate the pelvis.
The underlying pathology is now interpreted as evidence of neuroplasticity as a consequence of severe and repeating pain with changes in the functions of the dorsal horns of the spinal cord that results in altered function of visceral tissues and resultant somatic symptoms(3).
The importance of recognizing the condition lies in an awareness that this process may present coincidentally with the initiating condition or after it has been treated. It also permits the clinician to evaluate the situation from the perspective that alternative explanations for the pain may be present that may not require additional surgery.
Medicine, Issue 28, Chronic pelvic pain, cutaneous allodynia, trigger points, dysmenorrhea, endometriosis, dyspareunia