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Guest Editors: Michael Overduin, Alastair Barr
The study of protein-protein interactions is key to understanding the network of molecular interactions underlying biological effects, and defining how individual proteins perform their functional role within a cell. Detailed understanding of t… Show Morehese interactions often provides insight into aberrant signaling in disease. This collection brings together the wide range of techniques that have been developed to study interactions using in vitro, cell-based and in vivo approaches. Each technique has its own strengths, for example enabling systematic interrogation of the complete ‘interactome’, providing detailed atomic level information about a single interaction or insight into the sub-cellular localization of interacting proteins. Some techniques are easily adaptable to high-throughput screening of libraries or pulling out novel interacting partners while others have a more limited capacity. Readouts such as microscopy and mass-spectrometry are employed in protein-protein interaction techniques and some provide quantitative information on the affinity of an interaction.
Examples of methods to be included in this collection are: co-immunoprecipitation and antibody interference; pull-down assays using fusion proteins or modular protein domains; proximity-dependent labelling techniques (e.g. BioID); far western and receptor affinity probes; affinity purification coupled with mass-spectrometry; two-hybrid technologies (yeast and mammalian); fluorescence microscopy (e.g. immunofluoresence, FRET, super-resolution microscopy); protein complementation assays (e.g. BiFC); biophysical approaches (X-ray crystallography, NMR, AUC, SPR) and microarray based techniques (i.e. tissue and protein arrays). Show Less
Guest Editor: Katherine Stavropoulos
The neural reward system plays a key role in human learning, motivation, pleasure, and the ability to change one’s behavior to maximize desired outcomes. Given the wide range of behaviors affected by the reward system, it is unsurprising that a… Show Moreberrant functioning of this system is implicated in multiple psychiatric disorders.
Blunted reward anticipation and responsivity has been observed in individuals with major depressive disorder (MDD), and individuals with autism spectrum disorder (ASD) show attenuated reward-related brain activity. Conversely, individuals with bipolar disorder (BP) evidence hyper-activity of the reward system. Deficiencies in the reward system appear important for both the development and maintenance of drug addiction and pathological gambling. The reward system affects schizophrenia (SCZ)—with increased salience for irrelevant and meaningless stimuli, and decreased salience for relevant stimuli hypothesized as driving factors of positive and negative symptoms, respectively. Individuals with attention-deficit/hyperactivity disorder (ADHD) tend to discount the value of delayed rewards and over-value immediate rewards.
Given the importance of the reward system for behavior and its purported role in psychological disorders, neuroscience studies of typical and atypical reward functioning have the potential to improve our understanding of behavior and inform treatments. Considering the broad reach of the reward system, it is critical to highlight findings from a variety of neuroscience modalities—including functional magnetic resonance imaging (fMRI), electroencephalography (EEG), and magnetoencephalography (MEG). In the current collection, we present neuroscience studies that investigate the reward system both in neurotypical individuals and those diagnosed with MDD, ASD, BP, addiction, SCZ, and ADHD.
Guest Editor: Milo Lin
Complexity is a key obstacle in understanding and engineering biological systems.The details of this complexity rapidly accumulate due to advances in sequencing, imaging, and multiplexing. This leads to larger and more precise datasets. Longstanding … Show Moreand widely-applicable computational approaches, such as mutual information analysis, principal components analysis, and machine learning are standard tools currently used for data analysis. However, these standard tools are limited in matching a methodology to a data structure or biological process and typically do not lead to a mechanistic or causative understanding. Therefore, it is increasingly important to apply problem-specific yet unbiased methods to extract robust features from biological data and to fit the data within a predictive conceptual framework. This collection brings a wide range of computational and theoretical methods to extract underlying patterns and organizational principles in molecular biology that would otherwise be obscured by complexity. While these methods draw on insights from multiple disciplines, , they all aim to simplify complex phenomena by finding sparse and modular representations that take into account the constraints of molecular biology.
Guest Editor: Pavan Agrawal
Rapid advances in the field of neuroepigenetics and neurotranscriptomics have transformed our understanding of gene regulation in the nervous system. These discoveries have put to rest the mechanistic dichotomy between nature and nurture, effec… Show Moretively bridging the two concepts. Epigenetic mechanisms such as DNA methylation, post-translational histone modifications, non-coding RNA, and ATP dependent chromatin remodeling play a pivotal role in mediating environmental and psychological stress. These epigenetic changes alter neuronal transcription and lead to changes in animal behavior. Many of these epigenetic changes can last from several hours to several generations.
There is considerable diversity in the nervous system and emerging techniques make it possible to investigate transcriptional and epigenetic diversity in neurons and specific types of cells. Purification of genetically tagged specific cell types from the brain followed by genome-wide methods such as ChIP-seq and RNA-seq allows for comprehensive profiling of neuronal subsets. Furthermore, advances in molecular and imaging tools enable visualization of epigenetic modifications and transcriptional changes in both individual neurons and the entire brain. Emerging techniques in neuroepigenetic editing enable spatial changes in chromatin structure, allowing for causal connections between epigenetic modifications and transcriptional and behavioral outcomes. This collection will cover the latest advances in the field of neuroepigenetics and neurotranscriptomics that enable the study of the nervous system’s function from individual molecules to fully-developed animals across several model and non-model organisms.
Guest Editor: Matthew Boyko
Animal models are an important tool to evaluate the pathophysiology of cerebral injury and to advance the development of novel treatment options for improving neurological outcomes. Appropriate animal models are valued for their reproducibility and s… Show Moretandardization, and their ability to provide direct access to brain tissue. Historically, the two most common techniques for studying neurological damage have been focal insult through intramural middle cerebral artery occlusion and inducing traumatic brain injury by external force using a weight drop. Following cerebral insult, measured outcomes in determining the extent of neurological injury often include a behavioral neurological severity score or histologic examination of cerebral edema, infarct zone, and/or blood-brain barrier permeability. Over the last decade, however, a variety of innovative methods have been developed. The objective of this collection is to present new, state-of-the-art techniques available to induce cerebral insult and to investigate subsequent neurological damage. We believe these novel approaches provide more practical, accurate, cost-efficient, and ethically-favorable options to measure neurological injury, and may promote future studies to better understand and treat its associated consequences. Show Less
Guest Editor: Sergio Souza
Molecular imaging has the ability to non-invasively imaging functional aspects of different organs in the body and has been used as a bridge to translate preclinical results to the clinical scenario, playing a critical role in diagnosis and therapeut… Show Moreics. The evolving molecular functional imaging using specific molecular targets has great potential in translational research. In the last decades in vivo bioimaging techniques has revolutionized non-invasive evaluation.
For the non-invasive image of small animals, different modalities are available, all with inherent advantages and limitations. The choice of imaging mode depends on the parameter of interest under consideration, costs and measurement time per animal. Among the main modalities for the evaluation of functional aspects, we have nuclear medicine equipment such as positron emission tomography (PET) and single-photon emission computed tomography (SPECT). In comparison to molecules of other modalities like dyes or nanoparticles, conjugation of a radioisotope does not affect the properties of a biomolecule, and it is possible to make absolute quantification of different functional processes.
The aims of this special issue are to attract researchers in the field of pre-clinical development of new radiopharmaceuticals and those who have already translated it to a phase 1 clinical trial to show their results and share their achievements and difficulties.
Potential topics include but are not limited to the following:
New SPECT and PET radiopharmaceuticals for Cancer
New SPECT and PET radiopharmaceuticals for Cardiology
New SPECT and PET radiopharmaceuticals for Neurology
Translational results of pre-clinical studies
Guest Editor: Azad Eshghi
Knowledge of biochemical pathways is part of the foundation of basic and applied research in the field of biochemistry. Mass spectrometers are indispensable tools for the study of biomolecules due to the afforded specificity, multiplexing and quantit… Show Moreative capabilities for the analysis of a broad range of analytes, encompassing small molecules such as metabolites, and multicomponent macro-molecules, such as protein complexes. High mass accuracy and high resolving power are the two main factors that make mass spectrometers inherently selective, and when combined with an internal standard spike in, absolute specificity is achievable. Additionally, mass spectrometers can be utilized to scan mass to charge ratios on the order of milliseconds which enhances multiplexing. When coupled with upstream separation strategies, such as liquid chromatography, hundreds to thousands of compounds can be identified and/or quantified within a matter of hours.
Analyte quantification can be performed in relative terms, for example comparing healthy to disease subjects, and/or accurate concentrations can be measured using spiked in standards. Other proteomics specific applications of mass spectrometry include enzymatic treatment of samples and subsequent identification and quantification of site-specific post-translational modifications. Lastly, protein-protein interactions and protein complexes can be characterized using cross-linking strategies and structural proteomics performed utilizing deuterium exchange methodologies. Each of the aforementioned applications requires unique operational and instrumental setups and this collection aims to facilitate the dissemination of these methodologies to other researchers.
Guest Editor: Ramiro Almeida
Neurons are the primary cells of the nervous system and are responsible for the distinctive roles of the brain. Understanding the mechanisms that regulate their differentiation, development, and function is essential to understand the nervous system … Show Moreand how it regulates other organs. Neuronal cell cultures can be prepared from various regions of the central and peripheral nervous system. Although this model has some limitations, for example, the lack of interactions with other cell types and the absence of the layered organization of the brain, neuronal cultures reproduce most of the key properties and distinct characteristics observed in vivo and are invaluable tools to study the nervous system.
Culturing nerve cells is still a challenging task, due to technical difficulties in isolating specific brain regions and the precise requirements of each neuronal population. Primary neuronal cultures are excellent models to study fundamental aspects of neuronal development and function like axonal growth, synapse formation, neurotransmission, and neuronal plasticity; to investigate intracellular trafficking and subcellular localization of endogenous proteins; to uncover the effect of disease-related mutations; to perform drug screens, disease modeling, and toxicological studies. In this collection, we present a variety of neuronal cell culture models, which are the result of the expertise of experienced researchers in each specific neuronal type. These protocols will provide detailed information about the methodology and the most relevant procedures used to cultured neuronal cells from the mammalian nervous system.
Guest Editors: Mohd. Farooq Shaikh, Ayanabha Chakraborti
Neurological disorders have a high global prevalence. However, the molecular mechanisms mediating these disorders remain poorly understood. Animal models play a significant role in better understanding the pathophysiology of a variety of diseases aff… Show Morelicting humans and also help in the identification of potential therapeutic targets. Zebrafish (Danio rerio) has recently emerged as a powerful preclinical model for a wide range of brain disorders. Zebrafish based behavioral models have been successfully employed to better understand the neural and molecular aspects of a spectrum of CNS conditions i.e., epilepsy, schizophrenia, anxiety, Alzheimer’s disease, Parkinson’s disease, and substance abuse-related disorders. Both larval and adult zebrafish has proven to be valuable for translational neuropharmacology and drug discovery research. The zebrafish genome has orthologues corresponding to an extensive number of disease-related genes in humans. Zebrafish models are particularly useful for studying genotype-phenotype and genotype-drug-phenotype relationships. The high sensitivity of zebrafish to known antipsychotic and other CNS agents provides researchers with an ideal model organism capable of identifying molecular targets for therapy and empirical testing of their hypotheses. This collection aims to bring together the experimental protocols and approaches that have been developed using zebrafish to model a variety of neurological, neuropsychiatric and neurodegenerative conditions. Such models would provide important insights as to how zebrafish models have improved CNS disease modeling and the discovery of novel drug targets and candidates. The collection would serve as a valuable resource for researchers who aim to utilize such models in their laboratories to improve understanding of complex brain disorders. Show Less
Guest Editor: Giridhar Mudduluru
Cancer is a leading cause of death worldwide. Increased lethality of cancer is due to delayed diagnosis and especially due to the metastasis of the disease to different tissues. Secondly, the tumor microenvironment is different between the primary si… Show Morete and the metastasized locations, which could change the tumor characteristics. In addition, the origin or cause of malignancies is different in each individual based on many factors like genetics, ethnicity, age, environment, food and physical and social behavior. Due to different factors that influence the origins of different cancer entities and lots of complexity in stratifying the treatment strategies, the treatment procedures are challenging and treating patients with precision medicine is necessary. To understand the complexity in molecular functions and the specific phenotypes they result in, the cancer research field has developed and effectively used different methods both in vitro and in vivo to screen patient specimens, determine the aggressiveness of the tumor, and predict drug responses.
This method collection section brings some of the important methods which are designed to demonstrate 1) cancer prognostic aspects of cancer cell lines like cell migration and invasion in vitro, 2) Chorioallantoic Membrane (CAM) assay and mouse xenograft in vivo models, 3) angiogenesis in tumor and 4) drug efficiency determination in in vivo and ex-vivo explants. This collection of methods will bring together technical expertise for cell lines, tumor, and tumor specimen’s in vitro, in vivo and ex-vivo explants, to study cancer phenotypes and drug efficacy in cancer treatment.
Guest Editor: Daniel Vogt
The proper development of the brain requires multiple developmental events that eventually establish a balanced network of excitatory and inhibitory neurons that work in tandem with glia. Disruptions in molecular and cellular events during developmen… Show Moret are hypothesized to underlie some symptoms of neurological and neuropsychiatric disorders, including schizophrenia, autism spectrum disorder and epilepsy. However, much is still unknown about how different types of neurons are generated, assume their mature identity and establish a balance of excitation/inhibition by adulthood.
This proposed collection will present methods aimed at the study of neurons during developmental stages, with a focus on molecular and cellular assays. The ability to manipulate neurons at the molecular/genetic/cellular level is a valuable toolkit that has increased relevance with the advent of large-scale genomic data that need to be validated in model systems. Overall, these methods will provide a rich resource of novel and validated approaches that will enhance the study of developmental neuroscience.
Guest Editor: Sofia Sousa
Cancer research has focused on the cancer cell, its proliferation rates, apoptosis resistance, mutation rates, sensitivity to radio and chemotherapy. However, tumor metastatic progression involves many other cellular and molecular players, the tumor … Show Moremicroenvironment (TME). In the battle against cancer, to improve the survival and quality of life of metastatic patients, the key cancer supporting players must be identified and targeted. For a tumor to leave its primary site, it must invade the surrounding extracellular matrix, enter the blood or lymphatic circulation, survive in circulation, extravasate at the metastatic site, start proliferating in this new tissue, and during the entire process evade the immune system. In many instances, cancer cells co-opt physiologic niches and processes to survive and even thrive in hostile environments. This is not addressed in 2D monotypic cancer cell cultures. This collection aims to highlight the latest techniques in TME research and promote “multi-cellular” interdisciplinary approaches in the field of metastasis. The collection will cover but is not limited to:
- 3D co-culture methods of cancer and supporting cells and matrices (culture and imaging, e.g. Single Plane Illumination Microscopy)
- tumor microenvironment metabolomic studies
- tumor vasculature imaging techniques (animal models of angiogenesis, multiphoton confocal microscopy of vascularized primary tumors and metastases)
- immune cell modulation and exploitation by cancer cells (flow cytometry methods, adoptive transfer models, intra-vital microscopy)
- circulating and disseminating tumor cell (CTC and DTC) detection methods (including methods to detect the potential of dormant DTCs to reactivate)
- humanized and patient-derived animal models of metastasis and pre-metastatic niches.
Guest Editor: Karthik Krishnamurthy
Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease that ultimately leads to fatal paralysis due to impaired neuromuscular function. At the cellular level, ALS is caused by motor neuron degeneration either intrinsically or … Show Morethrough the contribution of toxic glial cells. About 10% of ALS cases are familial (fALS), and are associated with mutations in one or another of several genes, while the remaining are sporadic (sALS). fALS and sALS are clinically indistinguishable, suggesting that the different forms of the disease converge on common pathways. The molecular mechanisms contributing to the disease are beginning to be unraveled in recent years after the discovery of mutations in various genes including TDP-43, FUS, and C9ORF72.
Various model systems (Human, Mouse, Drosophila and Zebrafish) and cell biological techniques have been utilized recently to investigate important converging pathways in ALS. These include altered neuronal excitability, nucleocytoplasmic transport defects, aberrant phase transitions of RNA binding proteins, pathological stress granule assembly/disassembly, autophagy defects, and DNA damage. In this method collection, we present novel cell biology, electrophysiology and biochemical techniques to study the above pathways in ALS. These protocols will provide a visual platform for researchers in the ALS community to adopt a unified scientific approach.
Guest Editor: Matheus de Castro Fonseca
Imaging is an increasingly important tool in both basic and applied research in biology. An array of imaging techniques provides the unprecedented capability of visualizing organs and cell architecture and function. These techniques show a great reso… Show Morelution from the level of individual molecules to the whole structure. In neuroscience, the neuronal morphology, activity, and pattern of the network are the basis for understanding brain function in health and disease. Therefore, the deep understanding of these features, such as neurons, may help to uncover the whole functioning of the brain as a unit. From X-ray microtomography to optical microscopy in vivo or in vitro, researchers have been able to depict the cellular and molecular mechanisms behind brain functioning. In addition, several 3D imaging methods allow the visualization and understanding of neural networks, disease pathways, and more in whole tissues. In the past years, 3D neuroimaging (from computed tomography, serial block-face microscopy, MRI or ultrasonography) has increased in popularity bringing the ability to collect large-scale datasets. However, a good data collection obtained from images is largely dependent on a high-quality sample. Thereby, sample preparation is a key step to the acquisition of successful results from images. JoVE is an exceptional platform to broadcast, in a didactic manner, techniques and make them accessible to more researchers.
This methods collection focuses on the state-of-the-art 3D imaging techniques used in neuroscience. It covers points from sample preparation to data analysis, in order to showcase the strategies used to depict cellular and molecular mechanisms involved in health and in pathologies of the nervous system.
Guest Editor: Herbert Schneckenburger
Optical microscopy plays an increasing role in studies of 3-dimensional cell cultures as well as whole organisms. Since the sample thickness often exceeds the depth of focus considerably, confocal methods have been developed more than 30 years ago in… Show More order to select individual focal planes and to suppress out-of-focus images. After shifting of the focal plane in the axial direction a sharp image of the whole sample can be reconstructed. The potential of confocal microscopy has also been increased by advanced methods, e.g. Airy Scan Microscopy, Stimulated Emission Depletion Microscopy (STED) and related techniques. These techniques often provide super-resolution far below the Abbe criterion. However, wide-field methods, such as Super-localization techniques, Structured Illumination Microscopy (SIM), Light Sheet Microscopy or Total Internal Reflection (TIRF) microscopy gained considerable importance. These are of importance with respect to resolution (Super-localization, SIM), 3D imaging (Light Sheet) and selective detection of cell membranes (TIRF). Due to extremely low light exposure in Light Sheet Microscopy, this skill plays an important role in long-term observation, for example, in developmental biology. Further development of these methods often in combination with spectral or temporal resolution as well as their applications in a broad field of biomedical research, diagnostics or pharmacology is the subject of this JoVE methods collection. Show Less
Guest Editor: Manoj Muthukuru
Flow cytometry combines the dynamism and the versatility for a wide range of phenotypic and functional characterization of cells and molecules.. Flow cytometry plays vital role in clinical discovery which is exemplified in the areas of immunological … Show Moreresearch, hematological analysis, and oncology.
The evolution of flow cytometry is associated with improvements in the development of optical systems and the incorporation of multiple lasers that enable multi-parametric analysis. These aspects are complemented with the advances in generating monoclonal antibodies and designing fluorochrome tags. The combination of these enhancements can offer newer advantages for investigating intracellular compartments and organelles with in-depth cellular functionality. The intersection of robotics in handling multiple samples and utilizing plate formats, in conjunction with sophisticated algorithms and software, has further enabled rapid high-throughput quantitative analysis and extended flow cytometry applications in the areas of microbiology and drug discovery.
Recent advances and integrative technologies facilitate flow cytometry with single-cell imaging capabilities that combine the robust features of flow cytometry and high-resolution microscopy. he future of flow cytometry promises clinical advancements in non-invasive diagnostics and prognosis through in vivo flow cytometry. This collection aims to bring together a wide range of flow cytometry-based techniques and applications that are employed in research methodologies and are within the scope of current and future clinical practices. Show Less
Guest Editor: Gal Almogy
Mathematical models are widely used in the study of infectious diseases, providing valuable theoretical insights into the logic of pathogen spread, as well as guiding public health strategies. Digital technologies have the potential to transform heal… Show Morethcare, and “big data” approaches already show promise in improving the diagnosis and treatment of chronic diseases such as diabetes, heart conditions, cancer, and Alzheimer’s. Yet the large amounts of health data collected by hospitals and healthcare facilities are not used as effectively in the study and management of infectious diseases.
The objective of this collection is to explore new ways to analyze local clinical data, methodologies that will help unify various local data sources to reach more general conclusions, and novel approaches that may be useful in real-life clinical settings.
Examples of types of studies to be included in this collection are: characterizing transmission networks within healthcare facilities and the community they serve, methods to combine multiple local data sources while maintaining patient privacy, computational methods to diagnose the presence of specific pathogens in the absence of specific molecular test data, using non-traditional data sources in epidemiology (e.g. GPS or cellphone location data), and developing more accessible and intuitive methodologies that could be used by the general medical community.
Guest Editor: Yi Ma
Ca2+ is a ubiquitous secondary messenger involved in numerous plant signaling transduction cascades, such as defense responses to environmental cues including abiotic and biotic stresses, gravitropism, pollen tube growth, nodulation, etc. Ligand bind… Show Moreing to a cognate receptor triggers a cytosolic Ca2+ spike required for induction of downstream signaling. The two major ion channel families that have been identified in plants to conduct Ca2+ are cyclic nucleotide gated channels (CNGC) and ionic glutamate receptors (GLR). Members in both families have been characterized to conduct Ca2+ into the cytosol. A substantial aspect of CNGC signaling in plants involves CNGC-dependent cytosolic Ca2+ spikes that then are transduced into both long-term but also (often underappreciated) instantaneous signal responses. CNGCs are activated by cyclic nucleotides, cAMP and cGMP. Calmodulin and calmodulin-like proteins regulate CNGC channel function by binding to the calmodulin binding domains in CNGC. Cyclic nucleotides are also universal secondary messengers. Although evidence has suggested the existence of cyclic nucleotide signaling in plant cells, none of the enzymes responsible for their biosynthesis and degradation have been identified. Optogenetic approaches for the determination of Ca2+ elevation in response to stimuli and techniques of studying cyclic nucleotide, CNGC, and GLR associated signaling in plant cells have been developed and advanced in the past two decades. This collection of such cutting-edge techniques will provide practical, instructive, and experiential tools for researchers in this field. It will also encourage communication and collaboration between scientists studying Ca2+ signaling in plants. Show Less
Guest Editors: Sanne Lof, Frederique Vissers, Maarten Korrel, Amer Zureikat, Safi Dokmak, Ugo Boggi, Mohammed Abu Hilal, Marc Besselink
Minimally invasive pancreatic surgery is on the rise worldwide. Minimally invasive distal pancreatectomy (MIDP) has been reported to be superior to open distal pancreatectomy (ODP) in terms of blood loss and hospital stay. The oncological safety of M… Show MoreIDP for pancreatic cancer, however, remains uncertain and randomized controlled trials are needed (Riviere et al, Cochrane Database Syst Rev 2016). Interestingly, a recent study comparing open and minimally invasive pancreatoduodenectomy (MIPD), found similar morbidity and hospital stay, but higher rates of postoperative pancreatic fistula in the MIPD group (Klompmaker et al, Ann Surg 2018). These results may be related to the steep learning curve in MIPD suggesting the need of careful patients selection, access to a high number of patients and intense training for a safe implementation of this procedure.
In this collection, laparoscopic and robot-assisted procedures (both MIDP and MIPD) performed by world-renowned pancreatic surgeons are shown. Show Less
Guest Editor: Mingyi Wang
Aging populations are on the rise worldwide.
Aging is the major risk factor for the development of cardiac arrythmia.
Age-related alterations throughout the sinoatrial node (SAN), atrioventricular node (AVN), and His… Show More of bundle (HB) and its right and left branches (RBB and LBB) cardiac conduction system (CCS) (see illustration) predispose older individuals to bradyarrhythmia, conduction disturbances, and supraventricular and ventricular tachyarrhythmias, all of which increase in incidence and prevalence with advancing age. This is known as “geriatric CCS syndrome”. Age-associated adverse remodeling of CCS is the foundation of developing this syndrome due to either an increase in susceptibility or a decrease in the pathologic threshold. Hence, the opportunity for a visual, methodological catalog of CCS is helpful for clinical and basic research in molecular, cellular, tissue, and organismal levels for diagnosis, treatment, and prevention of this syndrome in the elderly population.
In this collection, methodologies for in situ studies of nodal cell-specific events, the number of pacemaker cells, morphology, cell-cell junction, action potential and current, sinoatrial extracellular matrix and arterial remodeling as well as cardiac Purkinje fiber cellular behavior of CCS bear important implications for a complete understanding of CCS biology in young and old cardiac conditions; these techniques will provide insight into the assessment and intervention of heartbeat aging and age-related CCS geriatric syndrome.
Guest Editor: Lynette Foo
In vitro techniques have been widely used to study neurodevelopment and neurodegeneration with great success. However, frequently these in vitro cultures contain only one or two cell types. Naturally, a next step in complexity prior to in vivo … Show Morestudies in suitable animal models are ex vivo cultures to examine how the full complement of cells within the nervous system parenchyma contribute to development or disease. Ex vivo slices have been performed in multiple species including human post-mortem tissue and rodent tissue. Ex vivo cultures provide the complexity yet accessibility of the nervous system parenchyma for manipulation, imaging and quantification. They provide a useful tool for understanding not only the molecular mechanisms underlying development but also how these mechanisms are altered in disease states. The cultures are also tremendously useful for drug development to demonstrate target engagement, efficacy on a cellular and molecular level, to investigate toxicity. This collection aims to showcase both the current and latest techniques commonly employed for ex vivo studies of neural tissue. These types of assays are frequently difficult to first establish and conduct, hence JoVE is an ideal platform to make these techniques accessible to more researchers.
Guest Editor: Isabel Garay
The cerebral cortex is the seat of higher brain function and plays a key role in memory, attention, thought, perception and language. Development of this cerebral structure is a complex process in which different steps need to be tightly regulated an… Show Mored coordinated in order to obtain a mature, six-layered structure. Progenitors at the ventricular zone generate specific types of neurons at the appropriate time and in correct numbers. These neurons then migrate into the nascent cortical plate while simultaneously extending their axons and, once they arrive at their final position in the right layer, they start to elaborate the dendritic arbor and establish synaptic contacts with specific targets. Defects in cortical development have long been associated with disorders displaying anatomical abnormalities, like microcephaly, lissencephaly or periventricular heterotopia. In addition, development of the cerebral cortex is also disrupted in other neurological conditions that do not manifest with obvious anatomical changes, such as ASD, schizophrenia or cognitive impairment.
Cortical development is therefore an area of very active research, which has progressed rapidly in recent times. This collection aims to include standard and novel methods in the study of corticogenesis, with a focus on approaches to target or label progenitors and neurons for lineage tracing and functional studies of neurogenesis, fate specification and neuronal migration.
Guest Editor: Chris Shaw
The proposed edition will focus on the value of zebrafish models for exploring early stages of neurological diseases such as in ALS, Parkinson's disease and Alzheimer's disease. Such stages are largely a mystery in curre… Show Morent research. In humans, the absence of effective biomarkers means that disease processes can only be studied once the disease state has been clinically diagnosed, a point at which an enormous amount of damage has already occurred in the particular regions of the central nervous system affected in each disease. Detailed histological analysis is performed only post mortem. In silico models have not proven useful in the search for early stages and in vitro methods suffer from a lack of interneuronal complexity that characterizes the diseases. In vivo methods in conventional rodent models also have a range of caveats, not least the inability to see emerging neurodegenerative cascades in real-time. Zebrafish models allow investigators to create high throughput studies and image areas of the nervous system in real-time. Needless to say, there are also disadvantages, but in our view, these are outweighed by the distinct advantages. One of these is the ability to create 'morpholinos' expressing gene variations. Such models are rapidly expanding in scope and sophistication with imaging techinques that are on the cutting edge. The proposed edition will examine some of these studies, the techniques they use and explore the potential for therapeutic development. Show Less
Guest Editor: Chiquito Crasto
Biomedical research related to olfactory receptors (OR) and their role in olfaction came into prominence after the discovery of these receptors in the early nineteen nineties and the subsequent Nobel Prize award to the discoverers of ORs in 2004. OR … Show Moregenes constitute overwhelmingly, the largest gene families in the human other mammalian genomes. Olfactory receptors have remarkably complex interactions with odorants. A few hundred olfactory receptors can discriminate between several thousand odorants and complex odors resulting in eliciting a sensory response. Research into olfactory receptor structure and function, which comes under the umbrella of neuroscience, is stymied. Olfactory receptors are Class A G(TPbinding) Protein Receptors (GPCRs) that line the nasal passages, each receptor associated with a single olfactory neuron. These membrane proteins, as one would expect are difficult to express and consequently, functionally assess. Functional analysis of ORs is thus, significantly slowed down. No experimentally determined structure of an OR is currently available. Computational structural studies though rigorous are speculative. Because of the vastness and complexity of this OR superfamily, the research that elucidates the structural and functional aspects of these genes will help clarify and rationalize the first in a multi-step process leading to a better understanding of olfaction, educating and eliciting a greater interest in this domain of research. Additionally, these efforts will aid in serving as a models to enhance our understanding of other GPCRs and membrane proteins. This methods collection will highlight methods including computational modeling, genomics analysis and current functional assays to study olfactory receptors. Show Less
Guest Editor: Kyuwan Lee
Cancer survivors remain distinctly at risk for cardiovascular diseases. Cardiovascular toxicity induced by cancer treatment such as radiation or chemotherapy has been strongly suggested to increase cardiovascular diseases in cancer survivors. While a… Show Morettempts have been made to measure cardiovascular toxicity in clinical oncology settings, the most ideal measure to diagnose cardiovascular toxicity is still understudied. Clinical or subclinical measures of cardiovascular function during and/or after cancer treatment may include ejection fraction, endothelial function, arterial stiffness, atherosclerosis, cardiopulmonary exercise testing and cardiac magnetic resonance.
This collection will focus on currently available methods which may precisely assess cardiovascular toxicity induced by cancer treatment. The goal of this collection is to demonstrate the standardized protocols for cardiovascular function before, during and/or after cancer treatment. This collection will provide a platform for the multiple approaches currently utilized in clinical setting and will establish a uniform procedure for the methodologies.
Guest Editor: David Hay
Pluripotent stem cells retain the ability to form all cell types in the human body. This provides the field with renewable forms of human tissue, from defined genetic backgrounds, to model physiology ‘in the dish’ and also for clinical ap… Show Moreplication in the future. While many protocols exist to generate somatic cell types from pluripotent stem cells, their phenotype is usually fetal in nature and can be unstable in simple 2-dimensional culture. The field has approached these limitations, using various strategies, such as 3-dimensional sphere formation, tissue microfabrication, genome editing, cell perfusion and scaffold driven tissue assembly.
In addition to these important considerations, the scale up of stem cell derived technologies requires significant input from other scientific disciplines. A good example of this is the use of bioprinting and encapsulation to generate functional tissue at scale to treat organ deficiencies in vivo. Vital to these endeavours are the quality control of stem cell-derived tissue. This requires the analyses of complex datasets to define the release criteria and tissue phenotyping. Complementary to these snapshot measurements of tissue performance is the use of label-free assessment. This allows the user to control for quality and tissue performance in real time.
Following prototype design and proof of concept, the next step is technology translation. A key step in this process is industrialisation and commercialisation. This promises the genesis of human tissue at scale, which can be banked and shipped on demand at an affordable cost. Because of the importance of interdisciplinary research in the field, this Collection will bring together techniques across disciplines for the generation of human tissue for application. Show Less
Guest Editors: Andrew Tosolini, Giampietro Schiavo, James Sleigh, Gerardo Morfini
Axons are the main line of information transmission among neurons. Accordingly, axonal pathology represents a common pathogenic feature associated with loss of neuronal connectivity in multiple unrelated neurodegenerative diseases including Alzheimer… Show More’s disease, Parkinson’s disease, Huntington’s disease, and motor neuron diseases. Therefore, the study of the mechanisms and molecular events within axons bears important implications for a complete understanding of neuronal cell biology in both healthy and disease conditions.
Even though axons represent the largest subcellular compartment of neurons, addressing axon-specific effects has remained a major challenge to the field of Cellular Neurobiology. As most cellular and animal models involve translation of recombinant DNA constructs in the neuronal soma, it is often difficult to discern whether effects triggered from the resulting recombinant proteins take place within axons, or are an epiphenomenon associated with alterations in the neuronal cell body itself. In this collection, we present a variety of experimental systems and methodologies for the study of cellular processes and molecular events within the axonal compartments, including the intracellular trafficking of membrane-bounded organelles by motor proteins (axonal transport), the regulation of axonal cytoskeleton dynamics, and the phosphorylation of axonal proteins.
Guest Editors: Zhaoxin Wang, Junling Gao, Qun Yang, Delin Sun
Emotions are tightly associated with human thoughts, feelings and behaviors in various ways. Abnormal emotion processing leads to excessive or insufficient reactions, and is often accompanied by mental disorders such as anxiety, depression, post… Show More-traumatic stress disorder (PTSD), and alcohol or drug abuse. Therefore, the study of the neural correlates of human emotion processing bear important implications for fully understanding the inner mechanisms of emotion, and may contribute to the diagnosis and treatment of emotion-related mental disorders. It remains a significant challenge to non-invasively capture the neural signals of emotion in vivo, which often vary temporally and change dramatically across contexts. It also becomes increasingly important to merge findings from different modalities and through diverse analyses approaches. In this collection, we present a variety of experimental studies and methodologies for the investigation of emotion-related dynamic brain responses. The multidimensional neural responses will be recorded through the most widely-used neuroimaging and neurophysiological techniques including functional magnetic resonance imaging (fMRI), electroencephalography (EEG), and magnetoencephalography (MEG). Show Less
Guest Editor: Wen Zhou
Metal-ceramic restoration has remained the benchmark of prosthetic restoration for many decades, especially in high-stress occlusion surface of clinical situations. Current trends in dental research and material development have shown a preference fo… Show Morer all-ceramic restorations, which is the most esthetic full-veneer restoration dentistry has to offer. As the demand for more natural-looking restorations increases, dentists and dental researchers have developed a variety of investigational methods (e.g. shear bonding assays) to explore new type of ceramics and new materials including high-strength dental resins.
Due to rapid technological developments in digital impression and recent advances in CAD/CAM technologies, these new approaches have the potential to increase our understanding of dental material science, including not only changes in the pace of fabrication of provisional restorations, but our assessments of repair/relining material and our work flow for fixed partial dentures. With all these new opportunities and challenges, dental research methods have been improved over time, but is still not completely standardized, and variations are currently in widespread use.
This special issue invites contributions which explore the different properties of dental materials, review the principle of the assays currently used in their labs and key steps in the tests, and share insights on the potential how these methodologies heralds a bright future for the dental research and work in related medical fields. The special issue will focus on new areas of dental material research, including manufacturing high strength dental ceramics and resins, optimizing their chemical composition and modifying their microstructure.
Guest Editor: Silvia Calusi
External beam radiotherapy consists of irradiating patients with photon, electron or particle beams that deliver a quantity of energy (radiation dose) sufficient to destroy tumor cells in a delineated target. Dose distributions are delivered in… Show More such a way that organs near the target organ are isolated from radiation exposure as much as possible. The dose distribution is patient-specific and it is designed by medical physicists using specialized commercial programs (treatment planning systems, TPS) on the patient CT or MR scans where radiation oncologists have delineated organs and target contours. The whole process is time-consuming and the quality of the result is highly dependent on user ability and time availability. Automation of both organ contouring and treatment planning steps provide high-quality standardized treatments where variability among patients and planners are considerably reduced. Automatic planning and automated segmentation solutions have been recently proposed and implemented by some research groups and several TPS or image processing platform vendors. In addition, automation can be utilized to perform aggregate analyses on large numbers of patients, expediting the knowledge of side effects due to doses delivered to healthy tissues and making these analyses part of the clinical practice. The spread of automation techniques employed in radiation therapy can help to standardize and optimize the quality of the treatment plans created in different centers and makes it possible to compare them directly. This collection aims to compile such automation techniques in one convenient location for physicians to view. Show Less
Guest Editor: Paul Jarvis
Chloroplasts are the organelles that define plants. Along with many other metabolic, developmental and signalling functions, chloroplasts are responsible for photosynthesis – the process whereby sunlight energy is harnessed to power the cellula… Show Morer activities of life. Consequently, chloroplasts are essential, not only for plants but also for the myriad ecosystems that depend on plants, and for agriculture. Chloroplasts are composed of thousands of different proteins, most of which are nucleus-encoded and imported from the cytosol before being routed internally to one of a number of clearly-distinct intraorganellar compartments. To achieve a more complete understanding of chloroplast development and functions, and to enable biotechnological strategies involving chloroplast manipulation that address global challenges linked to food security or bioenergy, it will be essential to determine the targeting, localization and interactions of important chloroplast proteins. This method collection describes a set of critically important and complementary techniques that may be used to accomplish these goals. The collection mostly focuses on the widely-used model plant Arabidopsis thaliana (thale cress), but the methods may also be adapted and applied to other organisms. Show Less
Guest Editors: Sanghee Lee, Christina Jamieson
Organoids are three-dimensional (3D) ex vivo cellular structures that are believed to recapitulate and maintain the in vivo conditions of cellular viability and responsiveness to therapeutic drugs. Matrigel or collagen have been utilized as an extrac… Show Moreellular matrix to enable patient-derived cells to form 3D/organoids including cyst-like structures and spheroids. 3D cultured organoids are considered to be excellent models to understand the mechanisms of tumorigenesis and to determine the effects of anticancer agents. Therefore, methods have been developed for a range of cancers such as brain, breast, lung and prostate, among others. Prostate, however, is somewhat unique in that current methods for culturing organoids have a lower success rate than for other cancers. Difficulty in establishing prostate organoid cultures may be driven in part by tumorigenic heterogeneity. Most prostate organoids are derived from patient-derived xenografts (PDXs) and are genomically representative of the patient’s prostate cancer. Therefore, PDX organoids models are invaluable for studying down-stream signaling of target molecules and the effects of anti-cancer agent treatment. Several groups have developed modified protocols that are optimized for their experimental objectives. The goal of this collection is to (1) provide several examples of optimized culture condition for patient-derived prostate cancer 3D organoids, (2) introduce follow-up methodologies for single cell methods such as RNA-FISH and flow cytometry, (3) share meticulous details of immunohistochemistry (IHC)/ immunofluorescent (IF) technique specific for organoids and (4) differential criteria to determine the effects of drug treatment on organoids. Show Less
Guest Editors: Huan Xu, Fubo Wang
This Methods Collection focuses on the early detection and diagnosis of prostate cancer (PCa). PCa is the second most frequently diagnosed cancer overall and is the fifth leading cause of cancer mortality in men. Prognosis is relatively positive if t… Show Morehe disease is diagnosed at an early stage and a radical prostatectomy in conjunction with androgen-deprivation therapy (ADT) can be performed. Measurement of blood levels of prostate-specific antigen (PSA) and MRI are currently the most widely used early stage PCa detection methods, however, early diagnosis is still incredibly challenging. Currently, biopsy is the gold standard for early stage PCa detection, however, it is an invasive technique that can result in complications.
Recently, an increasingly large array of PCa markers have been discovered and are being assayed using novel detection methods. Specifically, plasma levels of extracellular microRNAs, circulating exosomes, as well as circulating cancer cells can be measured as prognostic biomarkers for PCa. Plasma levels are typically assayed using either a plasmon sensor chip or flow cytometry, both of which yield extremely precise marker quantitation. Additionally, MRI in conjunction with hyperpolarized [1-13C] pyruvate detects PCa and other cancers with high accuracy. This Methods Collection aims to serve as a central repository of the most significant advances in PCa early detection. Show Less
Guest Editor: YoneJung Yoon
Stroke is the second leading cause of death and disability worldwide. Animal stroke models are important in studying the causative mechanisms of stroke as well as to investigate novel treatments. However, the different causes and biomarkers of stroke… Show More make it difficult to study and discover treatments. Animal models of stroke are essential in understanding stroke etiology and pathology and provide insight into the appropriate application for research of new therapeutic interventions.
This proposed collection will bring together a wide range of relevant animal stroke models, evaluated systematically based on outcomes, to help researchers develop appropriate applications for scientific and clinical discoveries. Stroke will be evaluated in animals using a variety of techniques including: behavior assays, MRI, and immunohistochemisty. This will provide qualitative and quantitative information about the mechanisms of stroke induced injuries as well as the therapeutic potential of identified drugs for treatment. Animal models will include; induced ischemic stroke using middle cerebral artery occlusion, intracerebral hemorrhage using autologous blood or collagenases injection, subarachnoid hemorrhage (SAH) using injection of autologous blood into the cisterna magna, aneurysmal SAH (aSAH) using artery puncture or blood injection and a blast induced multi-brain bleeding model. Intervention outcomes will be assessed using MRI, behavior testing, and immunohistochemistry.
Guest Editors: Giulia Corradetti, Enrico Borrelli
The major progress achieved in ocular imaging offered to ophthalmologists worldwide the opportunity to access and analyze subclinical features. Although, the development of millions of morphological datasets raised the necessity to identify a method … Show Moreto quantify these subclinical features with high reliability and applicability. Large scientific datasets can be analyzed in a fast and non-invasive manner by Artificial Intelligence (AI), using algorithms based on machine learning, especially deep learning. Convolutional neural networks reproduce the path of the human brain for object recognition through deep learning of pathological features learned from training sets. The integration of AI with clinical practice has become a necessity in order to improve prediction and prognostic data and to analyze sublinical data, empowering the ophthalmologists to reach a higher quality of care. Our purpose of this Methods Collection is to present a review of AI applications in ophthalmology.
Guest Editor: Ahmed Mahmoud
Cardiovascular diseases remain a major health and economic burden worldwide. This is a consequence to the poor regenerative ability of the adult mammalian heart to repair itself in response to injury. Understanding the cellular and molecular events t… Show Morehat control cardiomyocyte replenishment hold significant implications for treatment of human heart disease. In addition, employing the advancements in stem cell biology, cardiomyocyte differentiation and maturation represents an important therapeutic approach that remains to be harnessed for cardiac regeneration.
This collection will focus on currently established methods in the heart regeneration field. Both zebrafish and neonatal mice are model organisms to understand the mechanisms that guide endogenous heart regeneration following injury. In addition, embryonic stem (ES) and induced pluripotent stem (iPS) cell-derived cardiomyocytes have significant potential for disease modeling, as well as cellular therapy in heart regeneration. Furthermore, the direct reprogramming of cardiac fibroblasts into cardiomyocytes is a unique method for replacing the non-contractile scar with functional cardiomyocytes. The goal of this collection is to demonstrate the standardized protocols for cardiac injury, cardiomyocyte differentiation from pluripotent stem cells, and direct reprogramming. This collection will provide a platform for the multiple approaches currently used towards studying and promoting heart regeneration and to establish a unifying procedure for these methodologies.
Guest Editors: Carmen Nussbaum-Krammer, Cindy Voisine
Protein homeostasis, or ‘proteostasis’, is fundamental to cellular and organismal health. Conserved cellular processes cooperate to maintain a properly folded proteome by influencing protein synthesis, folding and clearance. Cells respond… Show More to proteostasis imbalances by activating the Heat shock response, the unfolded protein response (UPR) and the oxidative stress response (OxR) to readjust the cellular proteome. Alterations in proteostasis have been linked to age-related neurodegenerative diseases and cancers. Based on the high level of conservation, model systems such as C. elegans have been instrumental in understanding the role of proteostais in human health and disease. C. elegans possess many characteristics that make it an ideal experimental model including its short lifecycle, transparent cuticle and ease of chemical and genetic manipulations.
This collection will provide methodological strategies to monitor proteostasis at the cellular and organismal level in C. elegans. Stressors, such as misfolded disease proteins, can be experimentally introduced into specific tissues types while cellular and organismal response to proteotoxicity are monitored over time. Proteotoxicity leads to cellular dysfunction that can be quantitatively measured by phenotypic changes. Furthermore, activation of protective pathways in response to proteotoxic stress can be observed using fluorescent reporters. Genetic modifications and chemical treatments can then be employed to determine their effectiveness in readjusting proteostasis. The outcome of this series will provide a toolkit for probing and monitoring proteostasis, evaluating quality control systems that maintain a folded proteome, and investigating the dysregulation of these systems that lead to disease. Show Less
Guest Editor: Vania Cao
As the field of neuroscience continues to produce large and increasingly complex datasets of brain structure, physiology and function, the demand for accurate and targeted behavioral paradigms to model neurological dysfunctions continues to grow.&nbs… Show Morep; We face a significant interpretive challenge when relying on generalized animal behavioral tasks and metrics, often with human-imposed constraints, to lend meaning to functional neural datasets. A lack of novel, updated and innovative behavioral paradigms remains an experimental and informative bottleneck in disease-specific fields, particularly for advancing foundational neural circuit discoveries to translational applications.
This collection presents a diverse set of naturalistic behavioral paradigms and analysis methodologies tailored to neural circuit studies. Methods in this collection detail examples of how to appropriately conceive of, design, validate, modify, measure and interpret behavioral outputs from animal models of neurological dysfunctions and disease. These behavioral methodologies may be paired with neural circuit datasets obtained using a variety of techniques, including in vivo optical imaging, optogenetics, electrophysiology and physical or pharmacological perturbations.
These behavioral paradigms and analysis methodologies are sensitive to the natural environments, cognitive capabilities and behavioral tendencies of the animal model, and are careful to avoid or minimize human bias in task design and data interpretation to better inform the neural circuit data gathered in parallel.
Guest Editor: Irene Stefanini
The budding yeast Saccharomyces cerevisiae
plays major roles in a broad range of environments and has relevant impacts on several biotechnological applications. It has also recently been shown to have an impact on humans and other animals he… Show Morealth. Disclosing the natural niches hosting this yeast and the evolution of S. cerevisiae is fundamental in order to properly control the diffusion of this microorganism and its impact on the environment and on other organisms. Recent findings have highlighted the presence of this yeast in a previously unpredictable range of natural and human-related environments, spanning from the vineyard to human bodies and insect intestines. In addition, the intra-specific diversity of this yeast is increasingly recognized as being responsible for major changes in several functions of S. cerevisiae such as driving fermentations, colonizing human bodies, and attracting insects. New methods have allowed researchers to identify new natural niches for this microorganism, as well as to dissect the impact of its diversity on other microbes, animals’ health and behavior, and biotechnological applications. Thanks to these advancements, we are currently facing an unprecedented improvement in our understanding of yeast diffusion and ecology.
This Method Collection will include novel and improved approaches for the study of both yeast ecology and the impact of yeast diversity on other microorganisms or higher organisms.
Guest Editors: Haider Hassan, Milica Krstic
Breast cancer continues to be the most commonly diagnosed cancer worldwide and represents a leading cause of cancer-related mortality among women. Next-Generation Sequencing (NGS) has played an important role in providing mechanistic insights into th… Show Moree transcriptional, epigenetic and genomic nature of the disease, however, most of the work has focused on in vitro studies. The lack of in vivo characterization has led to an over-simplified account of molecular events, and limited our understanding of clonal variation and tumor heterogeneity in breast cancer. Hence, the combination of NGS with in vivo models provides an opportunity to bridge the gap between mechanistic models and the underlying pathophysiology of breast cancer. This collection will focus on emerging techniques in digital and molecular pathology, using lesion-specific tissue samples to assess transcriptomic and epigenetic aberrations in breast cancer. The goal of this collection is to provide protocols for execution of high-throughput genomic experiments, including: (1) Methods of assessing clonal variation and pathobiology of pre-cancerous lesions (RNA-seq), (2) Methods of validating genomic events such as transcription factor binding in tissue samples in vivo (ChIP-seq), (3) Methods of assessing defects in epigenetic regulation in breast cancer such as high-throughput methylase-assisted bisulfite sequencing (MAB-seq), and (4) Other high-throughput methods that allow for characterization of genomic aberrations in breast cancer tissue samples. Show Less
Guest Editor: Dan Ganea
Capturing large scale neuronal activity permits the functional analysis of inter and intra-areal neuronal networks in relation to perceptual decision-making under multi-faceted behaviors. From a technological perspective, recording the function of su… Show Morech large-scale neuronal activity has become permissible by using both electrophysiological recordings with neuropixel or multichannel silicon probes and imaging of genetically encoded calcium or voltage related activity under both head-fixed and freely moving behavioral paradigms. Such paradigms include multi alternative forced choice or Go/noGo tasks, where subjects are required to engage with the task in order to attain a reward.
Employing such paradigms enables precise control over stimulus statistics and reward contingencies, thus permitting extrinsic manipulations that drive and modulate intrinsic representations of perceptually driven decisions. Studying widescale neuronal function in correlation with various goal-oriented behaviors will permit to disentangle how network activity is shaped between various brain regions and within those regions how local activity is modulated in order to drive and support perceptual decisions.
However, as recording techniques improve and permit capturing widescale neuronal activity so do behavioral paradigms diversify and become more complex. This collection aims at presenting the variety of behavioral paradigms and experimental techniques accompanied to them, that are used in disentangling brain-wide perceptual decision-making components.
Guest Editors: Esther Zorio, Aitana Braza-Boils
miRNAs are the most studied non-coding RNAs, whose expression is tissue-specific. miRNAs post-transcriptionally modulate protein expression by degrading mRNA or blocking its translation. Increasing efforts are underway to characterize these molecules… Show More as biomarkers. However, recent findings have brought about a novel role for miRNAs as cell-to-cell communicators. miRNAs can be released by the emitting cell and packaged into extracellular vesicles protecting them from degradation by RNAses. Interestingly, miRNAs contained in extracellular vesicles are able to regulate the phenotype of their recipient cells by complex mechanisms which remain under investigation. Firstly, actively selected and packaged miRNAs are secreted. The lack of concordance between miRNA profiles in cells and exosomes, indicates that loading is not a passive process. Secondly, they may be transferred to their recipient cells as a novel mechanism of horizontal transfer of genetic material. And, finally, miRNAs suppress the translation of their target mRNAs into the recipient cell, modulating its fate. Nevertheless, little is known about the mechanisms regulating these stages although interesting experimental designs are being developed to shed light on this cellular mechanism. In light of these developments, this Methods Collection will compile methodologies, both in vitro and in vivo experiments, in order to assess the role of miRNAs as intercellular communicators, including both standard and groundbreaking methodologies.
Guest Editors: Clifford J. Tabin, Misty Rose Riddle
The Mexican Tetra, Astyanax mexicanus
is an excellent model organism to study the developmental and genetic basis of evolution. This species of fish consists of a river-dwelling (surface) population and a number of cave-dwelling po… Show Morepulations that evolved from surface fish trapped in caves. Cavefish have distinct morphology and physiology including: absence of eyes, reduced pigment, increased sensory structures, sensitive smell, reduced circadian rhythm, altered blood glucose regulation, and increased fat accumulation. Surface fish and all cavefish are interfertile allowing for genetic mapping studies that, in combination with a sequenced and annotated genome, have revealed the genetic changes leading to some of the unique cavefish traits. A. mexicanus have been bred in the lab for generations and numerous protocols exist for investigating their biology. The goal of this collection is to demonstrate standardized protocols for breeding, provide broader access to existing methods, and ultimately lower the threshold for researchers interested in working with this species.
Guest Editor: Charles E. Rupprecht
Rabies is an acute progressive encephalitis caused by a lyssavirus. The case fatality of this zoonosis is the highest for any infectious disease. Although laboratory techniques on rabies have progressed greatly over the last century, typically such i… Show Morenformation falls in disparate locations and is often unavailable for easy access by those with the greatest need. Hence, the opportunity for a visual, methodological catalog is critical for basic research in pathobiology, immunology and molecular biology, as well for applied diagnostics, prophylaxis, therapy and intervention. The time is opportune because a global program is underway for the elimination of human rabies transmitted by dogs through mass canine vaccination, requiring multiple tests for risk mitigation and measurement of programmatic success. Advances have occurred in pathogen discovery, characterization, novel biologics and anti-virals. Based on in vitro and animal models, hope has arisen not only for immunization after viral exposure but for potential treatment after illness. Besides direct application to humans and domestic animals, efficacious vaccines may also be distributed to free-ranging wildlife en masse via vaccine-laden baits. The objective of this collection is to: share standardized protocols for detection, characterization and response; promote broader global access to existing methods; produce open networks for research collaborations anew; and ultimately create a pathway for an evidence-based approach for additional knowledge on this neglected disease. We trust this approach will encourage a myriad of scientists to participate in this global endeavor in a trans-disciplinary manner, towards renewed biomedical progress in human and veterinary science and conservation biology. Show Less
Guest Editors: George J. Nelson, Leela M. R. Arava, Partha P. Mukherjee
Energy storage is a critical enabler for vehicle electrification and renewable energy integration into the electric grid. Primary challenges in lithium based intercalation (Li-ion) and conversion (e.g., Li-S) chemistries continue to remain in en… Show Moreergy and power density trade-off, life and safety. These energy storage systems consist of coupled physical processes at disparate spatio-temporal scales in porous electrodes, including species and charge transport in the solid and electrolyte phases, electrochemical reaction at active interfaces, heat generation and thermal transport, chemical reactions, and transport-mechanics manifestation. In recent years, there has been significant emphasis and progress in experimentally examining such coupled processes at different scales. This JoVE Methods Collection seeks contributions in multi-modal analytics, including but not limited to, electrochemical, spectroscopic, microscopic and tomographic analysis and diagnostics to examine coupled physicochemical processes in energy storage (e.g. Li-ion, and beyond Li-ion chemistries including Li-S, Li-air, Mg-ion, Na-ion, K-ion) systems. Show Less
Guest Editors: Trinidad García, Marisol Cueli, Débora Areces, Celestino Rodríguez
International classifications of mental disorders for the diagnosis of Learning Disabilities (LD) and Attention Deficit/Hyperactivity Disorders (ADHD) establish that they are not the result of other disabling conditions, although they might occur con… Show Morecurrently with other disabilities. Consequently, people who have a disability could also show limitations in learning process, professional or social development. These disorders are understood as intrinsic to the individual, assumed to be due to the dysfunction in the central nervous system, and may occur throughout the life cycle.
Taking into account the symptomatic complexity of these difficulties, as well as their high prevalence rates, it is necessary for professionals and researchers to have reliable and valid tools and methods for diagnosis and effective intervention design. These techniques would make it easier for professionals and researchers to develop their work in an appropriate and innovative way.
However, the main difficulty is the lack of correspondence between assessment and intervention proposals which is not always apparent. Assessment methods, tools, and diagnostic techniques are being currently developed, which provide objective information on the characteristics and needs of this population. This collection aims to bring together a wide range of techniques to provide insight into the assessment and intervention of LD and ADHD. Show Less
Guest Editors: Manu Sharma, Jacqueline Burré
Neurodegenerative diseases including Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, and frontotemporal dementia are devastating disorders with no known cure, mainly due to lack of unde… Show Morerstanding of the underlying disease mechanisms. Numerous mutant strains of mice have been generated as models to investigate these diseases. While fundamental insights into disease pathology and mechanism have been obtained, investigations at a subcellular resolution and high-throughput analyses such as drug screens are not feasible in animal models. Cellular model systems fill this niche. These systems have the advantage of addressing the underlying molecular and cellular disease mechanisms more acutely, enable fast and cost-effective genetic manipulations, and are ideal as a screening platform for libraries of new therapeutic compounds.
This methods collection includes a variety of cellular model systems to study neurodegenerative diseases, including yeast, non-neuronal models, primary neurons, and patient-derived neurons as 2D-induced neuron and 3D organoid cultures. Each model system has its particular strengths, for example, rapid genetic manipulation (yeast), active synapses (primary neurons), pure neuron types (induced neurons), and native-like neuronal connections and a glial milieu (organoids). Importantly, specific key aspects of neurodegenerative diseases have been reproduced in these cellular models, making them valuable for biochemical, cell biological, and high throughput studies, including those needed for early-stage drug discovery.
Guest Editors: Marcos Kulmann, Adriana Bos-Mikich
Renewed interest in parthenogenetic activation of mammalian oocytes has come about for scientific, medical, and economic reasons. Germinal vesicle (GV), MI oocytes and failed to fertilize (FF) oocytes are usually discarded in human IVF cycles. … Show MoreHowever, their developmental potential may be rescued by Artificial Oocyte Activation (AOA) to generate human embryonic stem cell (hESC) lines from blastocysts obtained after AOA. In the cattle industry AOA techniques have been developed to create ESC lines and to improve cloning efficiency. Various methods have been investigated for parthenogenetically activating oocytes and it seems that there is not a single method that activates oocytes from every species studied. Different groups have developed protocols optimized for their experimental objectives. The goal of this collection is to (1) provide an optimized AOA methodology for human immature and FF oocytes, to create parthenogenetic HESCs, (2) share meticulous details for manual isolation of the inner cell mass derivation in different substrates (3) provide information on AOA performed using a single or a combination of activating agents at different bovine oocyte maturation stages, as an important tool to achieve high efficiency on nuclear transfer experiments for cloning strategies. (4) share details on polimerase chain reaction for TNF-α gene expression and comet techniques to assess parthenote blastocyst viability and cell death for each activating treatment. Show Less
Guest Editors: Nathalie Giglioli-Guivarc'h, Bilal Abbasi, Christophe Hano
The use of plant in vitro
(bio)technologies offers an attractive alternative for the production of plant-based active ingredients. Plant cells are small factories able to synthesize an exceptional variety of commercially important phytochemi… Show Morecals used as flavors, dyes, pharmaceuticals, nutraceuticals, cosmeceuticals, etc. Conventional plant propagation techniques are subjected to variety of pests, environmental challenges and poor management practices that adversely affect the quality and yield of these valuable phytochemicals. Plant in vitro culture platform offers considerable advantages and advancements over conventional phytochemical production techniques. However, during culture development, the yield of metabolites is generally low due to poor understanding of various factors that affect the biosynthetic pathway. Therefore, various strategies have been developed and reported to enhance biosynthesis of these phytochemicals in plant cell cultures. The recent emergence of omics technologies, molecular biology, metabolic engineering and synthetic biology has revolutionized diverse fields in life sciences. These modern aspects of life sciences have considerably influencing plant in vitro technology and phytochemical production. An application of these technologies is expanding horizons for an understanding of metabolic pathways involved in the biosynthesis of valuable biologically active ingredients and their cost-efficient production. Show Less
Guest Editor: Antonio Simone Laganà
Gynecological surgery has gone through a radical evolution during recent years, due to technological improvements and a significant shift towards a minimally invasive approach. In the current scenario, both oncological and non-oncological conditions … Show Morecan be managed by endoscopic surgery, which has important advantages over classic open surgery including reduced hospital stay and post-operative pain, faster recovery, and better cosmetic results.
To date, laparoscopic and robotic surgery represent the gold standard management for several procedures, including myomectomy, adnexal surgery, hysterectomy, treatment of endometriosis, endometrial and ovarian cancers. In addition, hysteroscopy gained a key role for the evaluation of vaginal cavity, cervical canal, uterine cavity and tubal ostia and for the treatment of intracavitary diseases, such as polyps, submucous myomas, uterine septa/subsepta, T-shaped uterus, intrauterine adhesions, Asherman’s syndrome, and early diagnosis of endometrial cancer through targeted biopsies.
The development of augmented reality devices may further improve the feasibility and implementation of minimally invasive surgery in gynecology, allowing physicians to incorporate data visualization during surgical procedures in order to improve accuracy and safety, and to enhance surgical training. Considering all these factors, the purpose of the Collection is to offer a comprehensive overview of minimally invasive surgical techniques in gynecology, including laparoscopy, hysteroscopy and robotic surgery.
Guest Editors: Mario Serrano, Alexandre Tromas, Oswaldo Valdés-López
Plants constantly interact with different microorganisms; some of which could be potential pathogens or symbionts. Plants need to be able to distinguish between pathogens and symbionts to activate the correct molecular responses—either eng… Show Moreaging in symbiosis or acquiring resistance against them. To activate the correct response, plants rely on a battery of protein kinase receptors. These receptors perceive microbe-derived signalling molecules and activate different molecular and physiological responses. For example, most land plants detect the presence of bacterial pathogens through the detection of Pathogen Associated Molecular Patterns (PAMPs). The perception of PAMPs leads to the production of reactive oxygen species, activation of both calcium influx/efflux, and the expression of pathogen-related genes. These molecular responses are crucial to confer resistance against bacterial pathogens. For instance, legumes detect the presence of nitrogen-fixing soil bacteria by perceiving Nodulation Factors (NFs). Upon perception of the NFs, a series of molecular events, including calcium oscillations in the nuclear region, protein phosphorylation and the overexpression of symbiosis-related genes. These molecular events are required to establish a successful symbiosis.
The availability of experimental approaches yielding high resolution, reproducible, and reliable data is key to the successful study of plant-microbe interactions. This Method Collection will include novel and improved methods for the study of both plant-symbiont and plant-pathogen interactions.
Guest Editor: Dhrubajyoti Chowdhury
S-palmitoylation is the covalent attachment of the fatty acid palmitate to cysteine (Cys) residues of the substrate protein through thioester linkage. This universal post-translational lipid modification facilitates membrane association and therefore… Show More alters subcellular localization, stability and function of hundreds of proteins in cells. Among other lipid modifications of proteins, palmitoylation is unique because it is reversible and therefore can be regulated dynamically to influence various physiological processes within a cell. Palmitoylation occurs on diverse substrates, both soluble and transmembrane, and in a variety of different cell types and tissues and has crucial regulatory functions. Accumulating evidence indicates that malfunctions in palmitoylation are associated with cancer as well as a wide range of brain abnormalities, ranging from mild mental retardation to neurological disorders. Since its discovery 40 years ago, palmitoylation has been traditionally studied using metabolic radiolabeling that precluded the detection of dynamic as well as rare palmitoylation events. Over the last decade, a variety of different methods have been developed to study protein palmitoylation that overcome such limitations. Such methods broadly include chemical reporter-based metabolic labeling such as 17-octadecynoic acid (17-ODYA) labeling and biochemical assays such as and Acyl-PEG Exchange (APE), Acyl Biotin Exchange (ABE), and Acyl Resin-Assisted Capture (Acyl-RAC). Moreover, coupling these methods with other protein detection strategies has further expanded their applications. In this collection, we present the state-of-the-art techniques used to detect and enrich palmitoylated proteins that have opened up several exciting avenues for future research on dynamic palmitoylation. Show Less
Guest Editor: Cristiane Cunha Frota
Human papilloma virus (HPV) plays a fundamental role in carcinogenesis of several forms of cancer by subverting an effective immune response. There are more than 100 known HPV genotypes, of which, 13 can cause cancer of the cervix and are also associ… Show Moreated with various anogenital cancers and cancers of the head and neck. Breast cancer is the most common neoplasm among women throughout the world in both developed and developing countries. It has been demonstrated that HPV types 16 and 18 are capable of immortalizing normal breast epithelium. The identification of HPV as a major cause of breast cancer implies that implementation of interventions against the viral agent should prevent the disease(s). HPV positive tumors have a favorable prognosis, approximately 80% 5-year overall survival., but early diagnosis is crucial. There are many protocols to identify HPV, including commercial and in house assays. In this collection, we present the standardized protocols for the detection of HPV from several types of breast samples. We will share how to extract DNA and genotype from different samples, including formalin-fixed paraffin-embedded tissues and nipple aspirate. This collection will promote access to existing methods and encourage clinicians in the early identification of HPV. It will also facilitate a network for researchers to collaborate and compile knowledge on this disease. Show Less
Guest Editor: Ardian Hana
Performing brain surgery is a very demanding task for every neurosurgeon. In this discipline skull base surgery could be considered as the most difficult surgery to perform. Due to the geographical proximity of many vital structures of the brain… Show More in a very small space, skull base surgery can be very challenging for each neurosurgeon and is often perilous for the patient. Therefore the selection of the right approach for the various pathologies in this region of the brain is extremely important, in order to avoid as much damage as possible to these vital structures.
Currently, different approaches to the skull base have been proposed by the neurosurgeons worldwide. The type and the location of the pathology determines the exact approach to the skull base surgery. Approaches differ from surgeon to surgeon and country to country, as well. We aim to describe the different approaches for various pathologies like tumours and vascular pathologies to this critical region of the brain. In the process, we will gain further insight as to why a certain technique was chosen instead of an alternative.
Guest Editors: Tony Herrera Grao, Núria Bonada, Miguel Ca?edo-Argüelles Iglesias
We are facing a global biodiversity and environmental crisis that poses serious risks to human welfare. Among the different ecosystems on Earth, aquatic ecosystems are experiencing the fastest rates in species extinction and ecosystem degradation. Gi… Show Moreven that these ecosystems are highly valuable both in terms of biodiversity and ecosystem services, we need to take action to improve their management. Conservation and restoration are two powerful management tools that can help to prevent and reverse human impacts on aquatic ecosystems. During the last decades, remarkable methodological advances have been made in the field of conservation (e.g. systematic conservation planning) and restoration (e.g. bioengineering). This JoVE Methods Collection will visually explain some of the most relevant methods for conserving and restoring aquatic ecosystems and discuss their potential for reversing the global biodiversity and environmental crisis that aquatic ecosystems face. Show Less
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