JoVE Visualize What is visualize?
Stop Reading. Start Watching.
Advanced Search
Stop Reading. Start Watching.
Regular Search
Find video protocols related to scientific articles indexed in Pubmed.
Validation of a Molecular and Pathological Model for Five-Year Mortality Risk in Patients with Early Stage Lung Adenocarcinoma.
J Thorac Oncol
PUBLISHED: 11-15-2014
Show Abstract
Hide Abstract
The aim of this study was to validate a molecular expression signature [cell cycle progression (CCP) score] that identifies patients with a higher risk of cancer-related death after surgical resection of early stage (I-II) lung adenocarcinoma in a large patient cohort and evaluate the effectiveness of combining CCP score and pathological stage for predicting lung cancer mortality.
Related JoVE Video
Selective depletion of vascular EC-SOD augments chronic hypoxic pulmonary hypertension.
Am. J. Physiol. Lung Cell Mol. Physiol.
PUBLISHED: 10-19-2014
Show Abstract
Hide Abstract
Excess superoxide has been implicated in pulmonary hypertension (PH). We previously found that lung overexpression of the antioxidant extracellular superoxide dismutase (EC-SOD) attenuates PH and vascular remodeling. Though comprising only a small fraction of total SOD activity in most tissues, EC-SOD is abundant in arteries. We hypothesize that the selective loss of vascular EC-SOD promotes hypoxia-induced PH through redox-sensitive signaling pathways. EC-SOD(loxp/loxp)× Tg(cre/SMMHC) mice (SMC EC-SOD KO) received tamoxifen to conditionally deplete smooth muscle cell (SMC)-derived EC-SOD. Mice were exposed to hypobaric hypoxia for 35 days and PH was assessed by RV systolic pressure measurements and RV hypertrophy. Vascular remodeling was evaluated by morphometric analysis and two-photon microscopy for collagen. We examined lung cyclic GMP content, soluble guanylate cyclase expression and activity, phosphodiesterase5 expression and activity, expression of total and active endothelial nitric oxide synthase and GTP cyclohydrolase-1 (GTPCH-1), the rate-limiting enzyme in tetrahydrobiopterin synthesis. Knockout of SMC EC-SOD selectively decreased pulmonary artery EC-SOD without altering total lung EC-SOD. PH and vascular remodeling induced by chronic hypoxia was augmented in SMC EC-SOD KO. Depletion of SMC EC-SOD did not impact content or activity of lung sGC or PDE5, though it blunted the hypoxia-induced increase in cGMP. Though total eNOS was not altered, active eNOS and GTPCH-1 decreased with hypoxia only in SMC EC-SOD KO. We conclude that the localized loss of pulmonary artery EC-SOD augments chronic hypoxic PH. In addition to oxidative inactivation of NO, deletion of EC-SOD seems to reduce eNOS activity, further compromising pulmonary vascular function.
Related JoVE Video
Mitochondrial Cytochrome c Oxidase and Control of Energy Metabolism: Measurements in Suspensions of Isolated Mitochondria.
J. Appl. Physiol.
PUBLISHED: 10-18-2014
Show Abstract
Hide Abstract
Cytochrome c oxidase is the enzyme responsible for oxygen consumption by mitochondrial oxidative phosphorylation as well as coupling Site 3 of oxidative phosphorylation. In this role it determines the cellular rate of ATP synthesis by oxidative phosphorylation and is the key to understanding how energy metabolism is regulated. Four electrons are required for the reduction of oxygen to water, and these are provided by one-electron donor, cytochrome c. The rate of oxygen consumption (ATP synthesis) is dependent on the fraction of cyt c reduced (fred), oxygen pressure (pO2), energy state ([ATP]/[ADP][Pi]), and pH. In coupled mitochondria (high energy state) and pO2 > 60 torr, the rate increases in an exponential-like fashion with increasing fred. When the dependence on fred is fitted to the equation {rate = a(fred)(b)}, a decreased from 100 to near 20 and b increased from 1.3 to 4 as the pH of the medium increased from 6.5 to 8.3. During oxygen depletion from the medium fred progressively increases and the rate of respiration decreases. The respiratory rate falls to 1/2 (P50) by about 1.5 torr, at which point fred is substantially increased. The metabolically relevant dependence on pO2 is obtained by correcting for the increase in fred, in which case the P50 is 12 torr. Adding an uncoupler of oxidative phosphorylation eliminates the dependence of the cytochrome c oxidase activity on pH and energy state. The respiratory rate becomes proportional to fred and the P50 decreases to less than 1 torr.
Related JoVE Video
Trial of the route of early nutritional support in critically ill adults.
N. Engl. J. Med.
PUBLISHED: 10-01-2014
Show Abstract
Hide Abstract
Uncertainty exists about the most effective route for delivery of early nutritional support in critically ill adults. We hypothesized that delivery through the parenteral route is superior to that through the enteral route.
Related JoVE Video
MBD4 Interacts with and Recruits USP7 to heterochromatic foci.
J. Cell. Biochem.
PUBLISHED: 09-05-2014
Show Abstract
Hide Abstract
MBD4 is the only methyl-CpG binding protein that possesses a C-terminal glycosylase domain. It has been associated with a number of nuclear pathways including DNA repair, DNA damage response, the initiation of apoptosis, transcriptional repression, and DNA demethylation. However, the precise contribution of MBD4 to these processes in development and relevant diseases remains elusive. We identified UHRF1 and USP7 as two new interaction partners for MBD4. Both UHRF1, a E3 ubiquitin ligase, and USP7, a de-ubiquinating enzyme, regulate the stability of the DNA maintenance methyltransferase, Dnmt1. The ability of MBD4 to directly interact with and recruit USP7 to chromocenters implicates it as an additional factor that can potentially regulate Dnmt1 activity during cell proliferation. J. Cell. Biochem. © 2014 Wiley Periodicals, Inc.
Related JoVE Video
Why try to predict ICU outcomes?
Curr Opin Crit Care
PUBLISHED: 08-28-2014
Show Abstract
Hide Abstract
To describe why the prediction of ICU outcomes is essential to underpin critical care quality improvement programmes.
Related JoVE Video
Range-wide multilocus phylogeography of the red fox reveals ancient continental divergence, minimal genomic exchange and distinct demographic histories.
Mol. Ecol.
PUBLISHED: 08-19-2014
Show Abstract
Hide Abstract
Widely distributed taxa provide an opportunity to compare biogeographic responses to climatic fluctuations on multiple continents and to investigate speciation. We conducted the most geographically and genomically comprehensive study to date of the red fox (Vulpes vulpes), the world's most widely distributed wild terrestrial carnivore. Analyses of 697 bp of mitochondrial sequence in ~1000 individuals suggested an ancient Middle Eastern origin for all extant red foxes and a 400 kya (SD = 139 kya) origin of the primary North American (Nearctic) clade. Demographic analyses indicated a major expansion in Eurasia during the last glaciation (~50 kya), coinciding with a previously described secondary transfer of a single matriline (Holarctic) to North America. In contrast, North American matrilines (including the transferred portion of Holarctic clade) exhibited no signatures of expansion until the end of the Pleistocene (~12 kya). Analyses of 11 autosomal loci from a subset of foxes supported the colonization time frame suggested by mtDNA (and the fossil record) but, in contrast, reflected no detectable secondary transfer, resulting in the most fundamental genomic division of red foxes at the Bering Strait. Endemic continental Y-chromosome clades further supported this pattern. Thus, intercontinental genomic exchange was overall very limited, consistent with long-term reproductive isolation since the initial colonization of North America. Based on continental divergence times in other carnivoran species pairs, our findings support a model of peripatric speciation and are consistent with the previous classification of the North American red fox as a distinct species, V. fulva.
Related JoVE Video
The immune system in hypertension.
Trans. Am. Clin. Climatol. Assoc.
PUBLISHED: 08-16-2014
Show Abstract
Hide Abstract
Hypertension is generally attributed to perturbations of the vasculature, the kidney, and the central nervous system. During the past several years, it has become apparent that cells of the innate and adaptive immune system also contribute to this disease. Macrophages and T cells accumulate in the kidneys and vasculature of humans and experimental animals with hypertension, and likely contribute to end-organ damage. We have shown that mice lacking lymphocytes, such as recombinase-activating gene-deficient (RAG-1(-/-)) mice, have blunted hypertension in response to angiotensin II, increased salt levels, and norepinephrine. Adoptive transfer of T cells restores the blood pressure response to these stimuli. Others have shown that mice with severe combined immunodeficiency have blunted hypertension in response to angiotensin II. Deletion of the RAG gene in Dahl salt-sensitive rats reduces the hypertensive response to salt feeding. The central nervous system seems to orchestrate immune cell activation. We produced lesions of the anteroventral third ventricle and showed that these block T cell activation in response to angiotensin II. Likewise, we showed that genetic manipulation of reactive oxygen species in the subfornical organ modulates both hypertension and T cell activation. Current evidence indicates that production of cytokines including tumor necrosis factor alpha, interleukin 17, and interleukin 6 contribute to hypertension, likely by promoting vasoconstriction, production of reactive oxygen species, and sodium reabsorption in the kidney. We propose a working hypothesis linking the sympathetic nervous system, immune cells, the production of cytokines, and ultimately vascular and renal dysfunction, leading to augmentation of hypertension.
Related JoVE Video
Persistence and dose escalation of tumor necrosis factor inhibitors in US veterans with rheumatoid arthritis.
J. Rheumatol.
PUBLISHED: 08-15-2014
Show Abstract
Hide Abstract
Limited evidence exists comparing the persistence, effectiveness, and costs of biologic therapies for rheumatoid arthritis in clinical practice. Comparative effectiveness studies are needed to understand real-world experience with these agents. We evaluated treatment patterns, costs, and effectiveness of tumor necrosis factor inhibitor (TNFi) agents in patients enrolled in the Veterans Affairs Rheumatoid Arthritis (VARA) registry.
Related JoVE Video
Comparing mortality among adult, general intensive care units in England with varying intensivist cover patterns: a retrospective cohort study.
Crit Care
PUBLISHED: 08-14-2014
Show Abstract
Hide Abstract
Research has demonstrated that intensivist-led care of the critically ill is associated with reduced intensive care unit (ICU) and hospital mortality. The objective of this study was to evaluate whether a relation exists between intensivist cover pattern (for example, number of days of continuous cover) and patient outcomes among adult general ICUs in England.
Related JoVE Video
Superior mesenteric artery syndrome following sleeve gastrectomy: case report, review of the literature, and video on technique for surgical correction.
Surg Endosc
PUBLISHED: 08-13-2014
Show Abstract
Hide Abstract
Superior mesenteric artery (SMA) syndrome is a rare condition in which the duodenum is compressed between the SMA and aorta. This often occurs following extreme weight loss and has been reported in the bariatric population. We present the first reported case of SMA syndrome following sleeve gastrectomy. The patient underwent laparoscopic duodenojejunostomy and recovered uneventfully. The following is a review of the literature and detailed operative approach in the attached video.
Related JoVE Video
Novel flavonoids as anti-cancer agents: mechanisms of action and promise for their potential application in breast cancer.
Biochem. Soc. Trans.
PUBLISHED: 08-12-2014
Show Abstract
Hide Abstract
Flavonoids are a large group of ubiquitous polyphenolic secondary metabolites in plants with a wide range of properties, including a widely reported anti-cancer effect. The present review focuses on the different known mechanisms partaking in said anti-tumour effects, with particular emphasis on breast cancer. Their structure and reactivity allows flavonoids to work as antioxidant agents and phyto-oestrogens, modulating oestrogen signalling and metabolism to induce an overall anti-proliferative response. Other effects include the ability of flavonoids to modulate the CYP1 (cytochrome P450 1) and ABC (ATP-binding cassette) protein families, involved in carcinogenesis and drug delivery respectively. They can also induce apoptosis and cell cycle arrest and regulate other signalling pathways involved in the development and progression of cancer. In conclusion, there is accumulating evidence on the versatility of flavonoids and the numerous activities contributing to their anti-tumour effect. The complex, yet effective, mechanism of action of flavonoids, together with their interesting pharmacological properties, is the basis for their potential application in breast and other cancers. This rationale has led to the current interest in the application of flavonoids, including clinical trials currently underway and the development of novel flavonoids with improved properties, which hold great promise for tackling breast cancer.
Related JoVE Video
Oligoclonal CD8+ T Cells Play a Critical Role in the Development of Hypertension.
Hypertension
PUBLISHED: 08-04-2014
Show Abstract
Hide Abstract
Recent studies have emphasized a role of adaptive immunity, and particularly T cells, in the genesis of hypertension. We sought to determine the T-cell subtypes that contribute to hypertension and renal inflammation in angiotensin II-induced hypertension. Using T-cell receptor spectratyping to examine T-cell receptor usage, we demonstrated that CD8(+) cells, but not CD4(+) cells, in the kidney exhibited altered T-cell receptor transcript lengths in V?3, 8.1, and 17 families in response to angiotensin II-induced hypertension. Clonality was not observed in other organs. The hypertension caused by angiotensin II in CD4(-/-) and MHCII(-/-) mice was similar to that observed in wild-type mice, whereas CD8(-/-) mice and OT1xRAG-1(-/-) mice, which have only 1 T-cell receptor, exhibited a blunted hypertensive response to angiotensin II. Adoptive transfer of pan T cells and CD8(+) T cells but not CD4(+)/CD25(-) cells conferred hypertension to RAG-1(-/-) mice. In contrast, transfer of CD4(+)/CD25(+) cells to wild-type mice receiving angiotensin II decreased blood pressure. Mice treated with angiotensin II exhibited increased numbers of kidney CD4(+) and CD8(+) T cells. In response to a sodium/volume challenge, wild-type and CD4(-/-) mice infused with angiotensin II retained water and sodium, whereas CD8(-/-) mice did not. CD8(-/-) mice were also protected against angiotensin-induced endothelial dysfunction and vascular remodeling in the kidney. These data suggest that in the development of hypertension, an oligoclonal population of CD8(+) cells accumulates in the kidney and likely contributes to hypertension by contributing to sodium and volume retention and vascular rarefaction.
Related JoVE Video
DC isoketal-modified proteins activate T cells and promote hypertension.
J. Clin. Invest.
PUBLISHED: 08-04-2014
Show Abstract
Hide Abstract
Oxidative damage and inflammation are both implicated in the genesis of hypertension; however, the mechanisms by which these stimuli promote hypertension are not fully understood. Here, we have described a pathway in which hypertensive stimuli promote dendritic cell (DC) activation of T cells, ultimately leading to hypertension. Using multiple murine models of hypertension, we determined that proteins oxidatively modified by highly reactive ?-ketoaldehydes (isoketals) are formed in hypertension and accumulate in DCs. Isoketal accumulation was associated with DC production of IL-6, IL-1?, and IL-23 and an increase in costimulatory proteins CD80 and CD86. These activated DCs promoted T cell, particularly CD8+ T cell, proliferation; production of IFN-? and IL-17A; and hypertension. Moreover, isoketal scavengers prevented these hypertension-associated events. Plasma F2-isoprostanes, which are formed in concert with isoketals, were found to be elevated in humans with treated hypertension and were markedly elevated in patients with resistant hypertension. Isoketal-modified proteins were also markedly elevated in circulating monocytes and DCs from humans with hypertension. Our data reveal that hypertension activates DCs, in large part by promoting the formation of isoketals, and suggest that reducing isoketals has potential as a treatment strategy for this disease.
Related JoVE Video
Medication effectiveness with the use of tumor necrosis factor inhibitors among Texas medicaid patients diagnosed with rheumatoid arthritis.
J Manag Care Pharm
PUBLISHED: 06-27-2014
Show Abstract
Hide Abstract
Adalimumab (Humira [ADA]), etanercept (Enbrel [ETN]), and infliximab (Remicade [IFX]) are tumor necrosis factor (TNF) inhibitors indicated for the treatment of a variety of disorders. While their effectiveness has not been directly compared in a clinical trial, results from the majority of the indirect treatment comparisons suggest comparable efficacy and safety profiles. However, these TNF inhibitor agents differ in administration method and dosing flexibility, which may result in differences in medication use profiles (e.g., adherence, persistence, discontinuation, dose escalation, and switching to a new biologic rheumatoid arthritis drug) and effectiveness in clinical practice. 
Related JoVE Video
Effect of a perioperative, cardiac output-guided hemodynamic therapy algorithm on outcomes following major gastrointestinal surgery: a randomized clinical trial and systematic review.
JAMA
PUBLISHED: 05-21-2014
Show Abstract
Hide Abstract
Small trials suggest that postoperative outcomes may be improved by the use of cardiac output monitoring to guide administration of intravenous fluid and inotropic drugs as part of a hemodynamic therapy algorithm.
Related JoVE Video
Use of a validated algorithm to estimate the annual cost of effective biologic treatment for rheumatoid arthritis.
J Med Econ
PUBLISHED: 05-14-2014
Show Abstract
Hide Abstract
To estimate biologic cost per effectively treated patient with rheumatoid arthritis (RA) using a claims-based algorithm for effectiveness.
Related JoVE Video
Cost of biologics per treated patient across immune-mediated inflammatory disease indications in a pharmacy benefit management setting: a retrospective cohort study.
Clin Ther
PUBLISHED: 05-13-2014
Show Abstract
Hide Abstract
Pharmacy benefits management companies have emerged as the national standard for the management of prescription drugs in the United States. The objective of this study was to estimate the annual costs per treated patient of 8 biologics indicated for select immune-mediated inflammatory diseases: moderate to severe rheumatoid arthritis, moderate to severe plaque psoriasis, active psoriatic arthritis, and/or active ankylosing spondylitis.
Related JoVE Video
Development and validation of risk models to predict outcomes following in-hospital cardiac arrest attended by a hospital-based resuscitation team.
Resuscitation
PUBLISHED: 04-16-2014
Show Abstract
Hide Abstract
The National Cardiac Arrest Audit (NCAA) is the UK national clinical audit for in-hospital cardiac arrest. To make fair comparisons among health care providers, clinical indicators require case mix adjustment using a validated risk model. The aim of this study was to develop and validate risk models to predict outcomes following in-hospital cardiac arrest attended by a hospital-based resuscitation team in UK hospitals.
Related JoVE Video
Heterogeneity in tissue oxygenation: from physiological variability in normal tissues to pathophysiological chaos in malignant tumours.
Adv. Exp. Med. Biol.
PUBLISHED: 04-15-2014
Show Abstract
Hide Abstract
Heterogeneity is a feature of both normal oxygen supply to tissue and of a supply that is disturbed due to a wide range of different pathologies. Here, the physiological importance of heterogeneity of tissue oxygenation is revisited. The anatomical and functional basis for heterogeneity of blood flow, local and regional regulatory mechanisms in normal tissues and the pathophysiology of the failure of regulation will be examined.Under physiological conditions, regulation of blood flow distributions at global, regional and microregional levels play coordinated roles in ensuring adequate O2 supply to all tissue cells. How this is achieved may be organ-/organ layer-specific, depending on its function and priorities to match local O2 delivery to consumption. Examples where these regulatory mechanisms break down under conditions of ischaemia and shock will also be given.In contrast, pathologic heterogeneity in tissue oxygenation resulting from uncontrolled, chaotic growth as seen in malignant tumours represents a pathophysiological status that is not predictable which, in general, is associated with chronic and acute hypoxia. This can have fatal consequences due to hypoxia- induced (mal-)adaptive processes, malignant tumour progression and treatment resistance.
Related JoVE Video
Incidence and outcome of in-hospital cardiac arrest in the United Kingdom National Cardiac Arrest Audit.
Resuscitation
PUBLISHED: 04-03-2014
Show Abstract
Hide Abstract
To report the incidence, characteristics and outcome of adult in-hospital cardiac arrest in the United Kingdom (UK) National Cardiac Arrest Audit database.
Related JoVE Video
Systems analysis of drug-induced receptor tyrosine kinase reprogramming following targeted mono- and combination anti-cancer therapy.
Cells
PUBLISHED: 03-31-2014
Show Abstract
Hide Abstract
The receptor tyrosine kinases (RTKs) are key drivers of cancer progression and targets for drug therapy. A major challenge in anti-RTK treatment is the dependence of drug effectiveness on co-expression of multiple RTKs which defines resistance to single drug therapy. Reprogramming of the RTK network leading to alteration in RTK co-expression in response to drug intervention is a dynamic mechanism of acquired resistance to single drug therapy in many cancers. One route to overcome this resistance is combination therapy. We describe the results of a joint in silico, in vitro, and in vivo investigations on the efficacy of trastuzumab, pertuzumab and their combination to target the HER2 receptors. Computational modelling revealed that these two drugs alone and in combination differentially suppressed RTK network activation depending on RTK co-expression. Analyses of mRNA expression in SKOV3 ovarian tumour xenograft showed up-regulation of HER3 following treatment. Considering this in a computational model revealed that HER3 up-regulation reprograms RTK kinetics from HER2 homodimerisation to HER3/HER2 heterodimerisation. The results showed synergy of the trastuzumab and pertuzumab combination treatment of the HER2 overexpressing tumour can be due to an independence of the combination effect on HER3/HER2 composition when it changes due to drug-induced RTK reprogramming.
Related JoVE Video
Adhiron: a stable and versatile peptide display scaffold for molecular recognition applications.
Protein Eng. Des. Sel.
PUBLISHED: 03-25-2014
Show Abstract
Hide Abstract
We have designed a novel non-antibody scaffold protein, termed Adhiron, based on a phytocystatin consensus sequence. The Adhiron scaffold shows high thermal stability (Tm ca. 101°C), and is expressed well in Escherichia coli. We have determined the X-ray crystal structure of the Adhiron scaffold to 1.75 Å resolution revealing a compact cystatin-like fold. We have constructed a phage-display library in this scaffold by insertion of two variable peptide regions. The library is of high quality and complexity comprising 1.3 × 10(10) clones. To demonstrate library efficacy, we screened against the yeast Small Ubiquitin-like Modifier (SUMO). In selected clones, variable region 1 often contained sequences homologous to the known SUMO interactive motif (V/I-X-V/I-V/I). Four Adhirons were further characterised and displayed low nanomolar affinities and high specificity for yeast SUMO with essentially no cross-reactivity to human SUMO protein isoforms. We have identified binders against >100 target molecules to date including as examples, a fibroblast growth factor (FGF1), platelet endothelial cell adhesion molecule (PECAM-1; CD31), the SH2 domain Grb2 and a 12-aa peptide. Adhirons are highly stable and well expressed allowing highly specific binding reagents to be selected for use in molecular recognition applications.
Related JoVE Video
The immune system in hypertension.
Adv Physiol Educ
PUBLISHED: 03-04-2014
Show Abstract
Hide Abstract
While hypertension has predominantly been attributed to perturbations of the vasculature, kidney, and central nervous system, research for almost 50 yr has shown that the immune system also contributes to this disease. Inflammatory cells accumulate in the kidneys and vasculature of humans and experimental animals with hypertension and likely contribute to end-organ damage. We and others have shown that mice lacking adaptive immune cells, including recombinase-activating gene-deficient mice and rats and mice with severe combined immunodeficiency have blunted hypertension to stimuli such as ANG II, high salt, and norepinephrine. Adoptive transfer of T cells restores the blood pressure response to these stimuli. Agonistic antibodies to the ANG II receptor, produced by B cells, contribute to hypertension in experimental models of preeclampsia. The central nervous system seems important in immune cell activation, because lesions in the anteroventral third ventricle block hypertension and T cell activation in response to ANG II. Likewise, genetic manipulation of reactive oxygen species in the subfornical organ modulates both hypertension and immune cell activation. Current evidence indicates that the production of cytokines, including tumor necrosis factor-?, interleukin-17, and interleukin-6, contribute to hypertension, likely via effects on both the kidney and vasculature. In addition, the innate immune system also appears to contribute to hypertension. We propose a working hypothesis linking the sympathetic nervous system, immune cells, production of cytokines, and, ultimately, vascular and renal dysfunction, leading to the augmentation of hypertension. Studies of immune cell activation will clearly be useful in understanding this common yet complex disease.
Related JoVE Video
Role of vascular oxidative stress in obesity and metabolic syndrome.
Diabetes
PUBLISHED: 02-18-2014
Show Abstract
Hide Abstract
Obesity is associated with vascular diseases that are often attributed to vascular oxidative stress. We tested the hypothesis that vascular oxidative stress could induce obesity. We previously developed mice that overexpress p22phox in vascular smooth muscle, tg(sm/p22phox), which have increased vascular ROS production. At baseline, tg(sm/p22phox) mice have a modest increase in body weight. With high-fat feeding, tg(sm/p22phox) mice developed exaggerated obesity and increased fat mass. Body weight increased from 32.16 ± 2.34 g to 43.03 ± 1.44 g in tg(sm/p22phox) mice (vs. 30.81 ± 0.71 g to 37.89 ± 1.16 g in the WT mice). This was associated with development of glucose intolerance, reduced HDL cholesterol, and increased levels of leptin and MCP-1. Tg(sm/p22phox) mice displayed impaired spontaneous activity and increased mitochondrial ROS production and mitochondrial dysfunction in skeletal muscle. In mice with vascular smooth muscle-targeted deletion of p22phox (p22phox(loxp/loxp)/tg(smmhc/cre) mice), high-fat feeding did not induce weight gain or leptin resistance. These mice also had reduced T-cell infiltration of perivascular fat. In conclusion, these data indicate that vascular oxidative stress induces obesity and metabolic syndrome, accompanied by and likely due to exercise intolerance, vascular inflammation, and augmented adipogenesis. These data indicate that vascular ROS may play a causal role in the development of obesity and metabolic syndrome.
Related JoVE Video
Estimating effectiveness and cost of biologics for rheumatoid arthritis: application of a validated algorithm to commercial insurance claims.
Clin Ther
PUBLISHED: 02-14-2014
Show Abstract
Hide Abstract
The aim of this analysis was to implement a claims-based algorithm to estimate biologic cost per effectively treated patient for biologics approved for moderate to severe rheumatoid arthritis (RA).
Related JoVE Video
Quantification of tumour budding, lymphatic vessel density and invasion through image analysis in colorectal cancer.
J Transl Med
PUBLISHED: 02-13-2014
Show Abstract
Hide Abstract
Tumour budding (TB), lymphatic vessel density (LVD) and lymphatic vessel invasion (LVI) have shown promise as prognostic factors in colorectal cancer (CRC) but reproducibility using conventional histopathology is challenging. We demonstrate image analysis methodology to quantify the histopathological features which could permit standardisation across institutes and aid risk stratification of Dukes B patients.
Related JoVE Video
Patient-initiated second opinions: systematic review of characteristics and impact on diagnosis, treatment, and satisfaction.
Mayo Clin. Proc.
PUBLISHED: 02-13-2014
Show Abstract
Hide Abstract
The impact of second opinions on diagnosis in radiology and pathology is well documented; however, the value of patient-initiated second opinions for diagnosis and treatment in general medical practice is unknown. We conducted a systematic review of patient-initiated second opinions to assess their impact on clinical outcomes and patient satisfaction and to determine characteristics and motivating factors of patients who seek a second opinion. We searched PubMed, EMBASE, Cochrane, and Academic OneFile databases using Medical Subject Headings (MeSH) indexes and keyword searches. Search terms included referral and consultation, patient-initiated, patient preference, patient participation, second opinion, second review, and diagnosis. Multiple reviewers screened abstracts and articles to determine eligibility and extract data. We assessed risk of bias using the Cochrane Risk of Bias Tool and rated study quality using Cochrane's GRADE (Grades of Recommendation, Assessment, Development and Evaluation) approach. We screened 1342 abstracts and reviewed full text of 41 articles, identifying 7 articles that reported clinical agreement data and 10 that discussed patient characteristics, motivation, and satisfaction. We found that a second opinion typically confirms the original diagnosis or treatment regimen but that 90% of patients with poorly defined conditions remain undiagnosed. However, 10% to 62% of second opinions yield a major change in the diagnosis, treatment, or prognosis. A larger fraction of patients receive different advice on treatment than on diagnosis. Factors motivating a second opinion include diagnosis or treatment confirmation, dissatisfaction with a consultation, desire for more information, persistent symptoms, or treatment complications. Patients generally believed that second opinions were valuable. Second opinions can result in diagnostic and treatment differences. The literature on patient-initiated second opinions is limited, and the accuracy of the second opinion through follow-up is generally unknown. Standardized methods and outcome measures are needed to determine the value of second opinions, and the potential of second opinions to reduce diagnostic errors merits more rigorous evaluation.
Related JoVE Video
From ST segments to endothelial pathophysiology: hypercholesterolemia and endothelial superoxide production.
J. Clin. Invest.
PUBLISHED: 02-03-2014
Show Abstract
Hide Abstract
As a young medical resident, I encountered a patient suffering from spontaneous coronary vasospasm and was puzzled by these dramatic alterations in vasomotion. This encounter piqued my interest in understanding the drivers of vascular reactivity. In a paper published in the JCI, my colleagues and I revealed a role for superoxide production in the vascular dysfunction associated with hypercholesterolemia. Subsequent work by our group and others has unveiled complex associations between ROS generation and vascular disease.
Related JoVE Video
Related JoVE Video
GTP cyclohydrolase I gene polymorphisms are associated with endothelial dysfunction and oxidative stress in patients with type 2 diabetes mellitus.
PLoS ONE
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
The genetic background of atherosclerosis in type 2 diabetes mellitus (T2DM) is complex and poorly understood. Studying genetic components of intermediate phenotypes, such as endothelial dysfunction and oxidative stress, may aid in identifying novel genetic components for atherosclerosis in diabetic patients.
Related JoVE Video
Cost of tumor necrosis factor blockers per patient with rheumatoid arthritis in a multistate Medicaid population.
Clinicoecon Outcomes Res
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
The purpose of this study was to estimate the annual cost per treated patient for the tumor necrosis factor (TNF) blockers, etanercept, adalimumab, and infliximab in rheumatoid arthritis (RA) patients covered by Medicaid.
Related JoVE Video
Increased STAT1 signaling in endocrine-resistant breast cancer.
PLoS ONE
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Proteomic profiling of the estrogen/tamoxifen-sensitive MCF-7 cell line and its partially sensitive (MCF-7/LCC1) and fully resistant (MCF-7/LCC9) variants was performed to identify modifiers of endocrine sensitivity in breast cancer. Analysis of the expression of 120 paired phosphorylated and non-phosphorylated epitopes in key oncogenic and tumor suppressor pathways revealed that STAT1 and several phosphorylated epitopes (phospho-STAT1(Tyr701) and phospho-STAT3(Ser727)) were differentially expressed between endocrine resistant and parental controls, confirmed by qRT-PCR and western blotting. The STAT1 inhibitor EGCG was a more effective inhibitor of the endocrine resistant MCF-7/LCC1 and MCF-7/LCC9 lines than parental MCF-7 cells, while STAT3 inhibitors Stattic and WP1066 were equally effective in endocrine-resistant and parental lines. The effects of the STAT inhibitors were additive, rather than synergistic, when tested in combination with tamoxifen in vitro. Expression of STAT1 and STAT3 were measured by quantitative immunofluorescence in invasive breast cancers and matched lymph nodes. When lymph node expression was compared to its paired primary breast cancer expression, there was greater expression of cytoplasmic STAT1 (?3.1 fold), phospho-STAT3(Ser727) (?1.8 fold), and STAT5 (?1.5 fold) and nuclear phospho-STAT3(Ser727) (?1.5 fold) in the nodes. Expression levels of STAT1 and STAT3 transcript were analysed in 550 breast cancers from publicly available gene expression datasets (GSE2990, GSE12093, GSE6532). When treatment with tamoxifen was considered, STAT1 gene expression was nearly predictive of distant metastasis-free survival (DMFS, log-rank p?=?0.067), while STAT3 gene expression was predictive of DMFS (log-rank p<0.0001). Analysis of STAT1 and STAT3 protein expression in a series of 546 breast cancers also indicated that high expression of STAT3 protein was associated with improved survival (DMFS, p?=?0.006). These results suggest that STAT signaling is important in endocrine resistance, and that STAT inhibitors may represent potential therapies in breast cancer, even in the resistant setting.
Related JoVE Video
Customizing the therapeutic response of signaling networks to promote antitumor responses by drug combinations.
Front Oncol
PUBLISHED: 01-01-2014
Show Abstract
Hide Abstract
Drug resistance, de novo and acquired, pervades cellular signaling networks (SNs) from one signaling motif to another as a result of cancer progression and/or drug intervention. This resistance is one of the key determinants of efficacy in targeted anti-cancer drug therapy. Although poorly understood, drug resistance is already being addressed in combination therapy by selecting drug targets where SN sensitivity increases due to combination components or as a result of de novo or acquired mutations. Additionally, successive drug combinations have shown low resistance potential. To promote a rational, systematic development of combination therapies, it is necessary to establish the underlying mechanisms that drive the advantages of combination therapies, and design methods to determine drug targets for combination regimens. Based on a joint systems analysis of cellular SN response and its sensitivity to drug action and oncogenic mutations, we describe an in silico method to analyze the targets of drug combinations. Our method explores mechanisms of sensitizing the SN through a combination of two drugs targeting vertical signaling pathways. We propose a paradigm of SN response customization by one drug to both maximize the effect of another drug in combination and promote a robust therapeutic response against oncogenic mutations. The method was applied to customize the response of the ErbB/PI3K/PTEN/AKT pathway by combination of drugs targeting HER2 receptors and proteins in the down-stream pathway. The results of a computational experiment showed that the modification of the SN response from hyperbolic to smooth sigmoid response by manipulation of two drugs in combination leads to greater robustness in therapeutic response against oncogenic mutations determining cancer heterogeneity. The application of this method in drug combination co-development suggests a combined evaluation of inhibition effects together with the capability of drug combinations to suppress resistance mechanisms before they become clinically manifest.
Related JoVE Video
Inflammation and Mechanical Stretch Promote Aortic Stiffening in Hypertension Through Activation of p38 MAP Kinase.
Circ. Res.
PUBLISHED: 12-17-2013
Show Abstract
Hide Abstract
Aortic stiffening commonly occurs in hypertension and further elevates systolic pressure. Hypertension is also associated with vascular inflammation and increased mechanical stretch. The interplay between inflammation, mechanical stretch and aortic stiffening in hypertension remains undefined.
Related JoVE Video
The Protocolised Management in Sepsis (ProMISe) trial statistical analysis plan.
Crit Care Resusc
PUBLISHED: 12-03-2013
Show Abstract
Hide Abstract
The Protocolised Management in Sepsis (ProMISe) trial is an open, multicentre, randomised controlled trial (RCT) of the clinical effectiveness and cost-effectiveness of early, goaldirected, protocolised resuscitation compared with usual resuscitation for patients presenting to emergency departments (EDs) in the United Kingdom with early signs of severe sepsis or septic shock. The rationale for the ProMISe trial derives from a single-centre United States RCT that reported a reduction in hospital mortality from 46.5% to 30.5%.
Related JoVE Video
5-hydroxymethylcytosine profiling as an indicator of cellular state.
Epigenomics
PUBLISHED: 11-29-2013
Show Abstract
Hide Abstract
DNA methylation is widely studied in the context of cancer. However, the rediscovery of 5-hydroxymethylation of DNA adds a new layer of complexity to understanding the epigenetic basis of development and disease, including carcinogenesis. There have been significant advances in techniques for the detection of 5-hydroxymethylcytosine and, with this, greater insight into the distribution, regulation and function of this mark, which are reviewed here. Better understanding of the associated pathways involved in regulation of, and by, 5-hydroxymethylcytosine may give promise to new therapeutic targets. We discuss evidence to support the view of 5-hydroxymethylcytosine as a unique and dynamic mark of cellular state. These 5-hydroxymethylcytosine profiles may offer optimism for the development of diagnostic, prognostic and predictive biomarkers.
Related JoVE Video
Is the adult mouse striatum a hostile host for neural transplant survival?
Neuroreport
PUBLISHED: 11-22-2013
Show Abstract
Hide Abstract
Human donor cells, including neurally directed embryonic stem cells and induced pluripotent stem cells with the potential to be used for neural transplantation in a range of neurodegenerative disorders, must first be tested preclinically in rodent models of disease to demonstrate safety and efficacy. One strategy for circumventing the rejection of xenotransplanted human cells is to desensitize the host animal to human cells in the early neonatal period so that a subsequent transplant in adulthood is not immunorejected. This method has been robustly validated in the rat, but currently not in the mouse in which most transgenic models of neurodegeneration have been generated. Thus, we set out to determine whether this could be achieved through modification of the existing rat protocol. Mice were inoculated in the neonatal period with a suspension of human embryonic cortical tissue of varying cell numbers, and received a subsequent human embryonic cortical tissue cell transplant in adulthood. Graft survival was compared with those in mice immunosuppressed with cyclosporine A and those receiving allografts of mouse whole ganglionic eminence tissue. Poor survival was found across all groups, suggesting a general problem with the use of mouse hosts for testing human donor cells.
Related JoVE Video
Nox2-Induced Production of Mitochondrial Superoxide in Angiotensin II-Mediated Endothelial Oxidative Stress and Hypertension.
Antioxid. Redox Signal.
PUBLISHED: 10-30-2013
Show Abstract
Hide Abstract
Abstract Aims: Angiotensin II (AngII)-induced superoxide (O2(•-)) production by the NADPH oxidases and mitochondria has been implicated in the pathogenesis of endothelial dysfunction and hypertension. In this work, we investigated the specific molecular mechanisms responsible for the stimulation of mitochondrial O2(•-) and its downstream targets using cultured human aortic endothelial cells and a mouse model of AngII-induced hypertension. Results: Western blot analysis showed that Nox2 and Nox4 were present in the cytoplasm but not in the mitochondria. Depletion of Nox2, but not Nox1, Nox4, or Nox5, using siRNA inhibits AngII-induced O2(•-) production in both mitochondria and cytoplasm. Nox2 depletion in gp91phox knockout mice inhibited AngII-induced cellular and mitochondrial O2(•-) and attenuated hypertension. Inhibition of mitochondrial reverse electron transfer with malonate, malate, or rotenone attenuated AngII-induced cytoplasmic and mitochondrial O2(•-) production. Inhibition of the mitochondrial ATP-sensitive potassium channel (mitoK(+)ATP) with 5-hydroxydecanoic acid or specific PKC? peptide antagonist (EAVSLKPT) reduced AngII-induced H2O2 in isolated mitochondria and diminished cytoplasmic O2(•-). The mitoK(+)ATP agonist diazoxide increased mitochondrial O2(•-), cytoplasmic c-Src phosphorylation and cytoplasmic O2(•-) suggesting feed-forward regulation of cellular O2(•-) by mitochondrial reactive oxygen species (ROS). Treatment of AngII-infused mice with malate reduced blood pressure and enhanced the antihypertensive effect of mitoTEMPO. Mitochondria-targeted H2O2 scavenger mitoEbselen attenuated redox-dependent c-Src and inhibited AngII-induced cellular O2(•-), diminished aortic H2O2, and reduced blood pressure in hypertensive mice. Innovation and Conclusions: These studies show that Nox2 stimulates mitochondrial ROS by activating reverse electron transfer and both mitochondrial O2(•-) and reverse electron transfer may represent new pharmacological targets for the treatment of hypertension. Antioxid. Redox Signal. 00, 000-000.
Related JoVE Video
Costs of tumor necrosis factor blockers per treated patient using real-world drug data in a managed care population.
J Manag Care Pharm
PUBLISHED: 10-01-2013
Show Abstract
Hide Abstract
Several anti-inflammatory biologic medications are available in the United States for the treatment of moderate-to-severe rheumatoid arthritis, moderate-to-severe psoriasis, psoriatic arthritis, or ankylosing spondylitis. The tumor necrosis factor (TNF) blockers etanercept, adalimumab, and infliximab are approved for use in adults with any of these conditions, but predicting the annual costs of TNF-blocker treatment is complex due to differences in dosing schedules, treatment gaps, switching between TNF blockers, and dose escalation over time.
Related JoVE Video
The use of percutaneous lumbar fixation screws for bilateral pedicle fractures with an associated dislocation of a lumbar disc prosthesis.
Case Rep Orthop
PUBLISHED: 08-30-2013
Show Abstract
Hide Abstract
Study Design. Case report. Objective. To identify a safe technique for salvage surgery following complications of total disc replacement. Summary of Background Data. Lumbar total disc replacement (TDR) is considered by some as the gold standard for discogenic back pain. Revision techniques for TDR and their complications are in their infancy. This case describes a successful method of fixation for this complex presentation. Methods and Results. A 48-year-old male with lumbar degenerative disc disease and no comorbidities. Approximately two weeks postoperatively for a TDR, the patient represented with acute severe back pain and the TDR polyethylene inlay was identified as dislocated anteriorly. Subsequent revision surgery failed immediately as the polyethylene inlay redislocated intraoperatively. Further radiology identified bilateral pedicle fractures, previously unseen on the plain films. The salvage fusion of L5/S1 reutilized the anterior approach with an interbody fusion cage and bone graft. The patient was then turned intraoperatively and redraped. The percutaneous pedicle screws were used to fix L5 to the sacral body via the paracoccygeal corridor. Conclusion. The robust locking screw in the percutaneous screw allowed a complete fixation of the pedicle fractures. At 3-year followup, the patient has an excellent result and has returned to playing golf.
Related JoVE Video
Use of microarray analysis to investigate EMT gene signatures.
Methods Mol. Biol.
PUBLISHED: 07-23-2013
Show Abstract
Hide Abstract
The epithelial-to-mesenchymal transition (EMT) is a widely studied program of development of cells characterized by loss of cell adhesion, repression of E-cadherin expression, and increased cell mobility. Microarrays have become a well-established technique for simultaneously measuring the expression of thousands of transcripts encoded by the genome. In this chapter, we demonstrate how microarray analysis can be used to assess the role of EMT-genes associated with a collagen invading phenotype by generating a gene expression signature and relating this to cell line and tumor datasets from published microarray studies.
Related JoVE Video
Annual acquisition and administration cost of biologic response modifiers per patient with rheumatoid arthritis, psoriasis, psoriatic arthritis, or ankylosing spondylitis.
J Med Econ
PUBLISHED: 07-18-2013
Show Abstract
Hide Abstract
To estimate annual biologic response modifier (BRM) cost per treated patient with rheumatoid arthritis, psoriasis, psoriatic arthritis, and/or ankylosing spondylitis receiving etanercept, abatacept, adalimumab, certolizumab, golimumab, infliximab, rituximab, or ustekinumab.
Related JoVE Video
Simultaneous monitoring of brain and skin oxygenation during haemorrhagic shock in piglets.
Adv. Exp. Med. Biol.
PUBLISHED: 07-16-2013
Show Abstract
Hide Abstract
Phosphorescence quenching and visible lightguide spectrophotometry were used to measure brain cortex oxygen partial pressure and skin oxygen saturation, respectively, during stepwise haemorrhage and re-transfusion in four 4-7-day-old anaesthetised piglets. In three cases, the effect of administration of adrenalin (epinephrine) was investigated. Brain cortex partial pressure was measured using a conventional phosphorescence pO2 probe (bclocpO2) and using a self-contained phosphorescence microsensor (bcmicropO2). Peripheral tissue oxygen saturation was measured on the skin of the abdomen (abSsO2) and the distal right foreleg (flSsO2) using visible lightguide spectrophotometry. Haemorrhage of 65 ml reduced mean arterial blood pressure (MABP) from 75.5?±?11.0 mmHg (mean?±?standard deviation) to 42?±?2.6 mmHg. Mean bclocpO2 fell from 30.1?±?3.1 to 13.1?±?2.5 mmHg and mean bcmicropO2 fell from 33.8?±?11.4 to 13.3?±?9.5 mmHg. abSsO2 and flSsO2 values fell from 47.4?±?8.1 % and 43.6?±?10.9 %, respectively, to 21.9?±?5.5 % and 23.8?±?14.0 %. Infusion of adrenalin produced a mean transient increase in MABP to 137?±?2.6 mmHg, falling to 75.7?±?16.3 mmHg within 3 min. bclocpO2 also increased to 24.1?±?14.6 mmHg, but there were no significant changes in bcmicropO, abSsO2 or flSsO2. Following reinfusion all parameters returned to values that were not statistically different from their pre-haemorrhage values. The dynamic recordings of all the oxygenation parameters indicated that they were sensitive indicators of the degree of haemorrhage during the experiments.
Related JoVE Video
The presentation, recognition and management of bipolar depression in primary care.
J Gen Intern Med
PUBLISHED: 07-10-2013
Show Abstract
Hide Abstract
Bipolar disorder is a mood disorder characterized by episodes of major depression and mania or hypomania. Most patients experience chronic symptoms of bipolar disorder approximately half of the time, most commonly subsyndromal depressive symptoms or a full depressive episode with concurrent manic symptoms. Consequently, patients with bipolar depression are often misdiagnosed with major depressive disorder. Individual patient characteristics and population screening tools may be helpful in improving recognition of bipolar depression in primary care. Health risk behaviors including tobacco use, sedentary activity level and weight gain are highly prevalent in patients with bipolar disorder, as are the comorbid chronic diseases such as diabetes mellitus and cardiovascular disease. Patients with bipolar illness have about an eight-fold higher risk of suicide and a two-fold increased risk of death from chronic medical illnesses. Recognition of bipolar depression and its associated health risk behaviors and chronic medical problems can lead to the use of appropriate interventions for patients with bipolar disorder, which differ in important ways from the treatments used for major depressive disorder. The above topics are reviewed in detail in this article.
Related JoVE Video
A coaxial single fibre supercapacitor for energy storage.
Phys Chem Chem Phys
PUBLISHED: 06-21-2013
Show Abstract
Hide Abstract
An energy storage device--a coaxial single fibre supercapacitor--was developed using a dip coating method and characterised using electrochemical methods. The specific capacitance per unit area and length were calculated to be 3.18 mF cm(-2) and 0.1 mF cm(-1), respectively, for a 2.6 cm supercapacitor. A 70 cm long fibre supercapacitor was then woven into a piece of fabric. Following characterisation using cyclic voltammetry, we found that this device had the same capacitance as its original value before integration into the fabric.
Related JoVE Video
TMA Navigator: Network inference, patient stratification and survival analysis with tissue microarray data.
Nucleic Acids Res.
PUBLISHED: 06-12-2013
Show Abstract
Hide Abstract
Tissue microarrays (TMAs) allow multiplexed analysis of tissue samples and are frequently used to estimate biomarker protein expression in tumour biopsies. TMA Navigator (www.tmanavigator.org) is an open access web application for analysis of TMA data and related information, accommodating categorical, semi-continuous and continuous expression scores. Non-biological variation, or batch effects, can hinder data analysis and may be mitigated using the ComBat algorithm, which is incorporated with enhancements for automated application to TMA data. Unsupervised grouping of samples (patients) is provided according to Gaussian mixture modelling of marker scores, with cardinality selected by Bayesian information criterion regularization. Kaplan-Meier survival analysis is available, including comparison of groups identified by mixture modelling using the Mantel-Cox log-rank test. TMA Navigator also supports network inference approaches useful for TMA datasets, which often constitute comparatively few markers. Tissue and cell-type specific networks derived from TMA expression data offer insights into the molecular logic underlying pathophenotypes, towards more effective and personalized medicine. Output is interactive, and results may be exported for use with external programs. Private anonymous access is available, and user accounts may be generated for easier data management.
Related JoVE Video
Predicting response to the anti-estrogen fulvestrant in recurrent ovarian cancer.
Gynecol. Oncol.
PUBLISHED: 06-04-2013
Show Abstract
Hide Abstract
Anti-estrogen therapy appears to have efficacy in a subset of ovarian cancers, as demonstrated in multiple phase II studies. Identifying sensitive patients early in treatment may allow for targeted, low-toxicity primary therapy or prevention of recurrence. We have previously demonstrated that the likelihood of response to letrozole could be improved by patient selection based on estrogen-pathway marker expression. We sought to identify ovarian cancer biomarkers that might indicate sensitivity to fulvestrant, an estrogen receptor antagonist.
Related JoVE Video
Effect of early vs late tracheostomy placement on survival in patients receiving mechanical ventilation: the TracMan randomized trial.
JAMA
PUBLISHED: 05-23-2013
Show Abstract
Hide Abstract
Tracheostomy is a widely used intervention in adult critical care units. There is little evidence to guide clinicians regarding the optimal timing for this procedure.
Related JoVE Video
Dysfunctional resident lung mesenchymal stem cells contribute to pulmonary microvascular remodeling.
Pulm Circ
PUBLISHED: 05-11-2013
Show Abstract
Hide Abstract
Pulmonary vascular remodeling and oxidative stress are common to many adult lung diseases. However, little is known about the relevance of lung mesenchymal stem cells (MSCs) in these processes. We tested the hypothesis that dysfunctional lung MSCs directly participate in remodeling of the microcirculation. We employed a genetic model to deplete extracellular superoxide dismutase (EC-SOD) in lung MSCs coupled with lineage tracing analysis. We crossed (floxp)sod3 and mT/mG reporter mice to a strain expressing Cre recombinase under the control of the ABCG2 promoter. We demonstrated In vivo that depletion of EC-SOD in lung MSCs resulted in their contribution to microvascular remodeling in the smooth muscle actin positive layer. We further characterized lung MSCs to be multipotent vascular precursors, capable of myofibroblast, endothelial and pericyte differentiation in vitro. EC-SOD deficiency in cultured lung MSCs accelerated proliferation and apoptosis, restricted colony-forming ability, multilineage differentiation potential and promoted the transition to a contractile phenotype. Further studies correlated cell dysfunction to alterations in canonical Wnt/?-catenin signaling, which were more evident under conditions of oxidative stress. Our data establish that lung MSCs are a multipotent vascular precursor population, a population which has the capacity to participate in vascular remodeling and their function is likely regulated in part by the Wnt/?-catenin signaling pathway. These studies highlight an important role for microenviromental regulation of multipotent MSC function as well as their potential to contribute to tissue remodeling.
Related JoVE Video
Human tissue in systems medicine.
FEBS J.
PUBLISHED: 04-19-2013
Show Abstract
Hide Abstract
Histopathology, the examination of an architecturally artefactual, two-dimensional and static image remains a potent tool allowing diagnosis and empirical expectation of prognosis. Considerable optimism exists that the advent of molecular genetic testing and other biomarker strategies will improve or even replace this ancient technology. A number of biomarkers already add considerable value for prediction of whether a treatment will work. In this short review we argue that a systems medicine approach to pathology will not seek to replace traditional pathology, but rather augment it. Systems approaches need to incorporate quantitative morphological, protein, mRNA and DNA data. A significant challenge for clinical implementation of systems pathology is how to optimize information available from tissue, which is frequently sub-optimal in quality and amount, and yet generate useful predictive models that work. The transition of histopathology to systems pathophysiology and the use of multiscale data sets usher in a new era in diagnosis, prognosis and prediction based on the analysis of human tissue.
Related JoVE Video
Geographic disparities in patient travel for dialysis in the United States.
J Rural Health
PUBLISHED: 04-11-2013
Show Abstract
Hide Abstract
To estimate travel distance and time for US hemodialysis patients and to compare travel of rural versus urban patients.
Related JoVE Video
Immune cells control skin lymphatic electrolyte homeostasis and blood pressure.
J. Clin. Invest.
PUBLISHED: 04-05-2013
Show Abstract
Hide Abstract
The skin interstitium sequesters excess Na+ and Cl- in salt-sensitive hypertension. Mononuclear phagocyte system (MPS) cells are recruited to the skin, sense the hypertonic electrolyte accumulation in skin, and activate the tonicity-responsive enhancer-binding protein (TONEBP, also known as NFAT5) to initiate expression and secretion of VEGFC, which enhances electrolyte clearance via cutaneous lymph vessels and increases eNOS expression in blood vessels. It is unclear whether this local MPS response to osmotic stress is important to systemic blood pressure control. Herein, we show that deletion of TonEBP in mouse MPS cells prevents the VEGFC response to a high-salt diet (HSD) and increases blood pressure. Additionally, an antibody that blocks the lymph-endothelial VEGFC receptor, VEGFR3, selectively inhibited MPS-driven increases in cutaneous lymphatic capillary density, led to skin Cl- accumulation, and induced salt-sensitive hypertension. Mice overexpressing soluble VEGFR3 in epidermal keratinocytes exhibited hypoplastic cutaneous lymph capillaries and increased Na+, Cl-, and water retention in skin and salt-sensitive hypertension. Further, we found that HSD elevated skin osmolality above plasma levels. These results suggest that the skin contains a hypertonic interstitial fluid compartment in which MPS cells exert homeostatic and blood pressure-regulatory control by local organization of interstitial electrolyte clearance via TONEBP and VEGFC/VEGFR3-mediated modification of cutaneous lymphatic capillary function.
Related JoVE Video
Exercise attenuates neuropathology and has greater benefit on cognitive than motor deficits in the R6/1 Huntingtons disease mouse model.
Exp. Neurol.
PUBLISHED: 03-25-2013
Show Abstract
Hide Abstract
Huntingtons disease (HD) is a neurodegenerative disease caused by a mutation within the huntingtin gene that induces degeneration within the striatal nuclei, progressing to widespread brain atrophy and death. The neurodegeneration produces symptoms that reflect a corticostriatal disconnection syndrome involving motor, cognitive and psychiatric disturbance. Environmental enrichment has been demonstrated to be beneficial to patients with neurological disorders, with exercise being central to this effect. Rodent studies have confirmed exercise-induced neurogenesis and increased growth factor levels in the brain and improved behavioural function. The present study sought to determine whether an extended regime of exercise could retard disease progression in the R6/1 mouse model of HD. The study was designed specifically with a translational focus, selecting behavioural assessments with high clinical predictive validity. We found that exercise improved gait function in both control and HD mice and selectively improved performance in the R6/1 mice on a motor coordination aspect of the balance beam task. Exercise also retarded the progression of cognitive dysfunction on water T-maze procedural and reversal learning probes presented serially to probe cognitive flexibility. In addition, exercise reduced striatal neuron loss in the R6/1 mice but increased striatal neuronal intra-nuclear inclusion size and number relative to non-exercised R6/1 mice which demonstrated increased numbers of extra-neuronal inclusions, suggesting that the functional effects were striatally mediated. These results confirm and extend those from previous studies that demonstrate that HD may be amenable to exercise-mediated therapeutics, but suggest that the impact of such interventions may be primarily cognitive.
Related JoVE Video
Characteristics and outcome of children admitted to adult intensive care units in England, Wales and Northern Ireland (1996-2011).
Intensive Care Med
PUBLISHED: 03-22-2013
Show Abstract
Hide Abstract
Despite centralisation of paediatric intensive care units (PICU) in the UK, children continue to be admitted to adult intensive care units (AICU). We aimed to analyse trends in the admission of children to AICUs over a 16-year period from 1996, and describe their case mix, outcome and resource use in a recent cohort (2009-2011).
Related JoVE Video
Nitric oxide reduces Cl? absorption in the mouse cortical collecting duct through an ENaC-dependent mechanism.
Am. J. Physiol. Renal Physiol.
PUBLISHED: 03-20-2013
Show Abstract
Hide Abstract
Since nitric oxide (NO) participates in the renal regulation of blood pressure, in part, by modulating transport of Na? and Cl? in the kidney, we asked whether NO regulates net Cl? flux (JCl) in the cortical collecting duct (CCD) and determined the transporter(s) that mediate NO-sensitive Cl? absorption. Cl? absorption was measured in CCDs perfused in vitro that were taken from aldosterone-treated mice. Administration of an NO donor (10 ?M MAHMA NONOate) reduced JCl and transepithelial voltage (VT) both in the presence or absence of angiotensin II. However, reducing endogenous NO production by inhibiting NO synthase (100 ?M N(G)-nitro-L-arginine methyl ester) increased JCl only in the presence of angiotensin II, suggesting that angiotensin II stimulates NO synthase activity. To determine the transport process that mediates NO-sensitive changes in JCl, we examined the effect of NO on JCl following either genetic ablation or chemical inhibition of transporters in the CCD. Since the application of hydrochlorothiazide (100 ?M) or bafilomycin (5 nM) to the perfusate or ablation of the gene encoding pendrin did not alter NO-sensitive JCl, NO modulates JCl independent of the Na?-dependent Cl?/HCO?? exchanger (NDCBE, Slc4a8), the A cell apical plasma membrane H?-ATPase and pendrin. In contrast, both total and NO-sensitive JCl and VT were abolished with application of an epithelial Na(+) channel (ENaC) inhibitor (3 ?M benzamil) to the perfusate. We conclude that NO reduces Cl? absorption in the CCD through a mechanism that is ENaC-dependent.
Related JoVE Video
Differential expression of prognostic proteomic markers in primary tumour, venous tumour thrombus and metastatic renal cell cancer tissue and correlation with patient outcome.
PLoS ONE
PUBLISHED: 02-26-2013
Show Abstract
Hide Abstract
Renal cell carcinoma (RCC) is the most deadly of urological malignancies. Metastatic disease affects one third of patients at diagnosis with a further third developing metastatic disease after extirpative surgery. Heterogeneity in the clinical course ensures predicting metastasis is notoriously difficult, despite the routine use of prognostic clinico-pathological parameters in risk stratification. With greater understanding of pathways involved in disease pathogenesis, a number of biomarkers have been shown to have prognostic significance, including Ki67, p53, vascular endothelial growth factor receptor 1 (VEGFR1) and ligand D (VEGFD), SNAIL and SLUG. Previous pathway analysis has been from study of the primary tumour, with little attention to the metastatic tumours which are the focus of targeted molecular therapies. As such, in this study a tissue microarray from 177 patients with primary renal tumour, renal vein tumour thrombus and/or RCC metastasis has been created and used with Automated Quantitative Analysis (AQUA) of immunofluorescence to study the prognostic significance of these markers in locally advanced and metastatic disease. Furthermore, this has allowed assessment of differential protein expression between the primary tumours, renal vein tumour thrombi and metastases. The results demonstrate that clinico-pathological parameters remain the most significant predictors of cancer specific survival; however, high VEGFR1 or VEGFD can predict poor cancer specific survival on univariate analysis for locally advanced and metastatic disease. There was significantly greater expression of Ki67, p53, VEGFR1, SLUG and SNAIL in the metastases compared with the primary tumours and renal vein tumour thrombi. With the exception of p53, these differences in protein expression have not been shown previously in RCC. This confirms the importance of proliferation, angiogenesis and epithelial to mesenchymal transition in the pathogenesis and metastasis of RCC. Importantly, this work highlights the need for further pathway analysis of metastatic tumours for overcoming drug resistance and developing new therapies.
Related JoVE Video
Specific nuclear envelope transmembrane proteins can promote the location of chromosomes to and from the nuclear periphery.
Genome Biol.
PUBLISHED: 02-15-2013
Show Abstract
Hide Abstract
BACKGROUND: Different cell types have distinctive patterns of chromosome positioning in the nucleus. Although ectopic affinity-tethering of specific loci can be used to relocate chromosomes to the nuclear periphery, endogenous nuclear envelope proteins that control such a mechanism in mammalian cells have yet to be widely identified. RESULTS: To search for such proteins, 23 nuclear envelope transmembrane proteins were screened for their ability to promote peripheral localization of human chromosomes in HT1080 fibroblasts. Five of these proteins had strong effects on chromosome 5, but individual proteins affected different subsets of chromosomes. The repositioning effects were reversible and the proteins with effects all exhibited highly tissue-restricted patterns of expression. Depletion of two nuclear envelope transmembrane proteins that were preferentially expressed in liver each reduced the normal peripheral positioning of chromosome 5 in liver cells. CONCLUSIONS: The discovery of nuclear envelope transmembrane proteins that can modulate chromosome position and have restricted patterns of expression may enable dissection of the functional relevance of tissue-specific patterns of radial chromosome positioning.
Related JoVE Video
The use of reverse phase protein arrays (RPPA) to explore protein expression variation within individual renal cell cancers.
J Vis Exp
PUBLISHED: 02-06-2013
Show Abstract
Hide Abstract
Currently there is no curative treatment for metastatic clear cell renal cell cancer, the commonest variant of the disease. A key factor in this treatment resistance is thought to be the molecular complexity of the disease. Targeted therapy such as the tyrosine kinase inhibitor (TKI)-sunitinib have been utilized, but only 40% of patients will respond, with the overwhelming majority of these patients relapsing within 1 year. As such the question of intrinsic and acquired resistance in renal cell cancer patients is highly relevant. In order to study resistance to TKIs, with the ultimate goal of developing effective, personalized treatments, sequential tissue after a specific period of targeted therapy is required, an approach which had proved successful in chronic myeloid leukaemia. However the application of such a strategy in renal cell carcinoma is complicated by the high level of both inter- and intratumoral heterogeneity, which is a feature of renal cell carcinoma as well as other solid tumors. Intertumoral heterogeneity due to transcriptomic and genetic differences is well established even in patients with similar presentation, stage and grade of tumor. In addition it is clear that there is great morphological (intratumoral) heterogeneity in RCC, which is likely to represent even greater molecular heterogeneity. Detailed mapping and categorization of RCC tumors by combined morphological analysis and Fuhrman grading allows the selection of representative areas for proteomic analysis. Protein based analysis of RCC is attractive due to its widespread availability in pathology laboratories; however, its application can be problematic due to the limited availability of specific antibodies. Due to the dot blot nature of the Reverse Phase Protein Arrays (RPPA), antibody specificity must be pre-validated; as such strict quality control of antibodies used is of paramount importance. Despite this limitation the dot blot format does allow assay miniaturization, allowing for the printing of hundreds of samples onto a single nitrocellulose slide. Printed slides can then be analyzed in a similar fashion to Western analysis with the use of target specific primary antibodies and fluorescently labelled secondary antibodies, allowing for multiplexing. Differential protein expression across all the samples on a slide can then be analyzed simultaneously by comparing the relative level of fluorescence in a more cost-effective and high-throughput manner.
Related JoVE Video
The mosaic theory revisited: common molecular mechanisms coordinating diverse organ and cellular events in hypertension.
J Am Soc Hypertens
PUBLISHED: 01-17-2013
Show Abstract
Hide Abstract
More than 60 years ago, Dr. Irvine Page proposed the Mosaic Theory of hypertension, which states that many factors, including genetics, environment, adaptive, neural, mechanical, and hormonal perturbations interdigitate to raise blood pressure. In the past two decades, it has become clear that common molecular and cellular events in various organs underlie many features of the Mosaic Theory. Two of these are the production of reactive oxygen species and inflammation. These factors increase neuronal firing in specific brain centers, increase sympathetic outflow, alter vascular tone and morphology, and promote sodium retention in the kidney. Moreover, factors such as genetics and environment contribute to oxidant generation and inflammation. Other common cellular signals, including calcium signaling and endoplasmic reticulum stress are similarly perturbed in different cells in hypertension and contribute to components of Pages theory. Thus, Pages Mosaic Theory formed a framework for future studies of molecular and cellular signals in the context of hypertension, and has greatly aided our understanding of this complex disease.
Related JoVE Video
Annual costs of tumor necrosis factor inhibitors using real-world data in a commercially insured population in the United States.
J Med Econ
PUBLISHED: 12-08-2011
Show Abstract
Hide Abstract
To calculate annual cost per treated patient of tumor necrosis factor (TNF) inhibitors etanercept, adalimumab, and infliximab for common approved indications, based on actual TNF-inhibitor use in clinical practice.
Related JoVE Video
Tissue type is a major modifier of the 5-hydroxymethylcytosine content of human genes.
Genome Res.
PUBLISHED: 11-21-2011
Show Abstract
Hide Abstract
The discovery of substantial amounts of 5-hydroxymethylcytosine (5hmC), formed by the oxidation of 5-methylcytosine (5mC), in various mouse tissues and human embryonic stem (ES) cells has necessitated a reevaluation of our knowledge of 5mC/5hmC patterns and functions in mammalian cells. Here, we investigate the tissue specificity of both the global levels and locus-specific distribution of 5hmC in several human tissues and cell lines. We find that global 5hmC content of normal human tissues is highly variable, does not correlate with global 5mC content, and decreases rapidly as cells from normal tissue adapt to cell culture. Using tiling microarrays to map 5hmC levels in DNA from normal human tissues, we find that 5hmC patterns are tissue specific; unsupervised hierarchical clustering based solely on 5hmC patterns groups independent biological samples by tissue type. Moreover, in agreement with previous studies, we find 5hmC associated primarily, but not exclusively, with the body of transcribed genes, and that within these genes 5hmC levels are positively correlated with transcription levels. However, using quantitative 5hmC-qPCR, we find that the absolute levels of 5hmC for any given gene are primarily determined by tissue type, gene expression having a secondary influence on 5hmC levels. That is, a gene transcribed at a similar level in several different tissues may have vastly different levels of 5hmC (>20-fold) dependent on tissue type. Our findings highlight tissue type as a major modifier of 5hmC levels in expressed genes and emphasize the importance of using quantitative analyses in the study of 5hmC levels.
Related JoVE Video
Qualitative and quantitative MALDI imaging of the positron emission tomography ligands raclopride (a D2 dopamine antagonist) and SCH 23390 (a D1 dopamine antagonist) in rat brain tissue sections using a solvent-free dry matrix application method.
Anal. Chem.
PUBLISHED: 11-18-2011
Show Abstract
Hide Abstract
The distributions of positron emission tomography (PET) ligands in rat brain tissue sections were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI). The detection of the PET ligands was possible following the use of a solvent-free dry MALDI matrix application method employing finely ground dry ?-cyano-4-hydroxycinnamic acid (CHCA). The D2 dopamine receptor antagonist 3,5-dichloro-N-{[(2S)-1-ethylpyrrolidin-2-yl]methyl}-2-hydroxy-6-methoxybenzamide (raclopride) and the D1 dopamine receptor antagonist 7-chloro-3-methyl-1-phenyl-1,2,4,5-tetrahydro-3-benzazepin-8-ol (SCH 23390) were both detected at decreasing abundance at increasing period postdosing. Confirmation of the compound identifications and distributions was achieved by a combination of mass-to-charge ratio accurate mass, isotope distribution, and MS/MS fragmentation imaging directly from tissue sections (performed using MALDI TOF/TOF, MALDI q-TOF, and 12T MALDI-FT-ICR mass spectrometers). Quantitative data was obtained by comparing signal abundances from tissues to those obtained from quantitation control spots of the target compound applied to adjacent vehicle control tissue sections (analyzed during the same experiment). Following a single intravenous dose of raclopride (7.5 mg/kg), an average tissue concentration of approximately 60 nM was detected compared to 15 nM when the drug was dosed at 2 mg/kg, indicating a linear response between dose and detected abundance. SCH 23390 was established to have an average tissue concentration of approximately 15 ?M following a single intravenous dose at 5 mg/kg. Both target compounds were also detected in kidney tissue sections when employing the same MSI methodology. This study illustrates that a MSI may well be readily applied to PET ligand research development when using a solvent-free dry matrix coating.
Related JoVE Video
Heterogeneity mapping of protein expression in tumors using quantitative immunofluorescence.
J Vis Exp
PUBLISHED: 11-09-2011
Show Abstract
Hide Abstract
Morphologic heterogeneity within an individual tumor is well-recognized by histopathologists in surgical practice. While this often takes the form of areas of distinct differentiation into recognized histological subtypes, or different pathological grade, often there are more subtle differences in phenotype which defy accurate classification (Figure 1). Ultimately, since morphology is dictated by the underlying molecular phenotype, areas with visible differences are likely to be accompanied by differences in the expression of proteins which orchestrate cellular function and behavior, and therefore, appearance. The significance of visible and invisible (molecular) heterogeneity for prognosis is unknown, but recent evidence suggests that, at least at the genetic level, heterogeneity exists in the primary tumor(1,2), and some of these sub-clones give rise to metastatic (and therefore lethal) disease. Moreover, some proteins are measured as biomarkers because they are the targets of therapy (for instance ER and HER2 for tamoxifen and trastuzumab (Herceptin), respectively). If these proteins show variable expression within a tumor then therapeutic responses may also be variable. The widely used histopathologic scoring schemes for immunohistochemistry either ignore, or numerically homogenize the quantification of protein expression. Similarly, in destructive techniques, where the tumor samples are homogenized (such as gene expression profiling), quantitative information can be elucidated, but spatial information is lost. Genetic heterogeneity mapping approaches in pancreatic cancer have relied either on generation of a single cell suspension(3), or on macrodissection(4). A recent study has used quantum dots in order to map morphologic and molecular heterogeneity in prostate cancer tissue(5), providing proof of principle that morphology and molecular mapping is feasible, but falling short of quantifying the heterogeneity. Since immunohistochemistry is, at best, only semi-quantitative and subject to intra- and inter-observer bias, more sensitive and quantitative methodologies are required in order to accurately map and quantify tissue heterogeneity in situ. We have developed and applied an experimental and statistical methodology in order to systematically quantify the heterogeneity of protein expression in whole tissue sections of tumors, based on the Automated QUantitative Analysis (AQUA) system(6). Tissue sections are labeled with specific antibodies directed against cytokeratins and targets of interest, coupled to fluorophore-labeled secondary antibodies. Slides are imaged using a whole-slide fluorescence scanner. Images are subdivided into hundreds to thousands of tiles, and each tile is then assigned an AQUA score which is a measure of protein concentration within the epithelial (tumor) component of the tissue. Heatmaps are generated to represent tissue expression of the proteins and a heterogeneity score assigned, using a statistical measure of heterogeneity originally used in ecology, based on the Simpsons biodiversity index(7). To date there have been no attempts to systematically map and quantify this variability in tandem with protein expression, in histological preparations. Here, we illustrate the first use of the method applied to ER and HER2 biomarker expression in ovarian cancer. Using this method paves the way for analyzing heterogeneity as an independent variable in studies of biomarker expression in translational studies, in order to establish the significance of heterogeneity in prognosis and prediction of responses to therapy.
Related JoVE Video
Referral to an extracorporeal membrane oxygenation center and mortality among patients with severe 2009 influenza A(H1N1).
JAMA
PUBLISHED: 10-05-2011
Show Abstract
Hide Abstract
Extracorporeal membrane oxygenation (ECMO) can support gas exchange in patients with severe acute respiratory distress syndrome (ARDS), but its role has remained controversial. ECMO was used to treat patients with ARDS during the 2009 influenza A(H1N1) pandemic.
Related JoVE Video
Determining tamoxifen sensitivity using primary breast cancer tissue in collagen-based three-dimensional culture.
Biomaterials
PUBLISHED: 09-19-2011
Show Abstract
Hide Abstract
We developed a three-dimensional assay prepared from primary breast cancer tissue and quantified tumor response to tamoxifen therapy. Freshly harvested breast cancer biopsies obtained at the time of curative surgical resection were fragmented and embedded into collagen I cushions. Changes in proliferation, apoptosis and tumor volume in response to tamoxifen treatment were quantified using image analysis software and optical projection tomography. Individual and collective invasion of epithelial cells into the surrounding collagen I was observed over the course of the experiment using phase contrast light microscopy and histopathological methods. Addition of tamoxifen to preparations derived from ER+ tumors demonstrated a range of response as measured by proliferative and apoptotic markers. In keeping with published data, tamoxifen reduced the percentage of apoptotic cells expressing cleaved caspase-3 (p = 0.02, Poisson regression analysis). Tamoxifen also reduced residual epithelial volume in ER+ tumors (p = 0.001, Mann-Whitney test), but not in ER low/- tumors (p = 0.78). Changes in tumor volume, as measured by optical projection tomography, allowed stratification into responsive and non-responsive tumors. The model mirrors observations of breast cancer response and histopathological changes to tamoxifen in neo-adjuvant trials. This assay provides a method of screening a battery of therapeutics against individual cancers, informing subsequent design of neo-adjuvant trials.
Related JoVE Video
Medullary amyloidosis associated with apolipoprotein A-IV deposition.
Kidney Int.
PUBLISHED: 09-07-2011
Show Abstract
Hide Abstract
Amyloidosis is caused by extracellular deposition of proteins in an insoluble manner within tissues. In hereditary forms of amyloidosis, transthyretin, fibrinogen A-?, lysozyme, gelsolin, apolipoprotein A-I, and A-II accumulate in the tissue plaques. Here we describe a 52-year-old man with no family history of renal disease who presented with increased urinary frequency, gradual loss of renal function but no significant proteinuria. Renal biopsy found large amounts of amyloid restricted to the medulla with no involvement of glomeruli or vessels. Immunohistochemical analysis for transthyretin or serum amyloid A and tests for an underlying monoclonal gammopathy were negative. Although initially suspected to be amyloid light chain amyloidosis, laser microdissection and mass spectrometry showed that the amyloid was composed of large amounts of apolipoprotein A-IV. This was based on mass spectrometry studies that showed 100, 96, and 73 spectra in three microdissected samples that matched to apolipoprotein A-IV with 100% probability. DNA analyses detected three sequence variants representing common polymorphisms of the apolipoprotein A-IV gene. Thus, in this case, apolipoprotein A-IV deposition and renal involvement appear to be restricted to the medulla. A high degree of suspicion is required for the diagnosis of apolipoprotein A-IV amyloidosis as it may be missed if a renal biopsy consists only of cortex.
Related JoVE Video

What is Visualize?

JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

How does it work?

We use abstracts found on PubMed and match them to JoVE videos to create a list of 10 to 30 related methods videos.

Video X seems to be unrelated to Abstract Y...

In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.