Endobronchial ultrasound

During flexible fiberoptic bronchoscopy (FB), a solitary pulmonary nodule (SPN) is sampled by means of transbronchial needle aspiration (TBNA), brush, or transbronchial lung biopsy under fluoroscopy; and mediastinal lymph nodes are sampled using "blind" TBNA. Endobronchial ultrasound (EBUS) was developed to help visualize the lesion at the time of biopsy in order to improve the diagnostic yield. METHODS: There are two types of EBUS techniques: using a radial probe (RP) with a rotating transducer at the distal tip, which produces a 360 degrees image to the long axis of the bronchoscope; and using an EBUS bronchoscope with a linear transducer at its distal tip, producing a 50 degrees image parallel to its long axis. RESULTS: In biopsies of SPNs < 2 cm using an RP, EBUS demonstrates a higher diagnostic yield than conventional FB techniques. With mediastinal and hilar nodal stations, except for the subcarina, EBUS shows a higher yield over blind TBNA. The current procedural terminology code for EBUS is 31620, a "ZZZ" code submitted in addition to other performed procedures (31622-31638). In 2007, an estimate of physician Medicare reimbursement for EBUS is $70.49. Reimbursement is locality dependent and based on economic-exchange conversion factors. Incorporating an ultrasound image into the report substantiates the use of this technique. Limitations: The physician must learn ultrasound image interpretation and the EBUS technique, and be skilled in TBNA. Maintaining competency requires frequent performance of EBUS. CONCLUSION: EBUS-directed biopsy improves the yield over conventional FB for SPNs < 2 cm and for most mediastinal or hilar nodal stations. This reduces the need to conduct additional diagnostic procedures.     

Enhanced green fluorescence by the expression of an Aequorea victoria green fluorescent protein mutant in mono- and dicotyledonous plant cells.

The expression of the jellyfish green fluorescent protein (GFP) in plants was analyzed by transient expression in protoplasts from Nicotiana tabacum, Arabidopsis thaliana, Hordeum vulgare, and Zea mays. Expression of GFP was only observed with a mutated cDNA, from which a recently described cryptic splice site had been removed. However, detectable levels of green fluorescence were only emitted from a small number of protoplasts. Therefore, other mutations in the GFP cDNA leading to single-amino acid exchanges in the chromophore region, which had been previously studied in Escherichia coli, were tested in order to improve the sensitivity of this marker protein. Of the mutations tested so far, the exchange of GFP amino acid tyrosine 66 to histidine (Y66H) led to detection of blue fluorescence in plant protoplasts, while the exchange of amino acid serine 65 to cysteine (S65C) and threonine (S65T) increased the intensity of green fluorescence drastically, thereby significantly raising the detection level for GFP. For GFP S65C, the detectable number of green fluorescing tobacco (BY-2) protoplasts was raised up to 19-fold, while the fluorimetricly determined fluorescence was raised by at least 2 orders of magnitude.     

Characterizing asthma from a drop of blood using neutrophil chemotaxis

Asthma is a chronic inflammatory disorder that affects more than 300 million people worldwide. Asthma management would benefit from additional tools that establish biomarkers to identify phenotypes of asthma. We present a microfluidic solution that discriminates asthma from allergic rhinitis based on a patient’s neutrophil chemotactic function. The handheld diagnostic device sorts neutrophils from whole blood within 5 min, and generates a gradient of chemoattractant in the microchannels by placing a lid with chemoattractant onto the base of the device. This technology was used in a clinical setting to assay 34 asthmatic (n = 23) and nonasthmatic, allergic rhinitis (n = 11) patients to establish domains for asthma diagnosis based on neutrophil chemotaxis. We determined that neutrophils from asthmatic patients migrate significantly more slowly toward the chemoattractant compared with nonasthmatic patients (P = 0.002). Analysis of the receiver operator characteristics of the patient data revealed that using a chemotaxis velocity of 1.55 μm/min for asthma yields a diagnostic sensitivity and specificity of 96% and 73%, respectively. This study identifies neutrophil chemotaxis velocity as a potential biomarker for asthma, and we demonstrate a microfluidic technology that was used in a clinical setting to perform these measurements.Asthma is a chronic inflammatory disorder of the lungs that is associated with airway hyperresponsiveness (AHR) and obstructed airflow (1), affecting more than 300 million people worldwide (2). Over the past 30 y, asthma prevalence has increased significantly in many populations, with some indications that prevalence may be reaching a plateau in the developed world. Significant progress has been made in identifying primary mediators involved in the pathophysiology of asthma. Several cell types, such as T helper cells (T_H1/T_H2), dendritic cells, mast cells, macrophages, eosinophils, and neutrophils play central roles in the pathology of asthma (4–7). Additionally, various cytokines that regulate the leukocyte trafficking, such as interleukins, IFN-γ, and TNF-α, have been identified and targeted in drug therapies. The recruitment of leukocytes to the lungs, particularly eosinophils and neutrophils, is central to the pathogenesis of asthma. Increased numbers of eosinophils are prominently observed in the lung tissue and bronchoalveolar lavage (BAL) fluid for most asthmatics (5). Neutrophils play a more critical role in severe asthma, where elevated counts of neutrophils are often observed in the BAL fluid (7). An overview of the role of neutrophils in asthma is shown in Fig. 1A. Although significant progress has been made in uncovering mediators in the pathology of asthma, these gains have not yet greatly improved our ability to define clinically relevant phenotypes of asthma in patients.Open in a separate windowFig. 1.Overview of different diagnostic techniques and the role of neutrophils in the pathology of asthma. (A) Summary of the role of neutrophils in the pathology of asthma, showing neutrophil adhesion and transendothelial migration; chemotaxis mediated by macrophages and T-helper cells; and neutrophilia in the lung tissue that leads to airway remodeling and airflow obstruction. (B) Proposed microfluidic method (more details in Fig. S1) for phenotyping asthma patients by measuring upstream of the asthma pathology with rapid neutrophil sorting on a P-selectin–coated surface (1); neutrophil chemotaxis monitored with high-throughput microscopy and automatically tracked with software (2); and asthma characterization on the basis of chemotaxis outputs (3). (C) Traditional clinical asthma diagnostic methods occur downstream of the asthma pathophysiology by measuring the effect of leukocyte inflammation on airway obstruction, nitric oxide output, or clinical symptoms.Asthma is diagnosed clinically by physicians, informed by the patient’s medical history, spirometry tests that measure lung function, reversibility of AHR, and several other potential metrics (8). These diagnostic techniques measure the effects of the inflammatory response in the lung by assessing airway constriction, nitric oxide production, and the resulting clinical symptoms. However, all of these diagnostic tests require patient compliance, which can be challenging when diagnosing children or the elderly (9). Additionally, many asthma diagnostic tests partially rely on the patient experiencing clinical symptoms that are variable during or around the visit to the physician. Perhaps these common characteristics of current diagnostic techniques contribute to difficulties in diagnosing asthma, particularly in certain subpopulations. For example, in a recent Canadian study involving ∼500 obese and nonobese subjects, Aaron et al. (10) found that ∼30% of the test subjects had been falsely diagnosed with asthma by physicians. Additionally, it is well established that the elderly are consistently underdiagnosed for asthma (11, 12). Therefore, additional tools are needed to improve the diagnosis of asthma. Furthermore, current asthma assessments do not inform the clinician of disease severity, expected clinical course, and risk of exacerbations.To improve characterization of asthma in the clinic, we have developed a handheld microfluidic chip that can identify functional measures of asthma from a drop of whole blood. Microfluidic systems have several characteristics that make them well-suited for clinical use, including low sample-volume requirements (13, 14); simple integration with automated fluid handling systems (15); and diffusion-dominant laminar fluidic phenomena that allow for precise control of a cell’s microenvironment (16–18). Indeed, microfluidic-based tools are increasingly being used in clinical research for diagnostic purposes (19–26). Neutrophils have been used to diagnose clinical conditions in human patients based on proteomic and genomic analysis (22) and chemotaxis behavior (23, 27), demonstrating that assays measuring cell function can be used for diagnostics. In this work, we assay the neutrophil chemotactic function in a blind study to identify quantitative domains that can be used to discriminate asthma from nonasthmatic allergic rhinitis. This approach of directly measuring the effector cell in the pathology of asthma differs from traditional diagnostic tests, which measure the variable effect of inflammation on airway constriction (Fig. 1 B and C and Table S1). Importantly, we developed methods to simplify the sample preparation, assay protocol, and data analysis that offer significant time savings over traditional macroscale (28–30) and microscale (18) chemotaxis techniques, allowing for the translation of the technology into the clinic. We analyzed 34 patients, and discovered that neutrophil chemotaxis can be used to discriminate asthma from nonasthmatic, allergic rhinitis patients with sensitivity and specificity of 96% and 73%, respectively. The results of the clinical application of our microfluidic device represent a first step demonstration of how asthma can potentially be diagnosed and managed based on cellular function, rather than largely by clinical observations.     

Genetic portrait study for 23 Y-STR loci in the population of Rajasthan,India

International Journal of Legal Medicine - This study was conducted to come up with data on Y-STR markers for the population of Rajasthan comprising of the western arid region of India. Y-STR...     

The N-terminal region containing the zinc finger domain of tobacco streak virus coat protein is essential for the formation of virus-like particles

Tobacco streak virus (TSV), a member of the genus Ilarvirus (family Bromoviridae), has a tripartite genome and forms quasi-isometric virions. All three viral capsids, encapsidating RNA 1, RNA 2 or RNA 3 and subgenomic RNA 4, are constituted of a single species of coat protein (CP). Formation of virus-like particles (VLPs) could be observed when the TSV CP gene was cloned and the recombinant CP (rCP) was expressed in E. coli. TSV VLPs were found to be stabilized by Zn²⁺ ions and could be disassembled in the presence of 500 mM CaCl_2. Mutational analysis corroborated previous studies that showed that an N-terminal arginine-rich motif was crucial for RNA binding; however, the results presented here demonstrate that the presence of RNA is not a prerequisite for assembly of TSV VLPs. Instead, the N-terminal region containing the zinc finger domain preceding the arginine-rich motif is essential for assembly of these VLPs.     

Transgene‐mediated suppression of dengue viruses in the salivary glands of the yellow fever mosquito,Aedes aegypti

Controlled sex‐, stage‐ and tissue‐specific expression of antipathogen effector molecules is important for genetic engineering strategies to control mosquito‐borne diseases. Adult female salivary glands are involved in pathogen transmission to human hosts and are target sites for expression of antipathogen effector molecules. The Aedes aegypti 30K a and 30K b genes are expressed exclusively in adult female salivary glands and are transcribed divergently from start sites separated by 263 nucleotides. The intergenic, 5′‐ and 3′‐end untranslated regions of both genes are sufficient to express simultaneously two different transgene products in the distal‐lateral lobes of the female salivary glands. An antidengue effector gene, membranes no protein (Mnp), driven by the 30K b promoter, expresses an inverted‐repeat RNA with sequences derived from the premembrane protein‐encoding region of the dengue virus serotype 2 genome and reduces significantly the prevalence and mean intensities of viral infection in mosquito salivary glands and saliva.     

Almanac 2012: Cell therapy in cardiovascular disease. The national society journals present selected research that has driven recent advances in clinical cardiology

The rapid translation from bench to bedside that has been seen in the application of regenerative medicine to cardiology has led to exciting new advances in our understanding of some of the fundamental mechanisms related to human biology. The first generation of cells used in phase I-II trials (mainly bone marrow mononuclear cells) are now entering phase III clinical trials with the goal of producing a cell based therapeutic that can change the outcome of cardiac disease. First generation cell therapy appears to have addressed safety concerns as well as showing ‘activity’ in numerous published meta-analyses. With the knowledge gained to date, the field is moving towards the next generation of cells-the ‘engineered’ cell-that have been developed to display a phenotype that will further enhance the myocardial repair/salvage process. This almanac review covers the latest basic research that may soon have application to humans as well as the results of the latest clinical trials.     

Mobility among youth in Rakai,Uganda: Trends,characteristics, and associations with behavioural risk factors for HIV

Mobility, including migration and travel, influences risk of HIV. This study examined time trends and characteristics among mobile youth (15–24 years) in rural Uganda, and the relationship between mobility and risk factors for HIV. We used data from an annual household census and population-based cohort study in the Rakai district, Uganda. Data on in-migration and out-migration were collected among youth (15–24 years) from 43 communities from 1999 to 2011 (N?=?112,117 observations) and travel among youth residents from 2003 to 2008 (N?=?18,318 observations). Migration and travel were more common among young women than young men. One in five youth reported out-migration. Over time, out-migration increased among youth and in-migration remained largely stable. Primary reasons for migration included work, living with friends or family, and marriage. Recent travel within Uganda was common and increased slightly over time in teen women (15–19 years old), and young adult men and women (20–24 years old). Mobile youth were more likely to report HIV-risk behaviours including: alcohol use, sexual experience, multiple partners, and inconsistent condom use. Our findings suggest that among rural Ugandan youth, mobility is increasingly common and associated with HIV-risk factors. Knowledge of patterns and characteristics of a young, high-risk mobile population has important implications for HIV interventions.     

The string phenomenon: an important cause of internal mammary artery graft failure

Internal mammary arteries are the grafts of choice for coronary artery bypass because they are generally free of atherosclerosis and they have high patency rates as grafts. There are, however, increasing reports of graft failure due to diffuse or distal narrowing, known as the string phenomenon. From June 1999 to October 1999, we prospectively reviewed all cases of internal mammary artery angiography at our institution. Twenty-eight cases of internal mammary artery graft failure were identified (M:F, 15:13; mean age, 62 years) out of 261 patients who had undergone internal mammary artery grafting at any institution, including ours. The mean interval between coronary artery bypass and re-angiography was 35 months. There was evidence of competitive flow in 81% (22 of 27) of patients whose internal mammary grafts had developed the string sign. This observational study supports the hypothesis that competitive flow predisposes internal mammary grafts to the string phenomenon. Given the high frequency of competitive flow situations amongst patients with internal mammary artery graft failure, caution should be exercised in the anastomosis of the internal mammary artery to recipient vessels that are less than severely stenosed.