Rewriting Nature’s Assembly Manual for a ssRNA Virus

Satellite Tobacco Necrosis Virus (STNV) is one of the smallest viruses known. Its genome encodes only its coat protein (CP) subunit relying on the polymerase of its helper virus TNV for replication. The genome contains a cryptic set of dispersed assembly signals in the form of stem-loops that each present a minimal CP binding motif -A.X.X.A- in the loops. The genomic fragment encompassing nucleotides 1-127 is predicted to contain five such Packaging Signals (PSs). We have used mutagenesis to determine the critical assembly features in this region. These include the CP binding motif, the relative placement of PS stem-loops, their number and their folding propensity. CP binding has an electrostatic contribution but assembly nucleation is dominated by the recognition of the folded PSs in the RNA fragment. Mutation to remove all –A.X.X.A- motifs in PSs throughout the genome yields an RNA that is unable to assemble efficiently. In contrast, when a synthetic 127nt fragment encompassing improved PSs is swapped onto the RNA otherwise lacking CP recognition motifs assembly is partially restored although the virus-like particles created are incomplete, implying that PSs outside this region are required for correct assembly. Swapping this improved region into the wild-type STNV1 sequence results in a better assembly substrate than the viral RNA, producing complete capsids and outcompeting the wild-type genome in head-to-head competition. These data confirm details of the PS-mediated assembly mechanism for STNV, and identify an efficient approach for production of stable viruslike particles encapsidating non-native RNAs or other cargoes

OMERACT Definitions for Ultrasonographic Pathology and Elementary Lesions Of Rheumatic Disorders Fifteen Years On

Objective. The Outcome Measures in Rheumatology (OMERACT) ultrasound (US) working group (WG) operates research activities for the validation of US as an outcome measurement instrument according to the Filter 2.0 framework Methods. From the onset of the WG research in 2005 through now, original publications on definitions and scoring systems for pathophysiological manifestations and elementary lesions of various rheumatic disorders were reviewed Results. Definitions and scoring systems according to new terminology are provided Conclusions. We have redefined OMERACT definitions of US pathology and elementary lesions as well as scoring systems which are now proposed for OMERACT approval for application in clinical trial

Practice of ultrasound-guided arthrocentesis and joint injection, including training and implementation, in Europe: results of a survey of experts and scientific societies

Objectives. To document the practice and training opportunities of US-guided arthrocentesis and joint injection (UGAJ) among rheumatologists in the member countries of the European League Against Rheumatism (EULAR). Methods. An English-language questionnaire, containing questions on demographics, clinical and practical aspects of UGAJ, training options in UGAJ for rheumatologists, UGAJ education in the rheumatology training curriculum and other structured education programmes in UGAJ was sent to three different groups: (i) all national rheumatology societies of EULAR; (ii) all national societies of the European Federation of Societies for Ultrasound in Medicine and Biology (EFSUMB); and (iii) 22 senior rheumatologists involved in EULAR musculoskeletal US training from 14 European countries, who were also asked to circulate the questionnaire among relevant colleagues. Results. Thirty-three (75%) of 44 countries responded to the questionnaire (61.3% of national rheumatology societies, 25% of the national US societies and 100% of expert ultrasonographers). In the majority of countries (85%) 80%) rate of rheumatologists performing conventional joint injection in most of the surveyed countries. The reported variations in practice and the lack of available structured training programmes for trainees in most countries indicates the need for standardization in areas including training guideline

Self-directed learning of basic musculoskeletal ultrasound among rheumatologists in the United States

Objective Because musculoskeletal ultrasound (MSUS) is highly user dependent, we aimed to establish whether non-mentored learning of MSUS is sufficient to achieve the same level of diagnostic accuracy and scanning reliability as has been achieved by rheumatologists recognized as international experts in MSUS. Methods A group of 8 rheumatologists with more experience in MSUS and 8 rheumatologists with less experience in MSUS participated in an MSUS exercise to assess patients with musculoskeletal abnormalities commonly seen in a rheumatology practice. Patients' established diagnoses were obtained from chart review (gout, osteoarthritis, rotator cuff syndrome, rheumatoid arthritis, and seronegative arthritis). Two examining groups were formed, each composed of 4 less experienced and 4 more experienced examiners. Each group scanned 1 predefined body region (hand, wrist, elbow, shoulder, knee, or ankle) in each of 8 patients, blinded to medical history and physical examination. Structural abnormalities were noted with dichotomous answers, and an open-ended answer was used for the final diagnosis. Results Less experienced and more experienced examiners achieved the same diagnostic accuracy (US-established diagnosis versus chart review diagnosis). The interrater reliability for tissue pathology was slightly higher for more experienced versus less experienced examiners (Κ = 0.43 versus Κ = 0.34; P = 0.001). Conclusion Non-mentored training in MSUS can lead to the achievement of diagnostic accuracy in MSUS comparable to that achieved by highly experienced international experts. Reliability may increase slightly with additional experience. Further study is needed to determine the minimal training requirement to achieve proficiency in MSUS.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/65036/1/20063_ftp.pd

Genome wide mapping reveals PDE4B as an IL-2 induced STAT5 target gene in activated human PBMCs and lymphoid cancer cells

IL-2 is the primary growth factor for promoting survival and proliferation of activated T cells that occurs following engagement of the Janus Kinase (JAK)1-3/and Signal Transducer and Activator of Transcription (STAT) 5 signaling pathway. STAT5 has two isoforms: STAT5A and STAT5B ( commonly referred to as STAT5) which, in T cells, play redundant roles transcribing cell cycle and survival genes. As such, inhibition of STAT5 by a variety of mechanisms can rapidly induce apoptosis in certain lymphoid tumor cells, suggesting that it and its target genes represent therapeutic targets to control certain lymphoid diseases. To search for these molecules we aligned IL-2 regulated genes detected by Affymetrix gene expression microarrays with the STAT5 cistrome identified by chip-on-ChIP analysis in an IL-2-dependent human leukemia cell line, Kit225. Select overlapping genes were then validated using qRT(2)PCR medium-throughput arrays in human PHA-activated PBMCs. Of 19 putative genes, one key regulator of T cell receptor signaling, PDE4B, was identified as a novel target, which was readily up-regulated at the protein level (3 h) in IL-2 stimulated, activated human PBMCs. Surprisingly, only purified CD8+ primary T-cells expressed PDE4B, but not CD4+ cells. Moreover, PDE4B was found to be highly expressed in CD4+ lymphoid cancer cells, which suggests that it may represent a physiological role unique to the CD8+ and lymphoid cancer cells and thus might represent a target for pharmaceutical intervention for certain lymphoid diseases

Musculoskeletal ultrasound including definitions for ultrasonographic pathology

Ultrasound (US) has great potential as an outcome in rheumatoid arthritis trials for detecting bone erosions, synovitis, tendon disease, and enthesopathy. It has a number of distinct advantages over magnetic resonance imaging, including good patient tolerability and ability to scan multiple joints in a short period of time. However, there are scarce data regarding its validity, reproducibility, and responsiveness to change, making interpretation and comparison of studies difficult. In particular, there are limited data describing standardized scanning methodology and standardized definitions of US pathologies. This article presents the first report from the OMERACT ultrasound special interest group, which has compared US against the criteria of the OMERACT filter. Also proposed for the first time are consensus US definitions for common pathological lesions seen in patients with inflammatory arthritis