Evaluation of COVID-19 coagulopathy; laboratory characterization using thrombin generation and nonconventional haemostasis assays.

INTRODUCTION: Patients with COVID-19 are known to have a coagulopathy with a thrombosis risk. It is unknown whether this is due to a generalized humoral prothrombotic state or endothelial factors such as inflammation and dysfunction. The aim was to further characterize thrombin generation using a novel analyser (ST Genesia, Diagnostica Stago, Asnières, France) and a panel of haematological analytes in patients with COVID-19. METHODS: Platelet poor plasma of 34 patients with noncritical COVID-19 was compared with 75 patients with critical COVID-19 (as defined by WHO criteria) in a retrospective study by calibrated automated thrombography and ELISA. Patients were matched for baseline characteristics of age and gender. RESULTS: Critical patients had significantly increased fibrinogen, CRP, interleukin-6 and D-dimer compared to noncritical patients. Thrombin generation, in critical patients, was right shifted without significant differences in peak, velocity index or endogenous thrombin potential. Tissue plasminogen activator (tPA), tissue factor pathway inhibitor (TFPI) and vascular endothelial growth factor (VEGF) were significantly increased in the critical versus noncritical patients. Critically ill patients were on haemodiafiltration (31%; heparin used in the circuit) or often received escalated prophylactic low-molecular weight heparin. CONCLUSION: These results confirm increased fibrinogen and D-dimer in critical COVID-19-infected patients. Importantly, disease severity did not increase thrombin generation (including thrombin-antithrombin complexes and prothrombin fragment 1 + 2) when comparing both cohorts; counter-intuitively critical patients were hypocoaguable. tPA, TFPI and VEGF were increased in critical patients, which are hypothesized to reflect endothelial dysfunction and/or contribution of heparin (which may cause endothelial TFPI/tPA release).

Image guidance used for large joint and bursa injections; a survey study.

Objectives: To determine the current rates of use of available image guidance modalities for large joint and bursal injections, in addition to their relationships to physician demographics.Methods: An electronic survey was sent to 3,400 members of the American Medical Society for Sports Medicine (AMSSM), examining types of guidance used for each large joint and bursal injection.Results: A total of 674 sports medicine physicians responded to the survey. Intra-articular hip and glenohumeral joint injections were more commonly performed with ultrasound guidance, while palpation-guidance was more common with all other injections. Physicians who specialized in Physical Medicine & Rehabilitation (PM&R) were more likely to use ultrasound for trochanteric bursa (p = 0.007, OR = 4.16 [1.46-11.8]), while internal medicine-, pediatrics-, and family medicine-trained physicians were more likely to use palpation guidance for at least one joint (p < 0.05). Physicians with fewer years of experience were more likely to use ultrasound for glenohumeral joint injections (p ≤ 0.002 for all age groups with less than 20 years of experience, ORs ranging from 6.3 to 9.2).Conclusion: Palpation-guidance is the most common technique used for large joint and bursal injections, other than for glenohumeral and hip joint injections. PM&R-trained physicians and those with less experience tend to use ultrasound more frequently.

Evaluating Physician Knowledge of Commonly Prescribed Inpatient Rehabilitation Unit Discharge Medication's Costs: An Observational Study.

The aim of the study was to assess physiatrists' knowledge of the cost of medications commonly prescribed at discharge from inpatient rehabilitation units across the spectrum of practice experience from residents to attending physicians. Investigators contacted 92 pharmacies across four major regions of the United States and averaged the cash price of each medication. An electronic survey was created highlighting 17 medications in which physicians estimated the cost of a medication per pill and per month for a 30-day supply. Surveys were sent to all Accreditation Council for Graduate Medical Education-accredited programs across the United States; 43 participants responded to the survey. Most respondents overestimated the cost of the medications chosen for the survey. There was no significant difference between medication cost knowledge and practice experience (P = 0.497) or postgraduate year of training (P = 0.593). This raises awareness that physiatrists may not know the cost of medications they commonly prescribe at discharge, which may have implications on patient medication compliance, quality of care, and patient satisfaction.

Intragenic complementation of hepatitis C virus NS5A RNA replication-defective alleles.

Hepatitis C virus NS5A has three structural domains, is required for RNA replication and virion assembly, and exists in hypo- and hyperphosphorylated forms. Accumulated data suggest that phosphorylation is involved in modulating NS5A functions. We performed a mutational analysis of highly conserved serine residues in the linker region between domains I and II of genotype 2a JFH1 NS5A. As with genotype 1b Con1 NS5A, we found that specific serine residues were important for efficient hyperphosphorylation of JFH1 NS5A. However, in contrast with Con1 replicons, we observed a positive correlation between hyperphosphorylation and JFH1 replicon replication. We previously demonstrated trans-complementation of a hyperphosphorylation-deficient, replication-defective JFH1 replicon. Our results suggested that the defective NS5A encoded by this replicon, while lacking one NS5A function, was capable of performing a separate replication function. In this report, we examined an additional set of replication-defective NS5A mutations in trans-complementation assays. While some behaved similarly to the S232I replicon, others displayed a unique trans-complementation phenotype, suggesting that NS5A trans-complementation can occur by two distinct modes. Moreover, we were able, for the first time, to demonstrate intragenic complementation of replication-defective NS5A alleles. Our results identified three complementation groups: group A, comprising mutations within NS5A domain I; group B, comprising mutations affecting serine residues important for hyperphosphorylation and a subset of the domain I mutations; and group C, comprising a single mutation within the C-terminal region of domain II. We postulate that these complementation groups define three distinct and genetically separable functions of NS5A in RNA replication.

In vitro activity of daclatasvir on hepatitis C virus genotype 3 NS5A.

The NS5A replication complex inhibitor daclatasvir (DCV; BMS-790052) inhibits hybrid replicons containing hepatitis C virus (HCV) genotype 3a (HCV3a) NS5A genes with 50% effective concentrations (EC(50)s) ranging from 120 to 870 pM. Selection studies with a hybrid HCV3a replicon identified NS5A residues 31 and 93 as sites for DCV-selected resistance. Our results support the potential use of DCV as a component in combination therapies for HCV3a chronic infection.

Urine analysis and protein networking identify met as a marker of metastatic prostate cancer.

PURPOSE: Metastatic prostate cancer is a major cause of death of men in the United States. Expression of met, a receptor tyrosine kinase, has been associated with progression of prostate cancer. EXPERIMENTAL DESIGN: To investigate met as a biomarker of disease progression, urinary met was evaluated via ELISA in men with localized (n = 75) and metastatic (n = 81) prostate cancer. Boxplot analysis was used to compare the distribution of met values between each group. We estimated a receiver operating characteristic curve and the associated area under the curve to summarize the diagnostic accuracy of met for distinguishing between localized and metastatic disease. Protein-protein interaction networking via yeast two-hybrid technology supplemented by Ingenuity Pathway Analysis and Human Interactome was used to elucidate proteins and pathways related to met that may contribute to progression of disease. RESULTS: Met distribution was significantly different between the metastatic group and the group with localized prostate cancer and people with no evidence of cancer (P < 0.0001). The area under the curve for localized and metastatic disease was 0.90, with a 95% confidence interval of 0.84 to 0.95. Yeast two-hybrid technology, Ingenuity Pathway Analysis, and Human Interactome identified 89 proteins that interact with met, of which 40 have previously been associated with metastatic prostate cancer. CONCLUSION: Urinary met may provide a noninvasive biomarker indicative of metastatic prostate cancer and may be a central regulator of multiple pathways involved in prostate cancer progression.

Post-collection, pre-measurement variables affecting VEGF levels in urine biospecimens.

Angiogenesis, the development and recruitment of new blood vessels, plays an important role in tumour growth and metastasis. Vascular endothelial growth factor (VEGF) is an important stimulator of angiogenesis. Circulating and urinary VEGF levels have been suggested as clinically useful predictors of tumour behaviour, and investigations into these associations are ongoing. Despite recent interest in measuring VEGF levels in patients, little is known about the factors that influence VEGF levels in biospecimens. To begin to address this question, urine samples were collected from patients with solid tumours undergoing radiotherapy and healthy volunteers. Four factors were examined for their effects on VEGF concentrations as measured by chemiluminescent immunoassay: time from sample collection to freezing, number of specimen freeze-thaw cycles, specimen storage tube type and the inclusion or exclusion of urinary sediment. The results of this study indicate that time to freeze up to 4 hrs, number of freeze-thaw cycles between one and five, and different types of polypropylene tubes did not have statistically significant effects on measured urinary VEGF levels. Urinary sediment had higher VEGF levels than supernatant in five of six samples from healthy patients. It is not clear whether there is an active agent in the sediment causing this increase or if the sediment particles themselves are affecting the accuracy of the assay.Therefore, we recommend centrifuging urine, isolating the supernatant, and freezing the sample in polypropylene microcentrifuge tubes or cryogenic vials within 4 hrs of collection.In addition, we recommend the use of samples within five freeze-thaw cycles.