Standardized direct oral anticoagulants prescription for treatment of acute venous thromboembolism in the emergency department: A quality improvement initiative.

INTRODUCTION: Direct oral anticoagulants (DOACs) are commonly used for the treatment and prevention of venous thromboembolism (VTE). However, prescription errors with DOACs can lead to patient dissatisfaction and harm. This study aimed to evaluate the impact of a standardized prescription for DOACs for VTE on prescription appropriateness. MATERIALS AND METHODS: The study included patients discharged from the Emergency Department (ED) with a DOAC prescription for an acute VTE. A standardized prescription tool was developed and implemented, and patients were divided into pre- and post-intervention groups. The appropriateness of prescriptions was assessed using the Medication Appropriateness Index (MAI). RESULTS: A total of 161 patients with VTE were included in the study. The post-intervention group showed a significant increase in prescriptions with an MAI rating of "appropriate" and a decrease in ratings of "inappropriate." Improvements were observed in loading dose duration, maintenance dose frequency and duration, and inclusion of necessary drug coverage codes. CONCLUSION: The implementation of a standardized prescription for DOACs in the management of VTE in the ED significantly improved medication appropriateness and reduced inappropriate prescriptions. Standardized prescriptions have the potential to enhance patient safety and optimize care by providing clear and uniform guidance to healthcare providers. Further research is needed to explore the effectiveness of medication prescription software systems in real-world clinical settings to improve prescribing practices.

Management of anticoagulation in pregnant women with venous thromboembolism: An international survey of clinical practice.

INTRODUCTION: Venous thromboembolism (VTE) is an important cause of maternal morbidity and mortality. During pregnancy, VTE is treated with low-molecular-weight-heparin (LMWH). Studies assessing the optimal duration and peripartum management of therapeutic anticoagulation are lacking. This survey aimed to assess clinician practices for the management of anticoagulation in pregnant women with acute VTE. METHODS: An electronic survey consisting of clinical scenarios addressing anticoagulation management for VTE in pregnancy was created. The target sample was clinicians likely to be involved in the management of pregnant women with acute VTE. The survey completion rate and proportion of individuals selecting a response were determined. RESULTS: 96 respondents completed the survey including general internists (56.3%), hematologists (21.9%), and obstetricians (6.3%). In the management of a VTE in first or second trimester, most respondents preferred therapeutic LMWH until 6 weeks postpartum. In the first and second trimester, 48.0% and 37.5% of respondents, respectively, opted to reduce the dose of anticoagulation after 3 or 6 months. 29.2% of physicians opted for bridging with intravenous heparin around delivery when treating a VTE in the third trimester. 73.0% perceived an increased risk of clinically relevant non-major bleeding associated with the use of therapeutic anticoagulation in the peripartum and postpartum periods. CONCLUSIONS: The survey highlights a wide variability of practice in the management of therapeutic anticoagulation in pregnancy. Larger scale studies with relevant clinical outcomes including thrombosis and bleeding risks are needed to inform clinical practice.

Both cIAP1 and cIAP2 regulate TNFalpha-mediated NF-kappaB activation.

The cellular inhibitor of apoptosis 1 and 2 (cIAP1 and cIAP2) proteins have been implicated in the activation of NF-kappaB by TNFalpha; however, genetic deletion of either cIAP1 or 2 did not support a physiologically relevant role, perhaps because of functional redundancy. To address this, we used combined genetic and siRNA knockdown approaches and report that cIAP1 and 2 are indeed critical, yet redundant, regulators of NF-kappaB activation upon TNFalpha treatment. Whereas NF-kappaB was properly activated by TNFalpha in cultured and primary cells deficient in either cIAP1 or 2, removal of both cIAPs severely blunted its activation. After treatment with TNFalpha, cIAP1 and 2 were rapidly recruited to the TNF receptor 1, along with the adapter protein TNF receptor associated factor 2. Importantly, either cIAP1 or 2 was required for proper TNF receptor 1 signalosome function. In their combined absence, polyubiquitination of receptor interacting protein 1, an upstream event necessary for NF-kappaB signaling, was attenuated. As a result, phosphorylation of the inhibitor of kappaB kinase beta was diminished, and signal transduction was severely blunted. Consequently, cells missing both cIAP1 and 2 were sensitized to TNFalpha-mediated apoptosis. Collectively, these data demonstrate that either cIAP1 or 2 is required for proper Rip1 polyubiquitination and NF-kappaB activation upon TNFalpha treatment.

The effect of anti-neoplastic drugs on murine acquired immunodeficiency syndrome.

The murine acquired immunodeficiency syndrome (MAIDS) is associated with proliferation of target cells that have been infected by a defective retrovirus. To control the growth of this primary neoplasia, virus-inoculated mice were treated with anti-neoplastic drugs. Paradoxically, cyclophosphamide, which is also immunosuppressive, was very effective in preventing the appearance and progression of the disease, in restoring a normal T cell function, and in depleting the number of infected target cells. This result suggests that the proliferating infected target cells were responsible for the immunodeficiency.

Immunodeficiency and clonal growth of target cells induced by helper-free defective retrovirus.

The murine acquired immunodeficiency syndrome is induced by a defective retrovirus. To study the role of virus replication in this disease, helper-free stocks of defective Duplan virus were produced. These stocks were highly pathogenic in absence of detectable replicating murine leukemia viruses (MuLVs) other than xenotropic MuLV. They induced expansion of the infected cell population (over 1000-fold), and this cell expansion was oligoclonal in origin and, most likely, arose through cell division. These results suggest that this defective virus is oncogenic, inducing a primary neoplasia associated with an acquired immunodeficiency syndrome as a paraneoplastic syndrome. These data emphasize the need to determine whether virus replication is necessary for the progression of other immunodeficiency diseases, including acquired immunodeficiency syndrome, and whether these diseases also represent paraneoplastic syndromes.

Novel approach for detecting prohibited species-specific central nervous system tissue contamination in meat by one-step real-time reverse transcriptase PCR.

The dissemination of prohibited species-specific central nervous system (CNS) tissue contamination in meat must be tracked to mitigate human health risk associated with bovine spongiform encephalopathy. The efficiency of compliance monitoring and risk control measures taken by concerned regulatory authorities at meat production facilities to avoid such contamination depends on the ability to detect CNS tissue with a reliable and adequately sensitive quantitative method. A rapid and convenient one-step real-time quantitative reverse transcriptase PCR (qRT-PCR) assay was developed based on the absolute quantification of glial fibrillary acidic protein (GFAP) mRNA as a marker for CNS tissue contamination in meat. The GFAP RNA quantity corresponding to a percentage of CNS tissue in artificially spiked meat was determined using an appropriate in vitro transcribed target GFAP RNA as a calibration standard in the assay. The assay had a linear dynamic range of 10(2) to 10(9) copies of target RNA and was able to detect 0.01% CNS contamination in meat. Further evaluation consisted of an analysis of 272 random meat cuts from carcasses and 109 ground meat samples received from a federally inspected abattoir and two meat processing facilities, respectively, over a 5-month period. The analyzed samples were all negative for CNS tissue contamination at an arbitrarily set lower threshold of 0.025%. Overall, the newly developed one-step qRT-PCR may be useful as an objective quantitative compliance monitoring tool and for setting an acceptable low tolerance threshold for such contamination in meat.

Specific expression of the human CD4 gene in mature CD4+ CD8- and immature CD4+ CD8+ T cells and in macrophages of transgenic mice.

The CD4 protein plays a critical role in the development and function of the immune system. To gain more insight into the mechanism of expression of the human CD4 gene, we cloned 42.2 kbp of genomic sequences comprising the CD4 gene and its surrounding sequences. Studies with transgenic mice revealed that a 12.6-kbp fragment of the human CD4 gene (comprising 2.6 kbp of 5' sequences upstream of the transcription initiation site, the first two exons and introns, and part of exon 3) contains the sequences required to support the appropriate expression in murine mature CD4+ CD8- T cells and macrophages but not in immature double-positive CD4+ CD8+ T cells. Expression in CD4+ CD8+ T cells was found to require additional regulatory elements present in a T-cell enhancer fragment recently identified for the murine CD4 gene (S. Sawada and D. R. Littman, Mol. Cell. Biol. 11:5506-5515, 1991). These results suggest that expression of CD4 in mature and immature T-cell subsets may be controlled by distinct and independent regulatory elements. Alternatively, specific regulatory elements may control the expression of CD4 at different levels in mature and immature T-cell subsets. Our data also indicate that mouse macrophages contain the regulatory factors necessary to transcribe the human CD4 gene.

Implementing a routine flow cytometry assay for nucleated red blood cell counts in cord blood units.

INTRODUCTION: As required by standards organizations, Héma-Québec Cord Blood Bank performs enumeration of nucleated red blood cells (NRBCs) in cord blood units (CBUs). This study presents the validation and implementation approaches developed to transfer the routine NRBC enumeration from the manual blood film method to a flow cytometric assay. METHODS: The flow cytometry method was adapted from Tsuji (Cytometry, 37, 1999, 291). This assay was validated to assess the specificity, detection limit, repeatability, and reproducibility of the method, including interoperator and interlaboratory testing. Finally, postimplementation follow-up and adjustments were performed for CBU over a 7-month period. RESULTS: Blood film and flow cytometry NRBC enumerations showed a strong correlation (n = 40; Pearson's r correlation = 0.90). Validation was successful as exemplified by the correlation in interlaboratory testing (n = 30; r = 0.98). During implementation, our routine laboratory analyses revealed that CBU with low NRBC content (≤2%), representing 26% of all CBU tested, resulted in 15% of repeated reading and/or staining and was the principal source of nonconformity. Small adjustments in the standard operating procedures (SOPs), including a fixed 200-event setting in the NRBC gate for the second reading of the replicates, have completely solved this issue. CONCLUSION: Flow cytometric NRBC enumerations, now implemented in Héma-Québec Public Cord Blood Bank, is an improvement in the efficiency of our operations by integrating the count for NRBC into our flow cytometry platform.

Use of a phage vector for rapid synthesis and cloning of single-stranded cDNA.

We have developed a technique for synthesis of single stranded complementary DNA (ss cDNA) using specifically designed phage ssDNA as vector primer. This vector (pPBS27) was constructed by introducing a poly(dT) tail adjacent to the XbaI site of pTZ18R, which can exist either as a plasmid in Escherichia coli or as a ssDNA phage. The pPBS27 phage vector is linearized with XbaI using a restriction-site-directed fragment and used to anneal a mixture of poly(A) + RNA for cDNA synthesis by reverse transcriptase. The RNA is then hydrolysed with NaOH and a poly(dG) tail added to the 3' end of the vector-cDNA with terminal transferase. The linear hybrid ssDNA is then closed by annealing with a 15-mer site-directed fragment oligodeoxynucleotide molecule and ligated with T4 DNA ligase. Almost 10(5) E. coli transformants per microgram of vector primer can be obtained in two days.

Contractile and histochemical properties of young and old medial gastrocnemius muscle after suspension hypokinesia/hypodynamia.

In order to determine if the atrophic process was different in the young and in the aged fast twitch medial gastrocnemius muscle, in which the aging process is clearly apparent, hindlimb hypokinesia/hypodynamia (H/H) was induced on rats. After 3 weeks, we measured speed-related indices, tension indices and tension producing capacities. Fiber type composition and fiber cross-sectional area were also determined. The body weight and muscle weight decreased. The ratio of muscle weight to body weight was greater in the 3-month-old control group than in the 22-month-old group. However, the ratio was not altered by H/H. In both groups, contraction time, half relaxation time and twitch tension did not vary with H/H but maximally developed tension and force generated per gram of muscle were decreased. For the histochemical analysis, the medial gastrocnemius muscle was divided in its red and white parts. Fiber type composition remained the same after H/H in both parts of the muscle and in both groups of age except for an increase of the FOG fiber type proportion in the red part of the old group. Fiber cross-sectional area increased with age and decreased after the suspension except for the fast fibers in the white part of the aged muscle. The most affected group in fiber cross-sectional (SO-FOG-FG) area after suspension was the white part of medial gastrocnemius in the young group. The suspension atrophy appeared to be independent of age in the medial gastrocnemius at the contractile level whereas atrophy was greater at the histochemical level in the young muscle. With age, fiber cross-sectional area increased particularly in the red part of the muscle.

Enzymatic adaptations to suspension hypokinesia in skeletal muscle of young and old rats.

Hindlimb hypokinesia was induced in young and old rats. After 3 weeks, the activities of five enzymes have been measured in soleus and medial gastrocnemius muscles. Soleus showed increased activity of hexokinase and decreased activity of phosphofructokinase, lactate dehydrogenase and malate dehydrogenase in both groups. The activity of 3-hydroxyacyl-CoA-dehydrogenase was decreased in the old muscle. Medial gastrocnemius showed decreased activity of phosphofructokinase and lactate dehydrogenase in both groups. The activity of 3-hydroxyacyl-CoA-dehydrogenase was decreased in the old muscle whereas hexokinase increased its activity in the young one. It is concluded that suspension hypokinesia results in changes at the enzymatic level. These changes appear to be related to the age of the muscle and to its fibre composition.

Influence of electrical stimulation on enzymatic activity of old rat muscles during hindlimb suspension.

1. The purpose of this study was to determine the effects of electrical stimulation on the muscles of older rats (21-24 months old) during suspension hypokinesia/hypodynamia. The rats' hindlimbs were suspended for three weeks, after which the activities of three enzymes were measured in the extensor digitorum longus, the medial gastrocnemius and the soleus muscles. 2. The activity of phosphofructokinase decreased in the soleus muscle and in the white part of the medial gastrocnemius muscle. Malate dehydrogenase and beta-hydroxyacyl-CoA-dehydrogenase showed decreased activity in the red part of the medial gastrocnemius muscle. 3. When electrical stimulation was applied, the activity of the glycolytic enzyme phosphofructokinase increased substantially in all studied muscles. The activity of the malate dehydrogenase and beta-hydroxyacyl-CoA-dehydrogenase enzymes was not influenced by electrical stimulation in the majority of the studied muscles. 4. These results suggest that our electrical stimulation model not only prevents loss of phosphofructokinase activity of aged muscles but increases the activity of anaerobic glycolysis metabolism without affecting the oxidative and fatty acid pathways.

Effects of hindlimb suspension on contractile properties of young and old rat muscles and the impact of electrical stimulation on the recovery process.

The purpose of this study was to investigate the effects of hindlimb suspension (HS) on contractile properties of skeletal muscles of young and old rats and to determine the impact of electrical stimulation (ES) on the quality and degree of recovery of these muscles. After 21 days of HS, young soleus (SOL) muscle became faster, but there was no impact on young extensor digitorum longus (EDL) muscle. Twitch tension (Pt) decreased 61% in young and 70% in old SOL muscles. Specific tetanic tension (Po) decreased 53% in young and 64% in old SOL muscles, but again there was no impact on EDL muscle. After a 14-day period of recovery, contraction time (CT), half-relaxation time (RT1/2), Pt and Po returned to control group values in both young and old SOL muscles. Measurements of the contractile properties of young and old skeletal rat muscles showed ES sometimes to be beneficial but also sometimes to be harmful. A 14-day period of recovery, with or without ES, seemed sufficient for many variables to return to control group values.

Expression kinetics of the transcript and product of the UL28 homologue of bovine herpesvirus 1.

We report that the bovine herpesvirus 1 (BHV1) UL28 ORF, a homologue of the herpes simplex virus (HSV) UL28 gene, represents a functional gene encoding a viral specific protein. The BHV1 UL28 ORF, located at positions 53058-->55538 of the viral genome, encodes a viral specific transcript of 3.4 kb detected at 6 h post-infection (p.i.) after which its levels accumulated up to 12 h p.i. and then remained constant up to 24 h p.i. Transcription of the BHV1 UL28 was determined to initiate 95 bases upstream from the ORF's initiating codon, which corresponds to 33 nucleotides downstream from a putative TATA box. A BHV1 UL28 specific antiserum, generated against a T7-Tag/UL28 fusion protein expressed in E. coli, specifically reacted with a 100 kDa protein in Western blots of BHV1-infected protein cell lysates. The expression kinetics of the protein was delayed by 6 h relative to that of its transcript suggesting that the gene is regulated at the translational level. In contrast to the HSV and pseudorabies virus UL28 genes, which belong to viral genes of the early (beta) class, that of BHV1 was unambiguously classified as a gamma2 gene. Further studies will be required to determine whether these kinetic differences have any functional implications.

The bovine herpesvirus type 1 major tegument protein VP8 expressed in recombinant vaccinia virus does not induce significant immunity in mice.

We previously reported the characterization of the gene encoding the bovine herpesvirus type 1 (BHV-1) major tegument protein VP8. With the aim of defining the immunological properties of this protein, we constructed a recombinant vaccinia virus (VV-VP8) in which expression of the VP8 gene was regulated by the P7.5 early/late promoter. Since the sequence of the VP8 gene contained a TTTTTNT motif known to serve as a transcription termination signal of vaccinia virus genes of the early class, a second recombinant (VV-VP8-Mut) in which this signal was modified by site-directed mutagenesis was created. Characterization of the recombinant viruses revealed that truncated VP8 mRNA and protein (69 kDa) were synthesized in VV-VP8 infected cells, whereas cells infected with VV-VP8-Mut produced a protein which was undistinguishable from that of the BHV-1 encoded protein (92-94 kDa). Immunization of BALB/c mice (H-2d) with VV-VP8-Mut induced a low VP8-specific antibody response whereas no specific response was induced in VV-VP8 inoculated mice. The low humoral response elicited was similar in C57BL/6 (H-2b) and C3H (H-2k) mice. Furthermore, immunization of mice with VV-VP8-Mut did not induce a BHV-1-specific lymphoproliferation in the three mice strains examined. Our results contrast with a recent study showing that immunization of calves with purified VP8 stimulated both T cell proliferation and antibody production.

Linkage results on 11Q21-22 in Eastern Quebec pedigrees densely affected by schizophrenia.

The 11q21-22 region is of interest for schizophrenia because several candidate genes are located in this section of the genome. The 11q21-22 region, including DRD2, was surveyed by linkage analysis in a sample (N = 242) made of four large multigenerational pedigrees densely affected by schizophrenia (SZ) and eight others by bipolar disorder (BP). These pedigrees were ascertained in a large area of Eastern Quebec and Northern New Brunswick and are still being extended. Family members were administered a "consensus best-estimate diagnosis procedure" (DSM-III-R criteria) blind to probands and relatives' diagnosis and to pedigree assignment (SZ or BP). For linkage analysis, 11 microsatellite polymorphism (CA repeat) markers, located at 11q21-22, and comprising DRD2, were genotyped. Results show no evidence of a major gene for schizophrenia. However, a maximum lod score of 3.41 at the D11S35 locus was observed in an affected-only analysis of one large SZ family, pedigree 255. Whether or not the positive linkage trend in pedigree 255 reflects a true linkage for a small proportion of SZ needs to be confirmed through the extension of this kindred and through replication.

Evidence that the amino acid region 124-203 of glycoprotein G from the respiratory syncytial virus (RSV) constitutes a major part of the polypeptide domain that is involved in the protection against RSV infection.

The first 230 residues of the 298-amino acid glycoprotein G of respiratory syncytial virus (RSV) are sufficient to confer complete resistance to challenge with live RSV, whereas the first 180 residues completely failed (Olmsted et al. (1989) J. Virol. 63, 411-420). The characterization of a protective epitope corresponding to the amino acid region 174-187 of the G protein (Trudel et al. (1991) Virology 185, 749-757) suggests that interruption of this region in the 180 residue truncated polypeptide may be responsible for its inability to confer protection and consequently that the 174-187 region may play a major role in the protection effected by the protein G. To support these hypotheses, we examined the ability of the amino acid region 124-203 of glycoprotein G to confer protection. The corresponding peptide was expressed as a non-fusion protein in a recombinant vaccinia virus designated VG27. Immunization of BALB/c mice with this recombinant efficiently induced the production of antibodies capable of recognizing both the parental glycoprotein G and peptide 174-187. Furthermore, upon challenge with RSV, a significant decrease of infectious particles was found in the lungs of mice immunized with VG27 as compared with non-immunized mice. Our results suggest that the 124-203 amino acid region of the RSV G protein constitutes a major part of the domain involved in protection.

Somatostatin and adrenocorticotrophic hormone like immunoreactivity in small cell carcinoma of the lung.

The immunocytological detection of adrenocorticotrophic hormone (ACTH) and somatotropin release inhibitor factor (SRIF) like immunoreactivity was carried out on tumour cells from bronchial brush smears in 39 cases of lung tumours. Results obtained were compared with the cytological and histological diagnosis and confirmed the high incidence of ACTH synthesis by malignant bronchial carcinoma cells: the same phenomenon also seems to occur for somatostatin. The concomitant detection of ACTH and SRIF like immunoreactivity seems to be highly suggestive of small cell carcinoma and indicates that the immunocytological detection of hormones carried out at the same time as cytological examination can improve the accuracy of the diagnosis.

Detection of horses infected naturally with equine infectious anemia virus by nested polymerase chain reaction.

A nested polymerase chain reaction (PCR) amplifying a region of the gag gene of equine infectious anemia virus (EIAV) was developed for the rapid and direct detection of proviral DNA from the peripheral blood of naturally infected horses and was compared with the Coggins test. DNA prepared from white blood cells of 122 field horses from 15 stables with reported cases of EIAV and one seronegative stable were analysed. Amplifications of expected size fragments were obtained by nested PCR for 88 horses using two different sets of primers targeting the gag region. The specificity of the amplified products was confirmed by hybridization using a digoxigenin-labeled probe. Gag-nested PCR-restriction fragment length polymorphism analysis distinguished two different subtypes of gag gene, A and B. Subtype A was found to be the most prevalent among the infected horses that were tested. The PCR-gag amplified sequence of subtype A shared 84.6% nucleotide and 93% deduced amino acid sequence identities with the prototype Wyoming strain whereas subtype B sequence was almost 100% identical to the prototype. Sequence analysis of gag subtype A suggests the presence of a novel EIAV variant among infected horses in Canada. The nested PCR assay developed in the present study detected more EIAV positive animals and was found as specific as the agar gel immunodiffusion (Coggins) assay and offers great potential a diagnostic test for the detection of EIAV infections in field horses.