Presenting Baseline Coagulation of Infra Renal Ruptured Abdominal Aortic Aneurysm: A Systematic Review and Pooled Analysis.

BACKGROUND: The incidence of coagulopathy in patients presenting with rAAA is not clear. The lack of high-quality evidence has led to various speculations, reliance on anecdotal experience, and suggestions about their appropriate haemostatic resuscitation. The aim of this systematic review is to establish the baseline coagulation status of infra renal ruptured abdominal aortic aneurysms (rAAA) against defined standards and definitions. METHODS: An electronic search of literature in Medline, CINHAL, Scopus Embase, and Cochrane library was performed in accordance with the PRISMA guidelines. Quality assessment of articles was performed using the Oxford critical appraisal skills programme (CASP) and their recommendation for practice was examined through National Institute for Health and Care Excellence (NICE). Information on platelet count, international normalisation ratio (INR), activated partial prothrombin time (aPTT), prothrombin time (PT) fibrinogen and D-dimer was extracted, and pooled analysis was performed in accordance with the definition of coagulopathy and its subtypes. Pooled prevalence of coagulopathies and 95% CI were estimated with a variance weighted random effects model. RESULTS: Seven studies, comprising 461 patients were included in this systematic review. Overall weighted prevalence of coagulopathy was 12.3% (95% CI 10.7-13.9), 11.7% for INR (95% CI 1-31.6), 10.1% for platelet count (95% CI 1-26.8), and 11.1% for aPTT (95% CI 0.78-31). Fibrinogen serum concentration level was normal in 97%, and 46.2% (n = 55) of patients had elevated D-dimer. Only 6% of the entire population demonstrated significant coagulopathy. DIC was noted in 2.4% of the population. CONCLUSION: This first systematic review of literature on baseline coagulation of rAAAs suggests that the majority of these patients do not present with coagulopathy and only a minor proportion of patients present with significant coagulopathy.

Does the Presence of Scrapie Affect the Ability of Current Statutory Discriminatory Tests To Detect the Presence of Bovine Spongiform Encephalopathy?

Current European Commission (EC) surveillance regulations require discriminatory testing of all transmissible spongiform encephalopathy (TSE)-positive small ruminant (SR) samples in order to classify them as bovine spongiform encephalopathy (BSE) or non-BSE. This requires a range of tests, including characterization by bioassay in mouse models. Since 2005, naturally occurring BSE has been identified in two goats. It has also been demonstrated that more than one distinct TSE strain can coinfect a single animal in natural field situations. This study assesses the ability of the statutory methods as listed in the regulation to identify BSE in a blinded series of brain samples, in which ovine BSE and distinct isolates of scrapie are mixed at various ratios ranging from 99% to 1%. Additionally, these current statutory tests were compared with a new in vitro discriminatory method, which uses serial protein misfolding cyclic amplification (sPMCA). Western blotting consistently detected 50% BSE within a mixture, but at higher dilutions it had variable success. The enzyme-linked immunosorbent assay (ELISA) method consistently detected BSE only when it was present as 99% of the mixture, with variable success at higher dilutions. Bioassay and sPMCA reported BSE in all samples where it was present, down to 1%. sPMCA also consistently detected the presence of BSE in mixtures at 0.1%. While bioassay is the only validated method that allows comprehensive phenotypic characterization of an unknown TSE isolate, the sPMCA assay appears to offer a fast and cost-effective alternative for the screening of unknown isolates when the purpose of the investigation was solely to determine the presence or absence of BSE.

Prions are secreted in milk from clinically normal scrapie-exposed sheep.

The potential spread of prion infectivity in secreta is a crucial concern for prion disease transmission. Here, serial protein misfolding cyclic amplification (sPMCA) allowed the detection of prions in milk from clinically affected animals as well as scrapie-exposed sheep at least 20 months before clinical onset of disease, irrespective of the immunohistochemical detection of protease-resistant PrP(Sc) within lymphoreticular and central nervous system tissues. These data indicate the secretion of prions within milk during the early stages of disease progression and a role for milk in prion transmission. Furthermore, the application of sPMCA to milk samples offers a noninvasive methodology to detect scrapie during preclinical/subclinical disease.

Unexpected diminished innervation of epidermis and dermoepidermal junction in lichen amyloidosus.

BACKGROUND: Lichen amyloidosus is a localized, chronic, pruritic skin disease characterized by deposition of amyloid in the papillary dermis. The pathogenesis of the pruritus of lichen amyloidosus is largely unknown. OBJECTIVES: To determine any change in the nerve fibre density in lichen amyloidosus lesions as an explanation for itch. METHODS: Using an antibody to protein gene product (PGP) 9.5, the immunohistochemical analysis of the skin biopsies of 30 Hispanic patients with clinicopathologically proven lichen amyloidosus and of 11 healthy Hispanic controls matched for age, sex and site was performed. RESULTS: Unexpectedly, the mean amount of PGP9.5 stain, a measure for nerve fibre amount, for the healthy controls was higher than the lichen amyloidosus group both in the epidermis (P < 0.0019) and dermoepidermal junction (P < 0.0064). No change was observed in the papillary dermis. Furthermore, the proportion of area covered by PGP9.5 showed a significant decrease in the epidermis (P < 0.0024) and dermoepidermal junction (P < 0.0075) in lichen amyloidosus compared with healthy controls. Age, gender and body site were found not to be influencing factors in nerve fibre amounts in lichen amyloidosus samples. CONCLUSIONS: We speculate that the severe pruritus observed in lichen amyloidosus might be the result of the hypersensitivity of the remaining nerve fibres as a response to an unexplained neurodegeneration of the absent nerve fibres.

An in vitro model for assessing effective scrapie decontamination.

Scrapie infectivity enters the environment via a multiplicity of routes from infected animals. Environmentally associated scrapie persists on farms when infected animals have been removed and is particularly resistant to disinfection. Infectivity within the farm is not adequately removed by current recommended guidelines for farm decontamination. We describe an in vitro method for modelling decontamination, specifically the removal of scrapie prions from the surface of concrete fomites within buildings that have housed scrapie infected animals. Concrete that had been spiked with low amounts of a diluted scrapie positive brain homogenate was sampled before and after decontamination. Extracts were used to seed a semi-quantitative serial protein misfolding cyclic amplification assay (sPMCA). We demonstrate that methods currently recommended for prion decontamination result in inadequate reduction of prion seeding activity within this in vitro assay. Effective treatment was achieved using repeat dosing of surfaces with 20,000ppm available chlorine for 4h.

HIV-1 Gag binds specifically to RNA stem-loops in the 5' leader sequence.

GST-Gag(p55) binds specifically to HIV-1 RNA sequences 1-406, in vitro, with a Kd of about 50 nM. This RNA transcript contains a number of stem loop (SL) structures. The binding is due to the Gag moiety of the fusion protein, not GST. There is a high affinity binding site for Gag in an RNA containing nucleotides 325-362. SL4 is predicted by both biochemical studies and computer folding to be located between nucleotides 335 and 358. An RNA transcript ending at nucleotide 335 does not bind Gag. The deletion of nucleotides 334-358 from HIV-1 RNAs does not affect Gag binding. Digestions with RNase V1 and T1 show that nucleotides 297-300 in SL2, 310, 312, 313, 315, 317, 318, 325 in SL3, and 342 and 343 in SL4 are protected in the presence of Gag. The cleavage of nucleotides 348-351 in SL4 by RNAse V1 is enhanced by Gag binding. At least two Gag binding sites are therefore located in the leader RNA. Those located 5' of nucleotide 335 require the presence of additional 3' sequences.

Generation and characterisation of monoclonal antibodies to Rainbow trout (Oncorhynchus mykiss) prion protein.

We report the production and characterisation of three monoclonal antibodies to the prion protein (PrP) of Rainbow trout (Oncorhynchus mykiss), a piscine protein with characteristic structural features common to mammalian prion protein. All of the antibodies were used to detect PrP in ELISA, Western blot and by immunohistochemistry. The antibodies showed specificity for certain genera of the Salmonidae, binding to PrP of Rainbow trout and Atlantic salmon (Salmo salar) but not to that from Arctic char (Salvelinus alpinus). Using the immunoreagents in Western blots, we demonstrated that O. mykiss PrP protein is a 64 kDa protein present in brain, spinal chord and optic nerve. PrP was not detected in a range of peripheral tissues: eye, heart, stomach, intestine, liver, kidney, spleen, muscle and skin. Furthermore, PrP could be detected in all brain regions studied: optic lobe, cerebrum/olfactory lobe, cerebellum, hypothalamus/pituitary and medulla oblongata and was widespread within these tissues as determined by immunohistochemistry. These immunoreagents provide specific tools to study the biology of Rainbow trout and Atlantic salmon PrP and any possible transmissible spongiform encephalopathy-like disease of these economically important fish species.

Detection of transgenic and endogenous plant DNA in rumen fluid, duodenal digesta, milk, blood, and feces of lactating dairy cows.

The objective was to determine the presence or absence of transgenic and endogenous plant DNA in ruminal fluid, duodenal digesta, milk, blood, and feces, and if found, to determine fragment size. Six multiparous lactating Holstein cows fitted with ruminal and duodenal cannulas received a total mixed ration. There were two treatments (T). In T1, the concentrate contained genetically modified (GM) soybean meal (cp4epsps gene) and GM corn grain (cry1a[b] gene), whereas T2 contained the near isogenic non-GM counterparts. Polymerase chain reaction analysis was used to determine the presence or absence of DNA sequences. Primers were selected to amplify small fragments from single-copy genes (soy lectin and corn high-mobility protein and cp4epsps and cry1a[b] genes from the GM crops) and multicopy genes (bovine mitochondrial cytochrome b and rubisco). Single-copy genes were only detected in the solid phase of rumen and duodenal digesta. In contrast, fragments of the rubisco gene were detected in the majority of samples analyzed in both the liquid and solid phases of ruminal and duodenal digesta, milk, and feces, but rarely in blood. The size of the rubisco gene fragments detected decreased from 1176 bp in ruminal and duodenal digesta to 351 bp in fecal samples.

Distinct localization and function of (1,4,5)IP(3) receptor subtypes and the (1,3,4,5)IP(4) receptor GAP1(IP4BP) in highly purified human platelet membranes.

Platelet activation is associated with an increase of cytosolic Ca(++) levels. The (1,4,5)IP(3) receptors [(1,4,5)IP(3)R] are known to mediate Ca(++) release from intracellular stores of many cell types. Currently there are at least 3 distinct subtypes of (1,4, 5)IP(3)R-type I, type II, and type III-with suggestions of distinct roles in Ca(++) elevation. Specific receptors for (1,3,4,5)IP(4) belonging to the GAP1 family have also been described though their involvement with Ca(++) regulation is controversial. In this study we report that platelets contain all 3 subtypes of (1,4,5)IP(3)R but in different amounts. Type I and type II receptors are predominant. In studies using highly purified platelet plasma (PM) and intracellular membranes (IM) we report a distinct localization of these receptors. The PM fractions were found to contain the type III (1,4,5)IP(3)R and GAP1(IP4BP) in contrast to IM, which contained type I (1,4,5)IP(3)R. The type II receptor exhibited a dual distribution. In studies examining the labeling of surface proteins with biotin in intact platelets only the type III (1,4,5)IP(3)R was significantly labeled. Immunogold studies of ultracryosections of human platelets showed significantly more labeling of the PM with the type III receptor antibodies than with type I receptor antibodies. Ca(++) flux studies were carried out with the PM to demonstrate in vitro function of inositol phosphate receptors. Ca(++) release activities were present with both (1,4,5)IP(3) and (1, 3,4,5)IP(4) (EC(50) = 1.3 and 0.8 micromol/L, respectively). Discrimination of the Ca(++)-releasing activities was demonstrated with cyclic adenosine monophosphate (cAMP)-dependent protein kinase (cAMP-PK) specifically inhibiting (1,4,5)IP(3) but not (1,3,4, 5)IP(4)-induced Ca(++) flux. In experiments with both PM and intact platelets, the (1,4,5)IP(3)Rs but not GAP1(IP4BP) were found to be substrates of cAMP-PK and cGMP-PK. Thus the Ca(++) flux property of (1,3,4,5)IP(4) is insensitive to cAMP-PK. These studies suggest distinct roles for the (1,4,5)IP(3)R subtypes in Ca(++) movements, with the type III receptor and GAP1(IP4BP) associated with cation entry in human platelets and the type I receptor involved with Ca(++) release from intracellular stores.