Nuclear factors enhance binding of T3 receptors (TR) to <em>D</em>NA, suggesting that T3 action may require a multicomponent complex bound to thyroid hormone response elements (TREs). We refer to the 65,000 <em>D</em>a nuclear protein in GH3 cells that enhances TR binding to <em>D</em>NA as the TR-auxiliary protein (TRAP) and have characterized its interaction with TR. Using a TRE-<em>D</em>NA affinity matrix we show that TRAP is able to bind to <em>D</em>NA, even in the absence of functional TR. We then used carboxyl-terminal truncations of rat TR alpha-1 and human TR beta in the avidin-biotin complex <em>D</em>NA-binding assay to identify regions that are important for interaction with TRAP. Removal of 34 residues of hTR beta abolishes T3-binding activity, but the ability to bind TRAP is retained. Further truncations and point mutations suggest that TRAP interacts with the ligand-binding domain of TR and with an independent region which overlaps a conserved sequence adjacent to the second Zn<em>2</em>+ finger (amino acids 1<em>2</em>0-149 in rTR alpha-1). A fragment of rTR alpha-1 (alpha C<em>2</em>91) which encompasses these two regions inhibits the ability of TRAP to enhance TR binding to <em>D</em>NA. This is due to binding of alpha C<em>2</em>91 to TR, demonstrating the ability of TR to form homo<em>dimers</em>. The inability of TRAP to interact with TR <em>dimers</em> and the similarity of the locations of the estradiol receptor dimerization domains with the TRAP interaction regions lead us to conclude that TRAP stabilizes TR binding to <em>D</em>NA by formation of TRAP-TR hetero<em>dimers</em> with both proteins bound to the <em>D</em>NA. TR bound to the estrogen response element is unable to respond to TRAP and unable to stimulate transcription, possibly due to the absence of TRAP in the TR-estrogen response element complex. In addition, TRAP may interact with a certain subset of the nuclear receptor superfamily, since human retinoic acid receptor-beta and vitamin <em>D</em> receptor show increased binding to TREs in the presence of nuclear extract, but c-erbA alpha-<em>2</em>, a variant TR, does not respond to TRAP.

The Escherichia coli Rep protein is a DNA helicase that is involve<em>d</em> in DNA replication. We have examine<em>d</em> the effects of DNA bin<em>d</em>ing on the assembly state of the Rep protein using small-zone gel permeation chromatography an<em>d</em> chemical crosslinking of the protein. Complexes of Rep protein were forme<em>d</em> with short single-stran<em>d</em>e<em>d</em> an<em>d</em> <em>d</em>uplex hairpin oligo<em>d</em>eoxynucleoti<em>d</em>es with lengths such that only a single Rep monomer coul<em>d</em> bin<em>d</em> per oligo<em>d</em>eoxynucleoti<em>d</em>e (i.e. <em>2</em> Rep monomers coul<em>d</em> not bin<em>d</em> contiguously on the oligo<em>d</em>eoxynucleoti<em>d</em>es). In the absence of DNA, Rep protein is monomeric (Mr 7<em>2</em>,800) up to concentrations of at least 8 microM (monomer), even in the presence of its nucleoti<em>d</em>e cofactors (ATP, ADP, ATP-gamma-S). However, the bin<em>d</em>ing of Rep monomers to single-stran<em>d</em>e<em>d</em> (ss) oligo<em>d</em>eoxynucleoti<em>d</em>es, <em>d</em>(pN)n (1<em>2</em> less than or equal to n less than or equal to <em>2</em>0), in<em>d</em>uces the Rep monomers to oligomerize. Upon treatment of the Rep-ss oligo<em>d</em>eoxynucleoti<em>d</em>e complexes with the protein crosslinking reagent <em>d</em>imethyl-suberimi<em>d</em>ate (DMS) an<em>d</em> subsequent removal of the DNA, crosslinke<em>d</em> Rep <em>dimers</em> are observe<em>d</em>, in<em>d</em>epen<em>d</em>ent of oligo<em>d</em>eoxynucleoti<em>d</em>e length (n less than or equal to <em>2</em>0). Furthermore, short <em>d</em>uplex oligo<em>d</em>eoxynucleoti<em>d</em>es also in<em>d</em>uce the Rep monomers to <em>dimer</em>ize. Formation of the Rep <em>dimers</em> results from an actual DNA-in<em>d</em>uce<em>d</em> <em>dimer</em>ization, rather than the a<em>d</em>ventitious crosslinking of Rep monomers boun<em>d</em> contiguously to a single oligo<em>d</em>eoxynucleoti<em>d</em>e. The purifie<em>d</em> DMS-crosslinke<em>d</em> Rep <em>dimer</em> shows increase<em>d</em> affinity for DNA an<em>d</em> retains DNA-<em>d</em>epen<em>d</em>ent ATPase an<em>d</em> DNA helicase activities, as shown by its ability to unwin<em>d</em> M13 RF DNA in the presence of the bacteriophage f1 gene II protein. On the basis of these observations an<em>d</em> since the <em>dimer</em> is the major species when Rep is boun<em>d</em> to DNA, we suggest that a DNA-in<em>d</em>uce<em>d</em> Rep <em>dimer</em> is the functionally active form of the Rep helicase.

BACKGROUND

Effects were compared in patients in the Bypass Angioplasty Revascularization Investigation <em>2</em> Diabetes (BARI <em>2</em>D) trial of <em>2</em> mechanistically different strategies for treatment of hyperglycemia, insulin-sensitizing and insulin-providing strategies, on biomarker profiles reflecting the balance between fibrinolysis and thrombosis and the intensity of inflammation implicated in diabetic vasculopathy.

RESULTS

A total of <em>2</em>368 patients with type <em>2</em> diabetes mellitus and clinically stable, angiographically documented coronary artery disease were randomized to treatment with 1 of the <em>2</em> strategies and followed for an average of 5 years. Plasminogen activator inhibitor type 1 antigen and activity, tissue plasminogen activator antigen, fibrinogen, D-dimer, C-reactive protein, insulin, and hemoglobin A(1c) were assayed in blood samples acquired at baseline and at 1<em>2</em> regular intervals throughout the follow-up interval. Higher baseline D-dimer, fibrinogen, and C-reactive protein portended a poor prognosis in patients in both groups. In contrast to the insulin-providing strategy, the insulin-sensitizing strategy led to (1) lower plasma insulin; (<em>2</em>) lower plasminogen activator inhibitor type 1 antigen and activity and lower tissue plasminogen activator antigen (known to track with plasminogen activator inhibitor type 1); and (3) lower C-reactive protein and fibrinogen at all intervals after baseline (P<0.001 for each).

CONCLUSIONS

The insulin-sensitizing treatment strategy led to changes in biomarker profiles indicative of decreased insulin resistance, an altered balance between thrombosis and fibrinolysis favoring fibrinolysis, and diminished intensity of the systemic inflammatory state, factors that have been associated with cardiovascular risk.

BACKGROUND

http://www.clinicaltrials.gov. Unique identifier: NCT00006305.

An engineered antibody fragment (minibody; scFv-C(H)3gamma(1) <em>dimer</em>, M(r) 80 000) specific for carcinoembryonic antigen (CEA) has previously demonstrated excellent tumor targeting coupled with rapid clearance in vivo. In this study, variable (V) genes from the anti- p185(HER-<em>2</em>) 10H8 antibody were similarly assembled and expressed. Four constructs were made: first, the V genes were assembled in both orientations (V(L)-linker-V(H) and V(H)-linker-V(L)) as single chain Fvs (scFvs). Then each scFv was fused to the human IgG1 C(H)3 domain, either by a two amino acid linker (ValGlu) that resulted in a non-covalent, hingeless minibody, or by IgG1 hinge and a GlySer linker peptide to produce a covalent, hinge-minibody. The constructs, expressed in NS0 mouse myeloma cells at levels of <em>2</em>0-60 mg/l, demonstrated binding to the human p185(HER-<em>2</em>) overexpressing breast cancer cell line, MCF7/HER<em>2</em>. Binding affinities (K(<em>D</em>) approximately <em>2</em>-4 nM) were equivalent to that for the parental 10H8 mAb (K(<em>D</em>) approximately 1.6 nM). Radioiodinated 10H8 hinge-minibody was evaluated in athymic mice, bearing MCF7/HER<em>2</em> xenografts. Maximum tumor uptake was 5.6 (+/-1.65)% injected dose/g (I<em>D</em>/g) at 1<em>2</em> h, which was lower than that of the anti-CEA minibody, whereas the blood clearance (beta-phase, 5.6<em>2</em> h) was similar. Thus, minibodies with different specificities display similar pharmacokinetics, while tumor uptake may vary depending on the antigen-antibody system.

The effect of estrogen on the synthesis of plasma very low <em>d</em>ensity lipoproteins (VL<em>D</em>L) in the cockerel was stu<em>d</em>ie<em>d</em> both in vivo an<em>d</em> in vitro. Synthesis was stu<em>d</em>ie<em>d</em> by immunoprecipitation techniques with antisera prepare<em>d</em> against VL<em>D</em>L an<em>d</em> a major VL<em>D</em>L protein. VL<em>D</em>L were isolate<em>d</em> from the plasma of white Leghorn hens an<em>d</em> estrogen-treate<em>d</em> white Leghorn cockerels by ultracentrifugal flotation at <em>d</em> 1.006 g/ml. After <em>d</em>elipi<em>d</em>ation, the lipi<em>d</em>-free proteins (apoproteins) were fractionate<em>d</em> on Sepha<em>d</em>ex G-150 an<em>d</em> <em>D</em>EAE-cellulose. Both the hen an<em>d</em> the estrogen-treate<em>d</em> cockerel VL<em>D</em>L were shown to contain an i<em>d</em>entical apoprotein with a mol wt of approximately 1<em>2</em>,000; the apoprotein is <em>d</em>esignate<em>d</em> fraction B. Re<em>d</em>uction an<em>d</em> S-carboxy-methylation of fraction B resulte<em>d</em> in a re<em>d</em>uction of the molecular weight by approximately one-half, in<em>d</em>icating a <em>dimer</em>-monomer relationship. Antiserum prepare<em>d</em> to the hen VL<em>D</em>L <em>dimer</em> protein gave precipitin lines of complete i<em>d</em>entity to both the hen an<em>d</em> cockerel <em>dimer</em>, monomer, VL<em>D</em>L, apoVL<em>D</em>L, low <em>d</em>ensity lipoproteins, an<em>d</em> plasma; no precipitin line was forme<em>d</em> with either hen or cockerel high <em>d</em>ensity lipoproteins. After a single subcutaneous injection of <em>d</em>iethylstilbestrol into the cockerel, plasma VL<em>D</em>L protein, cholesterol, an<em>d</em> triglyceri<em>d</em>e increase<em>d</em>, reaching a maximum <em>2</em>4--48 h after hormone a<em>d</em>ministration. Liver slices from similarly treate<em>d</em> animals were incubate<em>d</em> in vitro in culture me<em>d</em>ium in the presence of [3H]lysine for <em>2</em> h. Immunoprecipitable ra<em>d</em>ioactivity in VL<em>D</em>L increase<em>d</em> within <em>2</em> h of <em>d</em>iethylstilbestrol treatment an<em>d</em> reache<em>d</em> a maximum at <em>2</em>4 h; VL<em>D</em>L ra<em>d</em>ioactivity returne<em>d</em> to base-line levels by 7<em>2</em> h. At the peak of in<em>d</em>uction, newly synthesize<em>d</em> VL<em>D</em>L represente<em>d</em> 11% of the total soluble protein synthesize<em>d</em>. When actinomycin-<em>D</em> (5 mg/kg) was a<em>d</em>ministere<em>d</em> simultaneously with estrogen, the in<em>d</em>uction of VL<em>D</em>L synthesis was totally inhibite<em>d</em>. To <em>d</em>etermine whether the effect of estrogen on VL<em>D</em>L synthesis was me<em>d</em>iate<em>d</em> at the level of transcription, partially-purifie<em>d</em> cockerel liver mRNA was prepare<em>d</em> from estrogen-treate<em>d</em> animals an<em>d</em> the mRNA activity for fraction B was quantitate<em>d</em> in a wheat germ translation system. Fraction B mRNA was foun<em>d</em> to increase from a low base-line value to a maximum 16-<em>2</em>4 h after estrogen treatment, returning towar<em>d</em>s baseline values at 30 h. At the peak of in<em>d</em>uction, fraction B constitute<em>d</em> 1<em>2</em>% of the total protein synthesize<em>d</em>. The kinetics of in<em>d</em>uction of fraction B mRNA activity in the cell-free translation system is very similar to that observe<em>d</em> in liver slice experiments. This fin<em>d</em>ing suggests that estrogen stimulates VL<em>D</em>L synthesis, at least partially, by enhancing the accumulation of the mRNA for one of their major apoproteins.

Recently many attempts have been ma<em>d</em>e to <em>d</em>esign high-affinity DNA-bin<em>d</em>ing proteins by linking two <em>d</em>omains. Here a theory for gui<em>d</em>ing these <em>d</em>esigns is presente<em>d</em>. Flexible linkers may play three types of roles: (a) linking <em>d</em>omains which by themselves are unfol<em>d</em>e<em>d</em> an<em>d</em> bin<em>d</em> to DNA only as a fol<em>d</em>e<em>d</em> <em>dimer</em> (as in a <em>d</em>esigne<em>d</em> single-chain Arc repressor), (b) connecting <em>d</em>omains which can separately bin<em>d</em> to DNA (as in the Oct-1 POU <em>d</em>omain), an<em>d</em> (c) linking a DNA-bin<em>d</em>ing <em>d</em>omain with a <em>dimer</em>ization <em>d</em>omain (as in the lamb<em>d</em>a repressor). In (a), the linker keeps the protein as a fol<em>d</em>e<em>d</em> <em>dimer</em> so that it is always DNA-bin<em>d</em>ing-competent. In (b), the linker is pre<em>d</em>icte<em>d</em> to enhance DNA-bin<em>d</em>ing affinity over those of the in<em>d</em>ivi<em>d</em>ual <em>d</em>omains (with <em>d</em>issociation constants K(A) an<em>d</em> K(B)) by p(<em>d</em>(0))/K(B) or p(<em>d</em>(0))/K(A), where p(<em>d</em>(0)) = (3/4pil(p)bL)(3/<em>2</em>) exp(-3<em>d</em>(0)(<em>2</em>)/4l(p)bL)(1 - 5l(p)/4bL +...) is the probability <em>d</em>ensity for the en<em>d</em>-to-en<em>d</em> vector of the linker with L resi<em>d</em>ues to have a <em>d</em>istance <em>d</em>(0). In (c), the linker is pre<em>d</em>icte<em>d</em> to enhance the bin<em>d</em>ing affinity by K(<em>d</em>)(C)/p(<em>d</em>(0)), where K(<em>d</em>)(C) is the <em>dimer</em> <em>d</em>issociation constant for the <em>dimer</em>ization <em>d</em>omain. The pre<em>d</em>icte<em>d</em> affinity enhancements are foun<em>d</em> to be actually reache<em>d</em> by the Oct-1 POU <em>d</em>omain an<em>d</em> lamb<em>d</em>a repressor. However, there is room for improvement in many of the recently <em>d</em>esigne<em>d</em> proteins. The theoretical limits presente<em>d</em> shoul<em>d</em> provi<em>d</em>e a useful gui<em>d</em>e for current efforts of <em>d</em>esigning DNA-bin<em>d</em>ing proteins.

BACKGROUND

Severe injury induces an acute coagulopathy associated with increased mortality. This study compared the Thrombelastography (TEG) and biomarker profiles upon admission in trauma patients.

METHODS

Prospective observational study of 80 trauma patients admitted to a Level I Trauma Centre. <em>D</em>ata on demography, biochemistry including standard coagulation tests, hematology, transfusions, Injury Severity Score (ISS) and TEG were recorded. Retrospective analysis of thawed plasma/serum for biomarkers reflecting tissue injury (histone-complexed <em>D</em>NA fragments), sympathoadrenal activation (adrenaline, noradrenaline), coagulation activation/inhibition and fibrinolysis (sC<em>D</em>40L, protein C, activated Protein C, tissue-type plasminogen activator, plasminogen activator inhibitor-1, <em>D</em>-<em>dimer</em>, prothrombinfragment 1+<em>2</em>, plasmin/α<em>2</em>-antiplasmin complex, thrombin/antithrombin complex, tissue factor pathway inhibitor, antithrombin, von willebrand factor, factor XIII). Comparison of patients stratified according to ISS/TEG maximum clot strength. Linear regression analysis of variables associated with clot strength.

RESULTS

Trauma patients had normal (86%), hypercoagulable (11%) or hypocoagulable (1%) TEG clot strength; one had primary hyperfibrinolysis. Hypercoagulable patients had higher age, fibrinogen and platelet count (all p < 0.05), none had increased activated partial thromboplastin time (APTT) or international normalized ratio (INR) and none required massive transfusion >> 10 red blood cells the initial <em>2</em>4 h). Patients with normal or hypercoagulable TEG clot strength had comparable biomarker profiles, but the few patients with hypocoagulable TEG clot strength and/or hyperfibrinolysis had very different biomarker profiles.Increasing ISS was associated with higher levels of catecholamines, histone-complexed <em>D</em>NA fragments, sC<em>D</em>40L, activated protein C and <em>D</em>-<em>dimer</em> and reduced levels of non-activated protein C, antithrombin, fibrinogen and factor XIII (all p < 0.05). Fibrinogen and platelet count were associated independently with clot strength in patients with ISS ≤ <em>2</em>6 whereas only fibrinogen was associated independently with clot strength in patients with ISS>> <em>2</em>6. In patients with ISS>> <em>2</em>6, adrenaline and sC<em>D</em>40L were independently negatively associated with clot strength.

CONCLUSIONS

Trauma patients displayed different coagulopathies by TEG and variables independently associated with clot strength changed with ISS. In the highest ISS group, adrenaline and sC<em>D</em>40L were independently negatively associated with clot strength indicating that these may contribute to acute coagulopathy.

BACKGROUND

Platelet-derived growth factor (PDGF), which is a major mitogen for vascular smooth muscle cells and has been implicated in the pathogenesis of arteriosclerosis, is composed of <em>dimers</em> of PDGF-A and PDGF-B polypeptide chains, encoded by different genes. Here, we have analyzed the chromosomal localization, structure, and expression of <em>2</em> newly identified human genes of the PDGF family, called PDGFC and PDGFD.

RESULTS

We used fluorescence in situ hybridization to locate PDGFC and PDGFD in chromosomes 4q3<em>2</em> and 11q<em>2</em><em>2</em>.3 to <em>2</em>3.<em>2</em>, respectively. Exon structures of PDGFC and PDGFD were determined by sequencing from genomic DNA clones. The coding region of PDGFC consists of 6 and PDGFD of 7 exons, of which the last <em>2</em> encode the C-terminal PDGF cystine knot growth factor homology domain. An N-terminal CUB domain is encoded by exons <em>2</em> and 3 of both genes, and a region of proteolytic cleavage involved in releasing and activating the growth factor domain is located in exon 4 in PDGFC and exon 5 in PDGFD. PDGF-C was expressed predominantly in smooth muscle cells and PDGF-D in fibroblastic adventitial cells, and both genes were active in cultured endothelial cells and in a variety of tumor cell lines. Both PDGF-C and PDGF-D also stimulated human coronary artery smooth muscle cells.

CONCLUSIONS

PDGFC and PDGFD have similar genomic structures, which resemble those of the PDGFA and PDGFB genes. Their expression in the arterial wall and cultured vascular cells suggests that they can transduce proliferation/migration signals to pericytes and smooth muscle cells.

Increased risk of thrombotic events occurs in chronic obstructive pulmonary disease (COP<em>D</em>). Elevated fibrinogen and C-reactive protein (CRP), being common in COP<em>D</em>, are associated with formation of dense fibrin clots resistant to lysis. Statins have been found to display anti-inflammatory and antithrombotic effects. We investigated fibrin clot properties in COP<em>D</em> patients prior to and following statin therapy. Ex vivo plasma fibrin clot permeability, compaction, and fibrinolysis were assessed in 56 patients with stable COP<em>D</em>, aged 64.9 +/- 9.<em>2</em> years (mean FEV(1), 54.7 +/- 15.9% predicted), versus 56 controls matched for age, sex and cardiovascular risk factors. Patients were then randomly assigned to receive simvastatin 40 mg/day (n = <em>2</em>8) or to remain without statins for three months (n = <em>2</em>8). Patients with COP<em>D</em> had lower clot permeability (6.1+/- 1.07 versus 9.<em>2</em> +/- 0.9 10(-9) cm(<em>2</em>), p < 0.0001), decreased compaction (44.9 +/- 4.5 versus 63.9 +/- 6.1%, p < 0.0001), higher maximum <em>D</em>-<em>dimer</em> levels released from clots (4.<em>2</em>3 +/- 0.55 versus 3.53 +/- 0.31 mg/l, p < 0.0001) with a decreased rate of this release (75.0 +/- 8.3 versus 80.9 +/- 8.0 microg/l/min, p = 0.03) and prolonged lysis time (9.84 +/- 1.33 versus 8.0<em>2</em> +/- 0.84 min, p < 0.0001) compared with controls. Scanning electron microscopy confirmed denser clot structure in COP<em>D</em>. Multiple linear regression analysis after adjustment for age and fibrinogen showed that in the COP<em>D</em> patients, CRP was the only independent predictor of permeability (R(<em>2</em>) = 0.47, p < 0.001) and lysis time (R(<em>2</em>) = 0.43, p < 0.001). Simvastatin improved clot properties (p < 0.05) despite unaltered CRP and irrespective of cholesterol reduction. Our study shows that fibrin clots in COP<em>D</em> patients are composed of much denser networks that are more resistant to lysis, and these properties can be improved by statin administration.

Renal (pro)renin receptor (PRR) expression is increase<em>d</em> in <em>d</em>iabetes. The exact mechanisms involve<em>d</em> in this process are not well establishe<em>d</em>. We hypothesize<em>d</em> that high glucose up-regulates PRR through protein kinase C (PKC)-Raf-ERK an<em>d</em> PKC-c-Jun N-terminal kinase (JNK)-c-Jun signaling pathways. Rat mesangial cells expose<em>d</em> to 30 mm <em>d</em>-glucose <em>d</em>emonstrate<em>d</em> significant increase in PRR mRNA an<em>d</em> protein expression, intracellular phosphorylation of Raf-1 (Y340/341), ERK, JNK, nuclear factor-kappaB (NF-kappaB) p65 (S536) an<em>d</em> c-Jun (S63). By chromatin immunoprecipitation assay an<em>d</em> EMSA, high glucose in<em>d</em>uce<em>d</em> more functional NF-kappaB an<em>d</em> activator protein (AP)-1 <em>dimers</em> boun<em>d</em> to correspon<em>d</em>ing cis-regulatory elements in the pre<em>d</em>icte<em>d</em> PRR promoter to up-regulate PRR transcription. Conventional an<em>d</em> novel PKC inhibitors Chelerythrine an<em>d</em> Rottlerin, Raf-1 inhibitor GW5074, MEK1/<em>2</em> inhibitor U01<em>2</em>6, JNK inhibitor SP6001<em>2</em>5, NF-kappaB inhibitor Quinazoline, an<em>d</em> AP-1 inhibitor Curcumin, respectively, attenuate<em>d</em> glucose-in<em>d</em>uce<em>d</em> PRR up-regulation. Chelerythrine an<em>d</em> Rottlerin also inhibite<em>d</em> glucose-in<em>d</em>uce<em>d</em> phosphorylation of Raf-1 (Y340/341), ERK1/<em>2</em>, JNK, NF-kappaB p65 (S536), an<em>d</em> c-Jun (S63). GW5074 an<em>d</em> U01<em>2</em>6 inhibite<em>d</em> the phosphorylation of ERK1/<em>2</em> an<em>d</em> NF-kappaB p65 (S536). SP6001<em>2</em>5 inhibite<em>d</em> phosphorylation of NF-kappaB p65 (S536) an<em>d</em> c-Jun (S63). We conclu<em>d</em>e that high glucose up-regulates the expression of PRR through mechanisms <em>d</em>epen<em>d</em>ent on both PKC-Raf-ERK an<em>d</em> PKC-JNK-c-Jun signaling pathways. NF-kappaB an<em>d</em> AP-1 are involve<em>d</em> in high-glucose-in<em>d</em>uce<em>d</em> PRR up-regulation in rat mesangial cells.

OBJECTIVE

The literature suggests that the d -dimer is useful in patients suspected of having pulmonary embolism and who have a low pretest probability of disease. A previously defined clinical decision rule, the Wells Criteria, may provide a reliable and reproducible means of determining this pretest probability. We evaluate the interrater agreement and external validity of Wells Criteria in determining pretest probability in patients suspected of having pulmonary embolism.

METHODS

This was a prospective observational study. Trained research assistants enrolled patients during 120 random 8-hour shifts. Patients who underwent imaging for pulmonary embolism after a medical history, physical examination, and chest radiograph were enrolled. Treating providers and research assistants determined pretest probability according to Wells Criteria in a blinded fashion. Two d -dimer assays were run. Three-month follow-up for the diagnosis of pulmonary embolism was performed. Interrater agreement tables were created. kappa Values, sensitivities, and specificities were determined.

RESULTS

Of the 153 eligible patients, 3 patients were missed, 16 patients declined, and 134 (88%) patients were enrolled. Sixteen (12%) patients were diagnosed with pulmonary embolism. The kappa values for Wells Criteria were 0.54 and 0.72 for the trichotomized and dichotomized scorings, respectively. When Wells Criteria were trichotomized into low pretest probability (n=59, 44%), moderate pretest probability (n=61, 46%), or high pretest probability (n=14, 10%), the pulmonary embolism prevalence was 2%, 15%, and 43%, respectively. When Wells Criteria were dichotomized into pulmonary embolism-unlikely (n=88, 66%) or pulmonary embolism-likely (n=46, 34%), the prevalence was 3% and 28%, respectively. The immunoturbidimetric and rapid enzyme-linked immunosorbent assay d -dimer assays had similar sensitivities (94%) and specificities (45% versus 46%).

CONCLUSIONS

Wells Criteria have a moderate to substantial interrater agreement and reliably risk stratify pretest probability in patients with suspected pulmonary embolism.

Interleukin-10 (IL-10) has been found to inhibit lipopolysaccharide (LPS)-induced tissue factor expression by monocytes in vitro. To determine the effects of IL-10 on LPS-induced activation of the hemostatic mechanisms in vivo, we performed a placebo-controlled, cross-over study of human endotoxemia. Two groups of eight volunteers were challenged with LPS (4 ng/kg) on two occasions: once in conjunction with placebo, and once with recombinant human IL-10 (rhIL-10; <em>2</em>5 microg/kg). In group 1, placebo or rhIL-10 was given <em>2</em> minutes before LPS challenge, group <em>2</em> received placebo or rhIL-10 1 hour after LPS administration. Pretreatment with rhIL-10 reduced both LPS-induced activation of the fibrinolytic system (plasma concentrations of tissue type plasminogen activator, plasmin-alpha<em>2</em>-antiplasmin complexes, and <em>D</em>-<em>dimer</em>), and inhibition of fibrinolysis (plasma levels of plasminogen activator inhibitor 1), whereas posttreatment only inhibited the latter response. Both IL-10 pre- and posttreatment attenuated activation of the coagulation system (plasma levels of prothrombin fragment F1 + <em>2</em> and thrombin-antithrombin complexes). These results indicate that rhIL-10, besides its well-described inhibitory effects on cytokine release, potently modulates the fibrinolytic system and inhibits the coagulant responses during endotoxemia.

We previously reported that the natural hormone 1,<em>2</em>5dihydroxyvitamin <em>D</em>3 (1,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3)) protects human skin cells from ultraviolet radiation (UVR)-induced apoptosis. UVR-induced pre-mutagenic cyclobutane pyrimidine <em>dimers</em> are diminished in number from 0.5h after cessation of UVR in all skin cell types, by treatment with three different Vitamin <em>D</em> compounds: by 1,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3), by the rapid acting, low calcemic analog, 1alpha,<em>2</em>5(OH)(<em>2</em>)lumisterol(3) (JN) and by the low calcemic but transcriptionally active hybrid analog 1alpha-hydroxymethyl-16-ene-<em>2</em>4,<em>2</em>4-difluoro-<em>2</em>5-hydroxy-<em>2</em>6,<em>2</em>7-bis-homovitamin <em>D</em>3 QW-16<em>2</em>4F<em>2</em>-<em>2</em> (QW), which may explain the enhanced cell survival. The rapid response antagonist analog 1beta,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3) (HL) abolished the photoprotective effects of 1,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3) whilst a genomic antagonist, (<em>2</em>3S)-<em>2</em>5-dehydro-1alpha-hydroxyvitamin <em>D</em>(3)-<em>2</em>6,<em>2</em>3-lactone (TEI-9647), had no effect. UVR increased p53 expression in human skin cells, whilst concurrent treatment with 1,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3) further enhanced this effect several fold, at 3 and 6h after UVR. Combined with previously reported lower nitrite levels with 1,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3), this increased p53 expression may favor <em>D</em>NA repair over apoptosis. We now report that topical application of 1,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3) or QW also suppressed solar simulated UV (SSUVR-induced pyrimidine <em>dimers</em> in the epidermis of irradiated hairless Skh:HR1 mice, measured <em>2</em>4h after irradiation. Furthermore, UVR-induced immunosuppression in the mice was markedly reduced by topical application of either 1,<em>2</em>5(OH)(<em>2</em>)<em>D</em>(3) or QW. These preliminary results show, for the first time, a protective effect of Vitamin <em>D</em> compounds against <em>D</em>NA photodamage in vivo.

BACKGROUND

Prediction models combine patient characteristics and test results to predict the presence of a disease or the occurrence of an event in the future. In the event that test results (predictor) are unavailable, a strategy is needed to help users applying a prediction model to deal with such missing values. We evaluated 6 strategies to deal with missing values.

METHODS

We developed and validated (in 1<em>2</em>95 and 53<em>2</em> primary care patients, respectively) a prediction model to predict the risk of deep venous thrombosis. In an application set (<em>2</em>59 patients), we mimicked 3 situations in which (1) an important predictor (<em>D</em>-<em>dimer</em> test), (<em>2</em>) a weaker predictor (difference in calf circumference), and (3) both predictors simultaneously were missing. The 6 strategies to deal with missing values were (1) ignoring the predictor, (<em>2</em>) overall mean imputation, (3) subgroup mean imputation, (4) multiple imputation, (5) applying a submodel including only the observed predictors as derived from the development set, or (6) the "one-step-sweep" method. We compared the model's discriminative ability (expressed by the ROC area) with the true ROC area (no missing values) and the model's estimated calibration slope and intercept with the ideal values of 1 and 0, respectively.

RESULTS

Ignoring the predictor led to the worst and multiple imputation to the best discrimination. Multiple imputation led to calibration intercepts closest to the true value. The effect of the strategies on the slope differed between the 3 scenarios.

CONCLUSIONS

Multiple imputation is preferred if a predictor value is missing.

In the present study, detailed information is presented on the hetero-dimerization of the serotonin 5-HT(<em>2</em>A) receptor and the dopamine <em>D</em>(<em>2</em>) receptor. Biophysical approaches (fluorescence spectroscopy as well as fluorescence lifetime microscopy) were used to determine the degree of fluorescence resonance energy transfer (FRET) between cyan and yellow fluorescent protein labeled receptor variants co-expressed in human embryonic kidney <em>2</em>93 cells (HEK<em>2</em>93). Recorded data demonstrate the existence of energy transfer between the wild-type forms of 5-HT(<em>2</em>A)R and <em>D</em>(<em>2</em>)R, pointing toward the formation of hetero-5-HT(<em>2</em>A)R/<em>D</em>(<em>2</em>)R <em>dimers</em> and homo-5-HT(<em>2</em>A)R/5-HT(<em>2</em>A)R <em>dimers</em>. Moreover, the present study investigates the role of specific motifs (one containing adjacent arginine residues (<em>2</em>17RRRRKR<em>2</em><em>2</em><em>2</em>) in the third intracellular loop (ic3) of <em>D</em>(<em>2</em>)R, and the other consisting of acidic glutamate residues (454EE455) in the C-tail of (5-HT(<em>2</em>A)R) in the formation of noncovalent complexes between these receptors. Our results suggest that these regions of 5-HT(<em>2</em>A)R and <em>D</em>(<em>2</em>)R may be involved in the interaction between these two proteins. On the other hand, the above-mentioned motifs do not play an important role in the homo-dimerization of these receptors. Furthermore, we estimated the influence of specific receptor ligands on the dimerization processes. Agonists (<em>D</em>OI and quinpirole) and antagonists (ketanserin and butaclamol) cause different effects on FRET efficiency depending on whether homo- or hetero-complexes are present. These data may have therapeutic implications, since (using the immunofluorescence double labeling protocols) the co-localization of these two receptors was demonstrated in the medial prefrontal cortex and pars reticulate of the substantia nigra of the rat brain.

Innate immunity reacts to conserved bacterial molecules. The outermost lipopolysaccharide (LPS) of Gram-negative organisms is highly inflammatory. It activates responsive cells via specific C<em>D</em>14 and toll-like receptor-4 (TLR4) surface receptor and co-receptors. Gram-positive bacteria do not contain LPS, but carry surface teichoic acids, lipoteichoic acids and peptidoglycan instead. Among these, the thick peptidoglycan is the most conserved. It also triggers cytokine release via C<em>D</em>14, but uses the TLR<em>2</em> co-receptor instead of TLR4 used by LPS. Moreover, whole peptidoglycan is 1000-fold less active than LPS in a weight-to-weight ratio. This suggests either that it is not important for inflammation, or that only part of it is reactive while the rest acts as ballast. Biochemical dissection of Staphylococcus aureus and Streptococcus pneumoniae cell walls indicates that the second assumption is correct. Long, soluble peptidoglycan chains (approximately 1<em>2</em>5 k<em>D</em>a) are poorly active. Hydrolysing these chains to their minimal unit (<em>2</em> sugars and a stem peptide) completely abrogates inflammation. Enzymatic dissection of the pneumococcal wall generated a mixture of highly active fragments, constituted of trimeric stem peptides, and poorly active fragments, constituted of simple monomers and <em>dimers</em> or highly polymerized structures. Hence, the optimal constraint for activation might be 3 cross-linked stem peptides. The importance of structural constraint was demonstrated in additional studies. For example, replacing the first L-alanine in the stem peptide with a <em>D</em>-alanine totally abrogated inflammation in experimental meningitis. Likewise, modifying the <em>D</em>-alanine decorations of lipoteichoic acids with L-alanine, or deacylating them from their diacylglycerol lipid anchor also decreased the inflammatory response. Thus, although considered as a broad-spectrum pattern-recognizing system, innate immunity can detect very subtle differences in Gram-positive walls. This high specificity underlines the importance of using well-characterized microbial material in investigating the system.

COVI<em>D</em>-19 is an infection induced by the SARS-CoV-<em>2</em> coronavirus, and severe forms can lead to acute respiratory distress syndrome (AR<em>D</em>S) requiring intensive care unit (ICU) management. Severe forms are associated with coagulation changes, mainly characterized by an increase in <em>D</em>-<em>dimer</em> and fibrinogen levels, with a higher risk of thrombosis, particularly pulmonary embolism. The impact of obesity in severe COVI<em>D</em>-19 has also been highlighted.In this context, standard doses of low molecular weight heparin (LMWH) may be inadequate in ICU patients, with obesity, major inflammation, and hypercoagulability. We therefore urgently developed proposals on the prevention of thromboembolism and monitoring of hemostasis in hospitalized patients with COVI<em>D</em>-19.Four levels of thromboembolic risk were defined according to the severity of COVI<em>D</em>-19 reflected by oxygen requirement and treatment, the body mass index, and other risk factors. Monitoring of hemostasis (including fibrinogen and <em>D</em>-<em>dimer</em> levels) every 48 h is proposed. Standard doses of LMWH (e.g., enoxaparin 4000 IU/<em>2</em>4 h SC) are proposed in case of intermediate thrombotic risk (BMI < 30 kg/m^<em>2</em>, no other risk factors and no AR<em>D</em>S). In all obese patients (high thrombotic risk), adjusted prophylaxis with intermediate doses of LMWH (e.g., enoxaparin 4000 IU/1<em>2</em> h SC or 6000 IU/1<em>2</em> h SC if weight > 1<em>2</em>0 kg), or unfractionated heparin (UFH) if renal insufficiency (<em>2</em>00 IU/kg/<em>2</em>4 h, IV), is proposed. The thrombotic risk was defined as very high in obese patients with AR<em>D</em>S and added risk factors for thromboembolism, and also in case of extracorporeal membrane oxygenation (ECMO), unexplained catheter thrombosis, dialysis filter thrombosis, or marked inflammatory syndrome and/or hypercoagulability (e.g., fibrinogen > 8 g/l and/or <em>D</em>-<em>dimers</em> > 3 μg/ml). In ICU patients, it is sometimes difficult to confirm a diagnosis of thrombosis, and curative anticoagulant treatment may also be discussed on a probabilistic basis. In all these situations, therapeutic doses of LMWH, or UFH in case of renal insufficiency with monitoring of anti-Xa activity, are proposed.In conclusion, intensification of heparin treatment should be considered in the context of COVI<em>D</em>-19 on the basis of clinical and biological criteria of severity, especially in severely ill ventilated patients, for whom the diagnosis of pulmonary embolism cannot be easily confirmed.

Keywords: Anticoagulant; COVID-19; Coagulation; Heparin; Obesity; Thrombosis.

The cannabinoid CB(1) (CB(1)) and dopamine <em>D</em>(<em>2</em>) (<em>D</em>(<em>2</em>)) receptors are coexpressed in the basal ganglia, an area of the brain involved in such processes as cognition, motor function, and emotional control. Several lines of evidence suggest that CB(1) and <em>D</em>(<em>2</em>) receptors may oligomerize, providing a unique pharmacology in vitro and in vivo. However, limited information exists on the regulation of CB(1) and <em>D</em>(<em>2</em>) receptor <em>dimers</em>. We used a novel technique, multicolor bimolecular fluorescence complementation (MBiFC) to examine the subcellular localization of CB(1)-<em>D</em>(<em>2</em>L) hetero<em>dimers</em> as well as <em>D</em>(<em>2</em>L)-<em>D</em>(<em>2</em>L) homo<em>dimers</em> in a neuronal cell model, Cath. a differentiated cells. MBiFC was then used to explore the effects of persistent ligand treatment on receptor dimerization at the plasma membrane and intracellularly. Persistent (<em>2</em>0-h) agonist treatment resulted in increased formation of CB(1)-<em>D</em>(<em>2</em>L) hetero<em>dimers</em> relative to the <em>D</em>(<em>2</em>L)-<em>D</em>(<em>2</em>L) homo<em>dimers</em>. The effects of the <em>D</em>(<em>2</em>) agonist quinpirole were restricted to the intracellular compartment and may reflect increased <em>D</em>(<em>2</em>L) receptor expression. In contrast, treatment with the CB(1) receptor agonist (<em>2</em>)-cis-3-[<em>2</em>-hydroxy-4-(1,1-dimethylheptyl)phenyl]-trans-4-(3-hydroxypropyl) cyclohexanol (CP55, 940) produced increases in both membrane and intracellular CB(1)-<em>D</em>(<em>2</em>L) hetero<em>dimers</em> independently of alterations in CB(1) receptor expression. The effects of CB(1) receptor activation were attenuated by the CB(1) antagonist 1-(<em>2</em>,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-4-morpholinyl-1H-pyrazole-3-carboxamide (AM<em>2</em>81) and were both time- and dose-dependent. The effects of CB(1) activation were examined further by combining MBiFC with a constitutively active CB(1) receptor mutant, CB(1)T<em>2</em>10I. These studies demonstrated that the expression of CB(1)T<em>2</em>10I increased intracellular CB(1)-<em>D</em>(<em>2</em>L) heterodimer formation. In summary, agonist-induced modulation of CB(1)-<em>D</em>(<em>2</em>L) oligomerization may have physiological implications in diseases such as Parkinson's disease and drug abuse.

Great attention has been paid to endothelial dysfunction (E<em>D</em>) in coronavirus disease <em>2</em>019 (COVI<em>D</em>-19). There is growing evidence to suggest that the angiotensin converting enzyme <em>2</em> receptor (ACE<em>2</em> receptor) is expressed on endothelial cells (ECs) in the lung, heart, kidney, and intestine, particularly in systemic vessels (small and large arteries, veins, venules, and capillaries). Upon viral infection of ECs by severe acute respiratory syndrome coronarvirus <em>2</em> (SARS-CoV-<em>2</em>), ECs become activated and dysfunctional. As a result of endothelial activation and E<em>D</em>, the levels of pro-inflammatory cytokines (interleukin -1, interleukin-6 (IL-6), and tumor necrosis factor-α), chemokines (monocyte chemoattractant protein-1), von Willebrand factor (vWF) antigen, vWF activity, and factor VIII are elevated. Higher levels of acute phase reactants (IL-6, C-reactive protein, and <em>D</em>-<em>dimer</em>) are also associated with SARS-CoV-<em>2</em> infection. Therefore, it is reasonable to assume that E<em>D</em> contributes to COVI<em>D</em>-19-associated vascular inflammation, particularly endotheliitis, in the lung, heart, and kidney, as well as COVI<em>D</em>-19-associated coagulopathy, particularly pulmonary fibrinous microthrombi in the alveolar capillaries. Here we present an update on E<em>D</em>-relevant vasculopathy in COVI<em>D</em>-19. Further research for E<em>D</em> in COVI<em>D</em>-19 patients is warranted to understand therapeutic opportunities.

<strong class="sub-title"> Keywords: </strong> COVI<em>D</em>-19; SARS-CoV-<em>2</em>; coagulation; cytokines; endothelial dysfunction; thrombosis; von Willebrand factor.

The hyperthermophilic archaeon Thermococcus kodakaraensis KO<em>D</em>1 possesses chitinase (Tk-ChiA) and exo-beta-<em>D</em>-glucosaminidase (Tk-GlmA) for chitin degradation; the former produces diacetylchitobiose (GlcNAc<em>2</em>) from chitin, and the latter hydrolyzes chitobiose (GlcN<em>2</em>) to glucosamine (GlcN). To identify the enzyme that physiologically links these two activities, here we focused on the deacetylase that provides the substrate for Tk-GlmA from GlcNAc<em>2</em>. The deacetylase could be detected in and partially purified from T. kodakaraensis cells, and the corresponding gene (Tk-dac) was identified on the genome. The deduced amino acid sequence was classified into the LmbE protein family including N-acetylglucosaminylphosphatidylinositol de-N-acetylases and 1-<em>D</em>-myo-inosityl-<em>2</em>-acetamido-<em>2</em>-deoxy-alpha-<em>D</em>-glucopyranoside deacetylase. Recombinant Tk-<em>D</em>ac showed deacetylase activity toward N-acetylchitooligosaccharides (GlcNAc(<em>2</em>-5)), and the deacetylation site was revealed to be specific at the nonreducing GlcNAc residue. The enzyme also deacetylated GlcNAc monomer. In T. kodakaraensis cells, the transcription of Tk-dac, Tk-glmA, Tk-chiA, and the clustered genes were induced by GlcNAc<em>2</em>, suggesting the function of this gene cluster in chitin catabolism in vivo. These results have revealed a unique chitin catabolic pathway in T. kodakaraensis, in which GlcNAc<em>2</em> produced from chitin is degraded by the concerted action of Tk-<em>D</em>ac and Tk-GlmA. That is, GlcNAc<em>2</em> is site-specifically deacetylated to GlcN-GlcNAc by Tk-<em>D</em>ac and then hydrolyzed to GlcN and GlcNAc by Tk-GlmA followed by a second deacetylation step of the remaining GlcNAc by Tk-<em>D</em>ac to form GlcN. This is the first elucidation of an archaeal chitin catabolic pathway and defines a novel mechanism for <em>dimer</em> processing using a combination of deacetylation and cleavage, distinct from any previously known pathway.

BACKGROUND

Patients with chronic urticaria (CU) frequently show signs of thrombin generation as a result of the activation of the extrinsic pathway of coagulation and signs of fibrinolysis as shown by slightly increased mean D-dimer plasma levels. Here, we studied patients with severe CU to see whether the activation of coagulation and fibrinolysis parallels the severity of the disease.

METHODS

Eight consecutive patients with severe exacerbations of CU and 13 with slight CU were studied. Plasma prothrombin fragment F(1+2) as well as D-dimer were measured by ELISA. Serum histamine-releasing activity was assessed by basophil histamine release assay. Seventy-four normal subjects were used as controls.

RESULTS

In patients with severe CU, median levels of both D-dimer (11.20 nmol/l) and F(1+2) (592 pmol/l) largely exceeded those found in patients with slight CU [D-dimer: 2.66 nmol/l (P = 0.001) and F(1+2): 228 pmol/l (P = 0.003)] and in normal subjects [D-dimer: 1.41 nmol/l (P = 0.0001) and F(1+2): 159 pmol/l (P = 0.0001)]. Sera from 25% of patients with severe CU and 31% of those with slight CU, but from none of normal subjects, showed in vitro histamine-releasing activity. D-dimer and F(1+2) levels were significantly correlated each other (r = 0.64, P = 0.002) and with CU severity score (r = 0.80-0.90, P = 0.0001), but no correlation was observed between serum histamine-releasing activity and coagulation parameters or severity score.

CONCLUSIONS

Severe exacerbations of CU are associated with a strong activation of coagulation cascade and fibrinolysis. Whether this activation is the cause of CU or acts as an amplification system is still a matter of debate.

<strong class="sub-title"> Background: </strong> Since December <em>2</em>019, an outbreak of Coronavirus disease <em>2</em>019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus <em>2</em> (SARS-Cov-<em>2</em>) initially emerged in Wuhan, China, and has spread worldwide now. Clinical features of patients with COVID-19 have been described. However, risk factors leading to in-hospital deterioration and poor prognosis in COVID-19 patients with severe disease have not been well identified.

<strong class="sub-title"> Methods: </strong> In this retrospective, single-center cohort study, 1190 adult inpatients (≥ 18 years old) with laboratory-confirmed COVID-19 and determined outcomes (discharged or died) were included from Wuhan Infectious Disease Hospital from December <em>2</em>9, <em>2</em>019 to February <em>2</em>8, <em>2</em>0<em>2</em>0. The final follow-up date was March <em>2</em>, <em>2</em>0<em>2</em>0. Clinical data including characteristics, laboratory and imaging information as well as treatments were extracted from electronic medical records and compared. A multivariable logistic regression model was used to explore the potential predictors associated with in-hospital deterioration and death.

<strong class="sub-title"> Results: </strong> 1190 patients with confirmed COVID-19 were included. Their median age was 57 years (interquartile range 47-67 years). Two hundred and sixty-one patients (<em>2</em><em>2</em>%) developed a severe illness after admission. Multivariable logistic regression demonstrated that higher SOFA score (OR 1.3<em>2</em>, 95% CI 1.<em>2</em><em>2</em>-1.43, per score increase, p < 0.001 for deterioration and OR 1.30, 95% CI 1.11-1.53, per score increase, p = 0.001 for death), lymphocytopenia (OR 1.81, 95% CI 1.13-<em>2</em>.89 p = 0.013 for deterioration; OR 4.44, 95% CI 1.<em>2</em>6-15.87, p = 0.0<em>2</em>1 for death) on admission were independent risk factors for in-hospital deterioration from not severe to severe disease and for death in severe patients. On admission D-dimer greater than 1 μg/L (OR 3.<em>2</em>8, 95% CI 1.19-9.04, p = 0.0<em>2</em>1), leukocytopenia (OR 5.10, 95% CI 1.<em>2</em>5-<em>2</em>0.78), thrombocytopenia (OR 8.37, 95% CI <em>2</em>.04-34.44) and history of diabetes (OR 11.16, 95% CI 1.87-66.57, p = 0.008) were also associated with higher risks of in-hospital death in severe COVID-19 patients. Shorter time interval from illness onset to non-invasive mechanical ventilation in the survivors with severe disease was observed compared with non-survivors (10.5 days, IQR 9.<em>2</em>5-11.0 vs. 16.0 days, IQR 11.0-19.0 days, p = 0.030). Treatment with glucocorticoids increased the risk of progression from not severe to severe disease (OR 3.79, 95% CI <em>2</em>.39-6.01, p < 0.001). Administration of antiviral drugs especially oseltamivir or ganciclovir is associated with a decreased risk of death in severe patients (OR 0.17, 95% CI 0.05-0.64, p < 0.001).

Conclusions: High SOFA score and lymphocytopenia on admission could predict that not severe patients would develop severe disease in-hospital. On admission elevated D-dimer, leukocytopenia, thrombocytopenia and diabetes were independent risk factors of in-hospital death in severe patients with COVID-19. Administration of oseltamivir or ganciclovir might be beneficial for reducing mortality in severe patients.

Keywords: COVID-19; Development; Mortality; Risk factors; Severe.

Background: COVID-19 is a virus pandemic. According to the first obtained data, COVID-19 has defined with findings such as cough, fever, diarrhea, and fatigue although neurological symptoms of patients with COVID-19 have not been investigated in detail. This study aims to investigate the neurological findings via obtained face-to-face anamnesis and detailed neurological examination in patients with COVID-19.

Methods: Two hundred thirty-nine consecutive inpatients with COVID-19, supported with laboratory tests, were evaluated. Detailed neurological examinations and evaluations of all patients were performed. All evaluations and examinations were performed by two neurologists who have at least five-year experience.

Results: This study was carried out 239 patients (133 male + 106 female) with diagnosed COVID-19. Neurological findings were present in 83 of 239 patients (34.7%). The most common neurological finding was a headache (27.6%). D-dimer blood levels were detected to be significantly higher in patients with at least one neurological symptom than patients without the neurological symptom (p < 0.05). IL-6 level was found to be significantly higher in patients with headache than without headache (p < 0.05). Creatine kinase (CK) level was detected to be significantly higher in patients with muscle pain (p < 0.05).

Conclusion: Neurological symptoms are often seen in patients with COVID-19. Headache was the most common seen neurological symptom in this disease. Dizziness, impaired consciousness, smell and gustation impairments, cerebrovascular disorders, epileptic seizures, and myalgia were detected as other findings apart from the headache. It is suggested that determining these neurological symptoms prevents the diagnosis delay and helps to prohibit virus spread.

Keywords: COVID-19; Coronavirus disease 2019; Neurologic symptoms; Neurological manifestations; SARS-CoV-2.

The PROLONG ran<em>d</em>omize<em>d</em> trial showe<em>d</em> that a normal <em>D</em>-<em>dimer</em> (<em>D</em>-<em>d</em>) 1 month after anticoagulation suspension for unprovoke<em>d</em> venous thromboembolism (VTE) was associate<em>d</em> with a low risk of late recurrences (4.4% patient years). However, it is unknown whether <em>D</em>-<em>d</em> changes subsequently. The aim of this prospective multicenter stu<em>d</em>y was to assess <em>D</em>-<em>d</em> time course an<em>d</em> its relation with late recurrences in patients with normal <em>D</em>-<em>d</em> 1 month after anticoagulation suspension for a first episo<em>d</em>e of unprovoke<em>d</em> VTE. <em>D</em>-<em>d</em> was measure<em>d</em> with a qualitative metho<em>d</em> (Clearview Simplify <em>D</em>-<em>dimer</em>; Inverness Me<em>d</em>ical Professional <em>D</em>iagnostics). Patients with a normal <em>D</em>-<em>d</em> 1 month after stopping anticoagulation repeate<em>d</em> <em>D</em>-<em>d</em> testing every <em>2</em> months for 1 year. <em>D</em>-<em>d</em> was normal in 68% (<em>2</em>43/355) of patients 1 month after anticoagulation suspension. Patients in whom <em>D</em>-<em>d</em> became abnormal at the thir<em>d</em> month an<em>d</em> remaine<em>d</em> abnormal afterwar<em>d</em> ha<em>d</em> a higher risk of recurrence (7/31; <em>2</em>7% patient years; 95% confi<em>d</em>ence interval [CI]: 1<em>2</em>-48) than patients in whom <em>D</em>-<em>d</em> remaine<em>d</em> normal at the thir<em>d</em> month an<em>d</em> afterwar<em>d</em> (4/149; <em>2</em>.9% patient years; 95% CI: 1-7; a<em>d</em>juste<em>d</em> hazar<em>d</em> ratio: 7.9; 95% CI: <em>2</em>.1-30; P = .00<em>2</em>). Repeate<em>d</em> <em>D</em>-<em>d</em> testing after anticoagulation suspension for a first episo<em>d</em>e of unprovoke<em>d</em> VTE coul<em>d</em> help tailor the <em>d</em>uration of treatment. This trial is registere<em>d</em> at http://clinicaltrials.gov as NCT00<em>2</em>66045.