OBJECTIVE

The aim of this study was to investigate whether multiple biomarkers contribute to improved coronary heart disease (CHD) risk prediction in post-menopausal women compared with assessment using traditional risk factors (TRFs) only.

BACKGROUND

The utility of newer biomarkers remains uncertain when added to predictive models using only TRFs for CHD risk assessment.

METHODS

The Women's Health Initiative Hormone Trials enrolle<em>d</em> <em>2</em>7,347 post-menopausal women ages 50 to 79 years. Associations of TRFs an<em>d</em> 18 biomarkers were assesse<em>d</em> in a neste<em>d</em> case-control stu<em>d</em>y inclu<em>d</em>ing 3<em>2</em>1 patients with CHD an<em>d</em> 743 controls. Four pre<em>d</em>iction equations for 5-year CHD risk were compare<em>d</em>: <em>2</em> Framingham risk score covariate mo<em>d</em>els; a TRF mo<em>d</em>el inclu<em>d</em>ing statin treatment, hormone treatment, an<em>d</em> car<em>d</em>iovascular <em>d</em>isease history as well as the Framingham risk score covariates; an<em>d</em> an a<em>d</em><em>d</em>itional biomarker mo<em>d</em>el that a<em>d</em><em>d</em>itionally inclu<em>d</em>e<em>d</em> the 5 significantly associate<em>d</em> markers of the 18 teste<em>d</em> (interleukin-6, <em>d</em>-<em>dimer</em>, coagulation factor VIII, von Willebran<em>d</em> factor, an<em>d</em> homocysteine).

RESULTS

The TRF mo<em>d</em>el showe<em>d</em> an improve<em>d</em> C-statistic (0.7<em>2</em>9 vs. 0.699, p = 0.001) an<em>d</em> net reclassification improvement (6.4<em>2</em>%) compare<em>d</em> with the Framingham risk score mo<em>d</em>el. The a<em>d</em><em>d</em>itional biomarker mo<em>d</em>el showe<em>d</em> a<em>d</em><em>d</em>itional improvement in the C-statistic (0.751 vs. 0.7<em>2</em>9, p = 0.001) an<em>d</em> net reclassification improvement (6.45%) compare<em>d</em> with the TRF mo<em>d</em>el. Pre<em>d</em>icte<em>d</em> CHD risks on a continuous scale showe<em>d</em> high agreement between the TRF an<em>d</em> a<em>d</em><em>d</em>itional biomarker mo<em>d</em>els (Spearman's coefficient = 0.918). Among the 18 biomarkers measure<em>d</em>, C-reactive protein level <em>d</em>i<em>d</em> not significantly improve CHD pre<em>d</em>iction either alone or in combination with other biomarkers.

CONCLUSIONS

Moderate improvement in CHD risk prediction was found when an 18-biomarker panel was added to predictive models using TRFs in post-menopausal women.

We have used an immunofluorescence inhibition assay to identify <em>2</em> BALB/c plasmacytomas, TEPC-1017 and TEPC-1033, that secrete large quantitites of IgD. Both TEPC-1017 and TEPC-1033 myeloma proteins bound to anti-kappa as well as hybridoma and heterologous anti-<em>delta</em> antibodies, but not to anti-mu, gamma, alpha, or lambda antibodies. Both myeloma proteins were purified by (NH4)<em>2</em>SO4 precipitation, ion exchange chromatography, gel filtration, and Staphylococcus aureus Protein A absorption. These IgD kappa myeloma proteins were used to prepare affinity purified rabbit antibodies to <em>delta</em>-chain and the TEPC-1017 and TEPC-1033 idiotypes. Native TEPC-1017 and TEPC-1033 both had mobilities between those of mouse IgA kappa <em>dimers</em> and trimers when analyzed by polyacrylamide gradient gel electrophoresis. Both IgD myeloma proteins broke down under mild reducing conditions into subunits with electrophoretic mobilities slightly slower than those of an IgA kappa monomer. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of reduced TEPC-1017 and TEPC-1033 demonstrated kappa-chains and heavy chains that co-migrated with alpha chain. These data suggested that secreted IgD contains <em>2</em> <em>delta</em> <em>2</em> kappa <em>2</em> subunits that are linked by an easily reducible disulfide bond. The kappa-chains of IgD secreted by TEPC-1017 and TEPC-1033 have apparent m.w. of approximately 63,000 daltons, whereas the apparent m.w. of intracytoplasmic <em>delta</em>-chain, intracytoplasmic <em>delta</em>-chain synthesized in the presence of tunicamycin, and the cellfree translation product of TEPC-1017 <em>delta</em>-chain mRNA are 54,000, 43,000, and 44,000 daltons, respectively. This is compatible with the interpretation that the <em>delta</em>-chain peptide has a leader sequence and is N-glycosylated during or shortly after peptide synthesis and is glycosylated further shortly before IgD secretion.

DNA triplex an<em>d</em> qua<em>d</em>ruplex structures have been successfully <em>d</em>etecte<em>d</em> by electrospray ionization mass spectrometry (ESI-MS). Circular <em>d</em>ichroism an<em>d</em> UV-melting experiments show that these structures are stable in 150 mM ammonium acetate at pH 7 for the qua<em>d</em>ruplexes an<em>d</em> pH 5.5 for the triplexes. The stu<em>d</em>ie<em>d</em> qua<em>d</em>ruplexes were the tetramer [<em>d</em>(TGGGGT)](4), the <em>dimer</em> [<em>d</em>(GGGGTTTTGGGG)](<em>2</em>), an<em>d</em> the intramolecular fol<em>d</em>e<em>d</em> stran<em>d</em> <em>d</em>GGG(TTAGGG)(3), which is an analog of the human telomeric sequence. The absence of so<em>d</em>ium contamination allowe<em>d</em> <em>d</em>emonstration of the specific inclusion of n - 1 ammonium cations in the qua<em>d</em>ruplex structures, where n is the number of consecutive G-tetra<em>d</em>s. We also <em>d</em>etecte<em>d</em> the complexes between the qua<em>d</em>ruplexes an<em>d</em> the qua<em>d</em>ruplex-specific <em>d</em>rug mesoporphyrin IX. MS/MS spectra of [<em>d</em>(TGGGGT)](4) an<em>d</em> the complex with the <em>d</em>rug are also reporte<em>d</em>. As the <em>d</em>rug <em>d</em>oes not <em>d</em>isplace the ammonium cations, one can conclu<em>d</em>e that the <em>d</em>rug bin<em>d</em>s at the exterior of the tetra<em>d</em>s, an<em>d</em> not between them. For the triplex structure the ESI-MS spectra show the <em>d</em>etection of the specific triplex, at m/z values typically higher than those typically observe<em>d</em> for <em>d</em>uplex species. Upon MS/MS the antigene stran<em>d</em>, which is boun<em>d</em> into the major groove of the <em>d</em>uplex, separates from the triplex. This is the same <em>d</em>issociation pathway as in solution. To our knowle<em>d</em>ge this is the first report of a triplex DNA structure by electrospray mass spectrometry.

The <em>d</em>issociation kinetics of a series of complementary an<em>d</em> noncomplementary DNA <em>d</em>uplexes, (TGCA)(<em>2</em>) (3-), (CCGG)(<em>2</em>) (3-), (AATTAAT)(<em>2</em>) (3-), (CCGGCCG)(<em>2</em>) (3-), A(7)*T(7) (3-), A(7)*A(7) (3-), T(7)*T(7) (3-), an<em>d</em> A(7)*C(7) (3-) were investigate<em>d</em> using blackbo<em>d</em>y infrare<em>d</em> ra<em>d</em>iative <em>d</em>issociation in a Fourier transform mass spectrometer. From the temperature <em>d</em>epen<em>d</em>ence of the unimolecular <em>d</em>issociation rate constants, Arrhenius activation parameters in the zero-pressure limit are obtaine<em>d</em>. Activation energies range from 1.<em>2</em> to 1.7 eV, an<em>d</em> preexponential factors range from 10(13) to 10(19) s(-1). Dissociation of the <em>d</em>uplexes results in cleavage of the noncovalent bon<em>d</em>s an<em>d</em>/or cleavage of covalent bon<em>d</em>s lea<em>d</em>ing to loss of a neutral nucleobase followe<em>d</em> by backbone cleavage pro<em>d</em>ucing sequence-specific (a - base) an<em>d</em> w ions. Four pieces of evi<em>d</em>ence are presente<em>d</em> which in<em>d</em>icate that Watson-Crick (WC) base pairing is preserve<em>d</em> in complementary DNA <em>d</em>uplexes in the gas phase: i. the activation energy for <em>d</em>issociation of the complementary <em>dimer</em>, A(7)*T(7) (3-), to the single stran<em>d</em>s is significantly higher than that for the relate<em>d</em> noncomplementary A(7)*A(7) (3-) an<em>d</em> T(7)*T(7) (3-) <em>dimers</em>, in<em>d</em>icating a stronger interaction between stran<em>d</em>s with a specific base sequence, ii. extensive loss of neutral a<em>d</em>enine occurs for A(7)*A(7) (3-) an<em>d</em> A(7)*C(7) (3-) but not for A(7)*T(7) (3-) consistent with this process being shut <em>d</em>own by WC hy<em>d</em>rogen bon<em>d</em>ing, iii. a correlation is observe<em>d</em> between the measure<em>d</em> activation energy for <em>d</em>issociation to single stran<em>d</em>s an<em>d</em> the <em>dimer</em>ization enthalpy (-DeltaH(<em>d</em>)) in solution, an<em>d</em> iv. molecular <em>d</em>ynamics carrie<em>d</em> out at 300 an<em>d</em> 400 K in<em>d</em>icate that WC base pairing is preserve<em>d</em> for A(7)*T(7) (3-) <em>d</em>uplex, although the helical structure is essentially lost. In combination, these results provi<em>d</em>e strong evi<em>d</em>ence that WC base pairing can exist in the complete absence of solvent.

OBJECTIVE

To determine the long-term safety and efficacy of rilonacept, an anti-interleukin-1 fusion protein, in patients with active systemic juvenile idiopathic arthritis (JIA).

METHODS

In patients with systemic JIA, ages 4-20 years, the efficacy of rilonacept was evaluated using 30%, 50%, and 70% levels of improvement according to the adapted American College of Rheumatology (ACR) Pediatric 30, 50, and 70 response criteria, respectively. Efficacy and safety were evaluated during 23 months of open-label treatment (3 phases) after a 4-week, double-blind, placebo-controlled phase. Following double-blind treatment with 2.2 mg/kg or 4.4 mg/kg of rilonacept, patients were eligible to receive open-label treatment at their prior dose, with adjustments. Reductions in the median daily dose of oral prednisone and improvements in laboratory parameters of disease activity (i.e., decreased levels of D-dimer and myeloid-related proteins [MRPs]) were also evaluated.

RESULTS

Twenty-four patients entered the double-blind study and 23 entered the open-label period. Patients were predominantly white and female, and had a median age of 14.0 years at baseline. No significant differences in efficacy were observed between the rilonacept- and placebo-treated patients during the double-blind phase, but fever and rash completely resolved by month 3 in all patients during the open-label treatment period and did not recur. Adapted ACR Pediatric 30, 50, and 70 response rates at 3 months from the start of the study were 78.3%, 60.9%, and 34.8%, respectively; these responses were generally maintained over the study duration. Levels of D-dimer and MRP-8/MRP-14 dramatically improved during the study, and in 22 of 23 patients, the prednisone dose was decreased or prednisone therapy was discontinued. No serious treatment-related adverse events were observed.

CONCLUSIONS

Sustained improvements in clinical and laboratory measures of the articular and systemic manifestations of systemic JIA were achieved in >50% of rilonacept-treated patients over 2 years. Treatment with rilonacept had a substantial steroid-sparing effect and was generally well-tolerated.

Three-dimensional crystal structures of holo (ternary complex enzyme-NA<em>D</em>-azide) and apo NA<em>D</em>-dependent <em>dimer</em>ic formate dehydrogenase (F<em>D</em>H) from the methylotrophic bacterium Pseudomonas sp. 101 have been refined to R factors of 11.7% and 14.8% at <em>2</em>.05 and 1.80 A resolution, respectively. The estimated root-mean-square error in atomic co-ordinates is 0.11 A for holo and 0.18 A for apo. X-ray data were collected from single crystals using an imaging plate scanner and synchrotron radiation. In both crystal forms there is a <em>dimer</em> in the asymmetric unit. Both structures show essentially <em>2</em>-fold molecular symmetry. NA<em>D</em> binding causes movement of the catalytic domain and ordering of the C terminus, where a new helix appears. This completes formation of the enzyme active centre in holo F<em>D</em>H. NA<em>D</em> is bound in the cleft separating the domains and mainly interacts with residues from the co-enzyme binding domain. In apo F<em>D</em>H these residues are held in essentially the same conformation by water molecules occupying the NA<em>D</em> binding region. An azide molecule is located near the point of catalysis, the C4 atom of the nicotinamide moiety of NA<em>D</em>, and overlaps with the proposed formate binding site. There is an extensive channel running from the active site to the protein surface and this is supposed to be used by substrate to reach the active centre after NA<em>D</em> has already bound. The structure of the active site and a hypothetical catalytic mechanism are discussed. Sequence homology of F<em>D</em>H with other NA<em>D</em>-dependent formate dehydrogenases and some <em>D</em>-specific dehydrogenases is discussed on the basis of the F<em>D</em>H three-dimensional structure.

Glycation of proteins leads to the formation of advanced glycation endproducts (AGEs) of diverse molecular structure and biological function. Serum albumin derivatives modified to minimal and high extents by methylglyoxal and glucose in vitro have been used in many studies as model AGE proteins. The early and advanced glycation adduct contents of these proteins were investigated using the 6-aminoquinolyl-N-hydroxysuccinimidyl-carbamate (AQC) chromatographic assay of enzymic hydrolysates. AGEs derived from methylglyoxal, glyoxal and 3-deoxyglucosone, the hydroimidazolones N(<em>delta</em>)-(5-hydro-5-methyl-4-imidazolon-<em>2</em>-yl)-ornithine (MG-H1), N(<em>delta</em>)-(5-hydro-4-imidazolon-<em>2</em>-yl)ornithine (G-H1) and N(<em>delta</em>)-[5-(<em>2</em>,3,4-trihydroxybutyl)-5-hydro-4-imidazolon-<em>2</em>-yl]ornithine (3DG-H1), bis(lysyl)imidazolium cross-links methylglyoxal-derived lysine <em>dimer</em> (MOLD), glyoxal-derived lysine <em>dimer</em> (GOLD), 3-deoxyglucosone-derived lysine <em>dimer</em> (DOLD), monolysyl adducts N(epsilon)-(1-carboxyethyl)lysine (CEL), N(epsilon)-carboxymethyl-lysine (CML) and pyrraline, other AGEs, N(<em>delta</em>)-(4-carboxy-4,6-dimethyl-5,6-dihydroxy-1,4,5,6-tetrahydropyrimidin-<em>2</em>-yl)ornithine (THP), argpyrimidine and pentosidine, and fructosyl-lysine were determined. AGEs with intrinsic fluorescence (argpyrimidine and pentosidine) were assayed without derivatization. Human serum albumin (HSA) glycated minimally by methylglyoxal in vitro contained mainly MG-H1 with minor amounts of THP and argpyrimidine. Similar AGEs were found in prothrombin glycated minimally by methylglyoxal and in N(alpha)-t-butyloxycarbonyl-arginine incubated with methylglyoxal. HSA glycated highly by methylglyoxal contained mainly argpyrimidine, MG-H1 and THP, with minor amounts of CEL and MOLD. HSA glycated minimally by glucose in vitro contained mainly fructosyl-lysine and CML, with minor amounts of THP, MG-H1, G-H1, 3DG-H1, argpyrimidine and DOLD. HSA glycated highly by glucose contained these AGEs and pyrraline, and very high amounts ( approximately 8 mol/mol of protein) of fructosyl-lysine. Most AGEs in albumin glycated minimally by methylglyoxal and glucose were identified. Significant proportions of arginine and lysine-derived AGEs in albumin modified highly by methylglyoxal, and lysine-derived AGEs in albumin modified highly by glucose, remain to be identified.

The platelet-derived growth factor (P<em>D</em>GF) proteins are potent stimulators of cell proliferation/transformation and play a major role in cell-cell communication. For over two decades, P<em>D</em>GFs were thought to exist as three dimeric polypeptides (the homo<em>dimers</em> AA and BB and the heterodimer AB). Recently, however, the P<em>D</em>GF C and <em>D</em> chains were discovered in a BLAST search of the expressed sequence tag databases. The P<em>D</em>GF CC and <em>D</em><em>D</em> <em>dimers</em> have a unique two-domain structure with an NH(<em>2</em>)-terminal CUB (compliment subcomponents C1r/C1s, Uegf, and Bmp1) domain and a COOH-terminal P<em>D</em>GF/vascular endothelial growth factor domain. Whereas secreted P<em>D</em>GF AA, BB, and AB readily activate their cell surface receptors, it was suggested that extracellular proteolytic removal of the CUB domain is required for the P<em>D</em>GF/vascular endothelial growth factor domain of P<em>D</em>GF CC and <em>D</em><em>D</em> to activate P<em>D</em>GF receptors. In the present study, we examined the processing of latent P<em>D</em>GF <em>D</em> into its active form and the effects of P<em>D</em>GF <em>D</em> expression on prostate cancer progression. We show that LNCaP cells auto-activate latent P<em>D</em>GF <em>D</em><em>D</em> into the active P<em>D</em>GF domain, which can induce phosphorylation of the beta-P<em>D</em>GF receptor and stimulates LNCaP cell proliferation in an autocrine manner. Additionally, LNCaP-P<em>D</em>GF <em>D</em>-conditioned medium induces migration of the prostate fibroblast cell line 153<em>2</em>-FTX, indicating LNCaP-processed P<em>D</em>GF <em>D</em><em>D</em> is active in a paracrine manner as well. In a severe combined immunodeficient mouse model, P<em>D</em>GF <em>D</em><em>D</em> expression accelerates early onset of prostate tumor growth and drastically enhances prostate carcinoma cell interaction with surrounding stromal cells. These demonstrate a potential oncogenic activity of P<em>D</em>GF <em>D</em><em>D</em> in the development and/or progression of prostate cancer.

BACKGROUND

Thrombosis is a pivotal event in the pathogenesis of coronary disease. We hypothesized that the presence of blood factors that reflect enhanced thrombogenic activity would be associated with an increased risk of recurrent coronary events during long-term follow-up of patients who have recovered from myocardial infarction.

RESULTS

We prospectively enrolled 1045 patients <em>2</em> months after an index myocardial infarction. Baseline thrombogenic blood tests included 6 hemostatic variables (<em>D</em>-<em>dimer</em>, fibrinogen, factor VII, factor VIIa, von Willebrand factor, and plasminogen activator inhibitor-1), 7 lipid factors [cholesterol, triglycerides, H<em>D</em>L cholesterol, L<em>D</em>L cholesterol, lipoprotein(a), apolipoprotein (apo)A-I, and apoB], and insulin. Patients were followed up for an average of <em>2</em>6 months, with the primary end point being coronary death or nonfatal myocardial infarction, whichever occurred first. The hemostatic, lipid, and insulin parameters were dichotomized into their top and the lower 3 risk quartiles and evaluated for entry into a Cox survivorship model. High levels of <em>D</em>-<em>dimer</em> (hazard ratio, <em>2</em>.43; 95% CI, 1.49, 3.97) and apoB (hazard ratio, 1.8<em>2</em>; 95% CI, 1.10, 3.00) and low levels of apoA-I (hazard ratio, 1.84; 95% CI, 1.10, 3.08) were independently associated with recurrent coronary events in the Cox model after adjustment for 6 relevant clinical covariates.

CONCLUSIONS

Our findings indicate that a procoagulant state, as reflected in elevated levels of D-dimer, and disordered lipid transport, as indicated by low apoA-1 and high apoB levels, contribute independently to recurrent coronary events in postinfarction patients.

OBJECTIVE

Inflammatory stimuli profoundly increase the vulnerability of the vessel wall to atherogenesis. The endothelial glycocalyx, a layer of glycosaminoglycans and proteoglycans covering the luminal side of the vasculature, has recently emerged as an orchestrator of vascular homeostasis. In the present study, we investigated whether endotoxin-induced inflammatory reactions lead to a decrease of endothelial glycocalyx thickness in humans and whether tumor necrosis factor-alpha (TNFalpha) plays a role in this process.

METHODS

Healthy male volunteers received low-dose endotoxin (1ng/kg) intravenously, with (n=8) or without (n=13) pre-treatment with the soluble TNFalpha receptor etanercept. Endothelial glycocalyx thickness and related parameters were determined after endotoxin challenge.

RESULTS

En<em>d</em>otoxin resulte<em>d</em> in a profoun<em>d</em> re<em>d</em>uction in microvascular glycocalyx thickness (from 0.60+/-0.1 to 0.30+/-0.1microm, p<0.01). Concomitantly, plasma levels of the principal glycocalyx constituent hyaluronan (6<em>2</em>+/-18 to 85+/-<em>2</em>4ng/mL, p<0.05), monocyte activation an<em>d</em> coagulation activation increase<em>d</em> (F1+<em>2</em>; 0.3+/-0.1 to <em>2</em>.8+/-1.5nmol/L, p<0.05 an<em>d</em> <em>d</em>-<em>dimer</em>; from 0.<em>2</em>+/-0.1 to 0.4+/-0.1mg/L, p<0.05 compare<em>d</em> to baseline). Inhibition of TNFalpha by etanercept attenuate<em>d</em> loss of microvascular glycocalyx thickness (0.54+/-0.1 to 0.35+/-0.1mum, p<0.05). Changes in hyaluronan (58+/-13 to 46+/-10ng/mL, p<0.05) an<em>d</em> coagulation activation were also attenuate<em>d</em> (F1+<em>2</em>; 0.3+/-0.1 to <em>2</em>.1+/-0.9nmol/L an<em>d</em> <em>d</em>-<em>dimer</em>; from 0.<em>2</em>+/-0.1 to 0.3+/-0.1mg/L, p<0.05 compare<em>d</em> to baseline).

CONCLUSIONS

These data suggest that inflammatory activity, in part mediated by TNFalpha, leads to perturbation of the endothelial glycocalyx in humans. This may contribute to the vascular vulnerability induced by inflammation.

Creatine an<em>d</em> phosphocreatine were evaluate<em>d</em> for their ability to prevent <em>d</em>eath of culture<em>d</em> striatal an<em>d</em> hippocampal neurons expose<em>d</em> to either glutamate or 3-nitropropionic aci<em>d</em> (3NP) an<em>d</em> to inhibit the mitochon<em>d</em>rial permeability transition in CNS mitochon<em>d</em>ria. Phosphocreatine (PCr), an<em>d</em> to a lesser extent creatine (Cr), but not (5R,10S)-(+)-5-methyl-10,11-<em>d</em>ihy<em>d</em>ro-5H-<em>d</em>ibenzo[a,<em>d</em>]cyclohepten-5,10-imine hy<em>d</em>rogen maleate (MK801), <em>d</em>ose-<em>d</em>epen<em>d</em>ently ameliorate<em>d</em> 3NP toxicity when applie<em>d</em> simultaneously with the 3NP in Mg<em>2</em>+-free me<em>d</em>ia. Pre-treatment of PCr for <em>2</em> or 5 <em>d</em>ays an<em>d</em> Cr for 5 <em>d</em>ays protecte<em>d</em> against glutamate excitotoxicity equivalent to that achieve<em>d</em> by MK801 post-treatment. The combination of PCr or Cr pre-treatment an<em>d</em> MK801 post-treatment <em>d</em>i<em>d</em> not provi<em>d</em>e a<em>d</em><em>d</em>itional protection, in<em>d</em>icating that both prevente<em>d</em> the toxicity attributable to secon<em>d</em>ary glutamate release. To <em>d</em>etermine if Cr or PCr <em>d</em>irectly inhibite<em>d</em> the permeability transition, mitochon<em>d</em>rial swelling an<em>d</em> <em>d</em>epolarization were assaye<em>d</em> in isolate<em>d</em>, purifie<em>d</em> brain mitochon<em>d</em>ria. PCr re<em>d</em>uce<em>d</em> the amount of swelling in<em>d</em>uce<em>d</em> by calcium by <em>2</em>0%. Cr <em>d</em>ecrease<em>d</em> mitochon<em>d</em>rial swelling when inhibitors of creatine kinase octamer-<em>dimer</em> transition were present. However, in brain mitochon<em>d</em>ria prepare<em>d</em> from rats fe<em>d</em> a <em>d</em>iet supplemente<em>d</em> with <em>2</em>% creatine for <em>2</em> weeks, the extent of calcium-in<em>d</em>uce<em>d</em> mitochon<em>d</em>rial swelling was not altere<em>d</em>. Thus, the neuroprotective properties of PCr an<em>d</em> Cr may reflect enhancement of cytoplasmic high-energy phosphates but not permeability transition inhibition.

The location of the 1:<em>2</em> borate-diol ester cross-link in the <em>dimer</em> of the plant cell wall polysaccharide rhamnogalacturonan II (RG-II) has been determined. The ester cross-links the apiofuranosyl residue of the <em>2</em>-O-methyl-<em>D</em>-xylose-containing side chains in each of the subunits of the <em>dimer</em>. The apiofuranosyl residue in each of the two aceric acid-containing side chains is not esterified. The site of borate esterification is identical in naturally occurring and in in vitro synthesized <em>dimer</em>. Pb<em>2</em>+, La3+, and Ca<em>2</em>+ increase <em>dimer</em> formation in vitro in a concentration- and pH-dependent manner. Pb<em>2</em>+ is the most effective cation. The <em>dimer</em> accounts for 55% of the RG-II when the monomer (0.5 mM) is treated for 5 min at pH 3.5 with boric acid (1 mM) and Pb<em>2</em>+ (0.5 mM); at pH 5 the rate of conversion is somewhat slower. Hg<em>2</em>+ does not increase the rate of <em>dimer</em> formation. A cation's charge density and its ability to form a coordination complex with RG-II, in addition to steric factors, may regulate the rate and stability of <em>dimer</em> formation in vitro. Our data provide evidence that the structure of RG-II itself determines which apiofuranosyl residues are esterified with borate and that in the presence of boric acid and certain cations, two RG-II monomers self-assemble to form a <em>dimer</em>.

Presente<em>d</em> is an extension of the CHARMM a<em>d</em><em>d</em>itive carbohy<em>d</em>rate all-atom force fiel<em>d</em> to enable mo<em>d</em>eling of polysacchari<em>d</em>es containing furanose sugars. The new force fiel<em>d</em> parameters encompass 1 ↔ <em>2</em>, 1 → 3, 1 → 4, an<em>d</em> 1 → 6 pyranose-furanose linkages an<em>d</em> <em>2</em> → 1 an<em>d</em> <em>2</em> → 6 furanose-furanose linkages, buil<em>d</em>ing on existing hexopyranose an<em>d</em> furanose monosacchari<em>d</em>e parameters. The mo<em>d</em>el compoun<em>d</em>s were chosen to be monomers or glycosi<em>d</em>ic-linke<em>d</em> <em>dimers</em> of tetrahy<em>d</em>ropyran (THP) an<em>d</em> tetrahy<em>d</em>rofuran (THF) as to contain the key atoms in full carbohy<em>d</em>rates. Target <em>d</em>ata for optimization inclu<em>d</em>e<em>d</em> two-<em>d</em>imensional quantum mechanical (QM) potential energy scans of the Φ/Ψ glycosi<em>d</em>ic <em>d</em>ihe<em>d</em>ral angles, with geometry optimization at the MP<em>2</em>/6-31G(<em>d</em>) level followe<em>d</em> by MP<em>2</em>/cc-pVTZ single-point energies. All possible chiralities of the mo<em>d</em>el compoun<em>d</em>s at the linkage carbons were consi<em>d</em>ere<em>d</em>, an<em>d</em> for each geometry, the THF ring was constraine<em>d</em> to the favorable south or north conformations. Target <em>d</em>ata also inclu<em>d</em>e<em>d</em> QM vibrational frequencies an<em>d</em> pair interaction energies an<em>d</em> <em>d</em>istances with water molecules. Force fiel<em>d</em> vali<em>d</em>ation inclu<em>d</em>e<em>d</em> comparison of compute<em>d</em> crystal properties, aqueous solution <em>d</em>ensities, an<em>d</em> NMR J-coupling constants to experimental reference values. Simulations of infinite crystals showe<em>d</em> goo<em>d</em> agreement with experimental values for intramolecular geometries as well as for crystal unit cell parameters. A<em>d</em><em>d</em>itionally, aqueous solution <em>d</em>ensities an<em>d</em> available NMR <em>d</em>ata were repro<em>d</em>uce<em>d</em> to a high <em>d</em>egree of accuracy, thus vali<em>d</em>ating the hierarchically optimize<em>d</em> parameters in both crystalline an<em>d</em> aqueous con<em>d</em>ense<em>d</em> phases. The newly <em>d</em>evelope<em>d</em> parameters allow for the mo<em>d</em>eling of linear, branche<em>d</em>, an<em>d</em> cyclic pyranose/furanose polysacchari<em>d</em>es both alone an<em>d</em> in heterogeneous systems inclu<em>d</em>ing proteins, nucleic aci<em>d</em>s, an<em>d</em>/or lipi<em>d</em>s when combine<em>d</em> with existing a<em>d</em><em>d</em>itive CHARMM biomolecular force fiel<em>d</em>s.

L-<em>2</em>-Haloacid dehalogenase catalyzes the hydrolytic dehalogenation of L-<em>2</em>-haloalkanoic acids to yield the corresponding <em>D</em>-<em>2</em>-hydroxyalkanoic acids. The crystal structure of the homo<em>dimer</em>ic enzyme from Pseudomonas sp. YL has been determined by a multiple isomorphous replacement method and refined at <em>2</em>.5 A resolution to a crystallographic R-factor of 19.5%. The subunit consists of two structurally distinct domains: the core domain and the subdomain. The core domain has an alpha/beta structure formed by a six-stranded parallel beta-sheet flanked by five alpha-helices. The subdomain inserted into the core domain has a four helix bundle structure providing the greater part of the interface for <em>dimer</em> formation. There is an active site cavity between the domains. An experimentally identified nucleophilic residue, Asp-10, is located on a loop following the amino-terminal beta-strand in the core domain, and other functional residues, Thr-14, Arg-41, Ser-118, Lys-151, Tyr-157, Ser-175, Asn-177, and Asp-180, detected by a site-directed mutagenesis experiment, are arranged around the nucleophile in the active site. Although the enzyme is an alpha/beta-type hydrolase, it does not belong to the alpha/beta hydrolase fold family, from the viewpoint of the topological feature and the position of the nucleophile.

OBJECTIVE

To review the diagnostic accuracy of D-dimer testing in older patients (>50 years) with suspected venous thromboembolism, using conventional or age adjusted D-dimer cut-off values.

METHODS

Systematic review and bivariate random effects meta-analysis.

METHODS

We searched Medline and Embase for studies published before 21 June 2012 and we contacted the authors of primary studies.

METHODS

Primary studies that enrolled older patients with suspected venous thromboembolism in whom D-dimer testing, using both conventional (500 µg/L) and age adjusted (age × 10 µg/L) cut-off values, and reference testing were performed. For patients with a non-high clinical probability, 2 × 2 tables were reconstructed and stratified by age category and applied D-dimer cut-off level.

RESULTS

13 cohorts including 12,497 patients with a non-high clinical probability were included in the meta-analysis. The specificity of the conventional cut-off value decreased with increasing age, from 57.6% (95% confidence interval 51.4% to 63.6%) in patients aged 51-60 years to 39.4% (33.5% to 45.6%) in those aged 61-70, 24.5% (20.0% to 29.7% in those aged 71-80, and 14.7% (11.3% to 18.6%) in those aged >80. Age adjusted cut-off values revealed higher specificities over all age categories: 62.3% (56.2% to 68.0%), 49.5% (43.2% to 55.8%), 44.2% (38.0% to 50.5%), and 35.2% (29.4% to 41.5%), respectively. Sensitivities of the age adjusted cut-off remained above 97% in all age categories.

CONCLUSIONS

The application of age adjusted cut-off values for D-dimer tests substantially increases specificity without modifying sensitivity, thereby improving the clinical utility of D-dimer testing in patients aged 50 or more with a non-high clinical probability.

<strong class="sub-title"> Background: </strong> Tocilizumab, an IL-6 receptor antagonist, can be used to treat cytokine release syndrome (CRS), with observed improvements in a coronavirus disease <em>2</em>019 (COVI<em>D</em>-19) case series.

Research question: The goal of this study was to determine if tocilizumab benefits patients hospitalized with COVID-19.

<strong class="sub-title"> Study design and methods: </strong> This observational study of consecutive COVI<em>D</em>-19 patients hospitalized between March 10, <em>2</em>0<em>2</em>0, and March 31, <em>2</em>0<em>2</em>0, and followed up through April <em>2</em>1, <em>2</em>0<em>2</em>0, was conducted by chart review. Patients were treated with tocilizumab using an algorithm that targeted CRS. Survival and mechanical ventilation (MV) outcomes were reported for 14 days and stratified according to disease severity designated at admission (severe, ≥ 3 L supplemental oxygen to maintain oxygen saturation > 93%). For tocilizumab-treated patients, pre/post analyses of clinical response, biomarkers, and safety outcomes were assessed. Post hoc survival analyses were conducted for race/ethnicity.

<strong class="sub-title"> Results: </strong> Among the <em>2</em>39 patients, median age was 64 years; 36% and 19% were black and Hispanic, respectively. Hospital census increased exponentially, yet MV census did not. Severe disease was associated with lower survival (78% vs 93%; P < .001), greater proportion requiring MV (44% vs 5%; P < .001), and longer median MV days (5.5 vs 1.0; P = .003). Tocilizumab-treated patients (n = 153 [64%]) comprised 90% of those with severe disease; 44% of patients with nonsevere disease received tocilizumab for evolving CRS. Tocilizumab-treated patients with severe disease had higher admission levels of high-sensitivity C-reactive protein (1<em>2</em>0 vs 71 mg/L; P < .001) and received tocilizumab sooner (<em>2</em> vs 3 days; P < .001), but their survival was similar to that of patients with nonsevere disease (83% vs 91%; P = .11). For tocilizumab-treated patients requiring MV, survival was 75% (95% CI, 64-89). Following tocilizumab treatment, few adverse events occurred, and oxygenation and inflammatory biomarkers (eg, high-sensitivity C-reactive protein, IL-6) improved; however, <em>D</em>-<em>dimer</em> and soluble IL-<em>2</em> receptor (also termed C<em>D</em><em>2</em>5) levels increased significantly. Survival in black and Hispanic patients, after controlling for age, was significantly higher than in white patients (log-rank test, P = .00<em>2</em>).

Interpretation: A treatment algorithm that included tocilizumab to target CRS may influence MV and survival outcomes. In tocilizumab-treated patients, oxygenation and inflammatory biomarkers improved, with higher than expected survival. Randomized trials must confirm these findings.

Keywords: COVID-19; cytokine release syndrome; disease severity; mechanical ventilation; survival; tocilizumab.

Mevalonate kinase catalyzes the ATP-dependent phosphorylation of mevalonic acid to form mevalonate 5-phosphate, a key intermediate in the pathways of isoprenoids and sterols. <em>D</em>eficiency in mevalonate kinase activity has been linked to mevalonic aciduria and hyperimmunoglobulinemia <em>D</em>/periodic fever syndrome (HI<em>D</em>S). The crystal structure of rat mevalonate kinase in complex with MgATP has been determined at <em>2</em>.4-A resolution. Each monomer of this <em>dimer</em>ic protein is composed of two domains with its active site located at the domain interface. The enzyme-bound ATP adopts an anti conformation, in contrast to the syn conformation reported for Methanococcus jannaschii homoserine kinase. The Mg(<em>2</em>+) ion is coordinated to both beta- and gamma-phosphates of ATP and side chains of Glu(193) and Ser(146). Asp(<em>2</em>04) is making a salt bridge with Lys(13), which in turn interacts with the gamma-phosphate. A model of mevalonic acid can be placed near the gamma-phosphoryl group of ATP; thus, the C5 hydroxyl is located within 4 A from Asp(<em>2</em>04), Lys(13), and the gamma-phosphoryl of ATP. This arrangement of residues strongly suggests: 1) Asp(<em>2</em>04) abstracts the proton from C5 hydroxyl of mevalonate; <em>2</em>) the penta-coordinated gamma-phosphoryl group may be stabilized by Mg(<em>2</em>+), Lys(13), and Glu(193); and 3) Lys(13) is likely to influence the pK(a) of the C5 hydroxyl of the substrate. V377I and I<em>2</em>68T are the most common mutations found in patients with HI<em>D</em>S. Val(377) is located over 18 A away from the active site and a conservative replacement with Ile is unlikely to yield an inactive or unstable protein. Ile-<em>2</em>68 is located at the <em>dimer</em> interface, and its Thr substitution may disrupt <em>dimer</em> formation.

Galectin-1 is a member of the galectin family of glycan-bin<em>d</em>ing proteins an<em>d</em> occurs as an approximately <em>2</em>9.5-kDa noncovalent homo<em>dimer</em> (<em>d</em>Gal-1) that is wi<em>d</em>ely expresse<em>d</em> in many tissues. Here, we report that human recombinant <em>d</em>Gal-1 boun<em>d</em> preferentially an<em>d</em> with high affinity (apparent K(<em>d</em>) approximately <em>2</em>-4 microM) to immobilize<em>d</em> exten<em>d</em>e<em>d</em> glycans containing terminal N-acetyllactosamine (LN; Galbeta1-4GlcNAc) sequences on poly-N-acetyllactosamine (PL; (-3Galbeta1-4GlcNAcbeta1-)(n)) sequences, complex-type biantennary N-glycans, or novel chitin-<em>d</em>erive<em>d</em> glycans mo<em>d</em>ifie<em>d</em> to contain terminal LN. Although terminal Gal resi<em>d</em>ues are important for <em>d</em>Gal-1 recognition, <em>d</em>Gal-1 boun<em>d</em> similarly to alpha3-sialylate<em>d</em> an<em>d</em> alpha<em>2</em>-fucosylate<em>d</em> terminal LN, but not to alpha6-sialylate<em>d</em> an<em>d</em> alpha3-fucosylate<em>d</em> terminal LN. The bin<em>d</em>ing specificity of human recombinant <em>d</em>Gal-1 was similar to that observe<em>d</em> with purifie<em>d</em> bovine heart-<em>d</em>erive<em>d</em> <em>d</em>Gal-1. Unexpecte<em>d</em>ly, <em>d</em>Gal-1 boun<em>d</em> free ligan<em>d</em>s in solution with relatively low affinity an<em>d</em> <em>d</em>isplaye<em>d</em> no preference for exten<em>d</em>e<em>d</em> glycans, in<em>d</em>icating that <em>d</em>Gal-1 preferentially recognizes exten<em>d</em>e<em>d</em> glycans only when they are surface-boun<em>d</em>, such as foun<em>d</em> on cell surfaces. Human <em>d</em>Gal-1 also boun<em>d</em> to both native an<em>d</em> <em>d</em>esialylate<em>d</em> human promyelocytic HL-60 cells with similar affinity as observe<em>d</em> for immobilize<em>d</em> long chain PL. Bin<em>d</em>ing to these cells was re<em>d</em>uce<em>d</em> upon treatment with en<em>d</em>o-beta-galactosi<em>d</em>ase, which cleaves PL sequences, in<em>d</em>icating that cell-surface PLs are ligan<em>d</em>s. To test the role of <em>dimer</em>ization in <em>d</em>Gal-1 bin<em>d</em>ing, we examine<em>d</em> the bin<em>d</em>ing of a mutate<em>d</em> form of <em>d</em>Gal-1 that weakly <em>dimer</em>izes (monomeric Gal-1 (mGal-1)) an<em>d</em> a covalently <em>dimer</em>ize<em>d</em> (chemically cross-linke<em>d</em>) form of mGal-1 (c<em>d</em>-mGal-1). <em>d</em>Gal-1 an<em>d</em> c<em>d</em>-mGal-1 ha<em>d</em> similar affinities that were both approximately 3.5-fol<em>d</em> higher for immobilize<em>d</em> PL than observe<em>d</em> for mGal-1, suggesting that <em>d</em>Gal-1 acts as a <em>dimer</em> to cross-link terminal LN units on immobilize<em>d</em> PL. These results in<em>d</em>icate that <em>d</em>Gal-1 functions as a <em>dimer</em> to recognize LN units on exten<em>d</em>e<em>d</em> PLs on cell surfaces.

Calsenilin/DREAM/KChIP3, a member of the recoverin branch of the EF-han<em>d</em> superfamily, interacts with presenilins, serves as a calcium-regulate<em>d</em> transcriptional repressor, an<em>d</em> interacts with A-type potassium channels. Here we report physicochemical characterization of calcium bin<em>d</em>ing, oligomerization, an<em>d</em> DNA bin<em>d</em>ing of human calsenilin/DREAM/KChIP3. Equilibrium Ca(<em>2</em>+) bin<em>d</em>ing measurements in<em>d</em>icate that the protein bin<em>d</em>s 3 Ca(<em>2</em>+) with a <em>d</em>issociation constant of 14 microM an<em>d</em> a Hill coefficient of 0.7. Dynamic light scattering an<em>d</em> size exclusion chromatography show that the Ca(<em>2</em>+)-boun<em>d</em> protein exists as a <em>dimer</em> at protein concentrations lower than 150 microM an<em>d</em> forms a tetramer at concentrations above <em>2</em>00 microM. The Ca(<em>2</em>+)-free protein is a tetramer in the concentration range <em>2</em>0-450 microM. Isothermal titration calorimetry an<em>d</em> <em>d</em>ynamic light scattering in<em>d</em>icate that the Ca(<em>2</em>+)-free protein tetramer bin<em>d</em>s en<em>d</em>othermically (DeltaH = +<em>2</em>5 kcal/mol) to four molecules of DNA <em>d</em>erive<em>d</em> from the <em>d</em>ownstream regulatory element (DRE) of either the pro<em>d</em>ynorphin or c-fos genes. One DRE molecule bin<em>d</em>s tightly to the protein with a <em>d</em>issociation constant (K(<em>d</em>)) of 75 nM, an<em>d</em> the other three bin<em>d</em> more weakly (K(<em>d</em>) = 640 nM). No significant DNA bin<em>d</em>ing was observe<em>d</em> for the Ca(<em>2</em>+)-boun<em>d</em> protein. The N-terminal protein fragment (resi<em>d</em>ues 1-70) bin<em>d</em>s nonspecifically to DRE in a Ca(<em>2</em>+)-in<em>d</em>epen<em>d</em>ent manner, whereas a C-terminal fragment containing the four EF-han<em>d</em>s (resi<em>d</em>ues 65-<em>2</em>56) bin<em>d</em>s DRE (K(<em>d</em>) = <em>2</em>00 nM) in a Ca(<em>2</em>+)-regulate<em>d</em> an<em>d</em> sequence-specific fashion. The C-terminal fragment is a tetramer in the Ca(<em>2</em>+)-free state an<em>d</em> <em>d</em>issociates into <em>dimers</em> at saturating Ca(<em>2</em>+) levels.

We analyse<em>d</em> an<em>d</em> compare<em>d</em> the functioning of UV-B screening pigments in plants from marine, fresh water an<em>d</em> terrestrial ecosystems, along the evolutionary line of cyanobacteria, unicellular algae, primitive multicellular algae, charophycean algae, lichens, mosses an<em>d</em> higher plants, inclu<em>d</em>ing amphibious macrophytes. Lichens were also inclu<em>d</em>e<em>d</em> in the stu<em>d</em>y. We were intereste<em>d</em> in the following key aspects: (a) <em>d</em>oes the water column function effectively as an 'external UV-B filter'?; (b) <em>d</em>o aquatic plants nee<em>d</em> less 'internal UV-B screening' than terrestrial plants?; (c) what role <em>d</em>oes UV screening play in protecting the various plant groups from UV-B <em>d</em>amage, such as the formation of thymine <em>dimers</em>?; an<em>d</em> (<em>d</em>) since early lan<em>d</em> 'plants' (such as the pre<em>d</em>ecessors of present-<em>d</em>ay cyanobacteria, lichens an<em>d</em> mosses) experience<em>d</em> higher UV-B fluxes than higher plants, which evolve<em>d</em> later, are primitive aquatic an<em>d</em> lan<em>d</em> organisms (cyanobacteria, algae, lichens, mosses) better a<em>d</em>apte<em>d</em> to present-<em>d</em>ay levels of UV-B than higher plants? Furthermore, polychromatic action spectra for the in<em>d</em>uction of UV screening pigments of aquatic organisms have been <em>d</em>etermine<em>d</em>. This is relevant for translating 'physical' ra<em>d</em>iation measurements of solar UV-B into 'biological' an<em>d</em> 'ecological' effects. From the action spectra, ra<em>d</em>iation amplification factors (RAFs) have been calculate<em>d</em>. These action spectra allow us to <em>d</em>etermine any mitigating or antagonistic effects in the ecosystems an<em>d</em> therefore qualify the <em>d</em>amage pre<em>d</em>iction for the ecosystems un<em>d</em>er stu<em>d</em>y. We summarize an<em>d</em> <em>d</em>iscuss the main results base<em>d</em> on three years of research of four European research groups. The central theme of the work was the investigation of the effectiveness of the various screening compoun<em>d</em>s from the <em>d</em>ifferent species stu<em>d</em>ie<em>d</em> in or<em>d</em>er to gain some perspective of the evolutionary a<em>d</em>aptations from lower to higher plant forms. The in<em>d</em>uction of mycosporine-like amino aci<em>d</em>s (MAAs) was stu<em>d</em>ie<em>d</em> in the marine <em>d</em>inoflagellate Gyro<em>d</em>inium <em>d</em>orsum, the green algal species Prasiola stipitata an<em>d</em> in the cyanobacterium Anabaena sp. While visible (400-700 nm) an<em>d</em> long wavelength UV-A (315-400 nm) showe<em>d</em> only a slight effect, MAAs were effectively in<em>d</em>uce<em>d</em> by UV-B (<em>2</em>80-315 nm). The growth of the lower lan<em>d</em> organisms stu<em>d</em>ie<em>d</em>, i.e. the lichens Cla<em>d</em>ina portentosa, Cla<em>d</em>ina foliacaea an<em>d</em> Cla<em>d</em>onia arbuscula, an<em>d</em> the club moss Lycopo<em>d</em>iumannotinum, was not significantly re<em>d</em>uce<em>d</em> when grown un<em>d</em>er elevate<em>d</em> UV-B ra<em>d</em>iation (simulating 15% ozone <em>d</em>epletion). The growth in length of the moss Tortula ruralis was re<em>d</em>uce<em>d</em> un<em>d</em>er elevate<em>d</em> UV-B. Of the aquatic plants investigate<em>d</em> the charophytes Chara aspera showe<em>d</em> <em>d</em>ecrease<em>d</em> longitu<em>d</em>inal growth un<em>d</em>er elevate<em>d</em> UV-B. In the 'aquatic higher plants' stu<em>d</em>ie<em>d</em>, Ceratophyllum <em>d</em>emersum, Batrachium trichophyllum an<em>d</em> Potamogeton alpinus, there was no such <em>d</em>epresse<em>d</em> growth with enhance<em>d</em> UV-B. In Chara aspera, neither MAAs nor flavonoi<em>d</em>s coul<em>d</em> be <em>d</em>etecte<em>d</em>. Of the terrestrial higher plants stu<em>d</em>ie<em>d</em>, Fagopyrum esculentum, Deschampsia antarctica, Vicia faba, Calamagrostis epigejos an<em>d</em> Carex arenaria, the growth of the first species was <em>d</em>epresse<em>d</em> with enhance<em>d</em> UV-B, in the secon<em>d</em> species length growth was <em>d</em>ecrease<em>d</em>, but the shoot number was increase<em>d</em>, an<em>d</em> in the latter two species of a <em>d</em>une grasslan<em>d</em> there was no re<em>d</em>uce<em>d</em> growth with enhance<em>d</em> UV-B. In the <em>d</em>une grasslan<em>d</em> species stu<em>d</em>ie<em>d</em> out<em>d</em>oors, at least five <em>d</em>ifferent flavonoi<em>d</em>s appeare<em>d</em> in shoot tissue. Some of the flavonoi<em>d</em>s in the monocot species, which were i<em>d</em>entifie<em>d</em> an<em>d</em> quantifie<em>d</em> with HPLC, inclu<em>d</em>e<em>d</em> orientin, luteolin, tricin an<em>d</em> apigenin. A greenhouse stu<em>d</em>y with Vicia faba showe<em>d</em> that two flavonoi<em>d</em>s (aglycones) respon<em>d</em> particularly to enhance<em>d</em> UV-B. Of these, quercetin is UV-B in<em>d</em>ucible an<em>d</em> mainly locate<em>d</em> in epi<em>d</em>ermal cells, while kaempferol occurs constitutively. In a<em>d</em><em>d</em>ition to its UV-screening function, quercetin may also act as an antioxi<em>d</em>ant. Polychromatic action spectra were <em>d</em>etermine<em>d</em> for in<em>d</em>uction of the UV-absorbing pigments in three photosynthetic organisms, representing very <em>d</em>ifferent taxonomic groups an<em>d</em> <em>d</em>ifferent habitats. In ultraviolet photobiology, action spectra mainly serve two purposes: (1) i<em>d</em>entification of the molecular species involve<em>d</em> in light absorption; an<em>d</em> (<em>2</em>) calculation of ra<em>d</em>iation amplification factors for assessing the effect of ozone <em>d</em>epletion. Ra<em>d</em>iation amplification factors (RAFs) were calculate<em>d</em> from the action spectra. In a somewhat simplifie<em>d</em> way, RAF can be <em>d</em>efine<em>d</em> as the percent increase of ra<em>d</em>iation <em>d</em>amage for a 1% <em>d</em>epletion of the ozone layer. Central European summer con<em>d</em>itions were use<em>d</em> in the calculations, but it has been shown that RAF values are not critically <em>d</em>epen<em>d</em>ent on latitu<em>d</em>e or season. If only the ultraviolet spectral region is consi<em>d</em>ere<em>d</em>, the RAF values obtaine<em>d</em> are 0.7 for the green alga Prasiola stipitata, 0.4 for the <em>d</em>inoflagellate Gyro<em>d</em>inium <em>d</em>orsum, an<em>d</em> 1.0 for the cyanobacterium Anabaena sp. In the case of P. stipitata, however, the effect of visible light (PAR, photosynthetically active ra<em>d</em>iation, 400-700 nm) is sufficient to lower the RAF to about 0.4, while the PAR effect for G. <em>d</em>orsum is negligible. RAFs for some <em>d</em>amage processes, such as for DNA <em>d</em>amage (RAF=<em>2</em>.1 if protective effects or photorepair are not consi<em>d</em>ere<em>d</em> [1]), are higher than those above. Our interpretation of this is that if the ozone layer is <em>d</em>eplete<em>d</em>, increase<em>d</em> <em>d</em>amaging ra<em>d</em>iation coul<em>d</em> overrule increase<em>d</em> synthesis of protective pigments. In a<em>d</em><em>d</em>ition to investigating the functional effectiveness of the <em>d</em>ifferent screening compoun<em>d</em>s, <em>d</em>irect UV effects on a number of key processes were also stu<em>d</em>ie<em>d</em> in or<em>d</em>er to gain further insight into the ability of the organisms to withstan<em>d</em> enhance<em>d</em> UV-B ra<em>d</em>iation. To this en<em>d</em>, the temperature-<em>d</em>epen<em>d</em>ent repair of cyclobutane <em>dimers</em> (CPD) an<em>d</em> (6-4) photopro<em>d</em>ucts in<em>d</em>uce<em>d</em> by enhance<em>d</em> UV-B was stu<em>d</em>ie<em>d</em> in Nicotiana tabacum, an<em>d</em> the UV-B in<em>d</em>uction of CPD was stu<em>d</em>ie<em>d</em> in the lichen Cla<em>d</em>onia arbuscula. Also, photosynthesis an<em>d</em> motility were monitore<em>d</em> an<em>d</em> the response relate<em>d</em> to the potential function of the screening compoun<em>d</em>s of the specific organism.

Severe acute respiratory syndrome coronavirus <em>2</em> (SARS-CoV-<em>2</em>) infection can cause acute respiratory distress syndrome, hypercoagulability, hypertension, and multiorgan dysfunction. Effective antivirals with safe clinical profile are urgently needed to improve the overall prognosis. In an analysis of a randomly collected cohort of 1<em>2</em>4 patients with Corona Virus <em>D</em>isease <em>2</em>019 (COVI<em>D</em>-19), we found that hypercoagulability as indicated by elevated concentrations of <em>D</em>-<em>dimers</em> was associated with disease severity. By virtual screening of a U.S. Food and <em>D</em>rug Administration (F<em>D</em>A) approved drug library, we identified an anticoagulation agent dipyridamole (<em>D</em>IP) <i>in silico</i>, which suppressed SARS-CoV-<em>2</em> replication <i>in vitro</i>. In a proof-of-concept trial involving 31 patients with COVI<em>D</em>-19, <em>D</em>IP supplementation was associated with significantly decreased concentrations of <em>D</em>-<em>dimers</em> (<i>P</i><0.05), increased lymphocyte and platelet recovery in the circulation, and markedly improved clinical outcomes in comparison to the control patients. In particular, all 8 of the <em>D</em>IP-treated severely ill patients showed remarkable improvement: 7 patients (87.5%) achieved clinical cure and were discharged from the hospitals while the remaining 1 patient (1<em>2</em>.5%) was in clinical remission.

RNA recognition receptors are important for detection of and response to viral infections. RIG-I and M<em>D</em>A5 are cytoplasmic <em>D</em>EX(<em>D</em>/H) helicase proteins that can induce signaling in response to RNA ligands, including those from viral infections. LGP<em>2</em>, a homolog of RIG-I and M<em>D</em>A5 without the caspase recruitment domain required for signaling, plays an important role in modulating signaling by M<em>D</em>A5 and RIG-I, presumably through heterocomplex formation and/or by serving as a sink for RNAs. Here we demonstrate that LGP<em>2</em> can be coexpressed with RIG-I to inhibit activation of the NF-kappaB reporter expression and that LGP<em>2</em> protein produced in insect cells can bind both single- and double-stranded RNA (dsRNA), with higher affinity and cooperativity for dsRNA. Electron microscopy and image reconstruction were used to determine the shape of the LGP<em>2</em> monomer in the absence of dsRNA and of the <em>dimer</em> complexed to a <em>2</em>7-bp dsRNA. LGP<em>2</em> has striking structural similarity to the helicase domain of the superfamily <em>2</em> <em>D</em>NA helicase, Hef.

<b>Background:</b> The aim of the study was to document cardiovascular clinical findings, cardiac imaging and laboratory markers in children presenting with the novel multisystem inflammatory syndrome (MIS-C) associated with COVI<em>D</em>-19 infection. <b>Methods:</b> A real-time internet-based survey endorsed by the Association for European Paediatric and Congenital Cardiologists (AEPC) Working Groups for Cardiac Imaging and Cardiovascular Intensive Care. Inclusion criteria was children 0-18 years admitted to hospital between February 1 and June 6, <em>2</em>0<em>2</em>0 with diagnosis of an inflammatory syndrome and acute cardiovascular complications. <b>Results:</b> A total of <em>2</em>86 children from 55 centers in 17 European countries were included. The median age was 8.4 years (IQR 3.8-1<em>2</em>.4 years) and 67% were males. The most common cardiovascular complications were shock, cardiac arrhythmias, pericardial effusion and coronary artery dilatation. Reduced left ventricular ejection fraction was present in over half of the patients and a vast majority of children had raised cardiac troponin (cTnT) when checked. The biochemical markers of inflammation were raised in majority of patients on admission: elevated CRP, serum ferritin, procalcitonin, NT-proBNP, IL-6 level and <em>D</em>-<em>dimers</em>. There was a statistically significant correlation between degree of elevation in cardiac and biochemical parameters and need for intensive care support (p <0.05). Polymerase chain reaction (PCR) for SARS-CoV-<em>2</em> was positive in 33.6% while IgM and IgG antibodies were positive in 15.7% and IgG 43.6 % cases, respectively when checked. One child died in the study cohort. <b>Conclusions:</b> Cardiac involvement is common in children with multisystem inflammatory syndrome associated with Covid-19 pandemic. A majority of children have significantly raised levels of NT pro-BNP, ferritin, <em>D</em>-<em>dimers</em> and cardiac troponin in addition to high CRP and procalcitonin levels. Compared to adults with Covid-19, mortality in children with MIS-C is uncommon despite multi-system involvement, very elevated inflammatory markers and need for intensive care support.

<strong class="sub-title"> Keywords: </strong> MIS-C; PIMS-TS; SARS-COV-<em>2</em>.

Atypical hemolytic uremic syn<em>d</em>rome (aHUS) is a genetic, life-threatening <em>d</em>isease characterize<em>d</em> by uncontrolle<em>d</em> complement activation, systemic thrombotic microangiopathy (TMA), an<em>d</em> vital organ <em>d</em>amage. We evaluate<em>d</em> the effect of terminal complement blocka<em>d</em>e with the anti-C5 monoclonal antibo<em>d</em>y eculizumab on biomarkers of cellular processes involve<em>d</em> in TMA in patients with aHUS longitu<em>d</em>inally, <em>d</em>uring up to 1 year of treatment, compare<em>d</em> with in healthy volunteers. Biomarker levels were elevate<em>d</em> at baseline in most patients, regar<em>d</em>less of mutational status, plasma exchange/infusion use, platelet count, or lactate <em>d</em>ehy<em>d</em>rogenase or haptoglobin levels. Eculizumab re<em>d</em>uce<em>d</em> terminal complement activation (C5a an<em>d</em> sC5b-9) an<em>d</em> renal injury markers (clusterin, cystatin-C, β<em>2</em>-microglobulin, an<em>d</em> liver fatty aci<em>d</em> bin<em>d</em>ing protein-1) to healthy volunteer levels an<em>d</em> re<em>d</em>uce<em>d</em> inflammation (soluble tumor necrosis factor receptor-1), coagulation (prothrombin fragment F1+<em>2</em> an<em>d</em> <em>d</em>-<em>dimer</em>), an<em>d</em> en<em>d</em>othelial <em>d</em>amage (thrombomo<em>d</em>ulin) markers to near-normal levels. Alternative pathway activation (Ba) an<em>d</em> en<em>d</em>othelial activation markers (soluble vascular cell a<em>d</em>hesion molecule-1) <em>d</em>ecrease<em>d</em> but remaine<em>d</em> elevate<em>d</em>, reflecting ongoing complement activation in aHUS <em>d</em>espite complete terminal complement blocka<em>d</em>e. These results highlight links between terminal complement activation an<em>d</em> inflammation, en<em>d</em>othelial <em>d</em>amage, thrombosis, an<em>d</em> renal injury an<em>d</em> un<em>d</em>erscore ongoing risk for systemic TMA an<em>d</em> progression to organ <em>d</em>amage. Further research regar<em>d</em>ing un<em>d</em>erlying complement <em>d</em>ysregulation is warrante<em>d</em>. This trial was registere<em>d</em> at www.clinicaltrials.gov as #NCT01194973.