Background-Risk stratification in troponin (cTn)-negative acute coronary syndrome (ACS) remains a clinical challenge.We investigated the predictive value of circulating pregnancy-associated plasma protein A (PAPP-A), a novel marker of atherosclerotic plaque activity, in these patients. Methods and Results-Two hundred consecutive hospitalized ACS patients were included, of whom 136 (69 men and 67 women; meanϮSD age, 66Ϯ16 years) remained cTnI-negative for up to 24 hours. PAPP-A was measured at admission, 6 to 12 hours, and 24 hours. During 6-month follow-up, 26 (19.1%) of the cTnI-negative patients reached a primary end point (cardiovascular death, myocardial infarction, or revascularization). At a cutoff level of 2.9 mIU/L, elevated PAPP-A was an independent predictor of adverse outcome (adjusted risk ratio [RR], 4.6; 95% confidence interval, 1.8 to 11.8; Pϭ0.002). Another independent predictor was admission CRP Ͼ2.0 mg/L (RR, 2.6; Pϭ0.03). Conclusions-Measurement of plasma PAPP-A, a zinc-binding matrix metalloproteinase, is a strong independent predictor of ischemic cardiac events and need of revascularization in patients who present with suspected myocardial infarction but remain troponin negative. (Circulation. 2003;108:1924-1926.)

Aspergillus flavus is one of the most important opportunistic pathogens of crops and animals. The carcinogenic mycotoxin, aflatoxins produced by this pathogen cause a health problem to human and animals. Since cyclic AMP signaling controls a range of physiological processes, like fungal development and infection when responding to extracellular stimuli in fungal pathogens, in this study, we investigated the function of adenylate cyclase, a core component of cAMP signaling, in aflatoxins biosynthesis and virulence on plant seeds in A. flavus. A gene replacement strategy was used to generate the deletion mutant of acyA that encodes the adenylate cyclase. Severe defects in fungal growth, sporulation and sclerotia formation were observed in the acyA deletion mutant. The defect in radical growth could be partially rescued by exogenous cAMP analog. The acyA mutant was also significantly reduced in aflatoxins production and virulence. Similar to the former studies in other fungi, The acyA mutant showed enhancing tolerance to oxidative stress, but more sensitive to heat stress. Overall, the pleiotropic defects of the acyA deletion mutant indicates that the cAMP-PKA pathway is involved in fungal development, aflatoxins biosynthesis and plant seed invasion in A. flavus.

In Aspergillus nidulans, the nitrogen metabolite repression (NMR) regulator NmrA plays a major role in regulating the activity of the GATA transcription factor AreA during nitrogen metabolism. However, the function of nmrA in A. flavus has not been previously studied. Here, we report the identification and functional analysis of nmrA in A. flavus. Our work showed that the amino acid sequences of NmrA are highly conserved among Aspergillus species and that A. flavus NmrA protein contains a canonical Rossmann fold motif. Deletion of nmrA slowed the growth of A. flavus but significantly increased conidiation and sclerotia production. Moreover, seed infection experiments indicated that nmrA is required for the invasive virulence of A. flavus. In addition, the ΔnmrA mutant showed increased sensitivity to rapamycin and methyl methanesulfonate, suggesting that nmrA could be responsive to target of rapamycin signaling and DNA damage. Furthermore, quantitative real-time reverse transcription polymerase chain reaction analysis suggested that nmrA might interact with other nitrogen regulatory and catabolic genes. Our study provides a better understanding of NMR and the nitrogen metabolism network in fungi.

Cyclic AMP signaling controls a range of physiological processes in response to extracellular stimuli in organisms. Among the signaling cascades, cAMP, as a second messenger, is orchestrated by adenylate cyclase (biosynthesis) and cAMP phosphodiesterases (PDEs) (hydrolysis). In this study, we investigated the function of the high-affinity (PdeH) and low-affinity (PdeL) cAMP phosphodiesterase from the carcinogenic aflatoxin producing fungus Aspergillus flavus, and found that instead of PdeL, inactivation of PdeH exhibited a reduction in conidiation and sclerotia formation. However, the ΔpdeL/ΔpdeH mutant exhibited an enhanced phenotype defects, a similar phenotype defects to wild-type strain treated with exogenous cAMP. The activation of PKA activity was inhibited in the ΔpdeH or ΔpdeL/ΔpdeH mutant, both of whom exhibited increasing AF production. Further analysis by qRT-PCR revealed that pdeH had a high transcriptional level compared to pdeL in wild-type strain, and affected pdeL transcription. Green fluorescent protein tagging at the C-terminus of PDEs showed that PdeH-GFP is broadly compartmentalized in the cytosol, while PdeL-GFP localized mainly to the nucleus. Overall, our results indicated that PdeH plays a major role, but has overlapping function with PdeL, in vegetative growth, development and AF biosynthesis in A. flavus.

PAPP-A is an additional marker for ACS, but does not seem to be a useful early marker for acute myocardial infarction (AMI). The possible clinical utility of PAPP-A calls for extensive studies of chest pain patients using serial sampling combined with short- and long-term outcome studies.

Background:In the blood of pregnant women, pregnancy-associated plasma protein A (PAPP-A) is present as a covalent complex with the proform of eosinophil major basic protein (proMBP). Recently, increased serum concentrations of PAPP-A have been found in acute coronary syndromes (ACS). The aim of this study was to investigate whether the circulating PAPP-A in ACS is the same as that in pregnancy. Methods: We developed two time-resolved immunofluorometric assays based on a relative epitope map constructed by the use of 17 monoclonal antibodies. One assay, which measured total PAPP-A, used two PAPP-A subunit-specific antibodies. The other assay, which measured PAPP-A/proMBP complex, used one proMBP subunit-specific antibody and one PAPP-A subunitspecific antibody. Serum samples from four patients with myocardial infarction (MI), three pregnant women in their first trimester, and one in her third trimester were fractionated by gel filtration on a Superose TM 6 precision column. The two assays were used to analyze fractions obtained by gel filtration as well as serum samples serially collected from four other MI patients. Results: Pregnancy-related PAPP-A was eluted as a single peak with a molecular mass of ϳ700 kDa, whereas ACS-related PAPP-A was also eluted as a single peak but with a molecular mass of ϳ530 kDa. Pregnancy-related PAPP-A was detected equally by the

Background:We recently reported that the pregnancyassociated plasma protein A (PAPP-A) form specifically related to acute coronary syndromes (ACS) is not complexed with the proform of eosinophil major basic protein (proMBP). The aim of this study was to develop rapid point-of-care immunoassays for the measurement of the noncomplexed PAPP-A. Methods: We developed immunofluorometric noncompetitive dry-reagent assays for total PAPP-A with 2 PAPP-A subunit-specific monoclonal antibodies and for PAPP-A/proMBP complex with 1 PAPP-A subunit-specific antibody and 1 proMBP subunit-specific antibody. The concentration of noncomplexed PAPP-A was determined as the difference of the results obtained with the 2 assays. Results: The assays were linear from 0.5 to 300 mIU/L. The analytical detection limit and functional detection limit (CV <20%) were 0.18 mIU/L and 0.27 mIU/L for total PAPP-A assay and 0.23 mIU/L and 0.70 mIU/L for PAPP-A/proMBP assay, respectively. The total assay imprecisions were <10%, and recoveries were 88%-107% for both assays. The mean difference (95% limits of agreement) between the new total PAPP-A assay and a previously reported total PAPP-A assay was ؊3.2% (؊45.7% to 39.3%; n ‫؍‬ 546; P ‫؍‬ 0.0019). In serum samples from 159 non-ACS individuals, median concentrations

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