Although alpha-cyano-4-hydroxycinnamic acid functions as an excellent matrix for the analysis of most peptides using matrix-assisted laser desorption/ionization time-of-flight (MALDI TOF) mass spectrometry, the ionization of phosphorylated peptides is usually suppressed by nonphosphorylated peptides. As an alternative matrix, 2',4',6'-trihydroxyacetophenone (THAP) with diammonium citrate was found to overcome this problem for the MALDI TOF mass spectrometric analysis of proteolytic digests of phosphorylated proteins. Specifically, the abundances of phosphorylated peptides in tryptic digests of bovine beta-casein and protein kinase C (PKC)-treated mouse cardiac troponin I were enhanced more than 10-fold using THAP during positive ion MALDI TOF mass spectrometry. The protonated molecules of phosphorylated peptides were sufficiently abundant that postsource decay TOF mass spectrometry was used to confirm the number of phosphate groups in each peptide. Finally, tryptic digestion followed by analysis using MALDI TOF mass spectrometry with THAP as the matrix facilitated the identification of a unique phosphorylation site in PKC-treated troponin I.

3D printing improves the understanding of surgical liver anatomy for surgical residents. The improved comprehension of liver anatomy may facilitate laparoscopy or open liver resection.

Supplemental Digital Content is available in the text

Supplemental Digital Content is available in the text.

MicroRNAs (miRNAs) have been found to act as key regulators in the pathogenesis of myocardial ischemic-reperfusion (I/R) injury. In this study, we explore the role and mechanism of microRNA-202-3p (miR-202-3p) in regulating cardiomyocyte apoptosis, in respective of the TGF-β1/Smads signaling pathway by targeting the transient receptor potential cation channel, subfamily M, member 6 (TRPM6). The targeting relationship between miR-202-3p and TRPM6 was verified by a dual-luciferase reporter gene assay. Sprague-Dawley rat models of myocardial I/R injury were initially established and treated with different mimics, inhibitors and siRNAs to test the effects of miR-202-3p and TRPM6 on myocardial I/R injury. The levels of inflammatory factors; IL-1β, IL-6, TNF-α as well as the degree of myocardial fibrosis and cardiomyocyte apoptosis were determined in rats transfected with different plasmids. TRPM6 was found to be the target of miR-202-3p. Upregulated miR-202-3p or knockdown of TRPM-6 alleviated oxidative stress and inflammatory response, reduced ventricular mass, altered cardiac hemodynamics, suppressed myocardial infarction, attenuated cell apoptosis, and inhibited myocardial fibrosis. MiR-202-3p overexpression activates the TGF-β1/Smads signaling pathway by negatively regulating TRPM6 expression. Taken together, these findings suggest that miR-202-3p offers protection against ventricular remodeling after myocardial I/R injury via activation of the TGF-β1/Smads signaling pathway. ARTICLE HISTORY

Redox and/or electrophilic metabolites formed during estrogen metabolism may play a role in estrogen carcinogenesis. 4-Hydroxyequilenin (4-OHEN) is the major phase I catechol metabolite of the equine estrogens equilenin and equilin, which are components of the most widely prescribed estrogen replacement formulation, Premarin. Previously, we have found that 4-OHEN rapidly autoxidized to an o-quinone in vitro and caused toxic effects such as the inactivation of human detoxification enzymes. 4-OHEN has also been shown to be a substrate for catechol-O-methyltransferase (COMT) in human breast cancer cells. In the present study, we demonstrated that 4-OHEN was not only a substrate of recombinant human soluble COMT in vitro with a K(m) of 2.4 microM and k(cat) of 6.0 min(-)(1) but it also inhibited its own methylation by COMT at higher concentrations in the presence of the reducing agent dithiothreitol. In addition, 4-OHEN was found to be an irreversible inhibitor of COMT-catalyzed methylation of the endogenous catechol estrogen 4-hydroxyestradiol with a K(i) of 26.0 microM and a k(2) of 1.62 x 10(-)(2) s(-)(1). 4-OHEN in vitro not only caused the formation of intermolecular disulfide bonds as demonstrated by gel electrophoresis, but electrospray ionization mass spectrometry and matrix-assisted laser desorption ionization time-of-flight mass spectrometry also showed that 4-OHEN alkylated multiple residues of COMT. Peptide mapping experiments further indicated that Cys33 in recombinant human soluble COMT was the residue most likely modified by 4-OHEN in vitro. These data suggest that inhibition of COMT methylation by 4-OHEN might reduce endogenous catechol estrogen clearance in vivo and further enhance toxicity.

Objective: The aim of this meta-analysis was to investigate retinal microvascular features in patients with Alzheimer's disease (AD) using optical coherence tomography angiography (OCTA).Methods: PubMed, Cochrane Library, Embase, and Web of Science databases were systematically searched for published articles comparing retinal microvascular characteristics in subjects with AD and controls. The mean difference (MD) with a 95% confidence interval (CI) was used to assess continuous variables. Review Manager Version (RevMan) 5.30, was employed to analyze the data.Results: Nine studies were included in the meta-analysis. The analysis revealed that the macular whole enface superficial and deep vessel density (VD) values measured by OCTA were significantly lower in patients with AD than in controls (MD = −1.10, P < 0.0001; MD = −1.61, P = 0.0001, respectively). The value measured by OCTA for parafoveal superficial VD in patients with AD was also remarkably lower than that in the control group (MD = −1.42, P = 0.001), whereas there was no significant difference in the value for parafoveal deep VD (MD = −3.67, P = 0.19), compared to the controls. In addition, the foveal avascular zone (FAZ) was larger in patients with AD than in the control group (MD = 0.08, P = 0.07), although it did not reach statistical significance.Conclusions: The present meta-analysis indicated that the macular whole enface and parafoveal vessel densities were reduced in patients with AD. Moreover, our pooled data revealed that FAZ is larger in patients with AD. Consequently, OCTA may be utilized as a diagnostic tool to identify and monitor patients with AD.

A rapid method to quantify levels of the β-thioglycoside N-hydroxyl sulfate, glucoraphanin, in dog and rat plasma to support pre-clinical toxicological and pharmacological studies has been developed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Glucoraphanin was extracted from plasma by protein precipitation with acetonitrile and separated via hydrophilic interaction liquid chromatography (HILIC) using a Luna 5 µm Silica (2) 100 Å column (50 × 2.0 mm) at a flow rate of 0.3 mL/min. Solvent A consisted of 200 mM ammonium acetate and formic acid (99:1, v/v) and Solvent B was acetonitrile. Initial conditions (90% Solvent B) were held for 0.01 min after injection, decreased to 40% in 0.5 min and held constant for 2.5 min, returning to initial conditions for 3 min (reequilibration time). Glucoraphanin was detected by MS/MS using a turbo ion spray interface as the ion source operating in negative ion mode. Acquisition was performed in multiple reaction monitoring mode at m/z 435.8 → 96.7. The method was validated for the calibration range 10 to 2000 ng/mL. Within-and between-run precision for the low, mid and high QC levels was 8 % R.S.D or less and accuracy ranged from 100 to 113%. The lower limit of quantification was 10 ng/mL; calibration curves encompassed the range of plasma concentrations expected to be found in bioavailability and pharmacokinetics studies with glucoraphanin. The method has successfully been applied to the determination of glucoraphanin in dog and rat plasma and should be extendable to other species as well.