The ambivalent effect of Fe3O4 nanoparticles on the urea-induced unfolding and dilution-based refolding of lysozyme.

Due to the numerous biological applications of magnetite (Fe3O4) nanoparticles (MNPs), it is essential to identify the influence of these nanoparticles on basic biological processes. Therefore, in this research, the effect of MNPs on the structure and activity of hen egg white lysozyme (HEWL) (EC 3.2.1.1) as a model protein was examined using tryptophan intrinsic fluorescence, UV/Vis, and circular dichroism spectroscopy. Moreover, enzyme activities were analyzed by a turbidometric approach in the presence of MNPs at concentrations providing MNPs/HEWL ratios in the range of 0.04-1.25. As-synthesized MNPS were characterized by Fourier transform infrared spectroscopy, x-ray diffraction, scanning electron microscopy, transmission electron microscopy, vibrating sample magnetometry and the zeta potential of MNPs was measured to be -29 mV. The goal of this work was investigating the ordering or disordering effect of MNPs on protein structure at ratios lower or higher than 0.918 as concentration ratio of threshold (CRT), respectively, in order to answer the question: 'How can the denaturation and refolding of a model protein (HEWL) be affected by MNPs?' As has been reported recently, the protein folding, helicity, and half-life were improved at CRT to make the protein more disordered upon interaction with MNPs. The disordering effect of urea at >CRT and even at

Modeling, simulation, and employing dilution-dialysis microfluidic chip (DDMC) for heightening proteins refolding efficiency.

Miniaturized systems based on the principles of microfluidics are widely used in various fields, such as biochemical and biomedical applications. Systematic design processes are demanded the proper use of these microfluidic devices based on mathematical simulations. Aggregated proteins (e.g., inclusion bodies) in solution with chaotropic agents (such as urea) at high concentration in combination with reducing agents are denatured. Refolding methods to achieve the native proteins from inclusion bodies of recombinant protein relying on denaturant dilution or dialysis approaches for suppressing protein aggregation is very important in the industrial field. In this paper, a modeling approach is introduced and employed that enables a compact and cost-effective method for on-chip refolding process. The innovative aspect of the presented refolding method is incorporation dialysis and dilution. Dilution-dialysis microfluidic chip (DDMC) increases productivity folding of proteins with the gradual reduction of the amount of urea. It has shown the potential of DDMC for performing refolding of protein trials. The principles of the microfluidic device detailed in this paper are to produce protein on the dilution with slow mixing through diffusion of a denatured protein solution and stepwise dialysis of a refolding buffer flowing together and the flow regime is creeping flow. The operation of DDMC was modeled in two dimensions. This system simulated by COMSOL Multiphysics Modeling Software. The simulation results for a microfluidic refolding chip showed that DDMC was deemed to be perfectly suitable for control decreasing urea in the fluid model. The DDMC was validated through an experimental study. According to the results, refolding efficiency of denaturant Hen egg white lysozyme (HEWL) (EC 3.2.1.17) used as a model protein was improved. Regard to the remaining activity test, it was increased from 42.6 in simple dilution to 93.7 using DDMC.

L-arginine modified magnetic nanoparticles: green synthesis and characterization.

In recent years, there has been considerable interest in Arg which is a unique, nontoxic, and biocompatible biomolecule since it can be utilized as an agent for the functionalization and subsequent stabilization of MNPs against oxidation and aggregation, during or after a synthesis procedure. Our studies demonstrate that Arg has great impacts on MNPs with the decreasing size of the particle. Also, saturation magnetization and electrostatic interactions of RMNPs have a direct impact on biological molecules such as proteins and nucleic acids. By controlling the concentration of Arg, it is possible to accurately control the above-mentioned characteristics, which are useful tools for applications such as connecting to antibodies, catalysis, drug loading, and modification of MNP stability. In the current study, three RMNPs with different Arg densities, i.e. 0.42, 1.62, and 2.29 µg per mg were successfully synthesized through a simple co-precipitation method (named p 0.5, p 1, and p 1.5, respectively) and verified by colorimetric determination. Also, the as-synthesized RMNP powders were characterized by XRD, SEM/EDAX, FTIR, VSM, and zeta potential analysis. The presence of a magnetic core was proved by XRD, FTIR, and EDAX. Colorimetric analysis showed the existence of Arg in the synthesized samples. According to the zeta potential and VSM results, increasing the cap of Arg on the MNP surface leads to an increase in the surface charge and decrease in the magnetization of the RMNPs, respectively.

Myocardial first-pass perfusion magnetic resonance imaging: a multicenter dose-ranging study.

BACKGROUND: MRI can identify patients with obstructive coronary artery disease by imaging the left ventricular myocardium during a first-pass contrast bolus in the presence and absence of pharmacologically induced myocardial hyperemia. The purpose of this multicenter dose-ranging study was to determine the minimally efficacious dose of gadopentetate dimeglumine injection (Magnevist Injection; Berlex Laboratories) for detecting obstructive coronary artery disease. METHOD AND RESULTS: A total of 99 patients scheduled for coronary artery catheterization as part of their clinical evaluation were enrolled in this study. Patients were randomized to 1 of 3 doses of gadopentate dimeglumine: 0.05, 0.10, or 0.15 mmol/kg. First-pass perfusion imaging was performed during hyperemia (induced by a 4-minute infusion of adenosine at a rate of 140 microg x kg(-1) x min(-1)) and then again in the absence of adenosine with otherwise identical imaging parameters and the same contrast dose. Perfusion defects were evaluated subjectively by 4 blinded reviewers. Receiver-operating curve analysis showed that the areas under the receiver-operating curve were 0.90, 0.72, and 0.83 for the low-, medium-, and high-contrast doses, respectively, compared with quantitative coronary angiography (diameter stenosis > or =70%). For the low-dose group, mean sensitivity was 93+/-0%, mean specificity was 75+/-7%, and mean accuracy was 85+/-3%. CONCLUSIONS: First-pass perfusion MRI is a safe and accurate test for identifying patients with obstructive coronary artery disease. A low dose of 0.05 mmol/kg gadopentetate dimeglumine is at least as efficacious as higher doses.

Lumbosacral spine: early and delayed MR imaging after administration of an expanded dose of gadopentetate dimeglumine in healthy, asymptomatic subjects.

PURPOSE: To determine normal magnetic resonance (MR) imaging enhancement characteristics of the lumbosacral spine after intravenous administration of an expanded dose of gadopentetate dimeglumine. MATERIALS AND METHODS: T1-weighted MR images of the lumbosacral spine were acquired before and after injection of 0.3 mmol/kg gadopentetate dimeglumine in 12 healthy subjects (eight men, four women; age range, 22-57 years). RESULTS: In 10 (91%) of the 11 subjects who completed the investigation, multifocal linear enhancement within the thecal sac that generally extended from the conus to the nerve-root sheaths was demonstrated. In all cases, enhancement was seen in the facet joints and the intervertebral disks parallel to the vertebral end-plates. CONCLUSION: After an expanded dose of intravenous gadopentetate dimeglumine, multiple intrathecal and extrathecal structures were enhanced. Recognition of normal enhancement patterns after an expanded gadopentetate dimeglumine dose is important because such enhancement after routine injection of 0.1 mmol/kg gadopentetate dimeglumine is often thought to indicate disease.

Serum analyte pattern characteristic of fulminant hepatic failure.

Fulminant hepatic failure (FHF) is a poorly understood condition in which total liver failure occurs and is thought to be caused by a variety of conditions including Reye's syndrome, hepatitis, drug overdoses, and vascular insufficiency. While this condition is an uncommon one, it carries with it a high fatality rate and must therefore be diagnosed as rapidly as possible. Six patients have been observed over a two-year period with biopsy and/or autopsy-confirmed FHF: one with acute hepatitis B-delta; three with histories of alcoholism, two of them with cirrhosis; one with acute tylenol overdose; and one with hepatic vascular insufficiency. All of these patients, except one, exhibited a rapid, fatal downhill course after onset of symptoms. In all of these patients, a consistent elevation was observed in serum levels of aspartate aminotransferase (AST) or serum glutamate oxaloacetate transaminase (SGOT) and alanine aminotransferase (ALT) or serum glutamate pyruvate transaminase (SGPT) such that the ratio of AST to ALT was significantly greater than 1 and in serum levels of ammonia. Other liver function tests were found to be abnormal but not in so consistent a pattern, although total protein and albumin were found to be significantly decreased in all of these patients. The stereotypical elevation of the transaminases with high AST-to-ALT ratios and the rise in ammonia appear to characterize this life-threatening illness most reliably.

Efficacy and safety of gadopentetate dimeglumine in the evaluation of patients with a suspected tumor of the extracranial head and neck.

The clinical efficacy and safety of gadopentetate dimeglumine as a paramagnetic contrast agent for magnetic resonance (MR) imaging of the extracranial head and neck was evaluated in a multicenter trial involving 60 patients. Patients with signs and/or symptoms of a tumor in the nasopharynx, oropharynx, hypopharynx, larynx, or neck were studied. T1-weighted images were obtained before and after injection of gadopentetate dimeglumine, 0.1 mmol/kg, at a rate of 10 mL/min. No lesions were seen on the pre- or postinjection images of five of the 60 patients. Postinjection lesion enhancement was present in 53 of the remaining 55 (96%) patients. The absence of postinjection lesion enhancement in one of the two remaining patients was useful information. Postinjection impressions differed from preinjection diagnosis in 22 of 60 (37%) patients. Additional information was obtained from postinjection relative to preinjection images in 38 of 60 (63%) patients. Four adverse experiences were reported in three of 60 (5%) patients. Two mild (chest wall pain and headache) and one moderate (nausea) adverse experiences were considered by the authors to be unrelated to the studied drug. One severe adverse experience was reported. This patient had a seizure, considered by the investigator to be remotely related to the study drug and attributed to the abrupt withdrawal of anticonvulsant medications. The data indicate that gadopentetate dimeglumine is safe and efficacious in the evaluation of patients with extracranial head and neck lesions.

Rapid bolus injection of gadopentetate dimeglumine: absence of side effects in normal volunteers.

The safety and tolerance of gadopentetate dimeglumine 0.1 mmol/kg when injected at a bolus rate (10 ml/15 sec; four times the recommended rate) was studied in 12 normal men between the ages of 20 and 36 years. Each of the subjects received a single injection of either gadopentetate dimeglumine (0.2 ml/kg) or a placebo (normal saline, 0.2 ml/kg), followed by a 7-day washout and a single injection of the alternative treatment (six subjects per sequence). Measurement parameters included blood pressure, pulse rate, ECG, cardiographic rhythm strips, hematology and blood chemistry evaluations, physical examinations, and adverse drug experiences. Variations of greater than or equal to +/- 15 mm Hg in diastolic blood pressure and greater than or equal to +/- 15 beats per min in heart rate were observed in some subjects after injection of either gadopentetate dimeglumine or the placebo, but no clinically significant changes from baseline were observed for any parameter. Post-gadopentetate dimeglumine results were not significantly different statistically from post-placebo results. No adverse experiences were reported for any subject. It is concluded that gadopentetate dimeglumine is safe and well tolerated when administered to normal men at a rapid injection rate of 10 ml/15 sec.

Multicenter study of gadopentetate dimeglumine as an MR contrast agent: evaluation in patients with spinal tumors.

In an open-label, multicenter study, the efficacy and safety of gadopentetate dimeglumine (0.1 mmol/kg) administered IV as an MR imaging contrast agent were evaluated in 113 patients with symptoms of spinal tumors. The examinations were performed with a variety of imagers at different field strengths. Scans with short and long TRs were obtained in all patients before and after IV administration of the contrast medium. Contrast enhancement was seen in 77% of patients. No enhancement was seen in 23%, but this absence was useful diagnostic information in all cases. In 66% of the cases, additional information regarding location, size, configuration, and/or characterization of the lesion was obtained from postcontrast scans. The investigators made a change from referral diagnosis to postinjection diagnosis in 30% of the cases. Postinjection images provided additional information in 96% (43/45) of intradural extramedullary and intramedullary tumors; it also provided additional information in 20 (53%) of 38 cases of extradural tumor. Gadopentetate dimeglumine demonstrated a high level of safety and tolerance, as evidenced by the lack of clinically significant trends toward abnormal changes from baseline evaluations for physical and neurologic examinations, vital signs, and hematologic and blood chemistry parameters and by the low prevalence and mild nature of adverse reactions. Gadopentetate dimeglumine was found to be efficacious in the evaluation of suspected spinal tumors. High levels of safety and tolerance were demonstrated.

Variation in concentration of lipids, lipoprotein lipids, and apolipoproteins A-I and B in plasma from healthy women.

The components of total variation--biological, inter- and intra-individual, and analytical--of plasma lipids, lipoprotein lipids, and apolipoproteins A-I and B have been determined in a population of 44 healthy women, ages 18-35 years. Blood was sampled three separate times at three-month intervals, with no restrictions on diet or physical activity during these periods. The greatest inter-individual variances were observed for high-density lipoprotein (HDL)-cholesterol, HDL2-cholesterol, and total plasma cholesterol. Triglycerides had the highest intra-individual variance (CV 22%). The percentage of inter- and intra-individual variances of apolipoprotein A-I concentrations were more reflective of total HDL- and HDL2-cholesterol content than of HDL3-cholesterol. Concentrations of calculated low-density-lipoprotein cholesterol showed a greater inter-individual variation (18.6%) than did apolipoprotein B (10.7%). The laboratory CVs for these analytes were similar to other reported values. From these data, we computed the expected variation for subjects in our study for single and repeated measurements. Such considerations can influence decision-making in the clinical setting or in designing epidemiological studies.

Safety assessment of gadopentetate dimeglumine in U.S. clinical trials.

Gadopentetate dimeglumine is an intravenous contrast medium used in magnetic resonance imaging. To determine its safety, the authors summarized data concerning adverse reactions, laboratory parameters, and other assessments for 1,068 adult patients who received gadopentetate dimeglumine in United States clinical trials. For all studies, 213 of 1,068 patients (19.9%) who received gadopentetate dimeglumine experienced one or more clinical adverse reactions. Most of these reactions were minor and short-lived. Hematologic, blood chemistry, and urinary evaluations showed no apparent drug-related effects, with the exception of a transient, asymptomatic rise in serum iron and bilirubin levels in some patients. Other safety assessments--electrocardiography, electroencephalography, neurologic examinations, and vital signs--showed no clinically significant trends in change from baseline results. It was concluded that gadopentetate dimeglumine demonstrated a high degree of safety and tolerance.

Neutrophil functional and arachidonic acid metabolic correlates and clinical disease activity in pirazolac-treated rheumatoid arthritis patients.

Neutrophils from 20 medication-free rheumatoid arthritis patients were chemotactically hyporesponsive to 2 x 10(-8) M formyl-methionyl-leucyl-phenylalanine (f-MLP) as compared to 20 normal controls. They were relatively high producers of 5-hydroxyeicosatetraenoic acid (5-HETE) and low producers of leukotriene C4 (LTC4). Neutrophils from 10 patients treated with pirazolac (4-(4-chlorophenyl)-1-(4-fluorophenyl)-5-pyrazole acetic acid) showed increased chemotactic responsiveness concomitantly with normalization of 5-HETE and LTC4 production and a decrease in the clinical parameters of disease activity. Clinically attainable (30-60 micrograms/ml) in-vitro doses of pirazolac further depressed the chemotactic responsiveness of normal neutrophils treated with 10(-5)M f-MLP. Effects on endothelial gating of migrating neutrophil populations may explain this apparent contradiction between in-vivo and in-vitro actions of pirazolac on neutrophil chemotaxis.