Quenched BODIPY dye-labeled casein substrates for the assay of protease activity by direct fluorescence measurement

We have prepared casein conjugates of two BODIPY dyes for use as fluorogenic protease substrates in homogeneous assays. Both conjugates are labeled to such an extent that the dyes are efficiently quenched in the protein, yielding virtually nonfluorescent substrate molecules. These fluorogenic substrates release highly fluorescent BODIPY dye-labeled peptides upon protease digestion, with fluorescence increases proportional to enzyme activity. These quenched substrates are suitable for the continuous assay of enzymatic activity using standard fluorometers, filter fluorometers, or fluorescence microplate readers using either fluorescein excitation and emission wavelengths to measure BODIPY FL casein hydrolysis or Texas Red wavelengths to detect proteolysis of BODIPY TR-X casein. Most current techniques for detecting protease activity, such as the fluorescein thiocarbamoyl casein (FTC-casein) protease assay, require extensive manipulation, including separation steps, and are therefore labor intensive and error-prone. In comparison, we found the BODIPY dye-labeled casein protease assays to be simple and precise and to have greater sensitivity and a broader dynamic range of detection than the FTC-casein assay. We were able to sensitively detect the activities of a wide variety of enzymes with these new substrates, including serine, acid, sulfhydryl, and metalloproteases. We also found the assay suitable for quantitating protease inhibitor concentrations and for real-time analysis of proteolysis.     

The visual simulation of amorphous phenomena

Numerical simulations of natural phenomena generate data of two basic types. One class can be characterized either as having a well-defined surface which can be approximated by planar or parametric patches or as having volume described by boolean operations on patonic solids. Other calculations yield data that is more amorphous in nature; its form changes markedly with time. The visualization of these results is less obvious, and, in fact, no single technique is appropriate. In this paper we will describe several techniques that we have found useful in representing fluid shapes.     

Synergism between malathion and low frequency sound to increase the mortality of adult Tribolium confusum

When adults of Tribolium confusum Jacquelin duVal were exposed to low frequency sound during and after exposure to residues of malathion, mortality increased. Eighteen Hz was the most effective sound treatment. However, no significant differences in mortality were found between insects that were or were not exposed to sound when the malathion was applied directly to the insect.     

The relationship between maternal and neonatal umbilical cord plasma homocyst(e)ine suggests a potential role for maternal homocyst(e)ine in fetal metabolism

OBJECTIVE: To determine whether increased total daily energy expenditure (TDEE) associated with repetitive, involuntary movements contributes to growth failure in girls with Rett syndrome (RS). STUDY DESIGN: Fourteen girls with RS and 11 healthy girls were studied for 10 days to obtain measurements of height, weight, body circumference, and skin-fold thickness with stadiometric and anthropometric methods; whole-body potassium by potassium 40 counting; 72-hour dietary energy intakes by test weighing; 24-hour activity patterns using observational methods; and TDEE using the doubly-labeled water technique. RESULTS: TDEE, when adjusted for differences in lean body mass, did not differ significantly between girls with RS and healthy girls. Although girls with RS spent more waking hours in physical activity than their healthy counterparts (85%+/-10% vs. 73%+/-11% awake time per day, p < 0.05), their repetitive movements were not sufficiently intense to increase TDEE. However, girls with RS had significantly less lean body mass, but not body fat, which contributed to their lower absolute TDEE in comparison with that of healthy girls (845+/-251 vs. 1453+/-534 kcal/day, p < 0.01). Dietary energy intake, when adjusted for differences in body weight, was not significantly different in girls with RS compared with healthy girls. CONCLUSIONS: Increased TDEE as a result of repetitive, involuntary movements does not explain the alterations in growth and body composition of girls with RS.     

Premature peripheral vascular disease: clinical profile and abnormal lipid peroxidation

Type 2 diabetes is extremely common and increasing in the United States. The pathophysiology of type 2 diabetes is a combination of increased insulin resistance and inadequate secretion. The main risk factors for diabetes are family history, obesity, sedentary lifestyle, ethnic background, age, and a history of gestational diabetes. Diet and exercise, the cornerstones of diabetes management, will improve insulin sensitivity and indirectly augment insulin secretion. Until recently, the only pharmacological approaches to diabetes were sulfonylureas and insulin, which either augment insulin secretion or replace insulin, thus acting only on the insulin side of the equation. Recently, a series of new drugs have become available that are capable of decreasing hepatic glucose output (metformin), slowing postprandial glucose absorption (acarbose), and improving peripheral insulin sensitivity (troglitazone). With these drugs, either alone or in combination, and behavioral therapies, it is now feasible to achieve good to outstanding glycemic control in most individuals with type 2 diabetes.     

Short-term folic acid supplementation induces variable and paradoxical changes in plasma homocyst(e)ine concentrations

Folic acid is presently the mainstay of treatment for most subjects with elevated plasma homocyst(e)ine concentrations [Plasma or serum homocyst(e)ine, or total homocysteine, refers to the sum of the sulfhydryl amino acid homocysteine and the homocysteinyl moieties of the disulfides homocystine and homocystein-cysteine, whether free or bound to plasma proteins.] Changes in homocyst(e)ine in response to folic acid supplementation are characterized by considerable interindividual variation. The purpose of this study was to identify factors that contribute to heterogeneity in short-term responses to folic acid supplementation. The effects of folic acid supplementation (1 or 2 mg per day) for 3 wk on plasma homocyst(e)ine concentrations were assessed in 304 men and women. Overall, folic acid supplementation increased mean plasma folate 31.5±98.0 nmol/L and decreased mean plasma homocyst(e)ine concentrations 1.2±2.4 μmol/L. There was evidence of substantial interindividual variation in the homocyst(e)ine response from ?18.5 to +7.1 μmol/L, including an increase in homocyst(e)ine in 20% of subjects (mean increase 1.5±1.4 μmol/L). Basal homocyst(e)ine, age, male gender, cigarette smoking, use of multivitamins, methylene tetrahydrofolate reductase, and cystathionine β-synthase polymorphisms accounted for 47.6% of the interindividual variability in the change in homocyst(e)ine after folic acid supplementation, but about 50% of variability in response to folic acid was not explained by the variables we studied.     

The application visualization system: a computational environmentfor scientific visualization

A software system for developing interactive scientific visualization applications quickly, with a minimum of programming effort, is described. This application visualization system (AVS) is an application framework targeted at scientists and engineers. The goal of the system is to make applications that combine interactive graphics and high computational requirements easier to develop for both programmers and nonprogrammers. AVS is designed around the concept of software building blocks, or modules, which can be interconnected to form visualization applications. AVS allows flow networks of existing modules to be constructed using a direct-manipulation user interface, and it automatically generates a simple user interface to each module     

Expansion by the introduction of new technology in a market characterized by overcapacity-the success of Urenco centrifuge technology

Urenco has successfully developed and deployed centrifuge technology for the enrichment of uranium in a period during which the market changed from a situation of ‘threatened undersupply’ in the 1970s to one of significant overcapacity. Five generations of centrifuges have been developed, introducing new materials as they became available. The sixth generation of centrifuge currently under development for deployment at the end of the century will be more than twice as fast and an order of magnitude longer than respectively the slowest and shortest pilot plant centrifuges, with an output approaching 50 times higher than that in the pilot plant. Plant operation has exceeded both design lifetimes and failure rates. Urenco has concluded that the economics of this sixth-generation centrifuge are better than any forseeable first generation AVLIS technology and thus, in Urenco's view the centrifuge represents the low risk, proven economic choice for the replacement of GDPs when this becomes necessary.