Glyceraldehyde-3-phosphate dehydrogenase activity as an independent modifier of methylglyoxal levels in diabetes

Methylglyoxal (MG) may be an important cause of diabetic complications. Its primary source is dihydroxyacetone phosphate (DHAP) whose levels are partially controlled by glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Using a human red blood cell (RBC) culture, we examined the effect of modifying GAPDH activity on MG production. With the inhibitor koningic acid (KA), we showed a linear, concentration-dependent GAPDH inhibition, with 5 microM KA leading to a 79% reduction of GAPDH activity and a sixfold increase in MG. Changes in redox state produced by elevated pH also resulted in a 2.4-fold increase in MG production at pH 7.5 and a 13.4-fold increase at pH 7.8. We found substantial inter-individual variation in DHAP and MG levels and an inverse relationship between GAPDH activity and MG production (R=0.57, P=0.005) in type 2 diabetes. A similar relationship between GAPDH activity and MG was observed in vivo in type 1 diabetes (R=0.29, P=0.0018).Widely varying rates of progression of diabetic complications are seen among individuals. We postulate that modification of GAPDH by environmental factors or genetic dysregulation and the resultant differences in MG production could at least partially account for this observation.     

Synthesis and conformation of an analog of the helix‐loop‐helix domain of the Id1 protein containing the O‐acyl iso‐prolyl‐seryl switch motif

Synthetic peptides reproducing the helix‐loop‐helix (HLH) domains of the Id proteins fold into highly stable helix bundles upon self‐association. Recently, we have shown that the replacement of the dipeptide Val‐Ser at the loop–helix‐2 junction with the corresponding O‐acyl iso‐dipeptide leads to a completely unfolded state that only refolds after intramolecular O → N acyl migration. Herein, we report on an Id HLH analog based on the substitution of the Pro‐Ser motif at the helix‐1–loop junction with the corresponding O‐acyl iso‐dipeptide. This analog has been successfully synthesized by solid‐phase Fmoc chemistry upon suppression of DKP formation. No secondary structure could be detected for the O‐acyl iso‐peptide before its conversion into the native form by O → N acyl shift. These results show that the loop–helix junctions are determinant for the folded/unfolded state of the Id HLH domain. Further, despite the high risk of DKP formation, peptides containing O‐acyl iso‐Pro‐Ser/Thr units are synthetically accessible by Fmoc chemistry. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.     

Standardization of the immunocytochemical detection of neuroblastoma cells in bone marrow.

Standard cytomorphological examination of bone marrow (BM) aspirates does not appear to be sensitive enough to detect single neuroblastoma cells. The SIOPEN Neuroblastoma Bone Marrow Committee developed a sensitive and reproducible anti-GD2 immunocytochemical assay and introduced morphological and immunocytological criteria for the interpretation of results. Fixed cytospins were incubated with a commercially available anti-GD2 monoclonal antibody and an APAAP kit. Cells fulfilling all morphological and immunocytological criteria were called criteria-positive cells (CPCs). Not convincingly interpretable cells fulfilled some, but not all, criteria, and negative cells displayed only exclusion criteria. The genetic profile of doubtful cells was checked by fluorescence in situ hybridization. Ideally, 3 x 10(6) cells were analyzed to reach a 95% probability of detecting one tumor cell in 1 x 10(6) mononuclear cells. Four quality control rounds were organized to validate the method. A total of 111 quality control samples were analyzed. Two main improvements were achieved: in discordant cases, the range between the lowest and highest reported result was reduced by half, and discordant results were only found in samples with less than 10 CPCs per 1 x 10(6). This article describes the first internationally standardized protocol to detect and quantify rare neuroblastoma cells by immunocytochemistry. This method is an indispensable tool for multicenter studies evaluating the clinical significance of minimal residual disease in neuroblastoma.     

Oxygen consumption-based evaluation of fungal activity

A novel oxygen-based microplate assay for studying fungal activity is described. Fungal activity results in a change of oxygen concentration in CVC-96 plates^® and thus produces a signal that enables continuous monitoring of fungal activity. In this study the oxygen consumption was different for three tested fungi, Fusarium oxysporum f. sp. lycopersici, Verticillium dahliae and Trichoderma longibrachiatum. The assay described is a highly sensitive and reliable method for monitoring fungal activity. This assay does not interfere with fungal development and there is no need to kill the cells to take a measurement. This test would be useful for studying reactions that consume oxygen so possible applications of this test could be physiological studies, testing of fungicidal or fungistatic compounds, and studying enzyme reactions. The system provides a valuable insight into the kinetics of fungal activity and is suitable to test other systems.     

X-linked dominant inherited diseases with lethality in hemizygous males

Summary X-linked dominant inheritance with lethality in hemizygous males is a rare mode of inheritance. The three best-known disorders which seem to be inherited in this way, are incontinentia pigmenti (IP) Bloch-Sulzberger, oral-facial-digital I (OFD I) syndrome, and focal dermal hypoplasia (FDH syndrome, Goltz syndrome). It is the purpose of this article to give a review of the clinical and genetic aspects of the abovementioned diseases and to add those disorders in which this mode of inheritance is discussed. These disorders are: X-linked chondrodysplasia punctata (CP), cervico-oculo-acusticus syndrome (Wildervanck syndrome, COA), congenital cataract with microcornea or slight microphthalmia, muscular dystrophy-hemizygous lethal, partial lipodystrophy with lipatrophic diabetes and hyperlipidemia, Aicardi syndrome, coxo-auricular syndrome, and Johanson-Blizzard syndrome. OTC defiency is included in the study, although there is no lethality in utero, only in the neonatal period.A critical evaluation of the current literature is carried out.     

Nocturnal activity and orientation behavior during spring migration and early summer in the indigo bunting,Passerina cyanea

Summary Indigo Buntings (Passerina cyanea) that had been trapped in August were kept during the following months under photoperiodic conditions simulating the ones they would normally experience at that time of the year. From April 24 to June 16, their nocturnal activity and their directional tendencies were recorded. It was found that the activity, although it varied greatly from night to night, continued to be high until the termination of the registration period in mid June, whereas the orientation deteriorated at the end of May, i.e., after the end of the normal spring migration (Fig. 1, Table 1). Thus directed spring migratory restlessness and undirected summer restlessness seem to represent two separate phenomena. A possible adaptive significance of summer restlessness is discussed.This work was supported by National Science Foundation grant BMS-72-02198-AO2 to S.T. Emlen and by a Deutsche Forschungsgemeinschaft grant to W. Wiltschko. The Hochschulrechenzentrum of the Universität Frankfurt a.M. provided computer time. We thank S. Bergman for his valuable help in conducting the experiments and quantifying the footprint records of the funnel cages, and R.J. Young and the Department of Poultry Science, Cornell University, for the use of the brooder houses as experimental rooms.     

Volatile metabolites in sera of normal and diabetic patients

The profiles of volatile metabolites in serum samples from normal individuals and from individuals with diabetes mellitus with varying degrees of polyneuropathy have been studied. The transevaporator procedure was used to obtain sample extracts which were chromatographed on a highly efficient glass column coated with Silar 10C (106 m × 0.25 mm I.D.). Differences in profiles between normal subjects and diabetic subjects on no drug therapy were noticed. However, correlations between the severity of the neuropathy and the concentrations of certain ketones could not be established. Compounds present both in diabetic and normal sera have been identified by mass spectrometry.