Insulin degrading enzyme is expressed in the human cerebrovascular endothelium and in cultured human cerebrovascular endothelial cells.

Insulin degrading enzyme (IDE) is found in the cytosol, peroxisomes and plasma membrane of many cells. Although it preferentially cleaves insulin it can also cleave many other small proteins with diverse sequences including the monomeric form of the amyloid beta peptide (A beta). In the brain, IDE has been reported to be expressed predominantly in neurons. In this study, IDE expression was detected in cultured human cerebrovascular endothelial cells. Using laser capture microdissection followed by PCR analysis, it was found that IDE mRNA is expressed in human brain blood vessels. Using immunofluorescence and multiphoton microscopy IDE was localized to the endothelium of the cerebrovascular blood vessels in human.

Inhibition of glycerol metabolism in hepatocytes isolated from endotoxic rats.

Sepsis or endotoxaemia inhibits gluconeogenesis from various substrates, the main effect being related to a change in the phosphoenolpyruvate carboxykinase transcription rate. In addition, sepsis has been reported to affect the oxidative phosphorylation pathway. We have studied glycerol metabolism in hepatocytes isolated from rats fasted and injected 16 h previously with lipopolysaccharide from Escherichia coli. Endotoxin inhibited glycerol metabolism and led to a very large accumulation of glycerol 3-phosphate; the cytosolic reducing state was increased. Furthermore glycerol kinase activity was increased by 33% (P<<0.01). The respiratory rate of intact cells was significantly decreased by sepsis, with glycerol or octanoate as exogenous substrates, whereas oxidative phosphorylation (ATP-to-O ratio or respirations in state 4, state 3 and the oligomycin-insensitive state as well as the uncoupled state) was unchanged in permeabilized hepatocytes. Hence the effect on energy metabolism seems to be present only in intact hepatocytes. An additional important feature was the observation of a significant increase in cellular volume in cells from endotoxic animals, which might account for the alterations induced by sepsis.

Biological studies of analogues of glucose iodinated in positions 1, 2, or 3.

Analogues of glucose labeled with 123 iodine in positions 1, 2 or 3 have been synthesized. The aim of this study was to examine their biological behavior in four experimental models in order to assess whether they could be used to evaluate the uptake of glucose with single photon emission computed tomography (SPECT). The results obtained have shown that none of these molecules enters the cell using the glucose transporter. Therefore, they cannot be used as tracers of glucose uptake.

Experimental models, protocols, and reference values for evaluation of iodinated analogues of glucose.

For an iodinated analogue of glucose to be useful for evaluating glucose uptake using single-photon emission computed tomography (SPECT), it must enter the cell via the same transporter as glucose and accumulate within the cell without being degraded. The biological behavior of the iodinated tracer must therefore be similar to that of 2-deoxy-D(-)[1-14C]-glucose (2-DG). In the present study, four experimental models (biodistribution in mouse, isolated rat heart, human erythrocytes in suspension and cultured neonatal rat cardiomyocytes) have been chosen and protocols have been set up which allow for the examination of small quantities of iodinated analogues of glucose. The uptakes of 2-DG and of L(-)[1-14C]-glucose have been measured in these models to establish reference values which will be compared with uptake values for iodinated analogues of glucose.

Comparative effects of halothane associated with verapamil and ischaemia on myocardial metabolism in isolated perfused rat hearts.

The association of verapamil with halothane causes ischaemic-like myocardial dysfunction. Using an isolated rat heart model perfused with a radiolabelled fatty acid (123I-labelled iodohexadecenoic acid) as a sensitive marker of ischaemia this study investigated whether or not this dysfunction is of ischaemic origin. Hearts were perfused with a control solution or with solutions containing either 1% of halothane or 150 ng ml-1 of verapamil or the association of 0.75% halothane + 120 ng ml-1 verapamil. The ischaemic group was perfused at a reduced perfusion rate (-50%). Intracellular fate of IHA was assessed, and its esterification ratio computed. Ischaemia and the drugs induced a similar depression of haemodynamics. The esterification ratio in the ischaemic group was significantly higher (0.723 +/- 0.04) than in controls (0.0526 +/- 0.03) and than in the treated groups: halothane (0.533 +/- 0.06), verapamil (0.411 +/- 0.027) or the association halothane+verapamil (0.408 +/- 0.05), suggesting a non-ischaemic origin for the dysfunction caused by halothane-verapamil.