Clinical impact and diagnostic accuracy of 2-[18F]-fluoro-2-deoxy-d-glucose positron-emission tomography/computed tomography (PET/CT) brain imaging in patients with cognitive impairment: a tertiary centre experience in the UK.

AIM: To evaluate the clinical impact of combined 2-[18F]-fluoro-2-deoxy-d-glucose (FDG) positron-emission tomography/computed tomography (PET/CT) brain imaging performed in selected patients with cognitive impairment at a tertiary referral centre in the UK, and to assess the accuracy of FDG PET/CT to correctly establish the diagnosis of Alzheimer's dementia (AD) in "real-world" clinical practice. METHODS AND MATERIALS: Using an institutional radiology database, 136 patients were identified for inclusion in the study. FDG PET/CT was performed using a standard technique and interpreted by dual-trained radiologists and nuclear medicine physicians. Standardised questionnaires were sent to the referring clinicians to establish the final clinical diagnosis and to obtain information about the clinical impact of FDG PET/CT. RESULTS: There was a 72% questionnaire return (98/136), with mean patient follow-up of 471 (standard deviation 205) days. FDG PET/CT had an impact on patient management in 81%, adding confidence to the pre-test diagnosis in 43%, changing the pre-test diagnosis in 35%, reducing the need for further investigations in 42%, and resulting in a change in therapy in 32%. There was substantial correlation between the PET/CT diagnosis and final clinical diagnosis with a correlation (k) coefficient of 0.78 (p<0.0001). The accuracy of FDG PET/CT in diagnosis of AD was 94% (95% confidence interval [CI]: 87-99), with a sensitivity of 87% (95% CI: 75-92) and a specificity of 97% (95% CI: 87-99). CONCLUSION: FDG PET/CT brain imaging has a significant clinical impact when performed selectively in patients with cognitive impairment and shows high accuracy in the diagnosis of AD in "real-world" clinical practice.

Vasodilator-stimulated phosphoprotein is involved in stress-fiber and membrane ruffle formation in endothelial cells.

Vasodilator-stimulated phosphoprotein (VASP) is highly expressed in vascular endothelial cells, where it has been implicated in cellular reorganization during angiogenesis, as well as in endothelial retraction and changes in vessel permeability. However, the cellular functions of VASP are not known. In this study, we have expressed wild-type and mutant forms of VASP in endothelial cells to determine in what aspects of cytoskeletal behavior this protein participates. Expression of wild-type VASP induces marked membrane ruffling and formation of prominent stress fibers in bovine aortic endothelial cells. Deletion of the proline-rich domain of VASP abolishes its ability to bind profilin but does not affect ruffling or stress fiber formation. Further deletions reveal a sequence within the carboxy-terminal domain that is responsible for in vivo bundle formation. Ruffling occurs only on the expression of forms of VASP that possess bundling activity and the capacity to bind zyxin/vinculin-derived peptide. The ability of distinct subdomains within VASP to bind adhesion proteins and induce F-actin bundling in vivo suggests that this protein could function in the aggregation and tethering of actin filaments during the formation of endothelial cell-substrate and cell-cell contacts. These data provide a mechanism whereby VASP can influence endothelial migration and organization during capillary formation and modulate vascular permeability via effects on endothelial cell contractility.

The focal adhesion phosphoprotein, VASP.

Vasodilator-stimulated phosphoprotein (VASP) is associated with focal adhesions and areas of dynamic membrane activity, where it is thought to have an important role in actin filament assembly and cell motility. VASP contains a central proline-rich sequence which recruits the G-actin binding protein profilin. Localization of VASP to the leading edge of a migrating cell can lead to local accumulation of profilin, which in turn can supply actin monomers to growing filament ends. VASP binds to the focal adhesion proteins vinculin and zyxin and this probably directs the phosphoprotein to focal adhesions and the leading edge of stimulated cells. VASP functions as a binding intermediate between profilin and focal adhesion proteins. Intracellular pathogens, including Listeria monocytogenes, have coat proteins which bind VASP. This is one way in which these pathogens use VASP, and other proteins from the host cell, to assemble the actin filaments they require to move around the cytoplasm of infected cells and enter neighbouring cells. Understanding the role of VASP and other proteins in cell and bacterial motility is likely to lead to development of new therapeutic strategies for diseases including atherosclerosis and tumour growth, and for limiting the spread of intracellular pathogens.

The endothelial receptor tyrosine kinase tie-1 is upregulated by hypoxia and vascular endothelial growth factor.

The receptor tyrosine kinase tie-1 is essential for angiogenesis where it appears to have a role in vessel maturation. Here we have examined the effects of hypoxia and vascular endothelial growth factor (VEGF) on the level of tie-1 protein expressed in bovine aortic endothelial cells. Both hypoxia (2% O2) and VEGF were found to increase tie-1 in a time-dependent manner. Hypoxic induction was direct and effects of hypoxia and VEGF were not additive. Experiments with actinomycin D indicate that these activators regulate tie-1 at the transcriptional level.

Inhibition of rabbit aortic smooth muscle cell proliferation by selective inhibitors of protein kinase C.

1. We studied the effect of two structurally-related, selective inhibitors of protein kinase C, Ro 31-8220 and Ro 31-7549, on the reinitiation of proliferation in quiescent first passage rabbit aortic smooth muscle cells in response to (a) the direct activator of protein kinase C, phorbol dibutyrate (PDBu), (b) platelet-derived growth factor (PDGF), (c) a combination of PDGF and 5-hydroxytryptamine (5-HT) or (d) serum. 2. Ro 31-8220 and Ro 31-7549 concentration-dependently inhibited proliferation in response to each mitogen. The inhibitory potency (IC50) of Ro 31-8220 and Ro 31-7549, respectively, was similar against proliferation induced by PDBu (0.55 and 1.1 microM), PDGF (0.6 and 0.9 microM), PDGF and 5-HT (0.68 and 1.1 microM), although slightly less against serum (1.7 and 5 microM). The effects of the protein kinase C inhibitors on proliferation could not be ascribed to cytotoxicity. Neither Ro 31-8220 nor Ro 31-7549 (0.3-3 microM) inhibited PDGF receptor tyrosine phosphorylation. 3. The results show that Ro 31-8220 and Ro 31-7549 are potent inhibitors of smooth muscle cell proliferation in response to a direct activator of protein kinase C, the defined growth factors, PDGF and 5-HT, and the complex mixture of mitogens in serum. Protein kinase C activation thus appears to be an important growth transducing mechanism for each of these agents.

Absence of association between Alzheimer disease and the -491 regulatory region polymorphism of APOE.

A novel polymorphism (-491 A/T) within the regulatory region on the apolipoprotein E gene has recently been reported to be associated with risk for Alzheimer disease (AD). To test this association in an independent data set, we have examined this polymorphism in a sample of 88 well-characterized AD cases and compared the allele frequency and genotype frequencies for this polymorphism with those observed in 112 cognitively normal subjects drawn from the same ethnic group. These results suggest that in the current data set at least, the -491 A/T polymorphism is not associated with risk for AD, but may be in partial linkage disequilibrium with the APOE epsilon2/epsilon3/epsilon4 polymorphism.

Schedule-dependent effects of haloperidol and amphetamine: multiple-schedule task shows within-subject effects.

A two-lever, multiple-schedule task was used to evaluate the effects of haloperidol (HA) and amphetamine (AM) on responding controlled by continuous reinforcement (CRF) and progressive ratio (PR) schedules of reinforcement. Rats were trained to press one lever for food delivered on a CRF schedule and the other lever for food delivered on a PR schedule. The operative schedule was signaled by the illumination of a cuelight mounted above the appropriate lever. Following 30 sessions of training, dose-response functions were determined for HA (0.0075, 0.015, 0.03, and 0.06 mg/kg) and AM (0.0625, 0.125, 0.25, 0.50, 0.75, and 1.00 mg/kg). Both drugs produced dose- and schedule-dependent effects. For example, administration of 0.03 mg/kg HA did not affect responding under the CRF schedule but did reduce responding during PR components, whereas administration of 0.06 mg/kg reduced responding under both schedules of reinforcement. Some doses of AM produced increased responding under the CRF schedule and, within the same session, decreased responding under the PR schedule. The results with HA are consistent with the view that interfering with dopaminergic function affects the allocation and maintenance of responding and that this effect depends on properties of the schedule of reinforcement. The results with AM emphasize that statements about the effects of the drug on positively reinforced behavior cannot be made without reference to specific schedules of reinforcement.

The genetics of Alzheimer's disease.

Since the first description of Alzheimer's disease (AD) at the beginning of the century until relatively recently, it was customary to define Alzheimer's disease as occurring in the presenium. The same neuropathological changes occurring in brains over the age of 65 were called "senile dementia." Because there have been no clinical or pathological features to separate the two groups, this somewhat arbitrary distinction has been abandoned. Although AD is currently considered to be a heterogeneous disease, the most consistent risk factor to be implicated other than advancing age is the presence of a positive family history. This potential genetic vulnerability to AD has been recognized for some time. Some of the earliest evidence suggestive of a genetic contribution to AD came from Kallmann's 1956 study (1) demonstrating a higher concordance rate in monozygotic twins for "parenchymatous senile dementia" compared with dizygotic twins and siblings. This monozygotic excess has been confirmed in studies applying more rigorous diagnostic criteria although there may be widely disparate ages of onset between twins (2). The most convincing evidence for a genetic contribution to AD has come form the study of pedigrees in which the pattern of disease segregation can be clearly defined. Thus, the abandonment of the early and late-onset dichotomy has occurred at a time when, at the genetic level, important differences have been identified through the discovery of specific gene defects in early onset cases.

Tie-1 receptor tyrosine kinase endodomain interaction with SHP2: potential signalling mechanisms and roles in angiogenesis.

The endothelial receptor tyrosine kinase plays an essential role in vascular development where it is thought to be required for vessel maturation and stabilization. The ligands responsible for activating Tie-1, its signalling pathways and specific cellular functions are however not known. As with some other receptor tyrosine kinases, Tie-1 is subject to extracellular proteolytic cleavage generating a membrane bound receptor fragment comprising the intracellular and transmembrane domains. Here we examine the signalling potential of this Tie-1 endodomain. We show that the Tie-1 endodomain has poor ability to induce tyrosine phosphorylation. However, on formation the endodomain physically associates with a number of tyrosine phosphorylated signalling intermediates including the tyrosine phosphatase and adaptor protein SHP2. The assembly of this multimolecular complex is consistent with the endodomain having a ligand-independent signalling role in the endothelial cell. The potential roles of ectodomain cleavage and cleavage activated signalling in regulating microvessel stability in angiogenesis, vessel remodelling and regression are considered.

Role of vascular endothelial cells in the allograft response.

Following transplantation endothelial cells lining an allograft come into contact with immune cells of the recipient. Activation of an immune response, by graft endothelial or other cells, will lead to local increases in cytokine production and cell-mediated lysis. Inflammatory cytokines have been shown, mainly in vitro, to have marked effects on endothelial function and act to produce a pro-thrombotic, pro-adhesive and promitogenic phenotype. These data are reviewed and ways in which these changes could lead to rejection due to graft lysis or vascular occlusion are discussed.

Suppression of triglyceride secretion by epinephrine in isolated rat hepatocytes.

The effect of epinephrine on triglyceride synthesis and secretion was examined in isolated rat hepatocytes. Epinephrine potently inhibited triglyceride secretion but did not affect cellular triglyceride content or the rate of incorporation of radiolabelled glycerol into cell triglyceride. The inhibitory effect of epinephrine was abolished by inclusion of the alpha-adrenergic antagonist prazosin but not the beta-antagonist propranolol.

Alpha-adrenergic suppression of very-low-density-lipoprotein triacylglycerol secretion by isolated rat hepatocytes.

The effect of adrenaline on triacylglycerol synthesis and secretion was examined in isolated rat hepatocytes. Cells were incubated with 0.5 mM-[1-14C]oleate, and the accumulation of triacylglycerol and [14C]triacylglycerol was measured in the incubation medium. Triacylglycerol appearing in the medium was present in a form with properties similar to very-low-density lipoproteins. Triacylglycerol, [14C]triacylglycerol and [14C]phospholipid contents of hepatocytes were also determined. Addition of 10 microM-(-)adrenaline decreased accumulation of glycerolipid in the incubation medium and also decreased cellular [14C]phospholipid content. Prazosin abolished these effects, whereas propranolol did not. The hormone did not affect cellular triacylglycerol content or rates of incorporation of [1-14C]oleate into cell triacylglycerol. The effect of adrenaline on the removal of newly secreted triacylglycerol and the secretion of synthesized glycerolipid was also examined. The catecholamine did not affect rates of removal of newly secreted triacylglycerol. Adrenaline did inhibit the secretion of pre-synthesized lipid by the cells, as assessed by the appearance of radiolabelled triacylglycerol from hepatocytes that had been preincubated with [1,2,3-3H]-glycerol. Adrenaline did not affect rates of fatty acid uptake by hepatocytes, but did stimulate oxidation of [1-14C]oleate, principally to 14CO2.

Regulation of carnitine palmitoyltransferase activity by malonyl-CoA in mitochondria from sheep liver, a tissue with a low capacity for fatty acid synthesis.

The characteristics of inhibition of carnitine palmitoyltransferase (CPT) I by malonyl-CoA were studied for the enzyme in mitochondria isolated from sheep liver, a tissue with a very low rate of fatty acid synthesis. Malonyl-CoA was as potent in inhibiting the sheep liver enzyme as in inhibiting the enzyme in rat liver mitochondria. CPT I in guinea-pig liver mitochondria was also similarly inhibited. The inhibition showed the same time-dependent characteristics previously established for the rat liver enzyme. Methylmalonyl-CoA was as effective an inhibitor of CPT I as malonyl-CoA in sheep liver mitochondria, but did not affect CPT I activity in mitochondria from rat or guinea-pig liver. The concentrations of malonyl-CoA required to inhibit CPT I in sheep liver mitochondria in vitro were similar to those found in freeze-clamped sheep liver samples (about 7 nmol of malonyl-CoA/g wet wt.). In sheep liver cells the content of malonyl-CoA was only one-tenth of that observed in vivo when glucose only was added to the incubation medium. Inclusion of acetate and/or insulin increased the malonyl-CoA content about 10-fold, to values similar to those observed in vivo. The rate of fatty acid synthesis in sheep liver cells was about 1% of that observed in rat liver, but was correlated with the concentrations of malonyl-CoA in the cells under various incubation conditions. These observations are discussed in relation to (i) the regulatory role of malonyl-CoA in tissues that have a low capacity for fatty acid synthesis, and (ii) the utilization by sheep liver of propionate as a gluconeogenic precursor.

Involvement of protein tyrosine kinases in regulation of endothelial cell organization by basement membrane proteins.

During capillary formation, endothelial cell migration and organization are critically dependent on surrounding basement membrane proteins. These proteins serve as a physical support and are likely to provide signals which regulate migration and organization of the cells. In this study the possible involvement of tyrosine kinase signalling pathways in basement membrane-induced organization of human endothelial cells is examined. Interaction of endothelial cells with reconstituted basement membrane Matrigel activates tyrosine phosphorylation of several proteins including focal adhesion kinase. Inhibition of this pathway with tyrosine kinase inhibitors impairs localization of paxillin to focal adhesions and organization of actin filaments, decreases motility and elongation of endothelial cells, and prevents their organization into cords or tubes on basement membrane. These data demonstrate that basement membrane-induced modulation of endothelial cell motility, shape, and organization is critically dependent on tyrosine kinase signalling pathway(s) involving cytoskeletal proteins.