Hindered proton collectivity in 16(28)S12: possible magic number at Z=16.

The reduced transition probability B(E2;0(gs)(+)→2(1)(+)) for (28)S was obtained experimentally using Coulomb excitation at 53 MeV/nucleon. The resultant B(E2) value 181(31) e(2)fm(4) is smaller than the expectation based on empirical B(E2) systematics. The double ratio |M(n)/M(p)|/(N/Z) of the 0(gs)(+)→2(1)(+) transition in (28)S was determined to be 1.9(2) by evaluating the M(n) value from the known B(E2) value of the mirror nucleus (28)Mg, showing the hindrance of proton collectivity relative to that of neutrons. These results indicate the emergence of the magic number Z=16 in the |T(z)|=2 nucleus (28)S.

Caloric restriction, aerobic exercise training and soluble lectin-like oxidized LDL receptor-1 levels in overweight and obese post-menopausal women.

BACKGROUND: Elevated circulating levels of soluble lectin-like oxidized low-density lipoprotein receptor-1 (sLOX-1) have been observed in obese persons and are reduced by weight loss. However, it is not known whether combining caloric restriction (CR) with exercise training is better in reducing sLOX-1 levels than CR alone. OBJECTIVE: We examined whether the addition of aerobic exercise to a weight loss intervention differentially affects sLOX-1 levels in 61 abdominally obese post-menopausal women randomly assigned to a CR only (n = 22), CR+moderate-intensity exercise (n = 22) or CR+vigorous-intensity exercise (n = 17) intervention for 20 weeks. The caloric deficit was ~2800 kcal per week for all groups. RESULTS: The intervention groups were similar at baseline with respect to body weight, body composition, lipids and blood pressure. However, plasma sLOX-1 levels were higher in the CR-only group (99.90 ± 8.23 pg ml(-1)) compared with both the CR+moderate-intensity exercise (69.39 ± 8.23 pg ml(-1), P = 0.01) and the CR+vigorous-intensity exercise (72.83 ± 9.36 pg ml(-1), P = 0.03) groups. All three interventions significantly reduced body weight (~14%), body fat and waist and hip circumferences to a similar degree. These changes were accompanied by a 23% reduction in sLOX-1 levels overall (-19.00 ± 30.08 pg ml(-1), P < 0.0001), which did not differ among intervention groups (P = 0.13). Changes in body weight, body fat and maximal oxygen consumption (VO(2) max) were not correlated with changes in sLOX-1 levels. In multiple regression analyses in all women combined, baseline sLOX-1 levels (ß = -0.70 ± 0.06, P < 0.0001), age (ß = 0.92 ± 0.43, P = 0.03) and baseline body mass index (BMI) (ß = 1.88 ± 0.66, P = 0.006) were independent predictors of the change in sLOX-1 with weight loss. CONCLUSIONS: Weight loss interventions of equal energy deficit have similar effects on sLOX-1 levels in overweight and obese post-menopausal women, with the addition of aerobic exercise having no added benefit when performed in conjunction with CR.

Noradrenergic excitation of a subpopulation of GABAergic cells in the basolateral amygdala via both activation of nonselective cationic conductance and suppression of resting K+ conductance: a study using glutamate decarboxylase 67-green fluorescent protein knock-in mice.

GABAergic interneurons play central roles in the regulation of neuronal activity in the basolateral nucleus of the amygdala (BLA). They are also suggested to be the principal targets of the brainstem noradrenergic afferents which are involved in the enhancement of the BLA-related memory. In addition, behavioral stress has been shown to impair noradrenergic facilitation of GABAergic transmission. However, the noradrenaline (NA) effects in the BLA have not been differentiated among medium- to large-sized GABAergic neurons and principal cells, and remain to be elucidated in terms of their underlying mechanisms. Glutamate decarboxylase 67 (GAD67) is a biosynthetic enzyme of GABA and is specifically expressed in GABAergic neurons. To facilitate the study of the NA effects on GABAergic neurons in live preparations, we generated GAD67-green fluorescent protein (GFP) knock-in mice, in which GFP was expressed under the control of an endogenous GAD67 gene promoter. Here, we show that GFP was specifically expressed in GABAergic neurons in the BLA of this GAD67-GFP knock-in mouse. Under whole-cell patch-clamp recordings in vitro, we identified a certain subpopulation of GABAergic neurons in the BLA chiefly on the basis of the electrophysiological properties. When depolarized by a current injection, these neurons, which are referred to as type A, generated action potentials at relatively low frequency. We found that NA directly excited type-A cells via alpha1-adrenoceptors, whereas its effects on the other types of neurons were negligible. Two ionic mechanisms were involved in this excitability: the activation of nonselective cationic conductance and the suppression of the resting K+ conductance. NA also increased the frequency of spontaneous IPSCs in the principal cells of the BLA. It is suggested that the NA-dependent excitation of type-A cells attenuates the BLA output for a certain period.

Respiratory activity in brainstem of fetal mice lacking glutamate decarboxylase 65/67 and vesicular GABA transporter.

The respiratory neural network in the mammalian medulla oblongata shows rhythmic activity before birth. GABA and glycine are considered to be involved in control of respiratory rhythm. Recently we have demonstrated respiratory failure in glutamic acid decarboxylase (GAD) 67-deficient mice [Tsunekawa N, Arata A, Obata K (2005) Development of spontaneous mouth/tongue movement and related neural activity, and their repression in mouse fetus lacking glutamate decarboxylase 67. Eur J Neurosci 21:173-178]. To further evaluate the involvement of GABA and glycine in fetal respiratory function, we studied neural activities in brainstem-spinal cord blocks prepared from GAD65-/-:67-/- and vesicular GABA transporter (VGAT)-/-mice on embryonic day 14 (E14)-E15 and E18. In these knockout mice, the synthesis of GABA and the vesicular release of GABA and glycine are completely absent, respectively. Spontaneous respiratory discharges were observed in the ventral roots at the cervical cord (C) 4 level from wild-type mice but not from the knockout mice on E18. Administration of substance P induced C4 discharges in GAD65-/-:67-/- preparations but not in VGAT-/- preparations. C4 discharges were observed in the knockout mice on E14-E15, although the frequency was lower than that in the wild-type. Neuronal activities in the respiratory network of the E18 brainstem were recorded using a "blind" patch-clamp technique. Expiratory and inspiratory neurons with their characteristic firing patterns were observed in the wild-type fetuses. Strychnine reversed inspiratory-phase hyperpolarization to large depolarization in expiratory neurons. On the other hand, neurons in the same area of the knockout mice fired spontaneously without any rhythm. Substance P induced hyperpolarizing potentials in medullary neurons of GAD65-/-:67-/- mice. Further administration of strychnine induced large depolarizing potentials. Rhythmic activities were not observed in VGAT-/- mice even in the presence of substance P and strychnine. These results indicate that the lack of GABA and glycine impairs the function of the respiratory network in mouse fetuses and the impairment progresses with fetal age.

Altered conditioned fear behavior in glutamate decarboxylase 65 null mutant mice.

We investigated the involvement of the 65 kDa isoform of glutamic acid decarboxylase (GAD65) and GAD65-mediated gamma-aminobutyric acid (GABA) synthesis in the formation and expression of Pavlovian fear memory. To this end, behavioral, endocrine and autonomic parameters were examined during conditioned fear retrieval of mice with targeted ablation of the GAD65 gene (GAD65-/- mice). These mutant mice were found to display specific fear behavior (freezing, escape), as well as autonomic (increased defecation) and endocrine activation (increased plasma corticosterone) during fear memory retrieval. However, freezing was reduced and flight and escape behavior were increased in GAD65-/- mice compared to their wild type and heterozygous littermates, while corticosterone levels and defecation rates did not differ between genotypes. Active defensive behavior of GAD65-/- mice was observed during both auditory cued and contextual retrieval of fear memory, as well as immediately after conditioning. These data indicate a selectively altered behavioral fear response in GAD65-/- mice, most likely due to deficits in threat estimation or the elicitation of appropriate conditioned fear behavior, and suggest that GAD65 is a genetic determinant of conditioned fear behavior. GAD65-/- mice provide a valuable tool to further dissect the GABAergic mechanisms involved in fear and anxiety and to model GABA-related neurological and psychiatric disorders.

Expression of SR-PSOX, a novel cell-surface scavenger receptor for phosphatidylserine and oxidized LDL in human atherosclerotic lesions.

Receptor-mediated endocytosis of oxidized low density lipoprotein (Ox-LDL) by macrophages and the subsequent foam cell transformation in the arterial intima are key events in early atherogenesis. Recently, we have identified a novel macrophage cell-surface receptor for Ox-LDL by expression cloning from a cDNA library of phorbol 12-myristate 13-acetate-stimulated THP-1 cells, designated as the scavenger receptor for phosphatidylserine and oxidized lipoprotein (SR-PSOX). Here, we examined SR-PSOX expression in human atherosclerotic lesions. Total cellular RNA and fresh frozen sections were prepared from human carotid endarterectomy specimens (from 21 patients) and directional coronary atherectomy specimens (from 11 patients). Fragments of human aortas of 2 patients without visible atherosclerotic lesions served as negative controls. Quantitative reverse transcription-polymerase chain reaction demonstrated that SR-PSOX mRNA expression was prominent in atherosclerotic lesions but undetectable in normal aortas. Immunohistochemistry showed that SR-PSOX was predominantly expressed by lipid-laden macrophages in the intima of atherosclerotic plaques in carotid endarterectomy and directional coronary atherectomy specimens, although its expression was not detectable in normal arterial wall. Double-labeled immunohistochemistry confirmed that SR-PSOX is expressed by intimal macrophages. Taken together, SR-PSOX may be involved in Ox-LDL uptake and subsequent foam cell transformation in macrophages in vivo and thus may play important roles in human atherosclerotic lesion formation.

VCAM-1 expression precedes macrophage infiltration into subendothelium of vein grafts interposed into carotid arteries in hypercholesterolemic rabbits--a potential role in vein graft atherosclerosis.

Intimal hyperplasia and atherosclerosis are major causes of late vein graft failure after coronary artery bypass surgery. Hypercholesterolemia appears to be a key risk factor for atherosclerosis in vein grafts as well as in native arteries. We used a rabbit model of interposition jugular vein graft to the carotid artery and compared intimal thickening, macrophage accumulation, and VCAM-1 expression between hypercholesterolemic (H group) and normocholesterolemic (N group) rabbits. Fifty-nine rabbits were divided into H and N groups. Intimal thickening in vein grafts was approximately three times more prominent in the H group than in the N group. Macrophage accumulation progressively increased with time in H group vein grafts, although it was negligible in the N group. In the H group, moreover, macrophages were initially more abundant in deep intima, and subsequently accumulated in subendothelium of the grafted vein. VCAM-1 expression in luminal endothelial cells of the grafted veins was time-dependently increased after the vein graft surgery in both the H and N groups, and was more prominent in the H group. Comparison of the time-courses between macrophage accumulation and VCAM-1 expression revealed that VCAM-1 expression in luminal endothelium preceded subendothelial accumulation of macrophages. VCAM-1 expression and macrophage accumulation may be key factors which regulate progression of vein graft atherosclerosis.

Relationship between thallium-201 kinetics and proliferative activity assessed by monoclonal antibody MIB-1 in brain tumours.

To clarify the relationship between thallium-201 chloride kinetics and proliferative activity in brain tumours. a single-photon emission tomographic (SPET) study was performed and the results correlated with monoclonal antibody MIB-1 staining of the tumour tissue. 201T1 SPET was performed 10 min (early scan) and 2 h (delayed scan) after intravenous administration of 201TI (111 MBq) in 34 intra-axial tumours including 24 malignant tumours, and in 27 extra-axial tumours including one malignant tumour. Tumour 201T1 kinetic parameters [early and delayed uptake ratios (ER and DR, respectively), retention index (RI), and the ratio of tumour delayed activity to early activity (Td/Te)] were compared with tumour tissue MIB-1 labelling indices (MIB-1 LI) representative of tumour cell proliferative activity. In the intra-axial tumours, ER and DR and MIB-1 LI were significantly higher in the malignant tumours than in the benign tumours. ER and DR were significantly correlated with MIB-1 LI (P<0.01 and P<0.05, respectively), but RI and Td/Te were not. In the extra-axial benign tumours, ER was as high as that in the intra-axial malignant tumours, while MIB-1 LI was equal to that in the intra-axial benign tumours. There were no significant correlations between any 201T1 kinetic parameters and MIB-1 LI. This study indicates that 201T1 ER may be the most reliable parameter for predicting the proliferative activity of intra-axial tumours.

Roles of lectin-like oxidized LDL receptor-1 and its soluble forms in atherogenesis.

Lectin-like oxidized LDL receptor (LOX)-1 is a type II membrane protein that belongs to the C-type lectin family of molecules, which can act as a cell-surface endocytosis receptor for atherogenic oxidized LDL. LOX-1 can support binding, internalization and proteolytic degradation of oxidized LDL, but not of significant amounts of acetylated LDL, which is a well-known high-affinity ligand for class A scavenger receptors and scavenger receptor expressed by endothelial cells (SR-EC). LOX-1 is initially synthesized as a 40-kDa precursor protein with N-linked high mannose-type carbohydrate, which is further glycosylated and processed into a 50-kDa mature form. LOX-1 expression is not constitutive, but can be induced by proinflammatory stimuli, such as tumour necrosis factor-alpha, transforming growth factor-beta and bacterial endotoxin, as well as angiotensin II, oxidized LDL itself and fluid shear stress. In addition, LOX-1 expression is detectable in cultured macrophages and activated vascular smooth muscle cells. In vivo, endothelial cells that cover early atherosclerotic lesions, and intimal macrophages and smooth muscle cells in advanced atherosclerotic plaques can express LOX-1. Cell-surface LOX-1 can be cleaved through some protease activities that are associated with the plasma membrane, and released into the culture media. Purification of soluble LOX-1 and the N-terminal amino-acid sequencing identified the two cleavage sites (Arg86-Ser87 and Lys89-Ser90), both of which are located in the membrane proximal extracellular domain of LOX-1. Measurement of soluble LOX-1 in vivo may provide a novel diagnostic tool for the evaluation and prediction of atherosclerosis and vascular disease.

Lectin-like oxidized low density lipoprotein receptor-1 (LOX-1) supports cell adhesion to fibronectin.

Lectin-like oxidized lipoprotein receptor-1 (LOX-1) is a specific receptor for atherogenic oxidized low density lipoprotein (OxLDL) which belongs to the scavenger receptor family. In the present report, we show that LOX-1 can also support cell adhesion to fibronectin (FN) in a divalent cation-independent fashion. CHO-K1 cells stably expressing bovine LOX-1 (BLOX-1-CHO), but not untransfected CHO-K1 cells, can adhere to FN-coated plates, but not to collagen-coated plates, in the presence of EDTA. BLOX-1-CHO adhesion to FN-coated plates can also be suppressed by scavenger receptor ligands, such as OxLDL, polyinosinic acid (poly I), and dextran sulfate, but not by native LDL, acetylated LDL, polycytidylic acid (poly C), or chondroitin sulfate. Cultured bovine aortic endothelial cells can similarly adhere to FN-coated plates, which was inhibited by OxLDL, poly I, and dextran sulfate in the presence of EDTA. LOX-1 may play an important role in cell adhesion to FN in an integrin-independent manner.

Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) in atherogenesis.

Lectin-like oxidized low density lipoprotein receptor-1 (LOX-1) is a type-II membrane protein belonging to the C-type lectin family molecules, which can act as a cell surface endocytosis receptor for atherogenic oxidized LDL (Ox-LDL). LOX-1 is synthesized as a 40 kDa precursor protein with N-linked high mannose-type carbohydrate, which is further glycosylated and processed into a 50 kDa mature form. LOX-1 expression is not constitutive but can be induced by proinflammatory, oxidative, and mechanical stimuli. In addition to endothelial cells, macrophages and activated vascular smooth muscle cells express LOX-1. In vivo, endothelial cells covering early atherosclerotic lesions and macrophages and smooth muscle cells accumulated in the intima of advanced atherosclerotic plaques express LOX-1. LOX-1 is cleaved at membrane proximal extracellular domain by some protease activities and released from the cell surface. Measurement of soluble LOX-1 in vivo may provide novel diagnostic strategy for the evaluation and prediction of atherosclerosis and vascular diseases.

Oxidized LDL modulates Bax/Bcl-2 through the lectinlike Ox-LDL receptor-1 in vascular smooth muscle cells.

Oxidized low density lipoprotein (Ox-LDL) induces apoptosis in vascular smooth muscle cells (VSMCs), which may increase atherosclerotic plaque instability. In this study, we examined the molecular mechanisms causing the Ox-LDL-induced apoptosis in VSMCs, especially focusing on the involvement of Bax/Bcl-2 and the lectinlike Ox-LDL receptor-1 (LOX-1). In cultured bovine aortic smooth muscle cells (BASMCs), Ox-LDL at high concentrations (>60 microg/mL) induced cell death as demonstrated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. DNA fragmentation was increased in BASMCs treated with high concentrations of Ox-LDL, indicating that the Ox-LDL-induced cell death in VSMCs was apoptosis. Ox-LDL upregulated LOX-1 expression through phosphorylation of extracellular signal-regulated kinase in BASMCs, and a neutralizing anti-LOX-1 monoclonal antibody, which can block LOX-1-mediated cellular uptake of Ox-LDL, prevented the Ox-LDL-induced apoptosis in BASMCs. This antibody also suppressed the increase in the Bax to Bcl-2 ratio induced by Ox-LDL in BASMCs. Furthermore, LOX-1 expression was well colocalized with Bax expression in the rupture-prone shoulder areas of human atherosclerotic plaques in vivo. LOX-1 may play an important role in Ox-LDL-induced apoptosis in VSMCs by modulating the Bax to Bcl-2 ratio. These molecular mechanisms may be involved in destabilization and rupture of atherosclerotic plaques.

Lysophosphatidylcholine induces early growth response factor-1 expression and activates the core promoter of PDGF-A chain in vascular endothelial cells.

Lysophosphatidylcholine (lyso-PC), a polar phospholipid that is increased in atherogenic lipoproteins and atherosclerotic lesions, has been shown to transcriptionally induce the expression of endothelial genes relevant to atherogenesis. In cultured bovine aortic endothelial cells (BAECs), we show that lyso-PC induces the expression of early growth response factor (Egr)-1 and thereby activates the proximal promoter of the platelet-derived growth factor (PDGF)-A chain located 55 to 71 bp upstream from the transcription start site, which has been shown to be crucial for PDGF-A chain expression induced by fluid shear stress and fibroblast growth factor-1. Northern blot analyses showed that lyso-PC (10 to 20 micromol/L) transiently (30 minutes to 1 hour) induced expression of Egr-1 mRNA. Induced expression of Egr-1 mRNA, which was associated with increased amounts of Egr-1 protein in nuclei, preceded PDGF-A chain mRNA induction in lyso-PC-activated BAECS: Nuclear runoff assay revealed that lyso-PC stimulates transcription of the Egr-1 gene. Transient transfection of the oligonucleotide corresponding to the proximal promoter of the PDGF-A chain (oligo A) linked to the luciferase reporter gene revealed that lyso-PC can activate the core promoter of the PDGF-A chain by 5-fold. Insertion of a guanine at 3 sites in the oligo A abolished the lyso-PC-induced increases in luciferase activities. Electrophoretic mobility shift assay with use of radiolabeled oligo A showed a lyso-PC-inducible shift band, which was suppressed by excess amounts of unlabeled oligo A or an anti-Egr-1 antibody. In addition, lyso-PC-induced Egr-1 expression was inhibited by PD98059, a specific inhibitor of mitogen-activated protein kinase kinase-1 (MEK1), suggesting that lyso-PC-induced expression of Egr-1 depends on the MEK1/extracellular signal-regulated kinase pathway. Taken together, transcriptional activation of Egr-1-dependent genes by this atherogenic lipid may be a key regulator of atherogenesis.

Assessment of leg oedema by dynamic lymphoscintigraphy with intradermal injection of technetium-99m human serum albumin and load produced by standing.

This study was a preliminary evaluation of the utility of dynamic lymphoscintigraphy with technetium-99m human serum albumin (HSA) and a load produced by standing in the assessment of lymphatic dysfunction in patients with leg oedema. The 71 subjects investigated included 53 patients with lymphoedema, six with venous occlusion alone and five with lymphovenous occlusion, as well as seven normal subjects. After intradermal injection of 99mTc-HSA into an interdigital space in each foot, dynamic scintigrams were recorded with the patient supine for 15 min. The subjects then stood in place and images were recorded for an additional 15 min. Relative changes in lymphatic tracer transport before and after standing were analysed on time-activity curves (TACs). This test was compared with a conventional test in a supine position in six patients with lymphoedema, and was repeated in five other patients with lymphoedema. It was found that in the normal limbs, a standing load activated tracer transport to the draining lymphatic vessels, resulting in a rapid stepwise increase in tracer activity, large spiking waves and a decreasing phase following a peak in tracer activity on TACs. In 59 lymphoedematous limbs, including some with a mild form of oedema without morphological abnormalities on scintigrams, this load failed to induce a sufficient activation of tracer transport, and the frequencies of each of the three normally appearing changes described above significantly decreased compared with those in the 14 normal limbs (P < 0.0001, P < 0.01 and P < 0.0001, respectively). In addition, there were significant reductions in the relative increases in maximum activity and clearance times after standing (both P < 0.0001). These abnormalities significantly correlated with the grade of severity of oedema. Six limbs with lymphovenous occlusion showed significant reductions in tracer transport compared to six limbs with venous occlusion. Lymphatic dysfunction was accentuated more by this test than by the conventional test, and repeated tests showed consistent results in the same individuals. It is concluded that under a standardized load, this quick test seems of value in providing a sensitive and objective assessment of lymphatic dysfunction in the lower limbs, and is also advantageous for image interpretation since accelerated tracer transport clearly visualizes compromised lymphatics. This test may also be helpful in distinguishing purely venous oedema from mixed lymphovenous disease.