New confining force solution of the QCD axion domain-wall problem.

The serious cosmological problems created by the axion-string-axion-domain-wall system in standard axion models are alleviated by positing the existence of a new confining force. The instantons of this force can generate an axion potential that erases the axion strings long before QCD effects become important, thus preventing QCD-generated axion walls from ever appearing. Axion walls generated by the new confining force would decay so early as not to contribute significantly to the energy in axion dark matter.

Obesity, pregnancy, inflammation, and vascular function.

Maternal obesity is associated with increased morbidity and mortality for both mother and offspring. The mechanisms underlying the increased risk associated with maternal obesity are not well understood. In non-pregnant populations, many of the complications of obesity are thought to be mediated in part by inflammation and its sequelae. Recent studies suggest that a heightened inflammatory response may also be involved in mediating adverse clinical outcomes during pregnancy. This review summarizes our current knowledge about adipose tissue biology, and its role as an endocrine and inflammatory organ. The evidence for inflammation as a key mediator of adverse pregnancy outcome is also presented, focusing on the role of inflammation in adipose tissue, systemic inflammation, the placenta, and vascular endothelium.

New type of seesaw mechanism for neutrino masses.

In a wide class of unified models there is an additional (and possibly dominant) term in the neutrino mass formula that under the simplest assumption takes the form M(nu)=(M(N)+M(T)(N))u/M(G), where M(N) is the neutrino Dirac mass matrix, and u=O(M(W)). This makes possible highly predictive models. A generalization of this form yields realistic neutrino masses and mixings more readily than the usual seesaw formula in some models. The conditions for resonant enhancement of leptogenesis can occur naturally in such models.

Ras-induced colony formation and anchorage-independent growth inhibited by elevated expression of Puralpha in NIH3T3 cells.

Levels of Puralpha, a conserved, sequence-specific single-stranded DNA and RNA binding protein, fluctuate during the cell cycle, declining at the onset of S-phase and peaking at mitosis. In early G1 Puralpha is associated with the hypophosphorylated form of the retinoblastoma protein, Rb. Microinjection of purified Puralpha into NIH3T3 cells arrests the cell cycle at either G1/S or G2/M checkpoints with distinct morphological consequences. Here we ask whether expression of Puralpha can affect colony formation and anchorage-independent growth in ras-transformed NIH3T3 cells. Two to five-fold elevated levels of Puralpha in stably-transfected cell lines retard entry into and progression through S phase in both ras-transformed and non-transformed cells. Puralpha significantly inhibits colony formation by ras-transformed cells but not by non-transformed cells. In addition, cells transfected to express Puralpha formed only about 1/5 the number of large colonies in soft agar as control-transfected cells, demonstrating a marked inhibition of anchorage-independent growth by Puralpha. Biochemical analysis of nuclear and cytoplasmic Puralpha proteins and confocal microscopic analysis of Puralpha location indicate that access of Puralpha to the nucleus is controlled by both protein modification and sequence domains within the protein. Analyses of deletion mutants identify Puralpha domains mediating nuclear exclusion, including several potential destruction motifs and a PEST sequence at aa's 215-231. In the nucleus Puralpha colocalizes with CDK2 and cyclin A. Puralpha and cyclin D1, however, do not colocalize in the nucleus. At mitosis Puralpha is visualized about the condensed chromosomes and in the cytoplasm, where it colocalizes with cyclin B1. The data indicate that the ability of Puralpha to interact with proteins regulating cell proliferation and transformation is controlled by signals that govern its intracellular localization.

Association of human Pur alpha with the retinoblastoma protein, Rb, regulates binding to the single-stranded DNA Pur alpha recognition element.

The retinoblastoma protein, Rb, is detected in extracts of monkey CV-1 cells complexed with Pur alpha, a sequence-specific single-stranded DNA-binding protein implicated in control of gene transcription and DNA replication. These complexes can be immunoextracted from cell lysates using monoclonal antibodies to either Pur alpha or Rb. The Pur alpha-Rb complexes contain a form of Pur alpha with extensive post-synthetic modification, as demonstrated following expression of Pur alpha cDNA fused to a 9-amino acid epitope tag. Human Pur alpha, expressed as a glutathione S-transferase fusion protein, specifically binds to the hypophosphorylated form of Rb with an affinity as high as that of SV40 large T-antigen. In the absence of DNA, glutathione S-transferase-Pur alpha binds to p56RB, an NH2-terminal-truncated Rb protein purified from Escherichia coli, containing the T-antigen binding domain, to form multimeric complexes. The single-stranded DNA Pur alpha recognition element disrupts these complexes. Conversely, high concentrations of p56RB prevent Pur alpha binding to DNA. Through use of a series of deletion mutants, the DNA binding activity of Pur alpha is localized to a series of modular amino acid repeats. Rb binding involves a Pur alpha region with limited homology to the Rb-binding region of SV40 large T-antigen. Binding of Pur alpha to p56RB, the COOH-terminal portion of Rb, is inhibited by a synthetic peptide containing the T-antigen Rb-binding motif.

Cooperative action of cellular proteins YB-1 and Pur alpha with the tumor antigen of the human JC polyomavirus determines their interaction with the viral lytic control element.

Human JC polyomavirus (JCV) is the etiologic agent of the neurodegenerative disease progressive mulifocal leukoencephalopathy. By using JCV as a model, we investigated the role of the viral early protein tumor antigen (TAg) in the binding of two cellular proteins, Pura alpha and YB-1, to JCV regulatory sequences. Results from band-shift assays with purified YB-1, Pur alpha, and TAg indicated that efficient binding of Pur alpha, a strong activator of early gene transcription, to a single-stranded target sequence corresponding to the viral lytic control element, is diminished in the presence of the late gene activator YB-1, which recognizes the opposite strand of the Pur alpha binding site. Of particular interest was the ability of Pur alpha and TAg to enhance binding of YB-1 to DNA molecules without being associated with this complex. Binding studies using a mutant peptide encompassing the N terminus of YB-1 indicated that the C terminus of YB-1 is important for its DNA binding activity. The ability of Pur alpha and TAg to increase binding of YB-1 to DNA is independent of the YB-1 C terminus. Similarly, results from band-shift assays using Pur alpha variants indicated that two distinct regions of this protein contribute either to its ability to bind DNA or to its ability to enhance YB-1 DNA binding activity. Based on the interaction of Pur alpha, YB-1, and TAg, and their binding to DNA, a model is proposed for the role of these proteins in transcription of viral early and late genes during the lytic cycle.

Metoprolol and ventricular repolarisation and refractoriness: lack of chronic adaptational class III effects in rabbit.

STUDY OBJECTIVE: The aim was to examine the right ventricular electrophysiological changes during metoprolol treatment and in conditions of stress in the adult rabbit. DESIGN: Metoprolol (6 mg.kg-1) was given twice daily for four weeks. Stress was induced by constant infusion of adrenaline (15.2 micrograms.kg-1.h-1) via osmotic pumps implanted in the left femoral vein. Control rabbits were treated with saline. Electrophysiological recordings were made weekly in conscious animals using bipolar pacing electrodes implanted in the right ventricular apex. SUBJECTS: Adult male New Zealand white rabbits, 2.8-3.3 kg, were used (n = 7 adrenaline/metoprolol treated, n = 7 adrenaline/saline treated, n = 7 saline/metoprolol treated, and n = 7 saline/saline treated). MEASUREMENTS AND MAIN RESULTS: Recordings were made of the effective refractory period and of the stimulus-T interval of the paced evoked response. Stimulus-T is used as an index of ventricular repolarisation time. No significant electrophysiological changes in these variables were observed throughout the study. CONCLUSIONS: These results indicate that chronic metoprolol therapy does not result in a class III antiarrhythmic effect in this in vivo rabbit model.

Selective urinary screening for mucopolysaccharidoses.

Two methods for analysis of urinary glycosaminoglycans (GAGs) have been modified and improved for efficient screening of mucopolysaccharidoses. Urinary excretion of GAGs was estimated by spectrophometric measurement of alcian blue complex formation and was used in conjunction with qualitative analysis by thin layer chromatography. After normal variation in GAG excretion was established using 120 urine samples, these screening methods were applied to a total of 2057 urine samples over 4 years. Six patients with abnormal urinary GAG excretion had a mean of 50.5 +/- 20.6 mg GAG/mmol creatinine compared to 3.4 +/- 2.9 for age-matched controls. Qualitative analysis by thin layer chromatography using alternating solvent systems identified the GAGs excreted in excess and facilitated selection of specific enzyme assays for final confirmation. Six cases were diagnosed prospectively and demonstrate these quantitative and qualitative methods to be economical, efficient, and suitable for clinical use.

Basal platelet intracellular free calcium concentrations in untreated hypertensive patients and normotensive controls.

Basal platelet intracellular free calcium concentrations were measured in 18 untreated essential hypertensive patients and 18 age and sex matched normotensive controls. No significant difference was observed between the mean platelet intracellular free calcium concentrations for these two groups. There was a weak, but significant positive correlation between both systolic and diastolic BP and basal platelet intracellular free calcium concentration. These results suggest there is a real, but quantitatively small relationship between platelet intracellular free calcium levels and BP in essential hypertension which may be of pathophysiological significance in the genesis and maintenance of hypertension.

Skeletal muscle beta 2-adrenoreceptors and the effect of adrenergic drugs on plasma potassium in perinephritis hypertension in rabbits.

1. It has been suggested that beta 2-adrenoreceptors in skeletal muscle regulate plasma potassium. The possibility that alterations in the function and/or density of these receptors occurs in perinephritis hypertension in rabbits was studied. 2. Intravenous infusion of adrenaline (0.2 micrograms kg-1 min-1) caused a fall in potassium while intravenous bolus injection of propranolol (0.75 mg kg-1) resulted in an increase in serum potassium which was of similar magnitude in both perinephritis hypertensive and sham-operated normotensive rabbits. 3. Binding studies with the radioligand [125I] cyanopindolol (ICYP) showed that there were no significant differences between the hypertensive and normotensive rabbits in the density (Bmax) or affinity (KD) of the skeletal muscle beta 2-adrenoreceptor. 4. The results suggest that function and density of skeletal muscle beta 2-adrenoreceptors are not altered in rabbits with perinephritis hypertension.

Platelet cytosolic free calcium before and after antihypertensive treatment in perinephritis hypertension of the rabbit.

Cytosolic free calcium concentration ([Ca2+]i) in platelets has been reported to be elevated in human essential hypertension, to be positively correlated with blood pressure and to decrease with blood pressure reduction. However, some groups have been unable to confirm these findings in either humans or hypertensive rats. We have examined the relationship between platelet [Ca2+]i and blood pressure in the perinephritis model of hypertension in the rabbit. In addition, the effects of both acute and chronic treatment with verapamil or prazosin were studied. Mean arterial pressure, heart rate and platelet [Ca2+]i were measured before and after treatment. Platelet [Ca2+]i was measured by the Quin 2 fluorescence technique. Platelet [Ca2+]i was similar for the normotensive and hypertensive rabbits, and no correlation between platelet [Ca2+]i and blood pressure was observed. None of the antihypertensive treatments produced a lowering of platelet [Ca2+]i. Therefore we conclude that platelet [Ca2+]i is unlikely to be a universally useful index of ([Ca2+]i in vascular smooth muscle of resistance vessels.

Platelet intracellular free calcium concentration in normotensive and hypertensive pregnancies in the human.

1. Basal and stimulated platelet intracellular free calcium concentrations were measured in non-pregnant women and in third trimester patients who were either normotensive or who had pregnancy-induced hypertension or pre-eclampsia. There were 15 subjects in each group. 2. A trend for a reduction of the maximal response of platelet calcium levels to stimulation by 5-hydroxytryptamine was seen in pregnant groups compared with nonpregnant subjects, but this was significant only in pre-eclampsia. 3. No significant differences in basal or adenosine 5'-pyrophosphate-stimulated levels of platelet intracellular free calcium concentration were observed between the four groups. 4. These results illustrate that basal platelet calcium levels are unchanged in hypertension of pregnancy. Alterations in basal platelet calcium levels may not be involved in the platelet activation that is a feature of pre-eclampsia.

Exercise and platelet intracellular free calcium concentration.

1. A positive correlation between blood pressure and platelet intracellular free calcium concentration ([Ca2+]i) has been reported. We examined the effect of acute changes in blood pressure associated with exercise on platelet [Ca2+]i. 2. Twenty-one subjects had blood pressure and heart rate readings taken after 60 min rest, and subjects were then exercised on a bicycle ergometer at 120 W for 30 min. Blood pressure and heart rate readings were repeated immediately after exercise, 30 min after exercise, and then after a further hour. Blood samples were taken after each set of blood pressure and heart rate readings for catecholamine, lactate and platelet [Ca2+]i estimations. 3. There were significant increases in systolic and diastolic blood pressure, heart rate, and plasma lactate and catecholamine levels during the course of the study. There were no significant changes in platelet [Ca2+]i. 4. These results suggest that the acute blood pressure changes related to exercise are not associated with a change in platelet [Ca2+]i.

Ethanol and platelet intracellular free calcium concentrations.

The effect of 0.8 g kg-1 absolute ethanol orally on platelet intracellular free calcium was assessed in a random order study in 24 normotensive subjects with an isocaloric control. Platelet calcium was measured 90 min and 12 h after treatment by the Quin 2 method. The study had 90% power of detecting a 16.5% change. After 90 min, breath ethanol was 37 +/- 9 micrograms 100 ml-1, blood pressure was unchanged and heart rate rose slightly. Platelet calcium was unchanged by ethanol after 90 min or 12 h.

Long-term amitriptyline treatment alters the affinity state of alpha 2 adrenoceptors in rabbit hindbrain.

The effects of 7 and 21 days amitriptyline treatment on brain stem alpha 2 adrenoceptor number and affinity state and on the in vivo depressor response to intracisternal clonidine were examined in male New Zealand white rabbits.

Cell-free assembly of a polyoma-like particle from empty capsids and DNA.

A polyoma-like particle (PLP) is formed when polyoma DNA and purified empty capsids are incubated in a cell-free system. The DNA of this new particle is protected against the action of pancreatic DNase. The density of the purified PLP in CsCl is 1.32 g/cm3, which is intermediate between that of polyoma virions (1.34 g/cm3) and empty capsids (1.29 g/cm3). Purified PLP sediments at 190 S in sucrose and is stable in solutions of high ionic strength. When the DNA is extracted from PLP by the use of detergent and phenol, it is found to be doublestranded with a molecular weight of approximately 1.1 x 10(6). The particles are stable in CsCl at 4 degrees for at least 5 months. Electron micrographs indicate that highly purified PLPs stained with 2% PTA have the same appearance as polyoma capsids. Neither aggregates nor complexes bound by loose ionic bonds appear reasonable to explain these results. The evidence indicates that the DNA of this new polyoma-like particle, made under cell-free conditions, is protected by the capsid.