The CRY1 blue light photoreceptor of Arabidopsis interacts with phytochrome A in vitro

Plants have at least two major photosensory receptors: phytochrome (absorbing primarily red/far-red light) and cryptochrome (absorbing blue/UV-A light); considerable physiological and genetic evidence suggests some form of communication or functional dependence between the receptors. Here, we demonstrate in vitro, using purified recombinant photoreceptors, that Arabidopsis CRY1 and CRY2 (cryptochrome) are substrates for phosphorylation by a phytochrome A-associated kinase activity. Several mutations within the CRY1 C terminus lead to reduced phosphorylation by phytochrome preparations in vitro. Yeast two-hybrid interaction studies using expressed C-terminal fragments of CRY1 and phytochrome A from Arabidopsis confirm a direct physical interaction between both photoreceptors. In vivo labeling studies and specific mutant alleles of CRY1, which interfere with the function of phytochrome, suggest the possible relevance of these findings in vivo.     

Familial neuroblastoma: report of a kindred with later age at diagnosis

The first reported effective adjuvant combination regimen for patients with operable breast cancer comprised oral cyclophosphamide (C) days 1-14 with intravenous methotrexate (M) and fluorouracil (F) on days 1 and 8, repeated every 28 days ('classical' CMF). These drugs have since been extensively used with or without endocrine therapies and/or other cytotoxics, as well as with radiation therapy to the chest wall yielding conflicting results. Although doses and schedules have varied widely, the combination of these three drugs has been generically referred to as CMF. Evidence exists that reducing the dose and/or altering the schedule of CMF ('modified' CMF) have compromised its efficacy in metastatic breast cancer. Reduction below standard dose of a similar regimen also gave inferior results in the adjuvant setting. In fact, the recently reported improved outcome of adding radiation therapy to CMF was only demonstrated in comparisons with a 'modified' CMF. Furthermore, trials in women with estrogen receptor-positive breast cancer, which did not demonstrate any significant benefit for the addition of adjuvant CMF to tamoxifen compared with tamoxifen alone, also used 'modified' CMF. Therefore, adherence to the 'classical' dose and schedule is recommended when CMF is used in adjuvant therapy.     

State-dependent NMDA receptor antagonism by Ro 8-4304, a novel NR2B selective, non-competitive, voltage-independent antagonist

1. Subunit-selective blockade of N-methyl-D-aspartate (NMDA) receptors provides a potentially attractive strategy for neuroprotection in the absence of undesirable side effects. Here, we describe a novel NR2B-selective NMDA antagonist, 4-?3-[4-(4-fluoro-phenyl)-3,6-dihydro-2H-pyridin-1-yl]-2-hydroxy-propoxy ?-benzamide (Ro 8-4304), which exhibits >100 fold higher affinity for recombinant NR1(001)/NR2B than NR1(001)/NR2A receptors. 2. Ro 8-4304 is a voltage-independent, non-competitive antagonist of NMDA receptors in rat cultured cortical neurones and exhibits a state-dependent mode of action similar to that described for ifenprodil. 3. The apparent affinity of Ro 8-4304 for the NMDA receptor increased in an NMDA concentration-dependent manner so that Ro 8-4304 inhibited 10 and 100 microM NMDA responses with IC50s of 2.3 and 0.36 microM, respectively. Currents elicited by 1 microM NMDA were slightly potentiated in the presence of 10 microM Ro 8-4304, and Ro 8-4304 binding slowed the rate of glutamate dissociation from NMDA receptors. 4. These results were predicted by a reaction scheme in which Ro 8-4304 exhibits a 14 and 23 fold higher affinity for the activated and desensitized states of the NMDA receptor, respectively, relative to the agonist-unbound resting state. Additionally, Ro 8-4304 binding resulted in a 3 4 fold increase in receptor affinity for glutamate site agonists. 5. Surprisingly, whilst exhibiting a similar affinity for NR2B-containing NMDA receptors as ifenprodil, Ro 8-4304 exhibited markedly faster kinetics of binding and unbinding to the NMDA receptor. This spectrum of kinetic behaviour reveals a further important feature of this emerging class of NR2B-selective compounds.     

Tuberculosis 2000: problems and solutions

The incidence of tuberculosis is expected to increase, from 8.8 million cases in 1995, to 10.2 million cases by the year 2000 and 11.9 million by 2005. Three million deaths due to tuberculosis occurred in 1995, and 3.5 million can be expected in the year 2000. The most important causes of the world-wide increase in tuberculosis are: 1) non-compliance with control programmes; 2) inadequate diagnosis and treatment; 3) migration; 4) endemic human immunodeficiency virus (HIV); 5) ambulatory and self-administered treatment. In the 1970s it was stated that treatment needed to be supervised-a recommendation that went unheeded. A number of fundamental changes should be introduced in order to make treatment effective, to cure patients and thus to arrest the transmission of the disease: 1) supervision during the whole period antituberculosis drugs are taken, and 2) hospitalization during the initial treatment stage for all groups at risk. It is already 50 years since the first antituberculosis drugs were discovered; effective treatments capable of curing all patients in 6 months have been available for the last 25 years, and the result is failure plus a growing mortality curve at the beginning of the twenty-first century. If we wish to alter this trend, we need trained doctors all over the world who possess enough clinical knowledge of tuberculosis; hospitalization for specific groups of tuberculosis patients; true supervision during the whole treatment period; fixed-dose combinations of drugs; and prophylaxis or preventive treatment whenever possible. We also need to take into account other factors such as drug resistance, endemic HIV, and migration.     

Negative regulation of Wee1 expression and Cdc2 phosphorylation during p53-mediated growth arrest and apoptosis

The G2 cell cycle checkpoint protects cells from potentially lethal mitotic entry after DNA damage. This checkpoint involves inhibitory phosphorylation of Cdc2 at the tyrosine-15 (Y15) position, mediated in part by the Wee1 protein kinase. Recent evidence suggests that p53 may accelerate mitotic entry after DNA damage and that the override of the G2 checkpoint may play a role in the induction of apoptosis by p53. To determine the biochemical mechanism by which p53 inactivates the G2 checkpoint, the effects of p53 activation on Wee1 expression, Cdc2-Y15 phosphorylation, and cyclin B1-associated Cdc2 kinase activity were examined. Under conditions of either growth arrest or apoptosis, p53 activation resulted in the down-regulation of Wee1 expression and dephosphorylation of Cdc2. A parallel increase in cyclin B1/Cdc2 kinase activity was observed during p53-mediated apoptosis. Negative regulation of the Wee1 expression and Cdc2 phosphorylation by p53 was also evident in thymus tissue from p53+/+ mice but not from p53-/- mice. Inactivation of the G2 checkpoint may contribute to the tumor suppressor activity of p53.     

Magnetic resonance imaging of the pelvic floor in the postpartum patient

Magnetic resonance imaging (MRI) was used to assess anatomical changes in the pelvic floor after childbirth. Six women underwent serial MRI examination within 30 hours and at 1 week, 2 weeks, 6 weeks and 6 months after delivery; 8 additional women were studied only within 30 hours of delivery. T-1 and T-2-weighted images of the pelvis in the transverse and sagittal planes with a 1.5-T MR imager were obtained. In the sagittal section we assessed the urethrovesical angle, urethral length, distance from the symphysis to the proximal and distal vagina, vaginal length, width and length of the sphincters, and the presence of sphincter defects. Axial sections were assessed for sphincter defects for the distance between the symphysis and midurethra, vagina and rectum. Only one parameter (distance between symphysis and distal vagina) changed significantly over time, without a clear trend in direction. Interobserver variation was reasonable (<15%) except for anal canal length, urethral length and distance between symphysis and anus. There were no significant correlations between birthweight and MRI parameters. There was a non-significant association (P = 0.09) between the sole combined sphincter defect and rectal injury, but not with episiotomy or parity. We concluded that it is feasible to determine multiple measurements on MR images to evaluate structures of the pelvic floor.     

Chronic administration of ORG 2766 for 6 months produces a subtle impairment in strategic learning by rats

Dysfunctions in serotonergic pathways may underlie several psychiatric disorders. The reuptake of serotonin (5-HT) from synaptic terminals is mediated by a specific transporter (5-HTT). Genetic variation in the gene coding for the 5-HTT protein might be involved in the predisposition to psychiatric disorders. A systematic screening of the whole coding sequence of the 5-HTT gene in mood disorder (MD) and obsessive-compulsive disorder (OCD) patients, as well as in healthy controls, using PCR and denaturing gradient gel electrophoresis (DGGE) revealed the presence of two mutations. The first was in intron 4, and the second was a C-->A transversion leading to an amino-acid exchange (Leu-->Met) in position 255 of the deduced protein sequence. No further occurrence of this substitution was found in an extended sample of patients and controls. Therefore, structural modifications of the 5-HTT gene do not seem to play either a major or minor role in the genetic predisposition to MD or OCD.     

Comparison of different oxygen exchange models

A functional distribution of coronary volume can be estimated from the response of arterio-venous O2 content difference (AVO2) to a flow step. However, the results depend on the assumed O2 exchange model. The previously used model consisted of a single mixed compartment with O2 exchange (reference model). The purpose of this study is to provide an estimate of the errors made in the volume estimations by not taking into account factors as flow heterogeneity, different mixing sites or Krogh-like O2 exchange. The approach is indirect: the response of the AVO2 to a flow step has been calculated with alternative O2 exchange models in which factors mentioned are incorporated. These transients are fitted with the reference model. The resulting estimated volumes are different from the volumes assumed in the alternative models. Large differences are obtained with some of the alternative models, e.g. the model with Krogh characteristics. However, these models seem unrealistic because capillary pO2 is higher than venous pO2. Only small differences in volume are obtained with the more realistic models. Therefore, these results indicate that the coronary volumes are approximated well by the estimations obtained with the reference model. These volume estimations were 9.9 and 3.8 ml 100 g-1 for the O2 exchange vessels and the distal venous volume, respectively.     

Effects of abstinence and family history for alcoholism on platelet adenylyl cyclase activity

Platelet adenylyl cyclase (AC) activity was measured in 32 alcohol-dependent subjects and 27 control subjects who were categorized as either family history-positive (FHP) or family history-negative (FHN) for alcoholism. The interview and blood sample collections were performed shortly after cessation of heavy drinking in the alcoholic group, and repeat blood samples were obtained at the end of the first and second weeks of monitored abstinence. Control subjects received the same interview and provided blood samples at the time of the interview. When subjects were not segregated for FHP or FHN status, there were no statistically significant differences in basal, cesium fluoride (CsF)-, or forskolin-stimulated mean AC activities between the controls and the alcoholics, at study entry or with 1 or 2 weeks of abstinence. On the other hand, over the 2-week course of sobriety from heavy drinking, the CsF-stimulated AC activity of FHP alcohol-dependent subjects decreased significantly (p = 0.03). FHP alcohol-dependent subjects after 2 weeks of sobriety had significantly lower mean CsF-stimulated AC activity than FHN controls (p = 0.04), whereas the FHN alcoholic subjects' CsF-stimulated AC activity did not differ significantly from FHN controls at this point in time. When all subjects were pooled and then categorized as either FHP or FHN, there was a significant difference in mean CsF-stimulated AC activity (p = 0.02) between the FHP and FHN subject groups. Genetic factors and abstinence appear to have roles in determining low platelet AC activity in alcoholic and nonalcoholic subjects. CsF-stimulated platelet AC activity, in particular, appears to act as a trait marker for a genetic vulnerability to developing alcoholism, but recent heavy drinking in male alcoholics is a factor that can mask differences between FHP and FHN subjects.