Partitioning the Apparent Temperature Sensitivity into Within- and Across-Taxa Responses: Revisiting the Difference between Autotrophic and Heterotrophic Protists.

AbstractConventional analyses suggest that the metabolism of heterotrophs is thermally more sensitive than that of autotrophs, implying that warming leads to pronounced trophodynamic imbalances. However, these analyses inappropriately combine within- and across-taxa trends. Our new analysis separates these, revealing that 92% of the difference in the apparent thermal sensitivity between autotrophic and heterotrophic protists does indeed arise from within-taxa responses. Fitness differences among taxa adapted to different temperature regimes only partially compensate for the positive biochemical relationship between temperature and growth rate within taxa, supporting the hotter-is-partially-better hypothesis. Our work highlights the importance of separating within- and across-taxa responses when comparing temperature sensitivities between groups, which is relevant to how trophic imbalances and carbon fluxes respond to warming.

Genetic and epigenetic regulation of the organic cation transporter 3, SLC22A3.

Human organic cation transporter 3 (OCT3 and SLC22A3) mediates the uptake of many important endogenous amines and basic drugs in a variety of tissues. OCT3 is identified as one of the important risk loci for prostate cancer, and is markedly underexpressed in aggressive prostate cancers. The goal of this study was to identify genetic and epigenetic factors in the promoter region that influence the expression level of OCT3. Haplotypes that contained the common variants, g.-81G>delGA (rs60515630) (minor allele frequency 11.5% in African American) and g.-2G>A (rs555754) (minor allele frequency>30% in all ethnic groups) showed significant increases in luciferase reporter activities and exhibited stronger transcription factor-binding affinity than the haplotypes that contained the major alleles. Consistent with the reporter assays, OCT3 messenger RNA expression levels were significantly higher in Asian (P<0.001) and Caucasian (P<0.05) liver samples from individuals who were homozygous for g.-2A/A in comparison with those homozygous for the g.-2G/G allele. Studies revealed that the methylation level in the basal promoter region of OCT3 was associated with OCT3 expression level and tumorigenesis capability in various prostate cancer cell lines. The methylation level of the OCT3 promoter was higher in 62% of prostate tumor samples compared with matched normal samples. Our studies demonstrate that genetic polymorphisms in the proximal promoter region of OCT3 alter the transcription rate of the gene and may be associated with altered expression levels of OCT3 in human liver. Aberrant methylation contributes to the reduced expression of OCT3 in prostate cancer.

Cultivation and serological characterization of a human Theiler's-like cardiovirus associated with diarrheal disease.

Cardioviruses (e.g., Theiler's murine encephalomyelitis virus [TMEV]) are members of the Picornaviridae family that cause myocarditis and encephalitis in rodents. Recently, several studies have identified human cardioviruses, including Saffold virus (SAFV) and a related virus named human TMEV-like cardiovirus (HTCV). At least eight cardiovirus genotypes are now recognized, with SAFV and most strains of HTCV belonging to genotypes 1 and 2, respectively; genotype 2 strains are the most common in the population. Although a genotype 3 cardiovirus has recently been cultured (SAFV-3), the genotype 1 and 2 cardioviruses have been difficult to propagate in vitro, hindering efforts to understand their seroprevalence and pathogenicity. Here we present the isolation and characterization of a genotype 2 human cardiovirus (HTCV-UC6). Notably, successful cultivation of HTCV-UC6 from stool required the addition of cytokine-blocking antibodies to interrupt downstream antiviral pathways. Unlike SAFV-3, HTCV-UC6 exhibited slow replication kinetics and demonstrated only a moderate cytopathic effect. Serologic assays revealed that 91% of U.S. adults carry antibodies to the genotype 2 cardioviruses, of which 80% generate neutralizing antibodies, in agreement with previous data showing that cardiovirus infection is widespread in humans. We also demonstrate an acute cardiovirus seroconversion event in a child with diarrhea and vomiting, thus reporting for the first time evidence linking cardiovirus infection to diarrheal disease in humans.

A new method for seed oil body purification and examination of oil body integrity following germination.

Plant seeds store triacylglycerols as energy sources for germination and postgerminative growth of seedlings. The triacylglycerols are preserved in small, discrete, intracellular organelles called oil bodies. A new method was developed to purify seed oil bodies. The method included extraction, flotation by centrifugation, detergent washing, ionic elution, treatment with a chaotropic agent, and integrity testing by use of hexane. These processes subsequently removed non-specifically associated or trapped proteins within the oil bodies. Oil bodies purified by this method maintained their integrity and displayed electrostatic repulsion and steric hindrance on their surface. Compared with the previous procedure, this method allowed higher purification of oil bodies, as demonstrated by SDS-PAGE using five species of oilseeds. Oil bodies purified from sesame were further analyzed by two-dimensional gel electrophoresis and revealed two potential oleosin isoforms. The integrity of oil bodies in germinating sesame seedlings was examined by hexane extraction. Our results indicated that consumption of triacylglycerols reduced gradually the total amount of oil bodies in seedlings, whereas no alteration was observed in the integrity of remaining oil bodies. This observation implies that oil bodies in germinating seeds are not degraded simultaneously. It is suggested that glyoxisomes, with the assistance of mitochondria, fuse and digest oil bodies one at a time, while the remaining oil bodies are preserved intact during the whole period of germination.

Thrombin-stimulated increases in cytosolic Ca2+ level and gonadotropin-releasing hormone release in GT1-7 neurons.

The effects of thrombin on cytosolic calcium levels ([Ca2+]cyt), and on gonadotropin-releasing hormone (GnRH) release, were characterized in cultured GT1-7 neurons. GnRH release from GT1-7 neurons was pulsatile with an average pulse amplitude of 14.3+/-5.8 pg x min x ml(-1) and an average pulse duration of 21.3+/-4.2 min. The [Ca2+]cyt response to 0.005 to 0.2 U/ml thrombin was saturable and concentration dependent (EC50 = 0.0268 U/ml). Ethyleneglycotetraacetic acid (EGTA) chelation of extracellular Ca2+ resulted in an approximately 70% attenuation of thrombin-stimulated increase in [Ca2+]cyt. By use of a special superfusion system, a 5-min exposure to 0.1 U/ml thrombin significantly increased the amplitude (193.2+/-67.8 pg x min x ml(-1); P = 0.001) but not the duration (22.5+/-2.4 min; P = 0.8) of GnRH release. These results suggest that thrombin increases [Ca2+]cyt and GnRH release from GT1-7 neurons via specific membrane-bound receptors.

Exercise and reproductive dysfunction.

OBJECTIVE: To provide an overview of our current understanding of exercise-induced reproductive dysfunction and an approach to its evaluation and management. DESIGN: A MEDLINE search was performed to review all articles with title words related to menstrual dysfunction, amenorrhea, oligomenorrhea, exercise, and athletic activities from 1966 to 1998. The pathophysiology, proposed mechanisms, clinical manifestations, evaluation, and management of exercise-associated reproductive dysfunction were compiled. CONCLUSION(S): Exercise-induced menstrual irregularity appears to be multifactorial in origin and remains a diagnosis of exclusion. The underlying mechanisms are mainly speculative. Clinical manifestations range from luteal phase deficiency to anovulation, amenorrhea, and even delayed menarche. Evaluation should include a thorough history and a complete physical plus pelvic examination. Most cases are reversible with dietary and exercise modifications. Hormonal replacement in cases of a prolonged hypoestrogenic state with evidence of increased bone loss is recommended, although the long-term consequences of prolonged hormonal deficiency are ill-defined.

Cocaine impairs follicular phase pulsatile gonadotropin secretion in rhesus monkeys.

OBJECTIVE: To assess cocaine's effect on follicular phase pulsatile gonadotropin secretion in normally cycling rhesus monkeys. METHODS: Sixteen monkeys were paired by body weight and randomized to receive intravenous saline (n = 8) or cocaine (4 mg/kg, n = 8) daily on cycle days 2 to 14. Monkeys were chronically cannulated to allow frequent blood collections without anesthesia. Blood samples were obtained every 15 minutes for 8 hours in early (EFP; cycle days 1 to 5), mid-(MFP; cycle days 6 to 10), and late (LFP; cycle days 11 to 15) follicular phase. Plasma concentrations of LH, FSH, and estradiol-17 beta (E2) were determined by radioimmunoassay. Pulses were identified by cluster analysis. Statistical differences were determined by analysis of variance (ANOVA) and Sidak's multiple comparison test. RESULTS: Seven out of eight monkeys in the control group demonstrated timely ovulation. Only one monkey in the cocaine-treated group ovulated. Similar gonadotropin pulse intervals (70 to 90 minutes) were observed throughout the follicular phase in both the controls and cocaine-treated monkeys. LH and FSH pulse amplitudes increased significantly from the EFP/MFP to the LFP in controls. In cocaine-treated monkeys, gonadotropin pulse amplitudes remained at EFP/MFP levels throughout the study period. The mean gonadotropin pulse amplitude and the mean E2 levels in the LFP were significantly greater in controls as compared with cocaine-treated monkeys (P < .001). CONCLUSION: These findings demonstrate that cocaine suppresses the normal increase in LH and FSH pulse amplitude seen in the LFP. Further studies are in progress to determine the mechanism of cocaine's disruption of the hypothalamic-pituitary-ovarian axis.

Molecular electron affinities and the calculation of the temperature dependence of the electron-capture detector response.

The use of the electron-capture detector (ECD) to measure molecular electron affinities and kinetic parameters for reactions of thermal electrons is reviewed. The advances of the past decade are emphasized and include the multistate electron-capture detector model and the use of semi-empirical self-consistent field quantum mechanical calculations and half wave reduction potential values to support gas phase experimental results. A procedure for the evaluation of the adiabatic electron affinities of the main group elements and the homonuclear diatomic molecules is presented. Potential excited states are identified for the magnetron (MGN) values for quinones, thermal charge transfer (TCT) values for CS2, C6F6, SF6 and photoelectron spectroscopy (PES) values for O2, NO, nitromethane, and the nucleic acids. Literature electron affinities are then evaluated. The temperature dependence of the electron-capture detector can be calculated using values for kinetic rate constants and electron affinities to optimize response and temperature sensitivity in analytical procedures. The temperature dependence for adenine, guanine, thymine and cytosine are predicted for reactions with thermal electrons. Using the recent advances, the new adiabatic electron affinities are: all in electron volts (eV), 4-F-benzaldehyde (0.57 +/- 0.05) and acetophenones (APs) 4-F-AP (0.52 +/- 0.05); 2-CF3-AP (0.79 +/- 0.05); 3-CF3-AP (0.79 +/- 0.05); 4-CF3-AP (0.89 +/- 0.05); 3-CI-AP (0.67 +/- 0.05); and 4-Cl-AP (0.64 +/- 0.05). The adiabatic electron affinities of chloro and fluorobenzenes range from 0.17 to 1.15 eV and 0.13 to 0.86 eV.

Molecular cloning of 11S globulin and 2S albumin, the two major seed storage proteins in sesame.

Insoluble 11S globulin and soluble 2S albumin, conventionally termed alpha-globulin and beta-globulin, are the two major storage proteins and constitute 80-90% of total seed proteins in sesame. Two full-length cDNA clones were sequenced and deduced to encode sesame 11S globulin and 2S albumin precursors, respectively. Deduced amino acid composition reveals that 2S albumin, but not 11S globulin, is a sulfur-rich protein. Three abundant polypeptides of 50-60 kDa were resolved on SDS-PAGE when seed-purified 11S globulin was prepared in nonreducing conditions. Immunological analysis suggests that these three polypeptides are encoded by homologous genes. Immunodetection on the overexpressed protein of the 11S globulin clone in Escherichia coli indicates that this clone encodes the precursor protein of one of the three purified 11S globulin polypeptides.

Insulin lispro: in-vivo potency determination by intravenous administration in conscious rabbits.

Insulin lispro is a monomeric analogue of human insulin, produced by genetic engineering, and has been reported to have a more rapid absorption following subcutaneous injection than insulin. Since it has been shown to have a similar hypoglycaemic action to insulin in clinical studies and comparable properties in radioimmunoassay, the feasibility of using a bioassay which was designed originally for insulin, to measure insulin lispro potency was evaluated in this investigation. A random-dose bioassay protocol, in which insulin lispro and two insulin standards were administered intravenously in a random sequence, was used and validated in nine conscious healthy rabbits. The decline in blood-glucose levels, following the intravenous injection of a dose of insulin or its lispro analogue, was monitored by a continuous glucose monitoring system. A glucose response curve was generated, from which various pharmacodynamic parameters were determined. Compared with the insulin standards, the potencies of insulin lispro determined from nadir, basal glucose normalized nadir, glycaemic reduction and ABGC (area of the blood-glucose response curve under baseline) were observed to have mean (95% confidence limits) values of 97.0 (69.5-124.6)%, 106.3 (72.4-140.2)%, 949 (51.8-138.0)% and 102.4 (76.3-128.5)%, respectively. In addition, the coefficients of variation for correspondent parameters were 36.9, 41.5, 59.1 and 33.2%, respectively. The results indicated that the hypoglycaemic potency calculated from the ABGC values was the most accurate (102.4%) with the least coefficient of variation (33.2%). In conclusion, the potency of insulin lispro can be determined accurately from the ABGC values measured by the random-dose bioassay used.

Profiling solute carrier transporters in the human blood-brain barrier.

The neuroprotective function of the blood-brain barrier (BBB) presents a major challenge for drug delivery to the central nervous system (CNS). Critical to this function, BBB membrane transporters include the ATP-binding cassette (ABC) transporters, which limit drug penetration across the BBB, and the less-well-studied solute carrier (SLC) transporters. In this work, expression profiling of 359 SLC transporters, comparative expression analysis with kidney and liver, and immunoassays in brain microvessels (BMVs) identified previously unknown transporters at the human BBB.

Phase I/II study of hypofractioned radiation with three-dimensional conformal radiotherapy for clinical T3-4N0-1M0 stage esophageal carcinoma.

Compared to conventional fractionated-dose radiotherapy, high hypofractionated-dose radiotherapy could yield tumoricidal effects. However, few clinical trials of hypofractionated radiotherapy in loco-regionally advanced incurable esophageal cancer at present have yet been performed. The purpose of the current study was to evaluate the efficacy and toxicity of hypofractioned radiation with three-dimensional conformal radiotherapy for clinical T3-4N0-1M0 stage esophageal carcinoma. From September 2003 to December 2005, 45 patients with locally advanced esophageal carcinoma were grouped and received three-dimensional conformal hypofractioned radiotherapy (3D-CRT) whose fractionated dose was gradually increase per group. Radiotherapy was administered to a total dose of from 50 to 54 Gy (fractionated dose of from 3.0 to 6.0 Gy, 3 times weekly), over a 3-4 week period. And patients received 4 cycles chemotherapy. The median follow-up period for survivors was 38 months. Treatment tolerance rate was 78.8% with daily dose of from 3 to 5 Gy. There are 21.2% patients occurring Grade ≥ 3 acute toxicities. But patients couldn't tolerate daily dose of 6 Gy (55.6%). The 1-year, 2-year and 3-year local control rates were 62%, 49% and 39% respectively. And the 1-year, 2-year and 3-year overall survival rates were 34%, 21% and 9% respectively. The median overall survival time was 17 months. At the time of following up, 13 patients (31.0%) had occurred esophageal late complications, with mainly esophageal perforation, hemorrhage or stenosis, including initial stenosis aggravation. Therefore hypofractionated irradiation was thought to be feasible for clinical T3-4N0-1M0 stage esophageal carcinoma. And daily dose of ≤5 Gy was comparatively suitable in hypofractionated irradiation for esophageal carcinoma, and the patients tolerated well. But further research was in need also.

The role of family planning communications--an agent of reinforcement or change.

PIP: Results are presented of a multiple classification analysis of responses to a 1972 KAP survey in Taiwan of 2013 married women aged 18-34 designed to determine whether family planning communication is primarily a reinforcement agent or a change agent. 2 types of independent variables, social demographic variables including age, number of children, residence, education, employment status, and duration of marriage; and social climate variables including ever receiving family planning information from mass media and ever discussing family planning with others, were used. KAP levels, the dependent variables, were measured by 2 variables each: awareness of effective methods and awareness of government supply of contraceptives for knowledge, wish for additional children and approve of 2-child family for attitude, and never use contraception and neither want children nor use contraception for practice. Social demographic and attitudinal variables were found to be the critical ones, while social climate and knowledge variables had only negligible effects on various stages of family planning adoption, indicating that family planning communications functioned primarily as a reinforcement agent. The effects of social demographic variables were prominent in all stages of contraceptive adoption. Examination of effects of individual variables on various stages of family planning adoption still supported the argument that family planning communications played a reinforcement role. Family planning communications functioned well in diffusing family planning knowledge and accessibility, but social demographic variables and desire for additional children were the most decisive influences on use of contraception.^ieng

The experimental hardness and electronegativity of the purines and pyrimidines in DNA and RNA supported by the AM1 calculation of the electron affinities and ionization potentials.

The ionization potential and electron affinities of the purines and pyrimidines in DNA and RNA were calculated with the AM1 semiempirical method. The values support the experimental values. The electron affinities are significant, and positive, such that donor-acceptor interactions can, and indeed should play a role in the stacking of bases in nucleic acids. Based on the confirmation of the experimental values by the theoretical calculations, reliable values of the experimental hardness and electronegativity were calculated.

Accuracy of mammographic appearances after breast fine-needle aspiration.

OBJECTIVE: The objective of this study was to document the observation that fine-needle aspiration of palpable breast masses by use of a modified technique performed shortly before mammography need not adversely interfere with mammographic interpretation nor produce falsely suspicious mammographic lesions that delay meaningful evaluation and management in this breast clinic. STUDY DESIGN: In a retrospective record review 1007 women who were seen in the Breast Diagnostic Center at Women's and Children's Hospital from January 1992 until April 1995 and who had fine-needle aspiration of a palpable solid breast mass within 2 weeks before mammography were analyzed overall and in 10-year age group subsets. The mammographic reports of "suspicious" lesions were correlated with having had a prior fine-needle aspiration (within 2 weeks). RESULTS: Of the 1007 women undergoing fine-needle aspirations, 91 had a cytologic or tissue biopsy specimen diagnosis of malignancy. Of these, 72 had "suspicious" mammograms and 19 had "nonsuspicious" mammograms. The calculated positive predictive value was 58%. The negative predictive value was 98%. Mammographic sensitivity was 79%. Specificity was 94%. Age stratification did not reveal any meaningful trends. Of the 916 patients with benign cytologic results of fine-needle aspiration specimens, 52 had "suspicious" mammograms and 864 had "nonsuspicious" mammograms. CONCLUSION: For patient convenience and expeditious diagnosis of a palpable breast mass, fine-needle aspiration can be performed on the initial visit and mammograms subsequently taken within 2 weeks without undue clinical confusion or misleading mammographic findings. Concordance of the diagnostic triad consisting of (1) clinical impression (by history and examination), (2) fine-needle aspiration, and (3) mammography gives a reliable conclusion and can appropriately be used as the basis for clinical management of a breast mass. However, when there is doubt or anxiety about the diagnosis either on the part of the patient or the physician, a definitive histologic tissue diagnosis should obtained.

The negative ion states of molecules: adenine and guanine.

Anions of cytosine and thymine predominate in radiation-damaged DNA. This is in contrast to the experimental order of adiabatic electron affinities: A, 0.95; G, 1.51; >T, 0.79, U, 0.80; C, 0.56 (+/-0.05 eV). Excited negative-ion states of adenine (A) and guanine (G) are identified using semiempirical AM1-MCCI quantum mechanical calculations. A planar G(-) has an excited state adiabatic electron affinity, AEA*, of 0.3 +/- 0.05 eV. This state and the unique Watson-Crick structure are responsible for the preponderance of charge on C(-) in radiation-damaged DNA. By analogy to the value for cytosine, the dipole-bound EA of G is estimated as 0.25 eV. New AEA values from literature reduction potentials for the ribose nucleotides are rC, 0.6; rU, 0.8; and rT, 0.8 (+/-0.1 eV). From literature photoelectron spectroscopy, AEA* vales for U are 0.15, 0.3, 0.5, and 0.6 eV. In GC(-2), stacked [GC:GC](-3), and [GC:GC:GC](-4), the charge moves to G. In [GC:GC:GC](-2 to -4), the charge moves from GC(1) to GC(3) through space without a bridge or bond. This is important to electron conduction, radiation damage and repair, and nanoscale devices.

The electron affinities of the radicals formed by the loss of an aromatic hydrogen atom from adenine, guanine, cytosine, uracil, and thymine.

The major ion formed in Negative Ion Chemical Ionization Mass Spectrometry of Adenine, Guanine, Cytosine, Uracil and Thymine is the dehydrogenated anion. The CURES EC procedure for optimizing Austin Model-1 Multiconfigurational Configuration Interaction semi-empirical calculations is applied to the electron affinities of the corresponding dehydrogenated bases and N-H bond dissociation energies. These calculated values will be compared with literature values of the gas phase acidities of the purines and pyrimidines. The N-H bond dissociation energies are about 3.95 eV for Guanine, Adenine, and Thymine and 4.08 eV for Cytosine and Uracil. The electron affinities of the radicals are AMinH = 3.50 eV, GMinH = 3.46 eV, CMinH = 3.38 eV, UMinH = 3.48 eV, TMinH = 3.46 eV.

The role of electron donors and acceptors in base stacking in DNA and RNA.

The role of donor-acceptor interactions in base pair stacking in DNA and RNA has been minimized because of the perceived low or negative electron affinities of the purines and pyrimidines. The use of the electron capture detector was among the first methods for measuring electron affinities in the gas phase. Recently, the experimental determination of electron affinities has been extended and improved. Now, there are data for similar compounds in the literature which enable us to estimate electron affinities for purines and pyrimidines. These values are significant, and positive, such that donor-acceptor interactions can, and indeed should play a role in the stacking of bases in nucleic acids.

Identification of three novel unique proteins in seed oil bodies of sesame.

Plant seeds store triacylglycerols in discrete organelles called oil bodies. An oil body preserves a matrix of triacylglycerols surrounded by a monolayer of phospholipids embedded with abundant structural proteins termed oleosins and probably some uninvestigated minor proteins of higher molecular mass. Three polypeptides of 27, 37, and 39 kDa (temporarily denominated as Sop1, Sop2, and Sop3) were regularly co-purified with seed oil bodies of sesame. Comparison of amino acid composition indicated that they were substantially less hydrophobic than the known oleosins, and thus should not be aggregated multimers of oleosins. The results of immuno-recognition to sesame proteins extracted from subcellular fractions of mature seeds, various tissues, and oil bodies purified from different stages of seed formation revealed that these three polypeptides were unique proteins gathered in oil bodies, accompanying oleosins and triacylglycerols, during the active assembly of the organelles in maturing seeds. Both in vivo and in intro, immunofluorescence labeling using secondary antibodies conjugated with FITC (fluorescein isothiocyanate) confirmed the localization of these three polypeptides in oil bodies.

Classification of organic molecules to obtain electron affinities from half-wave reduction potentials: cytosine, uracil, thymine, guanine and adenine.

A procedure for obtaining the adiabatic electron affinities (AEA) of organic molecules from half-wave reduction potentials in aprotic solvents is presented. Molecules are placed into groups according to their structure. Each group has a different solution energy difference. Calculations of AEA and charge distributions with AM1-multiconfiguration configuration interaction are used to support the intuitive classification of the molecules. The procedure is illustrated for Vitamins A and E, riboflavin, the azines, polyenes, hydroxy-pyrimidine, oxo-guanine, the hydrogen bonded cytosine-oxo-guanine as well as the AEA, and vertical EA (VEA) of Cytosine (C), Uracil (U), Thymine (T), Guanine (G) and Adenine (A). The latter values are: (VEA) G, 0.10; A, -0.49; U, 0.33; T, 0.31; C, -1.48 and (AEA) G, 1.51 +/- 0.05; A, 0.95 +/- 0.05; U, 0.80 +/- 0.05; T, 0.79 +/- 0.05; C, 0.56 +/- 0.05 in eV.