cAMP signaling in brain is decreased in unmedicated depressed patients and increased by treatment with a selective serotonin reuptake inhibitor.

Basic studies exploring the importance of the cyclic adenosine monophosphate (cAMP) cascade in major depressive disorder (MDD) have noted that the cAMP cascade is downregulated in MDD and upregulated by antidepressant treatment. We investigated cAMP cascade activity by using 11C-(R)-rolipram to image phosphodiesterase-4 (PDE4) in unmedicated MDD patients and after ~8 weeks of treatment with a selective serotonin reuptake inhibitor (SSRI). 11C-(R)-rolipram positron emission tomographic (PET) scans were performed in 44 unmedicated patients during a major depressive episode and 35 healthy controls. Twenty-three of the 44 patients had a follow-up 11C-(R)-rolipram PET scan ~8 weeks after treatment with an SSRI. Patients were moderately depressed (Montgomery-Åsberg Depression Rating Scale=30±6) and about half were treatment naïve. 11C-(R)-rolipram binding was measured using arterial sampling to correct for individual differences in radioligand metabolism. We found in unmedicated MDD patients widespread, ~20% reductions in 11C-(R)-rolipram binding compared with controls (P=0.001). SSRI treatment significantly increased rolipram binding (12%, P<0.001), with significantly greater increases observed in older patients (P<0.001). Rolipram binding did not correlate with severity of baseline symptoms, and increased rolipram binding during treatment did not correlate with symptom improvement. In brief, consistent with the results of basic studies, PDE4 was decreased in unmedicated MDD patients and increased after SSRI treatment. The lack of correlation between PDE4 binding and depressive symptoms could reflect the heterogeneity of the disease and/or the heterogeneity of the target, given that PDE4 has four subtypes. These results suggest that PDE4 inhibitors, which increase cAMP cascade activity, may have antidepressant effects.

Investing in prospective cohorts for etiologic study of occupational exposures.

Prospective cohorts have played a major role in understanding the contribution of diet, physical activity, medical conditions, and genes to the development of many diseases, but have not been widely used for occupational exposures. Studies in agriculture are an exception. We draw upon our experience using this design to study agricultural workers to identify conditions that might foster use of prospective cohorts to study other occupational settings. Prospective cohort studies are perceived by many as the strongest epidemiologic design. It allows updating of information on exposure and other factors, collection of biologic samples before disease diagnosis for biomarker studies, assessment of effect modification by genes, lifestyle, and other occupational exposures, and evaluation of a wide range of health outcomes. Increased use of prospective cohorts would be beneficial in identifying hazardous exposures in the workplace. Occupational epidemiologists should seek opportunities to initiate prospective cohorts to investigate high priority, occupational exposures.

Reduced cannabinoid CB1 receptor binding in alcohol dependence measured with positron emission tomography.

Brain cannabinoid CB1 receptors contribute to alcohol-related behaviors in experimental animals, but their potential role in humans with alcohol dependence is poorly understood. We measured CB1 receptors in alcohol dependent patients in early and protracted abstinence, and in comparison with control subjects without alcohol use disorders, using positron emission tomography and [(18)F]FMPEP-d2, a radioligand for CB1 receptors. We scanned 18 male in-patients with alcohol dependence twice, within 3-7 days of admission from ongoing drinking, and after 2-4 weeks of supervised abstinence. Imaging data were compared with those from 19 age-matched healthy male control subjects. Data were also analyzed for potential influence of a common functional variation (rs2023239) in the CB1 receptor gene (CNR1) that may moderate CB1 receptor density. On the first scan, CB1 receptor binding was 20-30% lower in patients with alcohol dependence than in control subjects in all brain regions and was negatively correlated with years of alcohol abuse. After 2-4 weeks of abstinence, CB1 receptor binding remained similarly reduced in these patients. Irrespective of the diagnostic status, C allele carriers at rs2023239 had higher CB1 receptor binding compared with non-carriers. Alcohol dependence is associated with a widespread reduction of cannabinoid CB1 receptor binding in the human brain and this reduction persists at least 2-4 weeks into abstinence. The correlation of reduced binding with years of alcohol abuse suggests an involvement of CB1 receptors in alcohol dependence in humans.

Addressing phase errors in fat-water imaging using a mixed magnitude/complex fitting method.

Accurate, noninvasive measurements of liver fat content are needed for the early diagnosis and quantitative staging of nonalcoholic fatty liver disease. Chemical shift-based fat quantification methods acquire images at multiple echo times using a multiecho spoiled gradient echo sequence, and provide fat fraction measurements through postprocessing. However, phase errors, such as those caused by eddy currents, can adversely affect fat quantification. These phase errors are typically most significant at the first echo of the echo train, and introduce bias in complex-based fat quantification techniques. These errors can be overcome using a magnitude-based technique (where the phase of all echoes is discarded), but at the cost of significantly degraded signal-to-noise ratio, particularly for certain choices of echo time combinations. In this work, we develop a reconstruction method that overcomes these phase errors without the signal-to-noise ratio penalty incurred by magnitude fitting. This method discards the phase of the first echo (which is often corrupted) while maintaining the phase of the remaining echoes (where phase is unaltered). We test the proposed method on 104 patient liver datasets (from 52 patients, each scanned twice), where the fat fraction measurements are compared to coregistered spectroscopy measurements. We demonstrate that mixed fitting is able to provide accurate fat fraction measurements with high signal-to-noise ratio and low bias over a wide choice of echo combinations.

Cytochrome P450 polymorphisms and the response of lupus nephritis to cyclophosphamide therapy.

BACKGROUND AND AIMS: The addition of cyclophosphamide to corticosteroids significantly improves the prognosis of severe kidney involvement in systemic lupus erythematosus (SLE). However, not all patients respond to cyclophosphamide. It has been suggested that genetic variations that reduce the metabolism of cyclophosphamide reduce its effectiveness. Cyclophosphamide is metabolized and activated by the cytochrome P450 (CYP) system and in particular CYP enzymes 2B6 and 2C19. Both CYP2B6 and CYP2C19 have variant alleles (CYP2B6*5 and CYP2C19*2) that attenuate or eliminate enzymatic activity. This investigation was done to determine the impact of CYP2B6*5 and CYP2C19*2 on the renal response in cyclophosphamide-treated lupus nephritis (LN) patients. METHODS: Patients with SLE (n = 237), unclassified autoimmune disease (n = 51), and healthy controls (n = 294) were genotyped for CYP2B6*5 and CYP2C19*2. Associations between these alleles and achievement of complete or partial response, development of end-stage renal disease, and time to remission were determined. RESULTS: The frequencies of the variant alleles CYP2B6*5 and CYP2C19*2 were 6.3 % and 15.9%, respectively. CYP2C19*2 genotypes were more frequent among African Americans than European Americans, and CYP2B6*5 genotypes were more frequent among European Americans than African Americans. Among LN patients treated with cyclophosphamide (n = 36), there were no differences between those with or without these genotypes relative to the frequency of complete or partial remissions or time to remission. CONCLUSION: This retrospective analysis failed to show an association between CYP2B6*5 and CYP2C19*2 and treatment outcomes in LN. This suggests that genotyping for these CYP450 variants may not be useful in individualizing treatment for severe LN.

Influence of exposure route and oral dosage regimen on 1, 1-dichloroethylene toxicokinetics and target organ toxicity.

The objective of this investigation was to elucidate the effects of route of exposure and oral dosage regimen on the toxicokinetics (TK) of 1,1-dichloroethylene (DCE). Fasted male Sprague-Dawley rats that inhaled 100 or 300 ppm for 2 h absorbed total systemic doses of (10 or 30 mg/kg DCE, respectively. Other groups of rats received 10 or 30 mg/kg DCE by intravenous injection, bolus gavage (by mouth), or gastric infusion (g.i.) over a 2-h period. Serial microblood samples were taken from the cannulated, unanesthetized animals and analyzed for DCE content by gas chromatography to obtain concentration versus time profiles. Inhalation resulted in substantially higher peak blood concentrations and area under blood-concentration time curves (AUC(0)(2)) than did gastric infusion of the same dose over the same time frame at each dosage level, although inhalation (AUC(0)(infinity)) values were only modestly higher. Urinary N-acetyl-beta-D-glucosaminidase (NAG) and gamma-glutamyltranspeptidase (GGT) activities were monitored as indices of kidney injury in the high-dose groups. NAG and GGT excretion were much more pronounced after inhalation than gastric infusion. Administration of DCE by gavage also produced much higher Cmax and AUC(0)(2) values than did 2-h g.i., although AUC(0)(infinity) values were not very different. The 30 mg/kg bolus dose produced marked elevation in serum sorbitol dehydrogenase, an index of hepatocellular injury. Administration of this dose by inhalation and gastric infusion was only marginally hepatotoxic. These findings demonstrate the TK and target organ toxicity of DCE vary substantially between different exposure routes, as well as dosage regimens, making direct extrapolations untenable in health risk assessments.

Ionic liquids based on azolate anions.

Compartmentalized molecular level design of new energetic materials based on energetic azolate anions allows for the examination of the effects of both cation and anion on the physiochemical properties of ionic liquids. Thirty one novel salts were synthesized by pairing diverse cations (tetraphenylphosphonium, ethyltriphenylphosphonium, N-phenyl pyridinium, 1-butyl-3-methylimidazolium, tetramethyl-, tetraethyl-, and tetrabutylammonium) with azolate anions (5-nitrobenzimidazolate, 5-nitrobenzotriazolate, 3,5-dinitro-1,2,4-triazolate, 2,4-dinitroimidazolate, 4-nitro-1,2,3-triazolate, 4,5-dinitroimidazolate, 4,5-dicyanoimidazolate, 4-nitroimidazolate, and tetrazolate). These salts have been characterized by DSC, TGA, and single crystal X-ray crystallography. The azolates in general are surprisingly stable in the systems explored. Ionic liquids were obtained with all combinations of the 1-butyl-3-methylimidazolium cation and the heterocyclic azolate anions studied, and with several combinations of tetraethyl- or tetrabutylammonium cations and the azolate anions. Favorable structure-property relationships were most often achieved when changing from 4- and 4,5-disubstituted anions to 3,5- and 2,4-disubstituted anions. The most promising anion for use in energetic ionic liquids of those studied here, was 3,5-dinitro-1,2,4-triazolate, based on its contributions to the entire set of target properties.

Synthesis and X-ray structure determination of highly active Pd(II), Pd(I), and Pd(0) complexes of di(tert-butyl)neopentylphosphine (DTBNpP) in the arylation of amines and ketones.

The air-stable complex Pd(η(3)-allyl)(DTBNpP)Cl (DTBNpP = di(tert-butyl)neopentylphosphine) serves as a highly efficient precatalyst for the arylation of amines and enolates using aryl bromides and chlorides under mild conditions with yields ranging from 74% to 98%. Amination reactions of aryl bromides were carried out using 1-2 mol % Pd(η(3)-allyl)(DTBNpP)Cl at 23-50 °C without the need to exclude oxygen or moisture. The C-N coupling of the aryl chlorides occurred at relatively lower temperature (80-100 °C) and catalyst loading (1 mol %) using the Pd(η(3)-allyl)(DTBNpP)Cl precatalyst than the catalyst generated in situ from DTBNpP and Pd(2)(dba)(3) (100-140 °C, 2-5 mol % Pd). Other Pd(DTBNpP)(2)-based complexes, (Pd(DTBNpP)(2) and Pd(DTBNpP)(2)Cl(2)) were ineffective precatalysts under identical conditions for the amination reactions. Both Pd(DTBNpP)(2) and Pd(DTBNpP)(2)Cl(2) precatalysts gave nearly quantitative conversions to the product in the α-arylation of propiophenone with p-chlorotoluene and p-bromoanisole at a substrate/catalyst loading of 100/1. At lower substrate/catalyst loading (1000/1), the conversions were lower but comparable to that of Pd(t-Bu(3)P)(2). In many cases, the tri-tert-butylphosphine (TTBP) based Pd(I) dimer, [Pd(µ-Br)(TTBP)](2), stood out to be the most reactive catalyst under identical conditions for the enolate arylation. Interestingly, the air-stable Pd(I) dimer, Pd(2)(DTBNpP)(2)(µ-Cl)(µ-allyl), was less active in comparison to [Pd(µ-Br)(TTBP)](2) and Pd(η(3)-allyl)(DTBNpP)Cl. The X-ray crystal structures of Pd(η(3)-allyl)(DTBNpP)Cl, Pd(DTBNpP)(2)Cl(2), Pd(DTBNpP)(2), and Pd(2)(DTBNpP)(2)(µ-Cl)(µ-allyl) are reported in this paper along with initial studies on the catalyst activation of the Pd(η(3)-allyl)(DTBNpP)Cl precatalyst.

Ionic liquid-based routes to conversion or reuse of recycled ammonium perchlorate.

New, potentially green, and efficient synthetic routes for the remediation and/or re-use of perchlorate-based energetic materials have been developed. Four simple organic imidazolium- and phosphonium-based perchlorate salts/ionic liquids have been synthesized by simple, inexpensive, and nonhazardous methods, using ammonium perchlorate as the perchlorate source. By appropriate choice of the cation, perchlorate can be incorporated into an ionic liquid which serves as its own electrolyte for the electrochemical reduction of the perchlorate anion, allowing for the regeneration of the chloride-based parent ionic liquid. The electrochemical degradation of the hazardous perchlorate ion and its conversion to harmless chloride during electrolysis was studied using IR and (35)Cl NMR spectroscopies.

Exposure to ethylene oxide at work increases sister chromatid exchanges in human peripheral lymphocytes.

Sister chromatid exchange rates increased significantly in the peripheral lymphocytes of a small group of hospital workers exposed to ethylene oxide for as little as 3.6 minutes per day regularly over a period of months. Results based on breathing zone exposure and task frequency estimates over a 6-month period for 14 workers suggest that sister chromatid exchanges are a sensitive indicator of exposure and that cumulative dose and dose rate are important predictors of sister chromatid exchange response.

Using ionic liquids to trap unique coordination environments: polymorphic solvates of ErCl3(OH2)(4).2([C2mim]Cl).

Two polymorphs of ErCl(3)(OH(2))(4).2([C(2)mim]Cl) solvates were isolated from the same solution of 1-ethyl-3-methylimidazolium chloride when HCl(aq) was added, while [C(2)mim](3)[ErCl(6)] was isolated without HCl addition, illustrating how ionic liquids can be used to trap unusual coordination environments in the solid state.

DNA structure and flexibility in the sequence-specific binding of papillomavirus E2 proteins.

The papillomavirus E2 proteins are transcriptional regulators that bind to a consensus DNA sequence ACCG NNNN CGGT. Multiple copies of this binding site are found in the viral genomes. The affinities of the naturally occurring binding sites for the E2 proteins are predominantly dependent upon the sequence of the NNNN spacer. The hierarchies of binding site affinities among the sites present in the viral genomes result in differential occupancy during the viral life-cycle. In turn, this differential binding regulates transcription from viral promoters, including those for the oncogenes E6 and E7. Structural and biochemical studies have shown that E2 proteins bend the DNA to which they specifically bind. Atomic resolution structures of complexes of the bovine papillomavirus strain 1 (BPV-1) E2 protein and DNA show that the protein does not contact the spacer DNA. A direct comparison of the binding of the DNA-binding domains of the E2 proteins from BPV-1 and human papillomavirus strain 16 (HPV-16) to a series of binding sites as a function of the sequence of their central spacer and/or the presence of a nick or gap in one strand of the spacer DNA is presented in this paper. The BPV-1 E2 DNA-binding domain is only moderately sensitive to the nature of the central spacer; less than several fold differences in affinity were observed when the DNA sequence of the spacer was varied and/or a nick or gap was introduced. In contrast, the HPV-16 E2 DNA-binding domain binds to sites containing A:T-rich central spacers with significantly increased affinity. The introduction of a nick or gap into the spacer of these high affinity sequences is very detrimental to HPV-16 E2 binding while comparable nicks or gaps have only small effects in the low affinity sequences. These results suggest that the HPV-16 E2 protein recognizes the structure of the DNA spacer and that the mechanism of DNA-sequence specific binding of the homologous HPV-16 E2 and BPV-1 E2 proteins is significantly different.

Cell cycle kinetic studies in 68 patients with myelodysplastic syndromes following intravenous iodo- and/or bromodeoxyuridine.

Sixty-eight patients with myelodysplastic syndromes (MDS) received sequential infusions of iodo- and/or bromodeoxyuridine for cell kinetic analysis. Bone marrow biopsy sections were treated by appropriate antibodies and a labeling index (LI), duration of S-phase (Ts), and total cell cycle time (Tc) of myeloid cells were determined. The mean LI was 28.4%, Ts was 11.8 hours and Tc was 40.7 hours. The %LI decreased as the disease evolved from refractory anemia toward transformation to acute leukemia (p = 0.04). Double-labeling of biopsy sections for apoptosis and proliferation showed that 30-90% of S-phase cells in MDS patients were simultaneously apoptotic or "antonymous." We conclude that MDS are highly proliferative disorders in which the ineffective hematopoiesis is probably the result of excessive apoptosis rather than slow proliferation.

Magnetic resonance imaging assessment of the ventricular system in the brains of adult and juvenile beagle dogs treated with posaconazole IV Solution.

INTRODUCTION: Noxafil® (posaconazole; POS) is a potent, selective triazole antifungal approved for use in adults as an oral suspension, oral tablet and intravenous (IV) Solution. In support of pediatric administration of POS IV Solution to childrentwo years of age, two studies were undertaken using magnetic resonance imaging (MRI) to monitor brain ventricle size longitudinally during three months administration of POS IV in adult and juvenile dogs. Necropsy was performed on all animals at the end of the studies. From the baseline MRI images, great variability in ventricle size was noted in both the adult and juvenile dogs; these images were used to distribute differently sized ventricles between treatment and vehicle groups as to not skew group means during the course of the study. RESULTS: POS IV Solution had no effect on ventricle volume at any timepoint during dosing in either the adult or the juvenile dogs. Further, no gross or histomorphologic differences between groups were observed in either study. Compared to juvenile dogs, MRI analysis showed that adult dogs had larger ventricles, lower variability in all ventricle volumes, and a greater rate of increase in total ventricle volume. DISCUSSION: Information on growth and development of brains is one of the few areas in which more detailed information is available about humans than about the standard laboratory animals used to model disease and predict toxicities. The use of MRI helped elucidate large natural variabilities in the dog brain, which could have altered the interpretation of this de-risking study, and provided a valuable noninvasive means to monitor the brain ventricles longitudinally.

Thyroid hormones and cytogenetic outcomes in backpack sprayers using ethylenebis(dithiocarbamate) (EBDC) fungicides in Mexico.

Ethylenebis(dithiocarbamate) (EBDC) fungicides are used heavily in the United States. EBDCs (e.g., mancozeb, maneb) are metabolized to ethylene thiourea (ETU). The EPA classifies ETU as a carcinogen, based on thyroid and other cancers in rodents, and has restricted the use of EBDCs, while requiring workers to use protective equipment. There are no data on the potential carcinogenicity of EBDCs in humans, and there is only one study on human genotoxicity. ETU is known to cause decreases of thyroxine (T4) and increases in thyroid-stimulating hormone (TSH) in rodents. We have studied cytogenetic outcomes and serum thyroid hormone levels among 49 heavily exposed workers without protective equipment spraying EBDC on tomatoes in Mexico. We also studied 14 lightly exposed landowners and 31 nonexposed controls. Urinary ETU was used to compare exposure between groups. We found an increase in TSH (p = 0.05) among applicators compared to controls, but no decrease in thyroid hormone (T4). We found increases in sister chromatid exchange (p = 0.03) and in chromosome translocations (chromosome aberrations that persist through cell division) for applicators compared to controls (p = 0.05). However, the subset of reciprocal translocations showed a lesser increase (p = 0.24). Our data suggest that EBDCs affect the thyroid gland and the lymphocyte genome among heavily exposed workers. However, our data are limited to subclinical outcomes, are of borderline statistical significance, and should be interpreted with caution.

Distribution of 2,4-D in air and on surfaces inside residences after lawn applications: comparing exposure estimates from various media for young children.

We collected indoor air, surface wipes (floors, table tops, and window sills), and floor dust samples at multiple locations within 11 occupied and two unoccupied homes both before and after lawn application of the herbicide 2,4-D. We measured residues 1 week before and after application. We used collected samples to determine transport routes of 2,4-D from the lawn into the homes, its subsequent distribution between the indoor surfaces, and air concentration as a function of airborne particle size. We used residue measurements to estimate potential exposures within these homes. After lawn application, 2,4-D was detected in indoor air and on all surfaces throughout all homes. Track-in by an active dog and by the homeowner applicator were the most significant factors for intrusion. Resuspension of floor dust was the major source of 2,4-D in indoor air, with highest levels of 2,4-D found in the particle size range of 2.5-10 microm. Resuspended floor dust was also a major source of 2,4-D on tables and window sills. Estimated postapplication indoor exposure levels for young children from nondietary ingestion may be 1-10 microg/day from contact with floors, and 0.2-30 microg/day from contact with table tops. These are estimated to be about 10 times higher than the preapplication exposures. By comparison, dietary ingestion of 2,4-D is approximately 1.3 microg/day.

Neurologic function among termiticide applicators exposed to chlorpyrifos.

Chlorpyrifos is a moderately toxic organophosphate pesticide. Houses and lawns in the United States receive a total of approximately 20 million annual chlorpyrifos treatments, and 82% of U.S. adults have detectable levels of a chlorpyrifos metabolite (3,5, 6-trichloro-2-pyridinol; TCP) in the urine. The U.S. Environmental Protection Agency has estimated that there are 5,000 yearly reported cases of accidental chlorpyrifos poisoning, and approximately one-fourth of these cases exhibit symptoms. Organophosphates affect the nervous system, but there are few epidemiologic data on chlorpyrifos neurotoxicity. We studied neurologic function in 191 current and former termiticide applicators who had an average of 2.4 years applying chlorpyrifos and 2.5 years applying other pesticides, and we compared them to 189 nonexposed controls. The average urinary TCP level for 65 recently exposed applicators was 629.5 microg/L, as compared to 4.5 microg/L for the general U.S. population. The exposed group did not differ significantly from the nonexposed group for any test in the clinical examination. Few significant differences were found in nerve conduction velocity, arm/hand tremor, vibrotactile sensitivity, vision, smell, visual/motor skills, or neurobehavioral skills. The exposed group did not perform as well as the nonexposed group in pegboard turning tests and some postural sway tests. The exposed subjects also reported significantly more symptoms, including memory problems, emotional states, fatigue, and loss of muscle strength; our more quantitative tests may not have been adequate to detect these symptoms. Eight men who reported past chlorpyrifos poisoning had a pattern of low performance on a number of tests, which is consistent with prior reports of chronic effects of organophosphate poisoning. Overall, the lack of exposure effects on the clinical examination was reassuring. The findings for self-reported symptoms raise some concern, as does the finding of low performance for those reporting prior poisoning. Although this was a relatively large study based on a well-defined target population, the workers we studied may not be representative of all exposed workers, and caution should be exercised in generalizing our results.

Evaluation of acute sensory--motor effects and test sensitivity using termiticide workers exposed to chlorpyrifos.

Sensory and motor testing was performed on a group of termiticide workers primarily using chlorpyrifos-containing products to evaluate both the acute effects from current exposure and sensitivity of the measures to detect effects. The study group comprised 106 applicators and 52 nonexposed participants. Current exposure was measured by urinary concentrations of 3,5,6-trichloro-2-pyridinol (TCP) collected the morning of testing. The mean TCP value for the 106 applicators was 200 microg/g creatinine. Participants received 4--5 h of testing and were evaluated using a sensory--motor test battery recommended by a National Institute for Occupational Safety and Health (NIOSH)-sponsored advisory panel to be appropriate for testing effects from pesticide exposures. Measurements testing olfactory dysfunction, visual acuity, contrast sensitivity, color vision, vibrotactile sensitivity, tremor, manual dexterity, eye--hand coordination, and postural stability were analyzed. Study results indicated limited acute effects from exposure to chlorpyrifos using urinary TCP as a measure of current exposure. The effects occurred primarily on measures of postural sway in the eyes closed and soft-surface conditions, which suggests a possible subclinical effect involving the proprioceptive and vestibular systems. Several other tests of motor and sensory functions did not show any evidence of acute exposure effects, although statistically significant effects of urinary TCP on the Lanthony color vision test scores and one contrast sensitivity test score were found. The visual measures, however, were not significant when a step-down Bonferroni correction was applied. Information also is presented on the sensitivity of the measures to detect effects in an occupationally exposed population using standard error of the parameter estimates.