The extensive metabolism and administration of low doses of ethinylestradiol (EE) in preclinical animal species necessitates a sensitive analytical method to quantify the drug at low picogram-per-milliliter concentrations in biological matrixes. A highly sensitive and accurate method based on the derivatization of EE with dansyl chloride coupled with liquid chromatography/tandem mass spectrometry is described. The dansyl derivatization of EE introduced a basic secondary nitrogen into the molecule that was readily ionized in commonly used acidic HPLC mobile phases. The derivative showed an intense protonated molecular ion at m/z 530 under positive turbo ion spray ionization. The collision-induced dissociation of this ion formed a distinctive product at m/z 171, corresponding to the protonated 5-(dimethylamino)naphthalene moiety. The selected reaction monitoring, based on the m/z 530 ->> 171 transition, was highly specific for EE, since no background signal was observed from blank plasma obtained from rhesus monkeys. The limit of detection, at a signal-to-noise ratio of 5, was 0.2 fg/mL EE spiked into blank plasma. This allowed for a lower limit of quantitation of 5 pg/mL using a 50-microL plasma sample and 10-microL injection of dansylated derivative into the CTC-PAL Leap autosampler coupled to a Sciex API 4000 mass spectrometer. Using fast-gradient liquid chromatography, the analyte peak eluted at 1.6 min. The validation results showed high accuracy (% bias < 4) and precision (% CV < 7.5) at broad linear dynamic ranges (0.005-20 ng/mL), using deuterated EE as internal standard. Therefore, the facile dansyl derivatization coupled with tandem mass spectral analysis allowed the development of a highly sensitive and specific method for quantitation of trace levels of EE in the plasma of rhesus monkeys dosed orally and intravenously with EE.

OBJECTIVE

To evaluate a telephone-based child mental health consult service for primary care providers (PCPs).

METHODS

Record review, provider surveys, and Medicaid database analysis.

METHODS

Washington State Partnership Access Line (PAL) program.

METHODS

A total of 2285 PAL consultations by 592 PCPs between April 1, 2008, and April 30, 2011.

METHODS

Primary care provider-initiated consultations with PAL service.

METHODS

The PAL call characteristics, PCP feedback surveys, and Medicaid claims between April 2007 and December 2009 for fee-for-service Medicaid children before and after a PAL call.

RESULTS

Sixty-nine percent of calls were about children with serious emotional disturbances, and 66% of calls were about children taking psychiatric medications. Primary care providers nearly always received new psychosocial treatment advice (87% of calls) and were more likely to receive advice to start rather than stop a medication (46% vs 24% of calls). Primary care provider feedback surveys reported uniformly positive satisfaction with the program. Among Medicaid children, there was significant increases in attention-deficit/hyperactivity disorder and antidepressant medication use after the PAL call but no significant change in reimbursements for mental health medications (P < .05). Children with a history of foster care experienced a 132% increase in outpatient mental health visits after the PAL call (P < .05).

CONCLUSIONS

Primary care providers used PAL for psychosocial and medication treatment assistance for particularly high-needs children and were satisfied with the service. Furthermore, PAL was associated with increased use of outpatient mental health care for some children.

CD4 binding on gp120 leads to the exposure of highly conserved regions recognized by the HIV co-receptor CCR5 and by CD4-induced (CD4i) antibodies. A covalent gp120-CD4 complex was shown to elicit CD4i antibody responses in monkeys, which was correlated with control of the HIV virus infection (DeVico, A., Fouts, T., Lewis, G. K., Gallo, R. C., Godfrey, K., Charurat, M., Harris, I., Galmin, L., and Pal, R. (2007) Proc. Natl. Acad. Sci. U.S.A. 104, 17477-17482). Because the inclusion of CD4 in a vaccine formulation should be avoided, due to potential autoimmune reactions, we engineered small sized CD4 mimetics (miniCD4s) that are poorly immunogenic and do not induce anti-CD4 antibodies. We made covalent complexes between such an engineered miniCD4 and gp120 or gp140, through a site-directed coupling reaction. These complexes were recognized by CD4i antibodies as well as by the HIV co-receptor CCR5. In addition, they elicit CD4i antibody responses in rabbits and therefore represent potential vaccine candidates that mimic an important HIV fusion intermediate, without autoimmune hazard.

In this paper, we present two complementary strategies for enrichment of glycoproteins on living cells that combine the desirable attributes of "robust enrichment" afforded by covalent-labeling techniques and "specificity for glycoproteins" typically provided by lectin or antibody affinity reagents. Our strategy involves the selective introduction of aldehydes either into sialic acids by periodate oxidation (periodate oxidation and aniline-catalyzed oxime ligation (PAL)) or into terminal galactose and N-acetylgalactosamine residues by galactose oxidase (galactose oxidase and aniline-catalyzed oxime ligation (GAL)), followed by aniline-catalyzed oxime ligation with aminooxy-biotin to biotinylate the glycans of glycoprotein subpopulations with high efficiency and cell viability. As expected, the two methods exhibit reciprocal tagging efficiencies when applied to fully sialylated cells compared with sialic acid-deficient cells. To assess the utility of these labeling methods for glycoproteomics, we enriched the PAL- and GAL-labeled (biotinylated) glycoproteome by adsorption onto immobilized streptavidin. Glycoprotein identities (IDs) and N-glycosylation site information were then obtained by liquid chromatography-tandem mass spectrometry on total tryptic peptides and on peptides subsequently released from N-glycans still bound to the beads using peptide N-glycosidase F. A total of 175 unique N-glycosylation sites were identified, belonging to 108 nonredundant glycoproteins. Of the 108 glycoproteins, 48 were identified by both methods of labeling and the remainder was identified using PAL on sialylated cells (40) or GAL on sialic acid-deficient cells (20). Our results demonstrate that PAL and GAL can be employed as complementary methods of chemical tagging for targeted proteomics of glycoprotein subpopulations and identification of glycosylation sites of proteins on cells with an altered sialylation status.

The alkaline pH-responsive Pal/Rim signal transduction pathway mediating regulation of gene expression by ambient pH has been extensively studied in Aspergillus nidulans and Saccharomyces cerevisiae. In A. nidulans, PalH, PalI, PalF, PalC, PalA and PalB are required for the proteolytic activation of the executing transcription factor PacC. Although necessary, Pal proteins are insufficient to transmit the signal, which additionally requires ESCRT-I, II and Vps20 with Snf7 in ESCRT-III. Although this initially suggested cooperation between a plasma membrane sensor and an ESCRT-containing Pal complex on endosomes, recent evidence convincingly indicates that pH signaling actually takes place in plasma membrane-associated foci in which Pal proteins and an ESCRT-III polymer scaffold cooperate for pH signaling purposes, representing another non-endosomal role of ESCRT components.

Trichoderma virens genome harbours genes encoding 22 non-ribosomal peptide synthetases (NRPSs) with at least one complete module (containing adenylation, thiolation and condensation domains) and four PKS/NRPS (polyketide synthase/NRPS) hybrid enzymes. After a primary screen for expression of these 26 genes when mycelia of T. virens are in contact with maize roots, seven genes that are upregulated were selected for further study. Using homologous recombination, loss-of-function mutants in six of these were obtained (the seventh, tex2, was acquired from our previous studies). Plant assays in a hydroponics system revealed that all seven mutants retained the ability to internally colonize maize roots. However, a mutation in one of the PKS/NRPS hybrid genes impaired the ability of T. virens to induce the defence response gene pal (phenylalanine ammonia lyase), suggesting a putative role for the associated metabolite product in induced systemic resistance. Interestingly, the mutant retained its ability to induce another defence response gene aos (allene oxide synthase). We thus provide evidence that a PKS/NRPS hybrid enzyme is involved in Trichoderma-plant interactions resulting in induction of defence responses.

BACKGROUND

Fruit color is one of the most important economic traits of the sweet cherry (Prunus avium L.). The red coloration of sweet cherry fruit is mainly attributed to anthocyanins. However, limited information is available regarding the molecular mechanisms underlying anthocyanin biosynthesis and its regulation in sweet cherry.

RESULTS

In this study, a reference transcriptome of P. avium L. was sequenced and annotated to identify the transcriptional determinants of fruit color. Normalized cDNA libraries from red and yellow fruits were sequenced using the next-generation Illumina/Solexa sequencing platform and de novo assembly. Over 66 million high-quality reads were assembled into 43,128 unigenes using a combined assembly strategy. Then a total of 22,452 unigenes were compared to public databases using homology searches, and 20,095 of these unigenes were annotated in the Nr protein database. Furthermore, transcriptome differences between the four stages of fruit ripening were analyzed using Illumina digital gene expression (DGE) profiling. Biological pathway analysis revealed that 72 unigenes were involved in anthocyanin biosynthesis. The expression patterns of unigenes encoding phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavanone 3'-hydroxylase (F3'H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS) and UDP glucose: flavonol 3-O-glucosyltransferase (UFGT) during fruit ripening differed between red and yellow fruit. In addition, we identified some transcription factor families (such as MYB, bHLH and WD40) that may control anthocyanin biosynthesis. We confirmed the altered expression levels of eighteen unigenes that encode anthocyanin biosynthetic enzymes and transcription factors using quantitative real-time PCR (qRT-PCR).

CONCLUSIONS

The obtained sweet cherry transcriptome and DGE profiling data provide comprehensive gene expression information that lends insights into the molecular mechanisms underlying anthocyanin biosynthesis. These results will provide a platform for further functional genomic research on this fruit crop.

Cell water permeability and cell wall properties are critical to survival of plant cells during freezing, however the underlying molecular mechanisms remain elusive. Here, we report that a specifically cold-induced nuclear protein, Tolerant to Chilling and Freezing 1 (TCF1), interacts with histones H3 and H4 and associates with chromatin containing a target gene, blue-copper-binding protein (BCB), encoding a glycosylphosphatidylinositol-anchored protein that regulates lignin biosynthesis. Loss of TCF1 function leads to reduced BCB transcription through affecting H3K4me2 and H3K27me3 levels within the BCB gene, resulting in reduced lignin content and enhanced freezing tolerance. Furthermore, plants with knocked-down BCB expression (amiRNA-BCB) under cold acclimation had reduced lignin accumulation and increased freezing tolerance. The pal1pal2 double mutant (lignin content reduced by 30% compared with WT) also showed the freezing tolerant phenotype, and TCF1 and BCB act upstream of PALs to regulate lignin content. In addition, TCF1 acts independently of the CBF (C-repeat binding factor) pathway. Our findings delineate a novel molecular pathway linking the TCF1-mediated cold-specific transcriptional program to lignin biosynthesis, thus achieving cell wall remodeling with increased freezing tolerance.

The aims of this study were to assess the periodontal health and oral hygiene status of pregnant women and non-pregnant controls and to evaluate the effect of socio-demographic and other variables on such periodontal status. A total of 400 women, of whom 200 were pregnant (mean age=30 years, s.e.m.=0.05) and 200 non-pregnant controls (mean age=32 years, s.e.m.=0.05) were chosen at random from four health centres. The clinical parameters used were the Silness and Loe plaque index (Pl.I), Loe and Silness gingival index (GI), probing pocket depth (PPD), probing attachment level (PAL), any relationship to socio-demographic (age, level of education and professional level) and clinical variables (gestation period, previous pregnancy and vomiting during pregnancy) was evaluated. The results showed that pregnant women had significantly higher GI and PPD scores (P < 0.005) but with no statistically significant differences in PAL or Pl.I (P>> 0.01) compared with non-pregnant controls. Increased age, lower level of education and non-employment were associated with significantly higher GI and PPD scores (P < 0.01). All these clinical parameters increased in parallel with the increase in the stage of pregnancy, reaching their maximum at the eighth month. Women with previous or multiple pregnancy had statistically significantly higher GI and PPD scores than those who were pregnant for the first time (P < 0.01), but with no statistically significant differences in Pl.I or PAL scores (P>> 0.05). Also, women who vomited during pregnancy had significantly higher GI and PPD scores compared with those who did not vomit (P < 0.05). It is concluded that gingival inflammatory symptoms are aggravated during pregnancy and are related to increased age, lower level of education and non-employment. Therefore periodontal preventive programmes are very important for pregnant women.

OBJECTIVE

Adult Still's disease is one of the febrile disorders of unknown etiology, characterized by high fever, transient cutaneous rash and leukocytosis. Liver dysfunction in adult Still's disease has been described in some case reports. The objective of this study was to analyze the pattern and the frequency of liver abnormalities in a monocenter series of adult Still's disease patients.

METHODS

Data of 17 patients with adult Still's disease (fulfilling Yamaguchi's diagnostic criteria) were retrospectively reviewed. These patients were followed in an Internal Medicine Department over a period of 7 years.

RESULTS

The median age was 27 years with a sex ratio M/F of 1.4. Fever was present in 100% of the cases and hepatomegaly occurred in 47% of the cases. Abnormalities in liver biochemistry, apparent in 76% of the subjects were characterized from moderate (elevation of transaminases between 2 and 5 N) (65%) to severe cytolysis (level of transaminases>> 5 N) (12%), cholestasis (elevation of gamma GT and/or PAL) (65%), and increase in the level of LDH (35%). All of these symptoms disappeared either spontaneously or under treatment (83%), within a median period of 18 days.

CONCLUSIONS

This study confirms the high frequency of liver dysfunction in adult Still's disease patients. Although it is moderate and asymptomatic in most cases, severe cytolytic hepatitis has been described. This study especially puts forward the need for exploring the possibility of adult Still's disease in the presence of a fever and hepatic cytolysis.

OBJECTIVE

In the current recommendations for energy intake of different countries as well as in the international WHO recommendations for energy intake it is assumed that the elderly are less physically active than young adults. Therefore, the aim of the present study was to compare physical activity patterns and physical activity level (PAL) of young and elderly subjects.

METHODS

In 178 female (age 67.8+/-5.7 y, BMI 26.4+/-3.7 kg/m(2)) and 107 male (age 66.9+/-5.1 y, BMI 26.3+/-3.1 kg/m(2)) participants of the longitudinal study on nutrition and health status in an aging population of Giessen, Germany as well as in a young age group consisting of 154 women (age 24.8+/-3.0 y, BMI 21.0+/-2.2 kg/m(2)) and 68 men (age 26.8+/-3.4 y, BMI 23.3+/-2.4 kg/m(2)) different activities like occupational work, housework, gardening, walking and sports were assessed by a questionnaire. Energy expenditure of the different activities was calculated using multipliers for resting metabolic rate (RMR) according to the WHO. The same multipliers were used for young and elderly subjects. RMR was measured by indirect calorimetry after an overnight fast. PAL of the subjects was calculated as total energy expenditure divided by RMR.

RESULTS

Young adults did more occupational work and performed more sports than elderly subjects. In contrast elderly women did more housework in comparison to young women, and elderly men walked more than young men. Both elderly women and men did more gardening than young women and men. In elderly women, PAL was significantly higher in comparison to young women, whereas PAL of young and elderly men did not differ significantly.

CONCLUSIONS

The results indicate that despite different activity patterns, the young-old do not necessarily show a lower PAL than young subjects.

The plant enzyme phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) shows homology to histidine ammonia-lyase (HAL) whose structure has been solved by X-ray crystallography. Based on amino-acid sequence alignment of the two enzymes, mutagenesis was performed on amino-acid residues that were identical or similar to the active site residues in HAL to gain insight into the importance of this residues in PAL for substrate binding or catalysis. We mutated the following amino-acid residues: S203, R354, Y110, Y351, N260, Q348, F400, Q488 and L138. Determination of the kinetic constants of the overexpressed and purified enzymes revealed that mutagenesis led in each case to diminished activity. Mutants S203A, R354A and Y351F showed a decrease in kcat by factors of 435, 130 and 235, respectively. Mutants F400A, Q488A and L138H showed a 345-, 615- and 14-fold lower kcat, respectively. The greatest loss of activity occurred in the PAL mutants N260A, Q348A and Y110F, which were 2700, 2370 and 75 000 times less active than wild-type PAL. To elucidate the possible function of the mutated amino-acid residues in PAL we built a homology model of PAL based on structural data of HAL and mutagenesis experiments with PAL. The homology model of PAL showed that the active site of PAL resembles the active site of HAL. This allowed us to propose possible roles for the corresponding residues in PAL catalysis.

BACKGROUND

The relative influence of diet and physical activity on body fatness remains uncertain.

OBJECTIVE

The objective of the study was to investigate associations of dietary variables and physical activity with body fatness in urban Chinese adults.

METHODS

We conducted a cross-sectional study in 130 weight-stable men and women aged 35-49 y. Subjects were selected from upper and lower tertiles of dietary fat and physical activity on the basis of screening questionnaires. Dietary intake was assessed by weighed food intake, physical activity level (PAL) was calculated as the ratio of predicted total energy expenditure (TEE) to predicted resting energy expenditure, and body composition was measured with the use of (2)H(2)O. Reported energy intake and predicted TEE were validated against TEE determined with the use of (2)H(2)(18)O (n = 73).

RESULTS

Body fatness was positively associated with dietary variety (ie, variety of ingredients) (partial r = 0.186, P = 0.039) and frequency of consuming restaurant foods (partial r = 0.237, P = 0.001) and negatively associated with PAL (partial r = -0.307, P = 0.001) in a multiple regression analysis that controlled for sex and confounders. The combined variance accounted for by dietary variety and restaurant food consumption (9.1%) was equivalent to that for PAL (9.4%). Neither dietary fat nor energy density predicted body fatness, but dietary energy density predicted within-subject day-to-day variation in reported energy intake (P < 0.001).

CONCLUSIONS

Dietary variety, frequency of restaurant food consumption, and PAL significantly predicted body fatness in urban Chinese adults, but dietary fat did not. These findings support previous studies in US adults and suggest that dietary variables other than fat have an important influence on adult body composition.

BACKGROUND

Schizophrenia (SZ) and schizoaffective disorders (SA) are associated with cognitive deficits. Generally, a schizoaffective diagnosis is associated with better prognosis on the level of social integration. It is also well established that cognition is an important factor for good social outcome in schizophrenia. We hypothesized that, although patients suffering from SA share symptoms with SZ, they can be differentiated on the basis of neurocognitive function and that SA perform better in several domains.

METHODS

Performances of two groups SA (N = 13) and SZ (N = 44) were compared on several visual-motor tasks using CANTAB [Motor Screening (MOT), Reaction Time (RTI), Paired Associates Learning Task (PAL), and Stockings of Cambridge items (SOC)]. The two groups were matched for symptom severity. ANOVA with repeated measures was employed to determine whether any difference in cognitive scores during a 2-year period was significantly related to the diagnostic status.

RESULTS

A significant and durable difference was observed between SZ and SA on motor screening and explicit memory tests where SA performed better.

CONCLUSIONS

Neurocognitive tests may be relevant for distinguishing schizoaffective from schizophrenia, chiefly via tests tapping into visuo-spatial and visuo-motor coordination abilities (e.g., paired associated learning and motor screening).

Swedish children and adolescents may be adopting a sedentary lifestyle with low energy expenditures and intakes, but no quantitative data are available. The purpose of the present study in 50 adolescents aged 15 y was to investigate whether assessment of total energy expenditure (TEE) and physical activity level (PAL) by the doubly labeled water method and indirect calorimetry and estimation of energy intake by a 7-d diet record would indicate physical inactivity. The boys' (n = 25) mean weight was 112% and the girls' (n = 25) was 109% of Swedish reference values from 1976; the mean height of both boys and girls was 102% of those reference values. Mean TEE in the boys and girls, 13.82 +/- 1.90 and 10.70 +/- 1.59 MJ/d, and mean PAL (TEE/basal metabolic rate), 1.89 +/- 0.16 and 1.79 +/- 0.22, respectively, were nonsignificantly higher than corresponding figures from other published studies. Mean energy intake as a percentage of TEE was 81.9 +/- 17.9% in the boys and 78.3 +/- 16.4% in the girls. Significant negative correlations were found both between energy intake as a percentage of TEE and percentage body fat and between energy intake as a percentage of TEE and body mass index. These results add to the evidence that 7-d diet records underestimate energy intake in adolescents, particularly those with a tendency for overweight and increased body fat. The results support indications of a trend of increasing body weight and height in Swedish adolescents, but conflict with the presumptions of low physical activity, low energy expenditure, and low energy intake. These results support the view that current recommendations for energy intake during adolescence are too low.

BACKGROUND

Experimental studies indicate that endogenous plasminogen activator inhibitor-1 (PAI-1, encoded by the gene SERPINE1) modulates the immune response to lipopolysaccharide (LPS). On the other hand, LPS induces PAI-1 secretion. Activation of individual cells by LPS is facilitated by CD14. The single nucleotide polymorphisms -675 4G/5G in SERPINE1 and C-159T in CD14 are major determinants of PAI-1 and CD14 expression, respectively.

OBJECTIVE

To evaluate the frequency of the -675 4G/5G SERPINE1 and C-159T CD14 polymorphisms in house dust mite (HDM) allergic asthma patients.

METHODS

The polymorphisms were evaluated in unrelated inhabitants of northeastern Poland, including 372 HDM-allergic asthmatic patients and 160 healthy nonatopic control subjects using polymerase chain reaction.

RESULTS

Both the C allele of CD14 and the 4G allele of SERPINE1 were more frequently encountered in HDM-allergic asthmatic patients than in healthy control individuals. When the 5G/5G-TT/CT genotype was considered as a nonrisk genotype, all other genotypes were associated with asthma. The odds ratios ranged from 3.96 (95% confidence interval, 1.56-10.1) for the 5G/5G-CC genotype to 10.7 (95% confidence interval, 5.1-24.9) for the 4G/4G-CC genotype. Bronchial reactivity to histamine and total serum immunoglobulin (Ig) E levels were predominantly associated with the 4G/5G SERPINE1 variants, while bronchial reactivity to Dermatophagoides pteronyssinus and serum concentrations of specific IgE against D pteronyssinus were predominantly associated with the C/T CD14 variants. Patients with 4G/4G-CC genotype had the lowest forced expiratory volume in 1 second and the highest bronchial reactivity.

CONCLUSIONS

The SERPINE1 and CD14 polymorphisms studied here are associated with different aspects of bronchial reactivity and IgE response. Our results indicate that PAl-1 and CD14 may interact to affect susceptibility to allergic asthma.

Phage display technology (PDT), a combinatorial screening approach, provides a molecular diversity tool for creating libraries of peptides/proteins and discovery of new recombinant therapeutics. Expression of proteins such as monoclonal antibodies (mAbs) on the surface of filamentous phage can permit the selection of high affinity and specificity therapeutic mAbs against virtually any target antigen. Using a number of diverse selection platforms (e.g. solid phase, solution phase, whole cell and in vivo biopannings), phage antibody libraries (PALs) from the start point provides great potential for the isolation of functional mAb fragments with diagnostic and/or therapeutic purposes. Given the pivotal role of PDT in the discovery of novel therapeutic/diagnostic mAbs, in the current review, we provide an overview on PALs and discuss their impact in the advancement of engineered mAbs.

The influence of salicylic acid (SA) doses of 50 and 250 microM, for a period of up to 7 days, on selected physiological aspects and the phenolic metabolism of Matricaria chamomilla plants was studied. SA exhibited both growth-promoting (50 microM) and growth-inhibiting (250 microM) properties, the latter being correlated with decrease of chlorophylls, water content and soluble proteins. In terms of phenolic metabolism, it seems that the higher SA dose has a toxic effect, based on the sharp increase in phenylalanine ammonia-lyase (PAL) activity (24 h after application), which is followed by an increase in total soluble phenolics, lignin accumulation and the majority of the 11 detected phenolic acids. Guaiacol-peroxidase activity was elevated throughout the experiment in 250 microM SA-treated plants. In turn, some responses can be explained by mechanisms associated with oxidative stress tolerance; these mitigate acute SA stress (which is indicated by an increase in malondialdehyde content). However, PAL activity decreased with prolonged exposure to SA, indicating its inhibition. Accumulation of coumarin-related compounds (umbelliferone and herniarin) was not affected by SA treatments, while (Z)- and (E)-2-beta-D: -glucopyranosyloxy-4-methoxycinnamic acids increased in the 250 microM SA-treated rosettes. Free SA content in the rosettes increased significantly only in the 250 microM SA treatment, with levels tending to decrease towards the end of the experiment and the opposite trend was observed in the roots.

The rice variety Tai06-1 is resistant to rice stripe disease and Xiushui63 is a highly susceptible rice variety to this disease. These two varieties were used to analyze the expression patterns of defense genes and antioxidant defense responses at the seedling stage, upon feeding with viruliferous small brown planthopper (SBPH) and nonviruliferous SBPH, respectively. The expression levels of CP (coat protein) gene of rice stripe virus (RSV) were higher upon feeding with viruliferous SBPH in Xiushui63 than in Tai06-1 throughout most of the experimental period, suggesting that RSV replicaiton is disturbed in Tai06-1 but not in Xiushui63, therefore, the resistance to RSV is higher in Tai06-1 than in Xiuhsui63. We found that defense genes PR1a (pathogenesis-related class 1a), PAL (phenylalanine ammonia-lyase), and CHS (chalcone synthase) may play roles in the defense responses to both RSV and SBPH in Tai06-1, and PR4 and PR10a may only participate in defending against SBPH attack but not against RSV infection in Tai06-1. Our data reveal that Gns1 (1,3; 1,4-β-glucanase) may participate in the defense responses to both RSV and SBPH in Xiushui63 but not in Tai06-1, and LOX (lipoxygenase) may only participate in defending against to SBPH in both Tai06-1 and Xiushui63. The antioxidant enzymes superoxide dismutase, peroxidase, catalase, hydrogen peroxide, and malondialdehyde coordinately participate in the resistance to RSV in Tai06-1, and that oxidative damage is less in Tai06-1 than in Xiushui63.

The expression of a lipid transfer protein (LTP) gene is regulated in Oryza sativa roots in response to colonization by the mycorrhizal fungus Glomus mosseae. Transcript levels increased when the fungus forms appressoria and penetrates the root epidermis and decreased at the onset of the intercellular colonization of the root cortex. The analysis of histochemical GUS staining in transgenic rice plants carrying the Ltp/Gus construct confirm the induction of LTP: gene associated with fungal appressoria formation and penetration area. The induction of Ltp gene expression coincided in time with a transient increase in the expression of a phenylalanine ammonia-lyase (PAL:) gene and a transient accumulation of salicylic acid (SA) in the mycorrhizal roots. The expression of LTP: and PAL: was induced in rice roots after treatment with SA and Pseudomonas syringae indicating that both genes could be implicated in the plant defence response. The exogenous application of SA to rice interacting with the mycorrhizal fungus did not affect appressoria formation but, instead, resulted in a transient delay of root mycorrhization. Nevertheless, although LTP: maintained a prolonged SA-induced expression level, mycorrhizal formation could still proceed.

Pyothorax-associated lymphoma (PAL) is a non-Hodgkin lymphoma of exclusively B-cell phenotype developing in the pleural cavity of patients after more than 20-year history of pyothorax resulting from an artificial pneumothorax for the treatment of pulmonary tuberculosis or tuberculous pleuritis. The most common symptoms on admission are chest pain and fever. Serum neuron-specific enolase level suggesting a diagnosis of small cell lung cancer is occasionally elevated. Histologically PAL usually shows a diffuse proliferation of large cells of B-cell type (diffuse large B-cell lymphoma [DLBL]). In PAL cells, representative B-cell markers other than CD20 are frequently negative with aberrant expression of T-cell markers such as CD2. A gene expression profile of PAL is distinct from nodal DLBL in its higher expression level of interferon-inducible genes. PAL is strongly associated with Epstein-Barr virus (EBV) infection with expression of EBV latent genes such as EBNA-2, LMP-1, together with EBNA-1. Taken together, PAL is a distinct entity both in its clinicopathologic presentation as well as its gene expression profile. Use of an artificial pneumothorax, EBV infection, and cytokines and reactive oxygen species produced in longstanding pyothorax might be important factors for PAL development.

The use of 'agonist therapy' for cocaine and methamphetamine addiction involves administration of stimulant-like medications (e.g. monoamine releasers) to reduce withdrawal symptoms and prevent relapse. A significant problem with this strategy is that many candidate medications possess abuse liability due to activation of mesolimbic dopamine (DA) neurons in the brain. One way to reduce DA-mediated abuse liability of candidate drugs might be to add in serotonin (5-HT)-releasing properties, since substantial evidence shows that 5-HT neurons provide an inhibitory influence over mesolimbic DA neurons. This chapter addresses several key issues related to the development of dual DA/5-HT releasers for the treatment of substance use disorders. First, we briefly summarize the evidence supporting a dual deficit in DA and 5-HT function during withdrawal from chronic cocaine or alcohol abuse. Second, we discuss data demonstrating that 5-HT release can dampen DA-mediated stimulant effects, and the 'anti-stimulant' role of 5-HT(2C) receptors is considered. Next, the mechanisms underlying potential adverse effects of 5-HT releasers are described. Finally, we discuss recently published data with PAL-287, a novel non-amphetamine DA/5-HT-releasing agent that suppresses cocaine self-administration but lacks positive reinforcing properties. It is concluded that DA/5-HT releasers could be useful therapeutic adjuncts for the treatment of cocaine and alcohol addictions as well as for obesity, attention deficit disorder and depression.

Cells of Daucus carota grown in a liquid medium produced large amounts of cyanidin as the only flavonoid aglycon. After inoculation in fresh medium a maximum activity of phenylalanine ammonia lyase (PAL; EC 4.3.1.5) was observed within 24 h. L-α-aminooxy-β-phenylpropionic acid (L-AOPP), thought to be a competitive inhibitor of PAL, inhibited cyanidin accumulation up to 80%. In order to study the regulatory role of PAL, the effects of L-AOPP and t-cinnamic acid, the product of the deamination of phenylalanine, were investigated. Cinnamic acid, applied in vivo (10(-4) M), was not able to compensate for the inhibition of cyanidin production caused by L-AOPP (10(-4) M) in the same sample. Carrot cells treated with L-AOPP exhibited a "super-induction" of PAL already described for gherkin hypocotyls (Amrhein and Gerhardt 1979). This effect was not influenced by t-cinnamic acid. L-AOPP seems to be a very specific inhibitor since it affected neither growth nor soluble protein content, whereas t-cinnamic acid inhibited both. Investigations on the content of soluble amino acids in L-AOPP-treated cells revealed a specific accumulation of soluble phenylalanine, whereas treatment with t-cinnamic acid led to an increase of amino acids in general, thus indicating that the latter compound has a rather unspecific effect on cellular metabolism. In vitro studies with PAL isolated from Daucus carota revealed that L-AOPP inhibited the enzyme at very low doses (K I=2.4·10(-9)), whereas t-cinnamic acid, by comparison, affected the enzyme at high concentrations (K I=1.8·10(-4)).