Hyperresponsiveness to inhaled but not intravenous methacholine during acute respiratory syncytial virus infection in mice

Background

To characterise the acute physiological and inflammatory changes induced by low-dose RSV infection in mice.

Methods

BALB/c mice were infected as adults (8 wk) or weanlings (3 wk) with 1 × 10⁵ pfu of RSV A2 or vehicle (intranasal, 30 μl). Inflammation, cytokines and inflammatory markers in bronchoalveolar lavage fluid (BALF) and airway and tissue responses to inhaled methacholine (MCh; 0.001 – 30 mg/ml) were measured 5, 7, 10 and 21 days post infection. Responsiveness to iv MCh (6 – 96 μg/min/kg) in vivo and to electrical field stimulation (EFS) and MCh in vitro were measured at 7 d. Epithelial permeability was measured by Evans Blue dye leakage into BALF at 7 d. Respiratory mechanics were measured using low frequency forced oscillation in tracheostomised and ventilated (450 bpm, flexiVent) mice. Low frequency impedance spectra were calculated (0.5 – 20 Hz) and a model, consisting of an airway compartment [airway resistance (Raw) and inertance (Iaw)] and a constant-phase tissue compartment [coefficients of tissue damping (G) and elastance (H)] was fitted to the data.

Results

Inflammation in adult mouse BALF peaked at 7 d (RSV 15.6 (4.7 SE) vs. control 3.7 (0.7) × 10⁴ cells/ml; p < 0.001), resolving by 21 d, with no increase in weanlings at any timepoint. RSV-infected mice were hyperresponsive to aerosolised MCh at 5 and 7 d (PC₂₀₀ Raw adults: RSV 0.02 (0.005) vs. control 1.1 (0.41) mg/ml; p = 0.003) (PC₂₀₀ Raw weanlings: RSV 0.19 (0.12) vs. control 10.2 (6.0) mg/ml MCh; p = 0.001). Increased responsiveness to aerosolised MCh was matched by elevated levels of cysLT at 5 d and elevated VEGF and PGE₂ at 7 d in BALF from both adult and weanling mice. Responsiveness was not increased in response to iv MCh in vivo or EFS or MCh challenge in vitro. Increased epithelial permeability was not detected at 7 d.

Conclusion

Infection with 1 × 10⁵ pfu RSV induced extreme hyperresponsiveness to aerosolised MCh during the acute phase of infection in adult and weanling mice. The route-specificity of hyperresponsiveness suggests that epithelial mechanisms were important in determining the physiological effects. Inflammatory changes were dissociated from physiological changes, particularly in weanling mice.     

Photoautotrophic culture of Coffea arabusta somatic embryos: development of a bioreactor for large-scale plantlet conversion from cotyledonary embryos

Somatic embryos were developed from in vitro-grown leaf discs of Coffea arabusta in modified Murashige and Skoog medium under 30 micromol m(-2) s(-1) photosynthetic photon flux (PPF). Cotyledonary stage embryos were selected from the 14-week-old cultures and were placed under a high (100 micromol m(-2) s(-1) PPF for 14 d. These pretreated embryos were grown photoautotrophically in three different types of culture systems: Magenta vessel; RITA-bioreactor (modified to improve air exchange); and a specially designed temporary root zone immersion bioreactor system (TRI-bioreactor) with forced ventilation. The aims of the study were to achieve large-scale embryo-to-plantlet conversion, and to optimize growth of plantlets under photoautotrophic conditions. The plantlet conversion percentage was highest (84 %) in the TRI-bioreactor and lowest in the modified RITA-bioreactor (20 %). Growth and survival of converted plantlets following 45 d of photoautotrophic culture in each of the three culture systems were studied. Fresh and dry masses of leaves and roots of plantlets developed in the TRI-bioreactor were significantly greater than those of plantlets developed in the modified RITA-bioreactor or Magenta vessel. The net photosynthetic rate, chlorophyll fluorescence and chlorophyll contents were also highest in plantlets grown in the TRI-bioreactor. Normal stomata were observed in leaves of plantlets grown in the TRI-bioreactor, whereas they could be abnormal in plantlets from the modified RITA-bioreactor. Survival of the plants after transfer from culture followed a similar pattern and was highest in the group grown in the TRI-bioreactor, followed by plants grown in the modified RITA-bioreactor and Magenta vessel. In addition, ex vitro growth of plants transferred from the TRI-bioreactor was faster than that of plants from the other culture systems.     

高温对烟叶品质的影响

研究了下列条件下烟叶的成熟和品质的变化.在烟草移栽后第90d进行7d人工高温处理(白天温度为35℃,晚上温度为29℃,光照强度为3×104lx,相对湿度为75%),同期对照的自然条件是3d阴天,日均温为19.7～23.9℃;4d晴天,日均温为25.9～30.8℃,日最强光照强度均在1×1051x以上.结果表明,人工高温处理后,烟叶叶色比对照更绿,成熟期推迟5～7d,没有出现整株烟叶快速变黄的"高温逼熟"现象.对烟叶的细胞结构和化学成分分析,也没有出现"高温逼熟"引起品质变差的变化.因此认为湘南烟区"高温逼熟"导致烟叶品质下降的原因除高温以外,还与强光辐射、"火南风"等因素所造成的危害有关.     

Rice octadecanoid pathway

Plant jasmonic acid (JA) and structurally similar animal prostaglandins play pivotal roles in regulating cellular responses against environmental cues, including the innate immune response(s). In plants, JA and its immediate precursor 12-oxo-phytodienoic acid (OPDA) are synthesized by the octadecanoid pathway, which employs at least five enzymes (lipase, lipoxygenase, allene oxide synthase and cyclase, and OPDA reductase), in addition to the enzymes involved in the beta-oxidation steps. Genetic, molecular, and biochemical analyses have led to the identification of almost all the genes of the octadecanoid pathway in Arabidopsis--a model dicotyledonous plant. In this regard, rice (Oryza sativa L.)--an important socio-economic monocotyledonous model research plant--remains poorly characterized. Until now, no gene has been specifically associated with this pathway. It is therefore of utmost importance to identify, characterize, and assign the pathway specific genes in rice. In this review, we have surveyed the rice genome, extracted a large number of putative genes of the octadecanoid pathway, and discussed their relationship with the known pathway genes from other plant species. Moreover, the achievements made so far on the rice octadecanoid pathway have also been summarized to reflect the contribution of rice towards extending our knowledge on this critical pathway in plants.     

Functional significance of glycogen synthase kinase-3 regulation by serotonin

Serotonin modulates brain physiology and behavior and has major roles in brain diseases involving abnormal mood and cognition. Enhancing brain serotonin has been found to regulate glycogen synthase Kinase-3 (GSK3), but the signaling mechanism and functional significance of this regulation remain to be determined. In this study, we tested the signaling mechanism mediating 5-HT1A receptor-regulated GSK3 in the hippocampus. Using mutant GSK3 knock-in mice, we also tested the role of GSK3 in the behavioral effects of 5-HT1A receptors and the serotonin reuptake inhibitor fluoxetine. The results showed that activation of 5-HT1A receptors by 8-hydroxy-N,N-dipropyl-2-aminotetralin (8-OH-DPAT) increased phosphorylation of the N-terminal serine of both GSK3α and GSK3β in several areas of the hippocampus. The effect of 8-OH-DPAT was accompanied by an increase in the active phosphorylation of Akt, and was blocked by LY294002, an inhibitor of phosphoinositide 3-kinases (PI3K). Phosphorylation of GSK3β, but not GSK3α, was necessary for 5-HT1A receptors to suppress the hippocampus-associated contextual fear learning. Furthermore, acute fluoxetine treatment up-regulated both phospho-Ser21-GSK3α and phospho-Ser9-GSK3β in the hippocampus. Blocking phosphorylation of GSK3α and GSK3β diminished the anti-immobility effect of fluoxetine treatment in the forced swim test, wherein the effect of GSK3β was more prominent. These results together suggest that PI3K/Akt is a signaling mechanism mediating the GSK3-regulating effect of 5-HT1A receptors in the hippocampus, and regulation of GSK3 is an important intermediate signaling process in the behavioral functions of 5-HT1A receptors and fluoxetine.     

Impulsivity, risk taking, and timing

This study examined the relations among measures of impulsivity and timing. Impulsivity was assessed using delay and probability discounting, and self-report impulsivity (as measured by the Barratt Impulsiveness Scale; BIS-11). Timing was assessed using temporal perception as measured on a temporal bisection task and time perspective (as measured by the Zimbardo Time Perspective Inventory). One hundred and forty three college students completed these measures in a computer laboratory. The degree of delay discounting was positively correlated with the mean and range of the temporal bisection procedure. The degree of delay and probability discounting were also positively correlated. Self-reported motor impulsiveness on the BIS-11 was positively correlated with present hedonism and negatively correlated with future orientation on the ZTPI. Self-reported non-planning on the BIS-11 was positively correlated with fatalism on the ZTPI. These results show that people who overestimate the passage of time (perceive time as passing more quickly) hold less value in delayed rewards. They also confirm previous results regarding the relation between delay and probability discounting, as well as highlight similarities in self-report measures of impulsivity and time perspective.     

Enhanced expression of human β-defensin 2 in peripheral lungs of patients with chronic obstructive pulmonary disease

Human β-defensin 2 (hBD-2) has antimicrobial activity and may play a role in airway mucosal defense, but studies have not yet examined its expression in lung tissue of patients with chronic obstructive pulmonary disease (COPD). Here we investigated hBD-2 levels in lung tissues of COPD patients and analyzed their correlations with IL-8, IL-1β, cigarette smoking and lung function in order to see whether the protein may be involved in pathogenesis of the disease. Peripheral lung tissue specimens were obtained from 51 patients who underwent lung resection for peripheral lung cancer: healthy non-smokers (n = 8), healthy current smokers (n = 7), non-smokers with COPD (n = 11), and current smokers with COPD (n = 25). RT-PCR and immunohistochemical staining were used to detect expression levels of hBD-2, IL-8 and IL-1β. Expression of hBD-2 mRNA was significantly higher in COPD patients than in healthy controls, and significantly higher in current smokers than in non-smokers (p < 0.05). Among healthy controls, hBD-2 mRNA levels were similar between current smokers and non-smokers. Immunohistochemistry showed hBD-2 protein to be expressed mainly in epithelia of distal bronchioles and its expression pattern among our patient groups mirrored that of the mRNA. IL-8 mRNA levels were significantly higher in COPD patients than in healthy controls (p < 0.05), while IL-1β mRNA levels did not differ significantly among the groups. Levels of hBD-2 mRNA positively correlated with levels of IL-8 mRNA (r = 0.545, p = 0.002), and negatively correlated with FEV1/FVC ratios and with predicted FEV1% values (r = −0.406, p = 0.011). Our results indicate that hBD-2 expression is elevated in distal airways of COPD patients and that it may be involved in pathogenesis of the disease. Our data implicate cigarette smoking as a factor that may elevate hBD-2 levels in lung tissues of COPD patients.     

Time course of self-desensitization of oral irritation by nicotine and capsaicin

Nicotine contacting mucous membranes elicits irritation that decreases with repeated exposures (self-desensitization). We investigated the time course of nicotine self-desensitization and compared it with that of capsaicin. Nicotine (300 mM, 10 microl) was applied to one-half of the dorsal tongue and vehicle to the other. Following a rest period ranging from 0.5 to 48 h, nicotine (5 microl) was reapplied to each side of the tongue and subjects indicated on which side they experienced stronger irritation and separately rated the intensity of the sensation on each side. After intervals of 0.5, 1, and 24 h, a significant majority of subjects chose the vehicle-treated side as having stronger irritation and assigned significantly higher intensity ratings to that side, indicating self-desensitization. The effect was not present after 48 h. By comparison, 10 parts per million (ppm) (33 microM) capsaicin induced significant self-desensitization at 1 but not 24 h, whereas a higher concentration of capsaicin (100 ppm, 330 microM) induced significant self-desensitization at intervals of 1, 24, and 48 h. These results indicate that initial exposure to nicotine or capsaicin can markedly attenuate irritant sensations elicited by subsequent exposure to these irritants hours to days later.     

Expression,SNV identification,linkage disequilibrium,and combined genotype association analysis of the muscle-specific gene CSRP3 in Chinese cattle

The cysteine and glycine-rich protein 3 (CSRP3) plays an important role in the myofiber differentiation. Here, we identified five SNVs in all exon and intron regions of the CSRP3 gene using DNA sequencing, PCR-RFLP and forced-PCR-RFLP methods in 554 cattle. Four of the five SNVs were significantly associated with growth performance and carcass traits of the cattle. In addition, we evaluated haplotype frequency and linkage disequilibrium coefficient of five sequence variants. The result of haplotype analysis demonstrated 28 haplotypes present in Qinchuan and two haplotypes in Chinese Holstein. Only haplotypes 1 and 8 were being shared by two populations, haplotype 14 had the highest haplotype frequency in Qinchuan (17.4%) and haplotype 8 had the highest haplotype frequency in Chinese Holstein (94.4%). Statistical analyses of combined genotypes indicated that some combined genotypes were significantly or highly significantly associated with growth and carcass traits in the Qinchuan cattle population. qPCR analyses also showed that bovine CSRP3 gene was exclusively expressed in longissimus dorsi muscle and heart tissues. The data support the high potential of the CSRP3 as a marker gene for the improvement of growth performance and carcass traits in selection programs.