Effective Management of Citrus Melanose Based on Combination of Ecofriendly Chemicals.

Citrus melanose, caused by the ascomycete fungus Diaporthe citri, is one of the most important diseases in China that affects not only the production but also the quality of citrus. In China, mancozeb is recommended to control melanose disease at the dose of 1.34 g/liter. However, it is widely applied in practice at the dose of 2.66 g/liter or even 4 g/liter, because reduced efficacy of the recommended dose was observed in regions severely damaged by melanose. In this study, some ecofriendly chemicals for melanose management were evaluated. First, the sensitivity to fungicides was screened in the laboratory based on the inhibition of mycelial growth and conidial germination of D. citri. Results showed that both quinone outside inhibitor (QoI) fungicides kresoxim-methyl and trifloxystrobin inhibited conidial germination of D. citri up to 100% at 0.1 µg/ml. The in vivo control efficacy on detached fruit indicated that treatments with elastic nanocopolymer film at 2 g/liter, mancozeb at 1 g/liter, and kresoxim-methyl at 0.1 g/liter significantly inhibited the infection process compared with the control treatment of mineral oil alone. In field trials, the efficacy of kresoxim-methyl at 0.1 g/liter and elastic nanocopolymer film at 2 g/liter mixed with mancozeb at 1 g/liter was equal to that of mancozeb at 2.66 g/liter. The use of mancozeb could be reduced greatly, and the newly developed fungicide combinations are more environmentally friendly due to the low toxicity of both QoI fungicides and elastic nanocopolymer film. The newly developed method with ecofriendly chemicals should play an important role in the management of citrus melanose in the future.

[Early changes within the lymphocyte population are associated with the long term prognosis in severely injured patients].

OBJECTIVE: To investigate the relationship between early lymphocyte responses and the prognosis in severely injured patients. METHODS: Consecutive patients with severe trauma who were treated in Peking University People's Hospital Trauma Medical Center between June 2017 and June 2020 were enrolled in this restropective chart-review study. According to the responses of lymphocyte after severe injury, the patients were divided into three groups, group 1: lymphopenia-returned to normal; group 2: persistent lymphopenia; group 3: never lymphopenic, and the outcome of 28 d were recorded. Clinical data such as gender, age, base excess, mechanism of injury, Glasgow coma scale (GCS), injury severity score (ISS) and massive blood transfusion were collected. Perform statistical analysis on the collected clinical data to understand the trend of lymphocyte changes in early trauma and the relationship with prognosis. In order to eliminate the interference of age, stratification was carried out according to whether the age was ≥ 65 years old, in different age groups, they were grouped according to whether the length of stay was ≥ 28 d, and the relationship between lymphocyte trend and length of stay was discussed. RESULTS: A total of 83 patients were included, 66 males and 17 females. The main injury mechanisms were traffic accident injuries and high-altitude fall injuries. The average ISS was (30±11) points. 65 patients had lymphopenia on the day of injury, 32 of them returned to normal on the 5th day, and the rest did not recover; the other 18 patients had normal lymphocyte levels after injury. Patients which are failure to normalize lymphopenia within the first 5 days following admission was related with the long hospitalization time and higher 28 d mortality rate. After further stratification by age, failure to normalize lymphopenia within the first 5 days following admission in the elderly group (age ≥65 years) was a risk factor for prolonged hospital stay (≥28 d), P=0.04. While in younger group, a high level of neutrophils within the first 5 d following admission was a risk factor for bad outcome. CONCLUSION: A failure to normalize lymphopenia in severely injured patients is associated with significantly higher mortality and longer hospital stay. This study reveals lymphocytes can be used as a reliable indicator for the prognostic evaluation.

Correlations of CYP11B2 gene polymorphisms with eclampsia.

OBJECTIVE: The aim of this study was to explore the relationship between CYP11B2 gene polymorphisms and eclampsia. PATIENTS AND METHODS: A total of 400 pregnant women treated in our hospital were enrolled in this study, including 200 normal pregnant women (pregnancy group) and 200 pregnant women with eclampsia (eclampsia group). Peripheral blood was collected from subjects of the two groups. Subsequently, genomic deoxyribonucleic acids (DNAs) were extracted and amplified via polymerase chain reaction (PCR) for detection of CYP11B2 rs4543, rs3802228 and rs104894072 polymorphisms. The expression level of CYP11B2 gene was measured as well. Additionally, the correlations of CYP11B2 gene polymorphisms with blood pressure and coagulation and renal function indexes were analyzed. RESULTS: The distribution of alleles of rs4543 locus in CYP11B2 gene was significantly different between eclampsia group and pregnancy group (p=0.027). The frequency of the allele C was significantly lower in eclampsia group than that of pregnancy group (p<0.05). There was a statistically significant difference in the genotype distribution of CYP11B2 rs3802228 (p=0.000) and rs104894072 (p=0.000) between eclampsia group and pregnancy group (p<0.05). Meanwhile, the frequency of AA genotype of rs3802228 and TG genotype of rs104894072 was remarkably higher in eclampsia group than that in pregnancy group (p<0.05). The distribution of the locus rs104894072 (p=0.044) in dominant model and rs3802228 (p=0.002) in recessive model in eclampsia group was different from that in pregnancy group (p<0.05). Eclampsia group showed remarkably elevated frequency of TT + TG of the locus rs104894072 in dominant model and lowered frequency of AG + GG of the locus rs3802228 in recessive model (p<0.05). Similarly, a significant difference was observed in the distribution of the haplotypes CGG (p=0.001) and TGT (p=0.048) in CYP11B2 gene between eclampsia group and pregnancy group (p<0.05). The linkage disequilibrium of the loci rs3802228 and rs104894072 was relatively high (D'=0.382). The polymorphism of the locus rs104894072 in CYP11B2 gene had an evident relation to CYP11B2 gene expression (p<0.05). Meanwhile, the expression of CYP11B2 gene was markedly higher in patients with GG genotype in eclampsia group (p<0.05). The polymorphism of CYP11B2 rs4543 was notably associated with PT level of patients in eclampsia group (p=0.000). Conversely, rs3802228 polymorphism was correlated with 24 h urine protein level (p=0.000). Besides, the proportion of patients with CGG haplotype was significantly larger among patients with systolic blood pressure of 140-160 mmHg (p<0.05). In addition, the proportion of patients with TGT haplotype was evidently greater among patients with systolic blood pressure >180 mmHg in eclampsia group (p<0.05). CONCLUSIONS: CYP11B2 gene polymorphisms are significantly correlated with the development and progression of eclampsia.

[Risk factors for mechanical ventilation in patients with severe multiple trauma].

OBJECTIVE: To eludicate the risk factors of mechanical ventilation and prolonged mechanical ventilation in patients with severe multiple injuries. METHODS: Consecutive patients with severe multiple injures who were treated in Peking University People's Hospital Trauma Medical Center between December 2016 and December 2019 were enrolled in this restropective chart-review study. According to mechanical ventilation and ventilatory time, the patients were divided into mechanical ventilation (MV) group and non-mechanical ventilation (NMV) groups, prolonged mechanical ventilation (PMV) group and shortened mechanical ventilation (SMV) groups. Clinical data such as gender, age, base excess, mechanism of injury, Glasgow Coma Scale (GCS), abbreviated injury scale (AIS) and injury severity score (ISS) were collected. To indentify the risk factors of mechanical ventilation and prolonged mecha-nical ventilation, univariate and multivariate Logistic analyses were carried out. RESULTS: In the present study, 112 patients (82 male, 30 female) with severe multiple injuries having a median age of 52 (range: 16-89 years) and a median ISS of 34 (range: 16-66) were enrolled. The primary mechanism of injury was traffic accident injury and falling injury. In the study, 62 and 50 patients were assigned to MV and NMV groups, respectively. Logistic analysis showed that GCS (OR=0.72, 95%CI: 0.53-0.92, P=0.03), base excess (OR=0.56, 95%CI: 0.37-0.88, P=0.002) and multiple rib fracture (OR=1.72, 95%CI: 1.60-2.80, P=0.012) were independent significant risk factors for mechanical ventilation after severe multiple injuries. Within the mechanical ventilation group, 38 and 24 patients were assigned to PMV and SMVgroups, respectively. Compared with the SMV group, the PMV group had a higher ISS and higher rate of severe head trauma. The length of hospital stay of PMV group was longer than that of SMV groups. Meanwhile, the incidence of tracheotomy in PMV group was high. CONCLUSIONS: GCS, base excess and rib fracture might be independent risk factors for mechanical ventilation. Higher ISS and lower GCS might prolong the ventilatory time and the length of hospital stay. Meanwhile, the incidence of tracheotomy was high in PMV group because of the longer ventilatory time and poor consciousness.

[Application of multidisciplinary team (MDT) in the treatment of severe trauma].

OBJECTIVE: To explore the effect of multi-disciplinary team (MDT) in general hospitals on severe trauma patients. METHODS: This study reviewed the treatment of patients with severe trauma in trauma center of Peking University People's Hospital from March 2017 to April 2019. The baseline information: the patients' gender, age, injury mechanism, etc.; the start indicators: the Glasgow coma scale (GCS), trauma index (TI), injury severity score (ISS); the start related indicators: time for activation, time for MDT to arrive, time for CT scan, time for damage control surgery; patient treatment and prognosis: ICU (intensive care unit) length of stay, number of cured and discharged patients, number of dead cases, number of patients transferred to rehabilitation hospital, were all analyzed. It discussed the composition of MDT, the initiation scheme, the indicators of initiation of MDT for severe trauma, and analyzed the correlation between the application of MDT and the prognosis of patients. RESULTS: From March 2017 to April 2019, 112 trauma patients were treated by MDT in Peking University People's Hospital. There were 69 males and 43 females. The minimum age was 15 years, the maximum age was 89 years, most of them were 36-55 years old. The main injury mechanism was traffic accident injury. The GCS, TI, ISS were 13.0±2.9, 13.0±2.8, and 21.5±11.9, respectively. It took 3.7±0.8 minutes to start the call, 6.1±0.9 minutes for MDT personnel to arrive at the emergency rescue area, 23.8±3.0 minutes for fast CT and 92.6±15.4 minutes for injury control operation. All the hospitalized patients were treated effectively. ICU (Intensive care unit) hospitalization time was 12.6±6.7 days. 55 discharged patients were cured, 5 died (1 died of hemorrhagic shock, 4 died of severe brain injury) and 52 transferred to rehabilitation hospital. CONCLUSION: The treatment of severe trauma patients by MDT in trauma center of general hospitals can greatly improve the ability and level of treatment of severe trauma patients, make up for the lack of treatment of severe trauma especially multiple trauma patients in large general hospitals, and improve the treatment effect of severe trauma patients. It provides a reference model for large general hospitals to treat patients with severe trauma and multiple trauma and for the construction of trauma centers.

Cadherin 6 is activated by Epstein-Barr virus LMP1 to mediate EMT and metastasis as an interplay node of multiple pathways in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is an epithelial malignancy, which is notorious among head-and-neck cancers with its metastatic feature. Epstein-Barr virus (EBV) infection plays a fundamental role in NPC development with the mechanism is not well understood. Here we demonstrate that EBV oncoprotein LMP1 drives EMT and metastasis of NPC by reactivating the adhesion molecule, cadherin 6 (CDH6), which normally occurs in embryogenesis with unknown role in NPC. CDH6 was found to be upregulated in LMP1-positive NPC tissues, and was identified as a target of the epithelium-specific miR-203. LMP1-activated NF-κB transcriptionally repressed the miR-203 expression by binding to the promoter region of miR-203 gene. CDH6 activation in turn induced EMT and promoted metastasis in NPC. CDH6 depletion, NF-κB inhibitor and miR-203 overexpression were able to impair the EMT effects. The miR-203 downregulation in NPC tissues was strongly associated with metastasis clinically. The CDH6 activator, Runt-related transcription factor 2 (RUNX2), was also activated by EBV in the event. For both CDH6 and RUNX2 are components at TGF-ß downstream, CDH6 became a node protein for the interplay of multiple signalings including NF-κB and TGF-ß. Therefore, the switch-on of miR-203 was important for nasopharyngeal epithelial cells to maintain normal phenotype. This study demonstrates that EBV has evolved sophisticated strategies by driving epithelial cells to obtain malignant features, particularly in NPC metastasis, providing novel biomarkers for the therapy and prognosis of EBV-associated NPC.

[Analysis of risk factors associated with weaning from mechanical ventilation in critical patients with major abdominal surgery].

OBJECTIVE: To analyze the risk factors involved in the weaning from mechanical ventilation in critical patients who underwent major abdominal surgery. METHODS: This retrospective study was conducted at Department of Critical Care Medicine in Peking University People's Hospital. The subjects included all critical ill patients who underwent major abdominal surgery from January 2011 to December 2013. Clinical and laboratory parameters in perioperative period were investigated for the risk factors involved in the weaning from mechanical ventilation. RESULTS: In this study, 381 patients were included, of whom, 274 were successfully weaned. We found old age, lower left ventricular ejection fraction (LVEF) before surgery, with the complication of myocardial injury after noncardiac surgery (MINS) and lower serum albumin level after surgery were the independent risk factors of weaning from mechanical ventilation. And the days of intensive care unit and 28-day mortality of patients who successfully weaned were better than patients who failed to wean. CONCLUSION: Old age, lower LVEF before surgery, with the complication of MINS and lower serum albumin level after surgery were independent risk factors of weaning in critical patients who underwent major abdominal surgery.

First Report of Dimethachlon Resistance in Field Isolates of Sclerotinia sclerotiorum on Oilseed Rape in Shaanxi Province of Northwestern China.

Sclerotinia sclerotiorum (Lib.) de Bary is a necrotrophic fungal pathogen causing diseases in a wide range of plants, including oilseed rape (3). Substantial economic losses caused by S. sclerotiorum have been reported in the United States, Canada, Brazil, South Africa, Hungary, India, Nepal, and Japan (1). Application of fungicides is the principal tool for controlling S. sclerotiorum because of lack of high level of host resistance. Dicarboximide fungicides such as dimethachlon have been widely used to control S. sclerotiorum in recent years in China and field isolates with reduced sensitivity to dimethachlon have been reported in Jiangsu Province of eastern China (2). In order to understand the current status of dimethachlon resistance in S. sclerotiorum isolates of northwestern China, 196 and 344 isolates of S. sclerotiorum collected from oilseed rape fields in 10 counties throughout Shaanxi Province in 2011 and 2012, respectively, were assayed for sensitivity to dimethachlon using 5 µg ml-1 dimethachlon as a discriminatory dose. Mycelial plugs (6 mm in diameter) cut from the margin of a 48-h-old colony were placed in the center of petri dishes containing potato dextrose agar (PDA) amended with 5 µg ml-1 dimethachlon; PDA without fungicide served as the control. Cultures were incubated at 26°C and colony growth was measured after 72 h of incubation. Isolates that showed growth on PDA amended with fungicide were tentatively considered resistant to dimethachlon, whereas the completely inhibited isolates were considered sensitive. Results showed that 1.02% or 2 isolates of the 196 isolates collected in 2011 and 3.78% or 13 isolates of the 344 isolates collected in 2012 were resistant to dimethachlon. For all the isolates considered resistant and 42 randomly selected sensitive isolates, 50% effective concentrations (EC50) were determined on PDA amended with a series of dimethachlon concentrations. The average EC50 value of dimethachlon for sensitive isolates was 0.29 ± 0.02 µg ml-1 Resistance ratios (EC50 of resistant isolate / average EC50 of sensitive isolates) for the two resistant isolates detected in 2011 were 10.28 and 23.83, respectively, whereas resistance ratios for the 13 resistant isolates detected in 2012 ranged from 24.90 to 101.97. The average EC50 value of dimethachlon for the 13 resistant isolates detected in 2012 was 19.05 µg ml-1, and EC50 values for the two resistant isolates detected in 2011 were 2.98 and 6.91 µg ml-1, respectively. These results indicated that both resistance frequency and resistance level increased from 2011 to 2012. Bioassay results of three resistant isolates indicated that there was positive cross-resistance between dimethachlon and other dicarboximide fungicides such as iprodione and procymidone. To our knowledge, this is the first report of dimethachlon resistance in S. sclerotiorum in Shaanxi Province of northwestern China. The molecular mechanism of dimethachlon resistance in field isolates of S. sclerotiorum remains to be studied. Although resistance frequency is low at present, dimethachlon resistance should be kept in mind and fungicide resistance management tactics such as use of biological control agents, fungicide tank-mixing, or alternating dimethachlon with other fungicides having different modes of action is recommended in controlling S. sclerotiorum. References: (1) M. D. Bolton et al. Mol Plant Pathol. 7:1, 2006. (2) H. X. Ma et al. Plant Dis. 93:36, 2009. (3) L. H. Prudy. Phytopathology 69:875, 1979.

First Report of the ß-tubulin E198A Allele for Fungicide Resistance in Monilinia fructicola from South Carolina.

Resistance to methyl benzimidazole carbamates (MBCs) in Monilinia fructicola, the causal agent of brown rot of stone fruits, is known to be present in South Carolina peach orchards, but the molecular mechanism of resistance has not been investigated. Nine isolates were collected from peach in five counties in South Carolina and examined in petri dish assays on potato dextrose agar (PDA) for resistance to the MBC fungicide thiophanate-methyl (Topsin-M 70WP; Ceraxagri, King of Prussia, PA) at the discriminatory dose of 50 µg/ml. Isolates that grew on the fungicide-amended medium were considered highly resistant (HR). The ß-tubulin gene from four sensitive (S) and five HR M. fructicola isolates was PCR-amplified with primer pair TubA and TubR1 as described previously (1). Sequence analysis revealed several silent mutations in introns and exons in S and HR isolates and the presence of the previously described E198A allele in HR but not S isolates (1). Nucleotide sequences of the ß-tubulin gene from three S (BS, S2, MfEgpc1) and two HR isolates (MfPdt6 and BR2) were submitted to GenBank under accession numbers HM051379, HM051380, HM051381, HM051382, and HM051383, respectively. To our knowledge, this is the first report of the E198A in M. fructicola isolates from South Carolina and the East Coast. This allele is responsible for high levels of MBC resistance in M. fructicola (1). A previously reported PCR-based method using primers HRF+HRR designed to detect the E198A mutation in M. fructicola HR isolates (1) was improved by adding primer TR739 (5'-TCA CGA CGA ACA ACA TCA AGA-3') to the PCR cocktail. This additional internal primer amplified a 222-bp fragment from all S and HR isolates and therefore provided a useful, additional control. The confirmation of the E198A allele in M. fructicola isolates provides another useful tool to detect MBC resistance in commercial peach orchards in South Carolina. Reference: (1) Z. H. Ma et al. Appl. Environ. Microbiol. 69:7145, 2003.

[cDNA cloning and sequencing of human urokinase receptor].

Human urokinase receptor (uPAR), a 55 kD glycoprotein linked to the cell membrane by a glycosylphosphatidylinositol anchor, plays a central role in cell migration and tissue remodeling. The human uPAR cDNA was cloned from a highly metastatic human lung giant cell line PG by RT-PCR and then subcloned into pGEM-T vector and sequenced. The data indicate that there are three bases substitution (705, 746, 755) which subsequently leads to two amino acid mutation (249, 252) compared to that of previously reported. The cDNA sequence of uPAR was registered in GenBank with accession number AF257789.

Development and use of a polymerase chain reaction-based diagnostic test for the causal mutation of progressive retinal atrophy in Cardigan Welsh Corgis.

OBJECTIVE: To develop an allele-specific polymerase chain reaction (ASPCR)-based diagnostic test for the mutation in the cyclic guanosine monophosphate phosphodiesterase alpha subunit gene (PDE6A) that causes the rcd3 form of progressive retinal atrophy (PRA) in Cardigan Welsh Corgis. ANIMALS: 1 affected homozygote, 1 unaffected carrier, 1 genotypically normal dog, and 500 unknown-PRA status Cardigan Welsh Corgis. PROCEDURE: Control blood samples were collected from Cardigan Welsh Corgis of known PRA status (ie, affected homozygote, unaffected carrier, and a genotypically normal dog) for test development. Test blood samples were collected from 500 Cardigan Welsh Corgis of unknown PRA status. Genomic DNA was used as a template in ASPCR. One pair of primers was designed to specifically amplify only the mutant allele, and another set to amplify only the wildtype allele. The PCR conditions were adjusted to ensure each reaction was 100% specific. RESULTS: The PCR conditions were identified so that each ASPCR only amplified the allele it was designed to amplify. Of the 500 Cardigan Welsh Corgis tested using the newly developed ASPCR, 457 were homozygous for the normal allele (genotypically normal), 43 were heterozygous (phenotypically normal carriers), and none were homozygous for the mutant allele. CONCLUSION AND CLINICAL RELEVANCE: A rapid, ASPCR diagnostic test able to detect the PDE6A gene mutation responsible for the rcd3 form of PRA in Cardigan Welsh Corgis was developed. The test provides a useful service for Cardigan Welsh Corgi breeders and will enable them to prevent the birth of homozygote mutant dogs.

[Construction of fusion recombinant plasmid coding for RGD peptide and urokinase B chain, its expression in Escherichia coli and preliminary characterization of its biological activity].

A synthetic RGD (Arg-Gly-Asp) peptide-coding sequence was fused with urokinase B-chain cDNA and then cloned into the prokaryotic expression vector pBV220. The fused gene was expressed in E. coli DH5 alpha under the control of PRPL promoter by 42 degrees C induction. The expression level of the fusion protein was over 9.2% of the total bacterial proteins as a or of inactive inclusion body. The purified fusion protein was obtained with similar antigenicity as urokinase shown by Western blotting. Its in vitro fibrinolysis and anti-platelet aggregation activity was also evaluated by bioassay.

Identification of the immediate-early transcripts of Kaposi's sarcoma-associated herpesvirus.

In the immediate-early phase of reactivation or primary infection, herpesviruses express a small number of genes without requiring prior viral protein synthesis. Immediate-early genes usually encode regulatory proteins critical for the viral life cycle. Kaposi's sarcoma-associated herpesvirus (KSHV) gene transcription in the immediate-early stage of viral reactivation was examined by using a chemical induction combined with a gene expression screening method. RNA transcripts from at least four KSHV genomic loci accumulate when latently infected B-lymphoma cells are induced for reactivation in the presence of an inhibitor of protein synthesis (cycloheximide) and thus represent immediate-early class transcripts. Among them, a 3.6-kb mRNA encodes three putative open reading frames (ORFs), namely, ORF50, K8, and K8.2. ORF50 is a homologue of Rta, a transcription activator encoded by Epstein-Barr virus (EBV). The K8 gene codes for a 237-amino-acid protein with a basic-leucine zipper domain near its C terminus and an acidic domain near its N terminus and which closely resembles the ZEBRA protein of EBV and Jun/Fos family proteins. Other immediate-early mRNAs of KSHV include a 1. 7-kb mRNA encoding ORF45, a 2.0-kb mRNA encoding ORF K4.2, and a 4. 5-kb mRNA. Functional roles of products of these KSHV immediate-early transcripts remain to be studied.

Modulation of ACh release from airway cholinergic nerves in horses with recurrent airway obstruction.

To evaluate the functional status of neuronal alpha2-adrenoceptors (ARs) and beta2-ARs on ACh release in horses with recurrent airway obstruction (RAO), we examined the effects of the physiological agonists epinephrine (Epi) and norepinephrine (NE) and the beta2-agonists RR- and RR/SS-formoterol on ACh release from airway cholinergic nerves of horses with RAO. Because SS-formoterol, a distomer of the beta2-agonist, increases ACh release from airways of control horses only after the autoinhibitory muscarinic receptors are blocked by atropine, we also tested the hypothesis that if there is an M2-receptor dysfunction in equine RAO, SS-formoterol should increase ACh release even in the absence of atropine. ACh release was evoked by electrical field stimulation and measured by HPLC. Epi and NE caused less inhibition of ACh release in horses with RAO than in control horses. At the catecholamine concentration achieved during exercise (10(-7) M), the inhibition induced by Epi and NE was 10.8 +/- 13.2 and 3.4 +/- 6.8%, respectively, in equine RAO versus 41.0 +/- 6.4 and 27.1 +/- 5.6%, respectively, in control horses. RR- and RR/SS-formoterol (10(-8) to 10(-5) M) increased ACh release to a similar magnitude as that in control horses. These results indicate that neuronal beta2-ARs are functioning; however, the alpha2-ARs are dysfunctional in the airways of horses with RAO in response to circulating catecholamines. SS-formoterol (10(-8) to 10(-5) M) facilitated ACh release in horses with RAO even in the absence of atropine. Addition of atropine did not cause significantly more augmentation of ACh release over the effect of SS-formoterol alone. The magnitude of augmentation in horses with RAO in the absence of atropine was similar to that in control horses in the presence of atropine. The latter observations could be explained by neuronal muscarinic-autoreceptor dysfunction in equine RAO.

Mediators of anaphylaxis but not activated neutrophils augment cholinergic responses of equine small airways.

Neutrophilic inflammation in small airways (SA) and bronchospasm mediated via muscarinic receptors are features of chronic obstructive pulmonary disease in horses (COPD). Histamine, serotonin, and leukotrienes (LTs) are reported to be involved in the exacerbation of COPD, and currently, histamine has been shown to increase tension response to electrical field simulation (EFS) in equine SA. We tested the effects of these mediators and the effects of activated neutrophils on the cholinergic responses in SA. Histamine, serotonin, and LTD4 had a synergistic effect on EFS responses and only an additive effect on the tension response to exogenous ACh or methacholine. Atropine and TTX entirely eliminated the EFS-induced tension response in the presence of all three inflammatory mediators, indicating that augmentation of the EFS response applies only to the endogenous cholinergic response. Neutrophils isolated from control and COPD-affected horses were activated by zymosan, producing 18.1 +/- 2.3 and 25.0 +/- 2.3 nmol superoxide. 10(6) cells-1. 30 min-1, respectively. However, in contrast to the profound effect of mediators, incubation of SA for over 1 h in a suspension of up to 30 x 10(6) zymosan-treated neutrophils/ml did not significantly affect EFS responses of SA isolated from either control or COPD-affected horses. We conclude that in equine SA 1) the endogenous cholinergic responses are subject to strong facilitation by inflammatory mediators; 2) activated neutrophils do not affect cholinergic responses in SA; and 3) in acute bouts of equine COPD, histamine, LTD4, and serotonin (mediators primarily associated with type I allergic reaction) rather than mediators derived from neutrophils most likely contribute to increased cholinergic airway tone.

Role of cAMP and neuronal K+ channels on alpha 2-AR-induced inhibition of ACh release in equine trachea.

To investigate the effects of changes in intracellular cAMP on alpha 2-adrenoceptor (AR)-induced inhibition of airway acetylcholine (ACh) release, we examined the effects of the alpha 2-AR agonist clonidine on electrical field stimulation-evoked ACh release from equine tracheal parasympathetic nerves before and after treatment with 8-bromo-cAMP or forskolin. We also tested whether charybdotoxin (ChTX)- or iberiotoxin (IBTX)-sensitive Ca(2+)-activated K+ channels mediate alpha 2-AR-induced inhibition by examining the effect of clonidine in the absence and presence of ChTX or IBTX on ACh release. The amount of released ACh was measured by HPLC coupled with electrochemical detection. Clonidine (10(-7) to 10(-5) M) dose dependently inhibited ACh release before and after treatment with 8-bromo-cAMP (10(-3) M) or forskolin (3 x 10(-5) M). ChTX and IBTX, both at the concentration of 5 x 10(-7) M, significantly increased ACh release; however, they did not alter the magnitude of clonidine-induced inhibition. These results indicated that in equine tracheal parasympathetic nerves, alpha 2-AR-induced inhibition of ACh release is via an intracellular cAMP-independent pathway. Activation of both ChTX- and IBTX-sensitive Ca(2+)-activated K+ channels inhibits the electrical field stimulation-evoked ACh release, but these channels are not involved in the alpha 2-AR-induced inhibition of ACh release.

[Effect of diazepam on delayed nausea and vomiting caused by anticancer agents].

We conducted an evaluation of the usefulness of antiemetics (5-Hydroxy-tryptamine 3 receptor antagonism, 5HT3RA) combined with diazepam for delayed nausea and vomiting due to anticancer agents in 17 patients with various malignancies (such as lung Ca, breast Ca, esophagus Ca, gastric Ca, colon Ca, and non Hodgkin's disease) for whom chemotherapy was performed with different regimens in the Dept. of Oncologic Chemotherapy, People's Hospital, Beijing Medical University. Antiemetics (5HT3RA) combined with diazepam were given only to cases that had symptoms of nausea and vomiting induced by anticancer agents in the 1st course and invalidity with antiemetics (5HT3RA) alone in this study. Antiemetic (5HT3RA) agents + Dexamethasone were dosed before chemotherapy and also diazepam 5 mg orally after 24 hours (namely, when nausea was observed). Nausea was reduced and vomiting decreased after the antiemetic treatment with 5HT3RA + Dexamethasone and diazepam. These results indicated that 5HT3RA and diazepam combination therapies were more effective than 5HT3 RA + Dexamethasone alone for delayed nausea and vomiting. Further, the antiemetics had characters that a short adminiter time, few times and a take not over dose. The only side effect related to this antiemetic therapy was light somnolence. Antiemetics combined with diazepam might be a useful therapy against delayed nausea and vomiting induced by anticancer agents.

Effects of enantiomers of beta 2-agonists on ACh release and smooth muscle contraction in the trachea.

The beta 2-agonists currently used as bronchodilators are racemic mixtures of R- and S-enantiomers. In the present study, we examined the effects of enantiomers of the beta 2-agonists albuterol and formoterol on acetylcholine (ACh) release from equine trachealis parasympathetic nerves. ACh release was evoked by electrical field stimulation (20 V, 0.5 ms, 0.5 Hz) and measured by high-performance liquid chromatography coupled with electrochemical detection. We also tested the effects of enantiomers of albuterol and formoterol on equine tracheal smooth muscle (TSM) contraction in response to exogenous ACh. R- and RS-albuterol (10(-8) to 10(-5) M) and RR- and RR/SS-formoterol (10(-8) to 10(-5) M) augmented ACh release in a concentration-dependent manner. Beginning at 10(-6) M, SS-formoterol significantly increased ACh release, and at 10(-5) M, release increased by 71.9 +/- 8.7% over baseline. This effect was only observed, however, when the prejunctional muscarinic autoinhibitory effect of ACh was prevented with atropine. Both the RR- and SS-formoterol-induced increases in ACh release were abolished by the beta 2-antagonist ICI-118551 (3 x 10(-7) M). The effect of S-albuterol on ACh release was variable, and the mean increase induced by 10(-5) M was 30.8 +/- 16.1% in the presence of atropine. In the muscle tension study, R- and RS-albuterol and RR- and RR/SS-formoterol (10(-8) to 10(-5) M) but not the S-enantiomers inhibited TSM contraction. Even though R-enantiomers augment ACh release, they potently inhibit TSM contraction. Because racemic beta 2-agonists are bronchodilators on acute administration, the postjunctional spasmolytic effects of R-enantiomers predominate over the spasmogenic effect evoked via increased ACh release. The S-enantiomers, in contrast, do not inhibit TSM contraction and therefore would not contribute to the observed bronchodilation of the racemate. The S-enantiomers do prejunctionally facilitate ACh release when prejunctional muscarinic autoreceptors are dysfunctional, suggesting a potentially deleterious effect.