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Gene US9 of herpes simplex virus type 1 has been predicted, from DNA sequence analysis, to encode a protein of mol wt 10,026, designated 10K (D.J. McGeoch, A. Dolan, S. Donald, and F.J. Rixon (1985). J. Mol. Biol. 181, 1-13). We have investigated this protein by using a synthetic peptide corresponding to the 11 amino acids adjacent to the amino-terminal methionine and rasing antisera in rabbits. One antiserum was able to precipitate at least 12 electrophoretically distinct polypeptide species from extracts of BHK cells infected with HSV-1. The estimated molecular weights of these polypeptides ranged from 12K to 20K and immunoblotting showed them to be related proteins. The primary translation product has an apparent mol wt of 13K. The various forms of 10K differ in their relative abundance in the infected cell and also in their degree of phosphorylation. Lower molecular weight forms of the 10K protein can be precipitated from NP-40 extracts of HSV-1 virions, suggesting that these forms of 10K are contained in the virion tegument or envelope. An association between this protein and nucleocapsids has also been observed in the nuclei of infected cells by immunoelectron microscopy. These observations imply that the product of US9 is a tegument protein which becomes associated with nucleocapsids at, or soon after, their formation in the nuclei of infected cells.

Increasing doses of fenoterol and terbutaline were administered by inhaler to 12 stable asthmatics in a double-blind crossover trial. 100 microgram doses of fenoterol were shown to be equal in potency to 250 microgram doses of terbutaline, as measured by increases in forced expiratory volume in the first second (FEV1) and specific airways conductance (sGaw). FEV1 and sGaw remained significantly greater than pretreatment levels up to five hours after both drugs and terbutaline showed a significantly greater effect on FEV1 than fenoterol after four hours suggesting a small but significantly longer duration of activity. Fenoterol used at the dosage of 100 micrograms per puff (half the standard commercially available dose of 200 micrograms per puff) caused significantly more cardiac effects than terbutaline, as measured by the magnitude and duration of increases in heart rate, and the incidence of reported palpitations.

The relative prednisolone-sparing effects of inhaled budesonide 400 micrograms daily and beclomethasone dipropionate (BDP) 400 micrograms daily were compared in a double-blind crossover study of 26 patients with chronic asthma requiring treatment with BDP and oral prednisolone in a daily dose of 5 mg or greater. During each period of the trial budesonide and BDP were inhaled via conventional pressurized inhalers in a dose of 100 micrograms four times daily. Prednisolone was reduced by 1 mg per month from the patient's normal maintenance dose to zero or to the point at which asthmatic symptoms became unacceptable. The mean reduction in prednisolone dose during BDP treatment was 2.65 mg compared with 1.8 mg at the end of the budesonide period. The difference between the prednisolone-sparing properties of BDP and budesonide assessed in this way in this group of patients was statistically significant in favour of BDP. The mean minimum prednisolone doses at the end of the treatment periods were 3.46 mg for BDP and 4.3 mg for budesonide. Since inhaled steroids were not withdrawn the absolute prednisolone-sparing properties of the two drugs were not assessed, and thus a pharmacological potency ratio cannot be derived from the results. It is concluded that BDP is marginally more potent than budesonide in its prednisolone-sparing properties.

The efficacy and side-effects of a new corticosteroid, budesonide, was assessed by comparison with beclomethasone dipropionate in double-blind, cross-over study of 30 patients with chronic asthma. The treatment regimens were budesonide 200 micrograms twice daily by conventional pressurized aerosol with a tube spacer attached and beclomethasone 100 micrograms four times daily via a conventional inhaler. Each treatment was given for four weeks. Results were analyzed using Student's t-test for paired comparisons. No significant differences were found for morning and evening peak flow rates, symptom scores, bronchodilator inhaler usage or forced vital capacity. Forced expiratory volume in one second after 4 weeks on each treatment was significantly higher following budesonide therapy (p less than 0.05), but the absolute changes were small and unlikely to be of clinical relevance. There were no major side-effects during either treatment period, but compared with pre-treatment levels serum creatinine and lactic dehydrogenase levels rose significantly during treatment with budesonide (p less than 0.01 and p less than 0.05 respectively). None of these results reflected clinically important biochemical or haematological changes. There was a significant reduction in neutrophil counts following treatment with beclomethasone (p less than 0.05). In the short term treatment of chronic asthma, budesonide administered twice daily is as effective as four times daily beclomethasone. A twice daily dosage regimen should improve patient compliance with therapy.

A double-blind crossover study has been performed in 14 patients with moderately severe chronic asthma to compare the bronchodilator efficacy of two dosage regimens of intravenous enprofylline (high dose = 2 mg/kg bolus and 1 mg/kg/hour infusion; low dose = 1 mg/kg bolus and 500 micrograms/kg/hour infusion) with aminophylline (5 mg/kg/bolus and 500 micrograms/kg/hour infusion) and placebo. The bolus injections were given over 20 minutes and infusion over 160 minutes. Twelve subjects completed the study. High dose enprofylline was more effective than aminophylline in increasing PEF (P = 0.008) and FEV1 (P = 0.004). Low dose enprofylline and aminophylline were of similar efficacy. Side-effects, notably headaches and nausea, were more common with enprofylline; three out of 14 subjects receiving the high dose regimen developed severe nausea. The plasma enprofylline levels achieved with the high dose regimen were greater than anticipated. Further studies are required in acute severe asthma to clarify the therapeutic role of intravenous enprofylline and the most appropriate dosage regimen.

Topical steroid therapy is now widely used in the treatment of chronic asthma, but few controlled studies have been conducted to evaluate the most efficient dosage regimen. The present study was designed to assess, in a double-blind fashion, the relative efficacy of a new topical steroid, budesonide (Pulmicort) in controlling pulmonary function and asthma symptoms during twice-and 4-times daily dosing. Both active and placebo drug were administered via the tube spacer inhaler. Objective lung function measurements indicate that twice-daily dosing is at least as effective as 4-times daily dosing, and it is suggested that the twice-daily dosage regimen may lead to better patient compliance.

The resonance Raman spectrum of turnip cytochrome f is similar to that of other c-type cytochromes with the exception of a single band at 1532 cm-1 which is shifted to lower frequency relative to its position (1542-1545 cm-1) in other c-type cytochromes. Comparison of the frequency of this band with that in alkylated cytochrome c at high pH suggests that the sixth heme iron ligand in cytochrome f is a deprotonated lysine amino group rather than a methionine sulfur. Comparison of the amino-acid sequences of cytochromes f and c1 suggests lysine-145 as a likely candidate for the sixth heme iron ligand in cytochrome f.

We report the sequence of a 7800 base pair region of herpes simplex virus type 1 DNA, representing approximately 0.16 to 0.20 map units in the genome. This contains sequences transcribed into a leftward oriented set of five 3' coterminal mRNAs, together with two rightward transcribed flanking genes. One of the leftward genes encodes the virus's alkaline exonuclease, but the other gene products are uncharacterized. The amino acid sequence of one encoded protein suggested that it is a membrane embedded species. The DNA sequence is densely utilised, with two predicted out-of-frame overlaps of coding sequences, and probably six occurrences of promoter elements within coding sequences. Homologues of five of the genes were found for the distantly related Epstein-Barr virus, with a similar overall relative arrangement.

Transcription of HSV immediate early genes is stimulated by a virus structural protein, Vmw65, in a process that requires specific recognition of the sequence TAATGARAT (R = purine). Upon incubation of nuclear extracts of HSV-infected cells with a short DNA fragment containing TAATGARAT, a novel virus-induced protein-DNA complex (named IEC) was detected. Addition of virion extract, containing Vmw65, to nuclear extract from uninfected cells also resulted in the formation of IEC. Vmw65 is a component of IEC, which contains proteins bound specifically to TAATGARAT. Thus, Vmw65 and cellular factors combine to form a sequence-specific DNA-binding complex. This system provides a model for studies of the regulation of inducible cellular genes.

Gene US4 of herpes simplex virus type 1 (HSV-1) has been predicted, from DNA sequence analysis, to encode a protein of molecular weight 25237 and its properties suggest it to be a membrane-associated protein. We have investigated this protein by raising antiserum to a synthetic oligopeptide corresponding to a stretch of amino acids from an internal hydrophilic region of the predicted sequence. This antiserum immunoprecipitates three glycoprotein species of apparent mol. wt. 37 000, 48 000 and 56 000 from extracts of cells infected with HSV-1. These species are also specifically immunoprecipitated from purified virions. The in vitro translation product of gene US4 has an apparent mol. wt. of 35 000. Sequence comparisons of the short unique regions of the HSV-1 and HSV-2 genomes, in combination with published mapping data for glycoprotein G (gG) of HSV-2, has led to the conclusion that the product of gene US4 of HSV-1 is the equivalent of gG.

An open, cross-over trial was conducted on 25 asthmatic children, aged 6-13 years, who required inhaled steroids. They inhaled Budesonide 200 micrograms twice daily, either directly from the metered dose inhaler or via the pear spacer (PS), for 2 months on each, in randomized order. The effects of the treatment were monitored with diary cards recording peak expiratory flow rates twice daily, symptoms and treatment taken, and with monthly clinical assessments including more sensitive lung function studies (flow-volume loops and single breath nitrogen wash-out tests). There was no specifically PS-related improvement in symptoms or in the majority of tests, but the results showed improvement with time when using either method. The improvement was more distinct in some tests reflecting proximal airway calibre (i.e., PEFR) than in tests thought to reflect predominantly peripheral airway calibre (i.e., F50, RV). The bronchodilator responsiveness, as shown by the increase in lung function tests after a beta-agonist was given, was significantly greater for FVC during the periods when the PS was used, although there was no significant improvement in FEV2 or PEFR. The improvement in tests reflecting proximal airways may have been due to optimization of the inhalation technique, greater understanding of asthma, or better compliance with medication associated with regular attendance for the study. The greater bronchodilator response whilst children were inhaling budesonide by the PS may have been due to increased deposition or better distribution of the steroid but was probably related to a difference between the two groups in initial baseline function tests.

We have determined the DNA sequence of the herpes simplex virus type 1 (HSV-1) gene encoding the immediate early protein IE110, which is involved in transcriptional activation of later virus genes. The locations of the 5' and 3' termini of IE110 mRNA together with the positions of two introns, were identified. Examination of the DNA sequence suggested that translation starts at the first ATG after the 5' terminus of the mRNA, and that both introns occur in protein-coding sequence. The predicted IE110 polypeptide contains 775 amino acids, and has a molecular weight of 78452. It contains a cysteine-rich region resembling regions found in several proteins which interact functionally with DNA. An antiserum was raised to the predicted C terminal amino acid sequence of the IE110 polypeptide and was shown to immunoprecipitate the native protein from HSV-1-infected cell extracts. The functional importance of regions of the protein was evaluated by construction of frameshift and deletion mutants of a plasmid-borne IE110 gene. The mutants were tested for IE110 function by short-term transfection assays, and the results were correlated with the DNA sequence and RNA mapping studies.

The efficacy and side-effects of a new corticosteroid aerosol, budesonide and beclomethasone dipropionate were assessed in 30 patients with chronic asthma in a double-blind cross-over study. Budesonide was administered, 200 micrograms twice daily, from a conventional pressurized aerosol with a tube spacer extension attached and beclomethasone was given via a conventional inhaler in the recommended four times daily dose of 100 micrograms, each treatment being administered for one month. No clinically significant differences were found between the two treatments and no significant side-effects were observed.

Herpes simplex virus type 1 (HSV-1) encodes a polypeptide of apparent mol. wt. 136 000 (Vmw136) known to be a component of the virus-specified ribonucleotide reductase. Monoclonal antibodies that precipitate this polypeptide also precipitate a polypeptide of mol. wt. 38 000 (Vmw38) from extracts of HSV-1-infected cells. The basis for this co-precipitation has been investigated using a monoclonal antibody directed against Vmw136 and an oligopeptide-induced antiserum directed against the carboxy terminus of Vmw38. We have also made use of a temperature-sensitive (ts) mutant of HSV-1 which maps within the sequences encoding Vmw136 and which induces a thermolabile ribonucleotide reductase. Our experiments show (i) Vmw136 and Vmw38 form a complex in infected cells and (ii) the mutation in the ts mutant results in the two polypeptides being unable to form the complex at the non-permissive temperature. We speculate that association of the two polypeptides is necessary for ribonucleotide reductase activity. No evidence was found for involvement of host proteins in the proposed virus-induced ribonucleotide reductase complex. The terms RR1 and RR2 are suggested for the large and small subunits of the HSV-induced enzyme.

Increasing dose of budesonide, each dose given for a minimum of 1 month, were administered via a Nebuhaler to 14 patients. Two consecutive abnormally low fasting morning plasma cortisol values, taken 2 weeks apart, were accepted as evidence of hypothalamo-pituitary-adrenal (HPA) hypofunction. Data from nine of the 14 patients entering the study were available for analysis. One patient developed HPA hypofunction while inhaling 2.4 mg budesonide per day and another retained normal HPA function on a dose of 12 mg. In the remaining patients, intermediate doses resulted in suppression, or the patients were withdrawn unsuppressed for other reasons (n = 3). The results in this small patient sample suggest that budesonide administered via the Nebuhaler in doses up to 1.6 mg daily does not significantly affect plasma cortisol.