Subdural spinal haematoma after epidural anaesthesia in a patient with spinal canal stenosis.

A 60-year-old male with a past history of T12 fracture had epidural analgesia for a radical prostatectomy. It was unknown prior to epidural insertion that the patient had a canal stenosis at T12 from the previous injury. The patient developed severe bilateral buttock pain after epidural catheter removal. Magnetic resonance imaging demonstrated a spinal subdural haematoma from T10 to L2 with mild cord compression. The patient made a successful recovery with conservative management. Neuraxial blockade should be approached with caution in patients with previous back injury, and only after a thorough assessment has been obtained to exclude spinal canal stenosis.

Biomass and aggregation analysis of human embryonic kidney 293 suspension cell cultures by particle size measurement.

A method has been developed to monitor the cell growth of aggregated human embryonic kidney 293 (HEK293) suspension cultures by measuring cumulative particle volume and the particle size distribution. This method employs a particle size analyzer that determines the size of individual particles by detecting their light obscuration (blockage) or scattering. Cell counts derived from the cumulative volume of the cell particle correlate well with manual cell counts from a hemacytometer at different stages of growth. This correlation was further confirmed by quantifying total cellular protein of the samples. Simultaneously, the aggregation state of the samples can also be monitored and mathematically described. Results from this study demonstrate that this simple and reproducible method allows the direct measurement of cumulative cell volume and the degree of cell aggregation, as well as an indirect assessment of cell counts.

General practice on a new housing estate. 1956.

An analysis has been carried out of the records kept for the year 1953 by six doctors practising on a new housing estate near London. The population of the estate is described in terms of age, sex, and social class. 76 per cent. of the registered patients consulted a doctor at some time during the year, the consultation rate per person being 4.1 based on the average registered population; females had more consultations than males; 80 per cent. of all consultations were made by people under the age of 45. One-sixth of the patients accounted for about half of the consultations, and 30 per cent. of the consultations were made by the 7 per cent. of registered patients who consulted twelve or more times in the year. The consultation rates showed a slight, but statistically significant, social gradient, with more consultations among patients in the lower social classes. The illness rate was 26 per person, and was higher among males in infancy and old age, and among females between the ages of 5 and 64. A method of estimating the duration of sickness in terms of the period under medical care was employed. This showed that 70 per cent. of all illnesses were dealt with in single consultations, and that only 3 per cent. of illnesses were under care for more than 90 days. Over half of the practice population were under care for less than 5 days, and only 8 per cent. for more than 90 days. The proportion of patients having more than 30 days sickness generally increased with age, but there was a slight fall among the women aged 65 and over. Certificates were issued at the rate of about one for every five consultations, two-thirds of these being necessitated by the requirements of the National Insurance regulations. Prescriptions were issued at the rate of about one per consultation, the prescription rate being 41 per person. Only 5 per cent. of the practice population consulted the doctor but did not obtain a prescription. About 30 per cent. of the patients who consulted a doctor were referred outside the practices, 80 per cent. of all referrals being to hospitals, either as inpatients or as out-patients. The person-hospital referral rate was 20 per 100, and there were 31 referrals for every 100 registered patients. Males over 65 had the highest rates for consultations, illnesses, prescriptions, and referrals. The reason for this is discussed. The value of record-keeping by general practitioners is stressed, together with the need for a generally accepted method of expressing rates in studies of this kind.

Serum antibody response to recombinant major inner capsid protein following human infection with group B rotavirus.

Recombinant major inner capsid protein (VP6) of the IDIR strain of group B rotavirus (GBR) was incorporated in a solid-phase immunoassay to access antibody response to infection in humans. Expression of VP6 in insect cells permitted design of a highly sensitive assay that avoided the contaminants present in GBR antigens obtained from fecal specimens. Among patients infected with the ADRV strain of GBR in China, increased reactivity with recombinant VP6 was observed in convalescent-phase sera in comparison with sera obtained shortly after infection (P = 0.0084). Anti-VP6 antibodies were detectable as soon as 7 days after onset of gastrointestinal symptoms, and serum reactivity persisted in specimens drawn more than 1 year after infection. Solid-phase immunoassay with recombinant VP6 was next employed in order to assess anti-GBR antibody in 513 serum specimens obtained from 423 Maryland residents (ages, 7 months to 96 years; median age, 42 years). Four individuals (< 1%) exhibited serum antibodies directed against the recombinant VP6 (ages, 54 to 95 years; mean age, 77 years). Examination of 129 additional serum specimens including some from other geographic regions of the United States failed to reveal the presence of anti-GBR antibody. Anti-GBR antibody was also not detected in any of 131 serum specimens from 60 staff and residents of a nursing home in Switzerland. While infection of humans with GBR has been uncommon in these locations outside of China, the detection of serum antibodies in older individuals in the United States either indicated an unknown, age-related risk factor or may have indicated infection in the more distant past. The availability of these reagents should allow surveys for GBR infection among additional populations that have not previously been investigated.

Detection of rat, porcine, and bovine group B rotavirus in fecal specimens by solid-phase enzyme immunoassay.

An enzyme immunoassay that uses easily regenerated reagents was developed and evaluated for the ability to detect group B rotaviruses (GBR) in fecal specimens. Homologous rat GBR and heterologous porcine and bovine GBR were detected by this immunoassay, although a human GBR isolate was not. This immunoassay should prove useful in studies of GBR infections of animals.

A multicentre comparison of the efficacy of terbutaline Turbuhaler and salbutamol pressurized metered dose inhaler in hot, humid regions.

Twenty seven adults and 20 children with previously diagnosed stable asthma, using a salbutamol pressurized metered dose inhaler (P-MDI) and living in Cairns, Townsville and Southport, Queensland, Australia participated in a randomized, open-label cross-over comparison of terbutaline administered via Turbuhaler and salbutamol administered via P-MDI. The aim of the study was to compare the clinical effectiveness and patient acceptance of the two treatments in hot, humid regions. Terbutaline was administered via Turbuhaler and salbutamol via P-MDI on at least two occasions per day during each four week treatment period. Spirometry was performed at the start of the study, after the two week run-in and at the end of each treatment period. Patients used diary cards to record morning and evening peak expiratory flows, daily symptom scores and daily intake of beta 2 agonist medication. At the end of the study, patients answered a treatment preference question. Forty six patients completed the study. No statistically significant differences were observed between the two treatments in peak expiratory flow, change in morning peak expiratory flow pre- and post-beta-agonist, daily symptom scores, diurnal variability and spirometry. Forty four percent of patients preferred the terbutaline Turbuhaler and 39% preferred salbutamol P-MDI. Both agents were similarly tolerated. Terbutaline delivered by Turbuhaler is as clinically effective as salbutamol delivered by P-MDI in patients with asthma living in hot, humid regions.

Group B rotavirus VP2: sequence analysis, expression, and gene coding assignment.

The second largest genomic segment of the IDIR strain (infectious diarrhea of infant rats) of group B rotavirus (GBR) was completely sequenced, cloned, and expressed in insect cells, and its gene coding assignment was determined. The sequence of IDIR virus gene 2 (IDIRg2) contained 2847 bp with a single, long open reading frame that encoded a deduced polypeptide of 934 amino acids (M(r) 106 kDa, pI 5.325). BestFit homology indicated that the predicted amino acid sequence of IDIRg2 shared 46.5% similar and 19.8% identical sequences with VP2 of the SA11 strain of group A rotavirus. The polypeptide product encoded by this gene was synthesized in insect cells by means of a baculovirus expression vector and employed to elucidate the corresponding gene to protein coding assignment. Recombinant IDIRg2 product maintained virion antigenic epitopes as evidenced by reactivity with convalescent antisera from infant rat pups infected with the IDIR agent. Reactivity with antisera from heterologous human and porcine GBR strains was also observed. Antibody directed against recombinant IDIRg2 product specifically reacted with VP2 of IDIR virus and a human strain of GBR (ADRV) obtained from fecal specimens. These experiments identified the IDIRg2 product as the VP2 core protein of the IDIR virus strain of GBR.

The nonstructural glycoprotein of rotavirus affects intracellular calcium levels.

Rotavirus infection of monkey kidney cells has been reported to result in a significant increase in the concentration of intracellular calcium. This increase in intracellular calcium was associated with viral protein synthesis and cytopathic effects in infected cells. We tested the effect of individual rotavirus proteins on intracellular calcium concentrations in insect Spodoptera frugiperda (Sf9) cells. Insect cells were infected with wild-type baculovirus or baculovirus recombinants that contained an individual rotavirus gene. The cells were harvested at different times postinfection, and the intracellular calcium concentration was measured by using fura-2 as a fluorescent calcium indicator. We found that the concentration of intracellular calcium was increased nearly fivefold in infected Sf9 cells that expressed the nonstructural glycoprotein (NSP4) of group A rotavirus, and this increase in intracellular calcium concentration coincided with NSP4 expression. A similar result was observed in insect cells expressing NSP4 from a group B rotavirus, suggesting the conservation of this function among rotavirus groups. Expression of the other 10 rotavirus proteins or of wild-type baculovirus proteins in Sf9 cells did not significantly increase intracellular calcium levels. These results suggest that the nonstructural glycoprotein NSP4 is responsible for the increase in cytosolic calcium observed in rotavirus-infected cells.

Identification and expression of the outer capsid protein (VP4) of the IDIR strain of group B rotavirus.

The polypeptide encoded by gene 3 of the IDIR strain of group B rotavirus (GBR) was synthesized by means of a baculovirus expression system. Immunoblot analysis identified the IDIR virus gene 3 product as analogous to the outer capsid protein, VP4, encoded by gene 4 of group A rotaviruses (GAR). This coding assignment had previously been difficult to establish by sequence comparisons, since IDIR virus gene 3 shared < 20% identical amino acid sequences with any GAR protein. Trypsin digestion of the gene 3 protein resulted in the appearance of a product indistinguishable in size from the VP5* outer capsid protein of IDIR agent virions. The recombinant IDIR virus VP4 maintained at least some antigenic epitopes of the native virion protein, as evidenced by reactivity with convalescent antibody obtained following infection of infant rat pups with the IDIR agent. Reactivity was also demonstrated with antisera directed against bovine and porcine isolates of GBR as well as with ADRV, a human strain of GBR.

Baculovirus expression of gene 6 of the IDIR strain of group B rotavirus (GBR): coding assignment of the major inner capsid protein.

The polypeptide product of gene 6 of the IDIR strain of group B rotavirus was synthesized by means of a baculovirus expression system in order to confirm the coding assignment of the gene and to develop reagents broadly reactive with heterologous strains of Group B rotaviruses (GBR). Earlier experiments indicated that IDIR virus gene 6 encoded the group-specific, major inner capsid protein, but direct confirmation of this coding assignment was not previously reported. The expression of IDIR virus gene 6 from baculovirus recombinants resulted in production of a protein with an apparent molecular weight equivalent to that deduced from the gene sequence (44 kDa). In addition, larger recombinant proteins were also observed, and these appeared to be consistent in size with oligomers of the primary VP6 product. The expressed protein reacted with antibody directed against the IDIR agent and other strains of GBR, but no reaction was observed with antibody directed against group A rotavirus. Serologic reactivity was also observed between the gene 6 product and a monoclonal antibody directed against the GBR group-specific antigen. Antibody directed against the recombinant gene 6 product specifically reacted with the IDIR virus major inner capsid protein in an immunoblot format. These experiments conclusively demonstrated that IDIR virus gene 6 encoded the major inner capsid protein and confirmed the presence of group B-specific antigenic epitopes on the protein.

Quantification of cell culture factors affecting recombinant protein yields in baculovirus-infected insect cells.

An experimental study was undertaken to quantify the effects of infection cell density, medium condition, and surface aeration on recombinant protein yields in insect cells. In the absence of surface aeration and fresh medium, insect cells generated higher product yields (on a per cell basis) when infected with recombinant baculovirus at low cell densities, LCD (3 x 10(5)-4 x 10(5) cells/mL), than at high cell densities, HCD (>0.9 x 10(6) cells/mL), for two distinct baculovirus types. Surface aeration of a HCD culture infected in spent medium improved beta-galactosidase yields 5-fold over the nonaerated case. Surface aeration and medium replenishment improved beta-galactosidase yields of a HCD culture by 20-fold (compared to a 1.6-fold improvement for a LCD culture), resulting in cultures with productivties that were independent of the cell density at infection.

Biokinetics of and discrimination between dietary RRR- and SRR-alpha-tocopherols in the male rat.

The net rates of uptake of the natural (2R,4'R,8'R) diastereoisomer of alpha-tocopherol (alpha-T) and the biodiscrimination relative to its 2S-epimer (2S,4'R,8'R) have been measured, in two experiments, for the blood and 21 tissues of male Sprague-Dawley rats fed over a period of several months diets containing deuterium-substituted forms of the alpha-T acetates. Gas chromatography-mass spectrometry was used to measure the amount of deuterated tocopherols taken up relative to the amount of nondeuterated tocopherol remaining. The measurements were performed at different times after the rats, placed for one month on a basal diet containing nondeuterated, natural alpha-T acetate, were switched to a diet containing the same total quantity of deuterated forms of either natural alpha-T acetate or a mixture of the acetates of the 2R- and 2S-epimers (i.e., ambo-alpha-T acetate). In experiment 1 the source of vitamin E in the replacement diet was trideuterio-2R,4'R,8'R-alpha-T acetate. The data obtained provide the first direct measure of the rate at which natural vitamin E is replaced and augmented in the tissues of growing animals under normal laboratory dietary conditions. There are dramatic differences in the tissue kinetics; for example, the apparent half-life of vitamin E, i.e., the time at which the total amount of ingested trideuterio-alpha-T taken up is the same as the amount of nondeuterated alpha-T remaining, varies from ca. 1 wk for the lung to ca. 11 wk for the spinal cord. In experiment 2 the vitamin E in the replacement diet was an equimolar mixture of trideuterio-2S,4'R,8'R- and hexadeuterio-2R,4'R,8'R-alpha-T acetates. The results show that there is a preferential uptake of the natural diastereoisomer of alpha-T by all tissues (except the liver during the first month). Examination of fecal material reveals that the biodiscrimination begins in the gut; the incomplete hydrolysis of the acetates shows clearly that this reaction proceeds to a greater extent with the natural diastereoisomer. The greatest discrimination of all the tissues examined was found to occur in the brain. After five months, the level of the deuterated natural diastereoisomer was more than five times that of the deuterated 2S-epimer. These results have potential implications for human nutrition.

Sodium cromoglycate as a pressurised aerosol (vicrom) in exercise-induced asthma.

We investigated the effect of sodium cromoglycate (SCG), delivered as a pressurised aerosol, in a dose of 2 or 4 mg in 15 patients known to have exercise-induced asthma (EIA). All patients had a fall in peak expiratory flow rate (PEFR) greater than 20% of the pre-exercise level following placebo (Range 21.1 to 74.7%). When an identical exercise test was performed after 2 mg of SCG aerosol, nine of the 15 patients had falls in PEFR less than 50% of the value observed on placebo. Of the remaining six patients, four had greater than 50% protection afforded by 4 mg of SCG. Of the 15 patients, 12 had falls in PEFR less than 11% after either 2 or 4 mg of SCG aerosol. The dose of aerosol SCG required to inhibit EIA is only 10 to 20% of that delivered by conventional spinhaler. The apparent superiority of the pressurised aerosol may be due to its site of deposition in the airways.

A comparison of oral theophylline and oral salbutamol in exercise-induced asthma.

Ten patients with exercise-induced asthma were premedicated with oral salbutamol and oral theophylline on separate occasions before performing a standardised exercise test. Although both agents produced bronchodilatation, reflected by increases in peak expiratory flow rates (PEFR) at rest, they were generally ineffective in preventing an abnormal post-exercise fall in PEFR. Only one patient was afforded protection by salbutamol and another two patients by theophylline.

Inhaled and oral salbutamol in exercise-induced asthma.

Twenty-one asthmatic patients participated in a single blind, crossover study comparing the effects of aerosol and oral salbutamol (a beta-adrenergic bronchodilator), on exercise-induced bronchospasm. By both routes of administration, salbutamol (albuterol) produced significant bronchodilatation at rest, but only the aerosol effectively prevented post-exercise bronchospasm in most patients. Because exercise-induced bronchospasm is a common problem in children and adults with asthma, these observations are relevant to the drug therapy of these patients. The existence of separate mechanisms for bronchodilatation and the prevention of exercise-induced asthma is postulated. It is possible that differing tissue concentrations of salbutamol are responsible for the observed differences between the 2 routes of administration.

A trial of an alpha-adrenoreceptor blocking drug (indoramin) in exercise-induced bronchoconstriction.

In 10 patients with exercise-induced asthma oral indoramin, an alpha adrenoreceptor blocking drug, was ineffective in preventing the post-exercise fall in peak expiratory flow rate (PEFR). The values of PEFR at rest increased after administration of indoramin, providing indirect evidence for the existence of alpha receptors in the bronchial smooth muscle of asthmatic subjects.