Thermodynamic stability of wild-type and mutant p53 core domain

Some 50% of human cancers are associated with mutations in the core domain of the tumor suppressor p53. Many mutations are thought just to destabilize the protein. To assess this and the possibility of rescue, we have set up a system to analyze the stability of the core domain and its mutants. The use of differential scanning calorimetry or spectroscopy to measure its melting temperature leads to irreversible denaturation and aggregation and so is useful as only a qualitative guide to stability. There are excellent two-state denaturation curves on the addition of urea that may be analyzed quantitatively. One Zn²⁺ ion remains tightly bound in the holo-form of p53 throughout the denaturation curve. The stability of wild type is 6.0 kcal (1 kcal = 4.18 kJ)/mol at 25°C and 9.8 kcal/mol at 10°C. The oncogenic mutants R175H, C242S, R248Q, R249S, and R273H are destabilized by 3.0, 2.9, 1.9, 1.9, and 0.4 kcal/mol, respectively. Under certain denaturing conditions, the wild-type domain forms an aggregate that is relatively highly fluorescent at 340 nm on excitation at 280 nm. The destabilized mutants give this fluorescence under milder denaturation conditions.     

VS38: a new monoclonal antibody for detecting plasma cell differentiation in routine sections.

AIMS--To characterise a new mouse monoclonal antibody, VS38, which recognises an intracytoplasmic antigen of 64 kilodaltons present in normal and neoplastic plasma cells; and to establish its value as a diagnostic reagent for routine pathological practice. METHODS--A range of normal and neoplastic tissue sections, both frozen and routinely fixed, were immunostained, using the microwave method of antigen retrieval for routinely fixed specimens. The antibody was also tested on blood and bone marrow specimens and a range of human cell lines. The molecular weight of the antigen recognised by the antibody was obtained by western blot analysis. FACS analysis was used to demonstrate the cellular location of the antigen and its presence on tonsil cell suspensions and myeloma cases. RESULTS--VS38 recognised normal and neoplastic plasma cells in all of the tissues, including all routinely fixed plasma cell neoplasms tested. The antibody also weakly stained epithelial elements within the tissue but was absent from haemopoietic cells of other lineages. CONCLUSION--Antibody VS38 is of potential value in identifying myeloma or plasmacytoma in bone marrow or other tissues. It differentiates lymphoplasmacytoid lymphoma from lymphocytic and follicular lymphoma. It also subdivides large cell lymphomas into two groups which may be a more reliable method of separating these tumours than morphology alone.     

Nasal masks for domiciliary positive pressure ventilation: patient usage and complications.

BACKGROUND--Nasal mask discomfort is a major factor in compliance with treatment by nasal intermittent positive pressure ventilation (NIPPV) and nasal continuous positive airway pressure (CPAP). METHODS--A study of skin complications resulting from mask usage, with particular reference to predisposing factors, was carried out in 66 patients by means of a postal questionnaire. An effective means of managing ulceration at the nasal bridge while continuing therapy is described. RESULTS--Some disruption of treatment due solely to mask discomfort was experienced by 35 patients (53%), consisting of broken skin or open sores in 11 cases (17%). CONCLUSIONS-Although complications resulting from nasal mask usage are common, early identification of patients at risk of developing such complications and appropriate intervention will result in improved patient compliance.     

Antigenuria after immunization with Haemophilus influenzae oligosaccharide CRM197 conjugate (HbOC) vaccine

We tested the urine of 30 infants 6 weeks to 7 months of age after they received standard 10-micrograms (0.5-ml) doses of HbOC (HibTITER) Haemophilus influenzae b (Hib) conjugate vaccine for the presence of Hib antigenuria using a commercially available latex particle agglutination assay (Directigen). Urines were collected within 1 hour, from 1 to 3 hours, at 24 hours and at 3, 6 and 9 days after vaccine administration and reactions were quantitated from 0 to 3+. In contrast to previous studies in older children which showed little or no antigenuria following HbOC vaccination, our study shows that in infants intense Hib antigenuria is evident within 2 to 3 hours and persists 3 days after vaccine administration and that less intense antigenuria may be detected in some infants for several days. With efficacious vaccines now being used in 2- to 6-month-old infants, invasive Hib disease may soon be limited to infants of this age just before their seroconversion. It should be recognized that antigenuria occurs for several days after vaccination with Hib conjugate vaccines and that it could be erroneously interpreted as evidence of invasive Hib infection.     

Influence of the Compliance of the Pump Housing and Cannulas of a Paracorporeal Pneumatic Ventricular Assist Device on Transient Pressure Characteristics

Abstract: The dependence of transient pressure characteristics of a ventricular assist device (VAD) on the compliance of its housing and cannulas was investigated in a mock circulation. The peak rate of change of pressure ( dP/dt _max) values was greater in the cannulas than other compartments and was associated with valve closure-induced pressure oscillations. When cannula compliance was increased from 0.0057 to 0.0129 cm³/mm Hg, these values decreased by ˜20%, and outflow cannula pressure oscillation frequency decreased from 17.5 Hz by 35%. This trend was also apparent in the inflow. A VAD housing compliance increase from 0.0162 to 0.0483 cm³/mm Hg caused a dP/dt _max decrease of 30% in both the blood chamber and the outflow cannula. The effect of this change on the inflow was weaker implying that housing absorbs the energy associated with outflow deceleration more effectively than the inflow. These findings suggest that increasing VAD housing and cannulas compliance can improve hydrodynamic performance.     

Ritanserin in the treatment of alcohol dependence – a multi-center clinical trial

Four hundred and twenty-three alcohol dependent subjects were enrolled into a 12-week randomized, double-blind, placebo-controlled study to determine the safety and efficacy of the 5-HT₂ receptor antagonist, ritanserin (2.5 mg/day or 5 mg/day), in reducing alcohol intake and craving. All subjects received 1 week of single-blind placebo prior to randomization into the 11-week double-blind phase. Additionally, all subjects received weekly individual sessions of manual-guided cognitive-behavioral therapy. Comparing the single-blind period with endpoint, there was approximately a 23% reduction in drinks/day; 34% fall in the total number of drinking days/week; 22% decrease in drinks/drinking day; and a 37% diminution in alcohol craving for all treatment groups. All treatment groups experienced a beneficial clinical outcome as assessed by the Clinical Global Impression Scale. There was, however, no significant difference between treatment groups on any of these measures of alcohol drinking, craving, or clinical outcome. Subjects were of relatively high social functioning at baseline, and this did not change significantly during treatment. Treatment groups did not differ significantly on either medication compliance or reported adverse events. Ritanserin treatment was associated with a dose-related prolongation of subjects’ QTc interval recording on the electrocardiogram. These results suggest that alcohol dependent subjects can show marked clinical improvement within a structured alcohol treatment program. These findings do not support an important role for ritanserin in the treatment of alcohol dependence. Received: 30 April 1996/Final version: 3 July 1996     

Omeprazole: A Comprehensive Review

Omeprazole is a member of a new class of substituted benzimidazoles. These agents inhibit the proton pump in the gastric parietal cell, blocking the final step in the gastric acid secretory pathway. Omeprazole has been investigated for the treatment of gastric ulcer, duodenal ulcer, gastroesophageal reflux, and various hypersecretory states. The prolonged inhibition of gastric acid secretion allows for once-daily dosing in patients with peptic ulcer disease and gastroesophageal reflux, and once- or twice-daily dosing in patients with Zollinger-Ellison syndrome. Compared with currently available therapies, omeprazole is well tolerated and demonstrates a more rapid ulcer healing rate. It is superior to conventional therapies in the treatment of Zollinger-Ellison syndrome. Side effects are infrequent when the drug is used for the short-term management of ulcers.     

The human major histocompatibility complex class Ib molecule HLA-E binds signal sequence-derived peptides with primary anchor residues at positions 2 and 9

Human histocompatibility leukocyte antigen E (HLA-E) and mouse major histocompatibility complex (MHC) class Ib antigen, Qa-1, share the same substitutions at two normally conserved positions 143 and 147, which are likely to affect binding of the C terminus of peptides. Qa-1 is able to bind a peptide derived from the leader sequence of H-2 D and H-2 L molecules. We developed a peptide binding assay in vitro to compare the binding specificity of HLA-E with the mouse MHC class Ib molecule Qa-1. We demonstrate that HLA-E binds, although poorly, the peptide which binds to Qa-1 and that it also binds nonamer signal sequence-derived peptides from human MHC class I molecules. Using alanine and glycine substitutions, we could define primary anchor residues at positions 2 and 9 and secondary anchor residues at position 7 and possibly 3.     

Quantifying how location and dose of botulinum toxin injections affect muscle paralysis

Despite the widespread use of botulinum toxin to treat muscle dystonias, no method exists to quantify muscle paralysis in either human or nonhuman models. In this study we examined how the location, dose, and volume of botulinum injection affects paralysis in the rat tibialis anterior muscle. Paralysis was quantified by electrically stimulating the nerve to the tibialis anterior and then staining sections of the muscle for glycogen. The areas of glycogen-containing fibers represented regions of botulinum action. The results showed that the most important injection technique is to inject botulinum directly into the motor endplate region of a muscle. Injections only 0.5 cm from the motor endplate resulted in a 50% decrease in paralysis. Increases in dose increased paralysis, however, some of that increase was simply due to the increased volume of injection. Thus, delivering toxin in small volumes near the MEP band of a muscle should produce the most effectiveparalysis. © 1993 John Wiley & Sons, Inc.