Principles and practice of HIV-protease inhibitor pharmacoenhancement

Continually maintaining maximally suppressive drug concentrations represents a key defence against the emergence of resistance. If drug levels fall and replication occurs, the opportunity for mutant virus to be selected occurs. It has been increasingly recognized that variability in the pharmacokinetics of antiretrovirals, particularly protease inhibitors (PIs), means that drug exposure is not always optimal, giving the virus a chance to replicate. A significant number of patients receiving PIs two or three times daily will have trough (C_trough or C_min) plasma concentrations, which are close to, or below, the plasma protein binding‐corrected inhibitory concentration (IC₅₀ or IC₉₅) during the dosing interval. It is primarily in this context that therapeutic drug monitoring of PIs has been proposed as an aid to patient management, to ensure that patients maintain adequate drug concentrations throughout the dosing interval. Ideally, an antiretroviral drug will have a pharmacokinetic (PK) profile that maintains drug levels well above the viral inhibitory concentration throughout the entire dosing interval. Beneficial drug–drug interactions have been shown to improve PI pharmacokinetics. Ritonavir (RTV) inhibits the key enzymes that limit the bioavailability or speed the metabolism of other PIs. It is therefore increasingly used for boosting and maintaining PI plasma concentrations. At low (100 mg twice a day) doses it acts as a pharmacoenhancer of indinavir (IDV), amprenavir, saquinavir, lopinavir and to a more limited degree nelfinavir. Using a pharmacoenhancer with a PI results in increased exposure to the PI, higher C_min levels, and in most cases prolonged elimination half‐lives. The long‐term clinical benefits of PK enhancing are unknown as are the long‐term toxicities, although the incidence of nephrolithiasis with IDV appears increased when IDV is combined with low‐dose RTV in HIV‐infected patients. Head‐to‐head clinical comparisons of boosted PI regimens will help answer some of the questions that remain with regard to PK enhancement.     

Elastin point mutations cause an obstructive vascular disease, supravalvular aortic stenosis

Supravalvular aortic stenosis (SVAS) is an inherited obstructive vascular disease that affects the aorta, carotid, coronary and pulmonary arteries. Previous molecular genetic data have led to the hypothesis that SVAS results from mutations in the elastin gene, ELN. In these studies, the disease phenotype was linked to gross DNA rearrangements (35 and 85 kb deletions and a translocation) in three SVAS families. However, gross rearrangements of ELN have not been identified in most cases of autosomal dominant SVAS. To define the spectrum of ELN mutations responsible for this disorder, we refined the genomic structure of human ELN and used this information in mutational analyses. ELN point mutations co-segregate with the disease in four familial cases and are associated with SVAS in three sporadic cases. Two of the mutations are nonsense, one is a single base pair deletion and four are splice site mutations. In one sporadic case, the mutation arose de novo. These data demonstrate that point mutations of ELN cause autosomal dominant SVAS.      

Morphological analysis of degeneration and regeneration of syncytiotrophoblast in first trimester placental villi during organ culture

We have recently shown using dansyl-L-lysine exclusion studies that the release of human chorionic gonadotrophin (HCG) in conjunction with L- lactate dehydrogenase (LDH) from first trimester villi during organ culture is symptomatic of syncytiotrophoblast degeneration. The purpose of this study was to examine chorionic villi at the ultrastructural level in order to determine events occurring during organ culture. The tissue was sampled after 0, 24, 48 and 120 h in culture and processed for electron microscopy. In addition to confirming the previously recorded syncytial degeneration, the electron micrographs showed clearly the generation of a new syncytiotrophoblast layer. The new layer, derived from differentiating cytotrophoblast cells, was largely formed by 48 h and was maintained for at least 120 h in culture. This study demonstrates a model which provides an opportunity to study the differentiation of cytotrophoblast cells whilst they retain their anatomical relationships within the villous structure.      

The Scimitar syndrome: CT findings in partial anomalous pulmonary venous return

An anomalous pulmonary vein draining into the subdiaphragmatic inferior vena cava was initially demonstrated on computed tomographic (CT) scans. The diagnosis of scimitar syndrome was confirmed with digital subtraction angiography. In retrospect, the anomalous vein and dextroposition of the heart were shown on chest radiographs.     

Eosinophil autofluorescence and its use in isolation and analysis of human eosinophils using flow microfluorometry

Unstained human eosinophils exhibit unusually bright autofluorescence, which allows them to be distinguished from other leukocytes using fluorescence microscopy. Eosinophil fluorescence is associated with the cytoplasmic granules of the cells. Eosinophil granule extracts, containing an as-yet-undefined eosinophil fluorescence factor, exhibited excitation maxima at 370 nm and 450 nm, with maximum emission at 520 nm. Eosinophils adhering to opsonized parasites in vitro deposit fluorescent material onto the parasite surface. Eosinophil fluorescence was of sufficient intensity to allow the preparation of viable, highly enriched (greater than or equal to 98%), eosinophil suspensions from peripheral blood of normal and eosinophilic donors using a fluorescence- activated cell sorter. Quantitative studies of eosinophil autofluorescence were performed using flow microfluorometry. Fluorescence intensity of blood eosinophils from normal volunteers and eosinophilic patients varied inversely with the log of the donor's absolute eosinophil count regardless of clinical diagnosis.     

The lipoprotein-associated coagulation inhibitor that inhibits the factor VII-tissue factor complex also inhibits factor Xa: insight into its possible mechanism of action

Blood coagulation is initiated when plasma factor VII(a) binds to its essential cofactor tissue factor (TF) and proteolytically activates factors X and IX. Progressive inhibition of TF activity occurs upon its addition to plasma. This process is reversible and requires the presence of VII(a), catalytically active Xa, Ca2+, and another component that appears to be associated with the lipoproteins in plasma, a lipoprotein-associated coagulation inhibitor (LACI). A protein, LACI(HG2), possessing the same inhibitory properties as LACI, has recently been isolated from the conditioned media of cultured human liver cells (HepG2). Rabbit antisera raised against a synthetic peptide based on the N-terminal sequence of LACI(HG2) and purified IgG from a rabbit immunized with intact LACI(HG2) inhibit the LACI activity in human serum. In a reaction mixture containing VIIa, Xa, Ca2+, and purified LACI(HG2), the apparent half-life (t1/2) for TF activity was 20 seconds. The presence of heparin accelerated the initial rate of inhibition threefold. Antithrombin III alpha alone had no effect, but antithrombin III alpha with heparin abrogated the TF inhibition. LACI(HG2) also inhibited Xa with an apparent t1/2 of 50 seconds. Heparin enhanced the rate of Xa inhibition 2.5-fold, whereas phospholipids and Ca2+ slowed the reaction 2.5-fold. Xa inhibition was demonstrable with both chromogenic substrate (S-2222) and bioassays, but no complex between Xa and LACI(HG2) could be visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Nondenaturing PAGE, however, showed that LACI(HG2) bound to Xa but not to X or Xa inactivated by diisopropyl fluorophosphate. Thus, LACI(HG2) appears to bind to Xa at or near its active site. Bovine factor Xa lacking its gamma-carboxyglutamic acid-containing domain, BXa(-GD), through treatment with alpha-chymotrypsin, was used to further investigate the Xa requirement for VIIa/TF inhibition by LACI(HG2). LACI(HG2) bound to BXa(-GD) and inhibited its catalytic activity against a small molecular substrate (Spectrozyme Xa), though at a rate approximately sevenfold slower than native BXa. Preincubation of LACI(HG2) with saturating concentrations of BXa(-GD) markedly retarded the subsequent inhibition of BXa. The VII(a)/TF complex was not inhibited by LACI(HG2) in the presence of BXa(-GD), and further, preincubation of LACI(HG2) with BXa(-GD) slowed the inhibition of VIIa/TF after the addition of native Xa. The results are consistent with the hypothesis that inhibition of VII(a)/TF involves the formation of a VIIa-TF-XA-LACI complex that requires the GD of XA.(ABSTRACT TRUNCATED AT 400 WORDS)     

Currarino triad: Characteristic appearances on magnetic resonance imaging and plain radiography

The Currarino triad is a complex anomaly consisting of an anorectal malformation, a sacral bone defect and a presacral mass. It was first described in 1981 and since then, approximately 250 cases have been reported. Radiology has an important part to play in the diagnosis of this entity, as the imaging features are characteristic. We report a case of Currarino triad in an infant who presented with intractable constipation and discuss relevant MRI and plain radiography findings.     

Risk factors for deaths in under-age-five children attending a diarrhoea treatment centre

Few case-control studies have examined possible risk factors for diarrhoeal deaths in under-age-five children in the developing countries. We analysed data from the surveillance system of our diarrhoea treatment centre/hospital for the period 1990-94 on 928 children less than 5 years of age. In univariate analysis, 11 factors were significantly associated with death: lack of breastfeeding, severe malnutrition, complicated diarrhoea, pneumonia, xerophthalmia, duration of diarrhoea 7-14 days, moderate or severe dehydration, recent history of measles, Shigella flexneri infection, maternal illiteracy, and very low household income. Rotavirus diarrhoea was negatively associated with fatal outcome. In the assessment of severe malnutrition, weight-for-height measurement discriminated mortality risk better than weight-for-age or height-for-age indices. Only two factors retained their significance, severe malnutrition and non-breastfeeding in the multivariate analysis with adjusted odds ratio (95% confidence interval) of 84.2 (9.1, 775.9) and 4.2 (1.3, 13.2) respectively.     

Short stay facilities: the future of efficient paediatric emergency services

OBJECTIVE: To assess the reactions of parents and their children to the request for a blood sample and an attempt to take blood. METHODS: 1859 children aged 1.5-4.5 years took part in a national survey of diet and nutrition. A retrospective inquiry of the parents' and children's reported reactions was carried out six to 18 months later by postal questionnaire sent only to the 1157 who had given consent for an attempt to take blood. RESULTS: 866 questionnaires were returned; 790 were from parents of children in whom an attempt to take blood had been successful. Thirteen per cent said that their child had given blood previously. About 30% discussed the request with the family doctor or nurse. Some 90% said that they were given enough information and that the phlebotomist was sympathetic. Attempting to take blood caused upset in over 50%, which, in most, lasted for less than five minutes. A substantial minority were upset for up to 30 minutes and a few much longer. Bruising or bleeding occurred in 20-27%. Degree and duration of upset were both adversely associated with a failed attempt to obtain blood. CONCLUSION: The majority of preschool children experienced no more than a little upset of short duration after an attempt to take blood, but a substantial minority exhibited a greater degree of upset. These responses should be taken into account when assessing the benefits and risks of the procedure. The best equipment and expertise should be employed for taking blood as successful attempts are less upsetting.