A simple and inexpensive particle agglutination test to distinguish recent from established HIV-1 infection.

OBJECTIVES: We sought to modify the Serodia HIV-1/HIV-2 particle agglutination assay (PA), a simple and cost-effective HIV assay that is used globally for the detection of HIV antibodies, as a sensitive/less sensitive test (S/LS) to identify recently infected individuals and to estimate HIV incidence. METHODS: The Serodia PA test was modified as an S/LS test (PA-LS) by using HIV antigen-coated gelatin particles at a dilution of 1:68 and a specific diluent, and calibrated using 37 HIV clade B seroconversion panels (309 samples) from Trinidad and from a commercial source that were tested at dilution intervals from 1:10 to 1:80,000. The greatest sensitivity for correctly classifying samples from recent and established infections was determined by receiver operator curve (ROC) analysis. RESULTS: At a 1:40,000 sample dilution and a days post-seroconversion cutoff of 190 days, the PA-LS test yielded a 97% sensitivity for classifying recent and established infection samples. Furthermore, at a 1:20,000 dilution, the positive predictive value for correctly identifying recently infected individuals was 99%. The PA-LS test offers a 30-44-fold cost saving over currently available S/LS tests. CONCLUSION: A modified, low cost and simple-to-perform PA test is appropriate for use in resource-limited countries, and has exhibited excellence in distinguishing recent from established HIV infection.     

Regulation of muscarinic receptors by intrahippocampal injections of gallamine.

Multiple intrahippocampal injections of gallamine impair performance of a representational memory task in rats. The binding of [3H]-(-)-quinuclidinyl benzilate (QNB) to rat brain sections was measured to determine if changes in receptor binding were associated with the deleterious effects of gallamine. [3H]-(-)-QNB binding to sections taken from gallamine-injected animals was compared with binding in saline-injected control animals. Autoradiographic analyses indicated an increase in [3H]-(-)-QNB binding sites within all layers of the cerebral cortex and in the superior colliculus in gallamine-treated animals as compared to saline-injected controls. Significant increases were noted in cortical layers IV and V (P less than 0.025) in gallamine-treated animals. No significant changes (P greater than 0.05) in the number of binding sites were observed in the hippocampus, neostriatum or various thalamic nuclei. The ability of unlabeled pirenzepine, gallamine and carbamylcholine to inhibit 0.2 nM [3H]-(-)-QNB binding also was measured to determine changes in the distribution of receptor subtypes. No significant changes were observed in any brain region for the binding of the selective antagonists pirenzepine and gallamine or the agonist carbamyl-choline. Although other possibilities are considered, the data suggest that an increase in the number of muscarinic receptors may contribute to the observed behavioral deficits associated with long-term gallamine treatment.     

Toxicity, inflammation, and anti-human immunodeficiency virus type 1 activity following exposure to chemical moieties of C31G.

C31G, which has potent activity against the human immunodeficiency virus type 1 (HIV-1) and an established record of safety in animal studies and human trials, is a microbicidal agent comprised of a buffered equimolar mixture of two amphoteric, surface-active agents: an alkyl amine oxide (C14AO) and an alkyl betaine (C16B). Studies of long-term in vitro exposure to C31G and its constituents have suggested that the components of C31G may contribute differentially to its toxicity and efficacy. In the present studies, in vitro assays of cytotoxicity and anti-HIV-1 activity demonstrated that C16B was slightly less cytotoxic compared to either C31G or C14AO, whereas the anti-HIV-1 activities of C31G and its individual constituents were similar. In the murine model of cervicovaginal microbicide toxicity, in vivo exposure to C14AO resulted in severe cervical inflammation followed by a delayed disruption of the columnar epithelium. In contrast, exposure to C16B caused severe cervical epithelial disruption and a secondary, less intense inflammatory response. These results demonstrate that (i) there are both mechanistic and temporal differences in toxicity associated with the components of C31G not necessarily predicted by in vitro assessments of cytotoxicity and (ii) contributions of each component to the anti-HIV-1 activity of C31G appear to be equal. In addition, these findings indicate that direct and indirect mechanisms of in vivo toxicity can be observed as separate but interrelated events. These results provide further insight into the activity of C31G, as well as mechanisms potentially associated with microbicide toxicity.     

Mikrofrakturierung bei fokalem Knorpelschaden

The microfracture technique and spongiolization using Pridie drilling are the most commonly used and highly cost-efficient operative therapies for restoration of cartilage in Germany. Microfracturing can be performed arthroscopically in various joints including the knee, ankle, elbow, and shoulder joint. Specifically designed instruments are recommended for this procedure. Steadman recently reported excellent results in a series of 72 patients undergoing microfracturing with a mean follow-up of 11 years showing consistent and long-term reduction of weight-bearing pain and swelling as well as improvement of daily activities. Ideal indications for this easy arthroscopic procedure of microfracture treatment include focal lesions up to 4 cm². Some studies have demonstrated that microfracturing results not merely in the formation of fibrocartilage but also in the induction of hyaline cartilage with superior properties.     

Outcome of 60 neonates who had ARED flow prenatally compared with a matched control group of appropriate-for-gestational age preterm neonates.

OBJECTIVE: To describe the course and outcome of fetuses with absent or reversed end-diastolic (ARED) flow in the umbilical artery (UA) and to examine the influence of prematurity according to gestational age at delivery. METHODS: Sixty pregnancies complicated by ARED flow in the UA were monitored by repeat Doppler measurements of arterial and venous vessels, non-stress tests (cardiotocogram (CTG)) and maternal investigations, and were delivered between 24 and 34 weeks. Fetal outcome was investigated and compared to a control group of appropriate-for-gestational age (AGA) preterm neonates, matched for gestational age. Mortality, birth weight, Apgar scores, postnatal cord arterial pH and need for ventilation were all recorded, as were cases of respiratory distress syndrome, bronchopulmonary dysplasia, persistent ductus arteriosus, necrotizing enterocolitis, intraventricular hemorrhage, periventricular leukomalacia, abnormal neurological findings and those requiring surgical intervention. Additionally, the group of fetuses with ARED flow was divided into three subgroups of different degrees of prematurity (delivery between 24 + 0 and 28 + 6 weeks, delivery between 29 + 0 and 31 + 6 weeks, and delivery after 32 weeks) and compared according to the above parameters. RESULTS: Pre- or postnatal death occurred in 16 cases. Comparing the 44 (61%) that were born alive with the AGA neonates, significant differences were found in birth weight (P < 0.001), arterial pH value (P < 0.001), bronchopulmonary dysplasia (P = 0.002) and intestinal complications (P < 0.01). Prematurity-related complications were: need for ventilation (P = 0.001), respiratory distress syndrome (P < 0.0001), periventricular leukomalacia (P = 0.002) and pathological neurological testing (P = 0.005). CONCLUSIONS: Neonates displaying ARED flow before birth are growth restricted, acidemic at delivery and are at high risk of developing bronchopulmonary dysplasia and intestinal complications. While perinatal mortality seems to be related to abnormal fetal Doppler velocimetry, age at delivery has a significant impact on short-term morbidity. After 32 weeks, morbidity is low and delivery should be considered. It could be speculated from our data that prolongation of pregnancy with Doppler velocimetry monitoring could help to reduce morbidity, although prolongation remains limited in most cases.     

Quantitatively distinct requirements for signaling-competent cell spreading on engineered versus natural adhesion ligands.

To design synthetic microenvironments that elicit desired cell behaviors, we must better understand the molecular mechanisms by which cells interact with candidate biomaterials. Using cell lines with distinct alpha5beta1 integrin expression profiles, we demonstrate that this integrin mediates cell spreading on substrata coated with genetically engineered artificial extracellular matrix (aECM) proteins containing the RGD sequence (RGD-containing aECM protein [aRGD]) but lacking the PHSRN synergy site. Furthermore, aRGD-mediated adhesion stimulates an intracellular focal adhesion kinase (FAK) signal that is indicative of integrin tethering. Although both aRGD and the natural ECM protein fibronectin (FN) support alpha5beta1 integrin-mediated cell spreading, quantitative single-cell analysis revealed that aRGD-mediated spreading requires ten-fold greater threshold amount of integrin expression than FN-mediated spreading. Our analysis demonstrates that aRGD-based substrata mediate both biophysical (cell spreading) and biochemical (FAK signaling) events via the alpha5beta1 integrin, albeit with efficacy quantitatively distinct from that of natural ECM proteins that possess the full spectrum of adhesion and synergy domains.     

The influence of formulation components on the aerosolisation properties of spray-dried powders.

Dry powders suitable for inhalation containing beta-estradiol, leucine as a dispersibility enhancer and lactose as a bulking agent were prepared by spray-drying from aqueous ethanol formulations. The influence of formulation components on the characteristics of the resultant spray-dried powders was examined through the use of a range of ethanol concentrations (10-50% v/v) in the solvent used to prepare the initial formulations. Additionally, the amount of leucine required to act as a dispersibility enhancer was investigated by varying the amount of leucine added to the formulation prior to spray-drying. Following spray-drying, resultant powders were characterised using scanning electron microscopy, laser diffraction and tapped density measurements, and the aerosolisation performance determined using Twin Stage Impinger and Andersen Cascade Impactor analysis. We demonstrate that selection of appropriate solvent systems and leucine concentration allows the preparation of spray-dried powders that display enhanced aerosolisation properties, and would be predicted to exhibit high deposition in the lower regions of the respiratory tract.