A quantitative bioassay for nerve growth factor,using PC12 clones expressing different levels of trkA receptors

Nerve growth factor (NGF) is a neurotrophin required for differentiation, development, and survival of the sympathetic nervous system, with many of its biological effects being mediated via trkA receptors. There is a need for a standard quantitative bioassay for NGF, to be used in basic research and in pharmaceutical studies. The objective of the present research was to develop a selective, quantitative, and reliable bioassay for NGF, using a morphological criterion: neurite cell outgrowth. In addition, we aimed to apply the aforementioned bioassay to measure NGF administered to mice. Pheochromocytoma PC12 cell variants including wild-type cultures, and a trkA-overexpressing stable transfectant PC12-6.24-I, PC12nnr5, and PC12EN lacking trkA receptors, were used. Dose-response curves were generated with NGF β-subunit (2.5S) purified from mouse submaxillary glands. Our results demonstrated that the bioassay was sensitive to 0.3–20 ng/mL, and selective, as neurite outgrowth was not seen by any other growth factor other than NGF. In addition, variant clones PC12nnr5 and PC12EN, lacking trkA receptors, did not respond to NGF. The bioassay detected NGF in serum of mice injected with NGF. This novel developed bioassay can serve as a model system for various neuroscience purposes.     

The differential preference of scorpion alpha-toxins for insect or mammalian sodium channels: implications for improved insect control.

Receptor site-3 on voltage-gated sodium channels is targeted by a variety of structurally distinct toxins from scorpions, sea anemones, and spiders whose typical action is the inhibition of sodium current inactivation. This site interacts allosterically with other topologically distinct receptors that bind alkaloids, lipophilic polyether toxins, pyrethroids, and site-4 scorpion toxins. These features suggest that design of insecticides with specificity for site-3 might be rewarding due to the positive cooperativity with other toxins or insecticidal agents. Yet, despite the central role of scorpion alpha-toxins in envenomation and their vast use in the study of channel functions, molecular details on site-3 are scarce. Scorpion alpha-toxins vary greatly in preference for sodium channels of insects and mammals, and some of them are highly active on insects. This implies that despite its commonality, receptor site-3 varies on insect vs. mammalian channels, and that elucidation of these differences could potentially be exploited for manipulation of toxin preference. This review provides current perspectives on (i) the classification of scorpion alpha-toxins, (ii) their mode of interaction with sodium channels and pharmacological divergence, (iii) molecular details on their bioactive surfaces and differences associated with preference for channel subtypes, as well as (iv) a summary of the present knowledge about elements involved in constituting receptor site-3. These details, combined with the variations in allosteric interactions between site-3 and the other receptor sites on insect and mammalian sodium channels, may be useful in new strategies of insect control and future design of anti-insect selective ligands.     

Effects of age,period, and cohort on stroke mortality among the Lithuanian urban population during the 24-year follow-up period

The main purpose of this paper was to assess the effects of age, period, and cohort on stroke mortality among the urban Lithuanian population. Routine stroke mortality data among the Lithuanian urban population aged 25–64 years (1041 men and 724 women) between 1980 and 2004 were obtained from the official Kaunas region mortality register and classified by codes 430–438 and 160–169 in the 9^th and 10^th revisions of the International Classifications of Diseases (ICD), respectively. Mortality rates per 100,000 persons for men and women were age-adjusted using the age distribution of the European Standard Population. Goodness of fit of the Poisson regression models was evaluated using the Pearson and Freeman-Tukey residuals. During the study period, mortality rates decreased from 46.8 to 33.0 per 100,000 for men, and from 20.2 to 18.1 per 100,000 for women (average annual decrease of −1.3%, p<0.1 for men, and −1.6%, p<0.03 for women). An age effect was present in both sexes. The definite upward period effect was observed from 1990 to 1994 both among men and women, and was followed by a sharp fall during 2000–2004. Cohort and period effects have contained relevant information that partially explained trends in stroke mortality among 25–64 year-olds in the Lithuanian urban population. The Poisson regression models could be applied for the examination and explanation of the different causes of the population mortality.     

Detection of adrenocorticotropin-secreting pituitary adenomas by magnetic resonance imaging in children and adolescents with cushing disease

CONTEXT: We recently showed that pre- and postcontrast spoiled gradient-recalled acquisition in the steady-state (SPGR) was superior to conventional pre- and postcontrast T-1 weighted spin echo (SE) acquisition magnetic resonance imaging (MRI) for the diagnostic evaluation of pituitary tumors in adult patients. OBJECTIVE: The present investigation assessed the use of SPGR vs. SE-MRI in the diagnostic evaluation of ACTH-secreting tumors in children and adolescents with Cushing disease. DESIGN: Data were analyzed retrospectively from a series of patients seen over 7 yr (1997-2004). SETTING: The setting for this study was a tertiary care referral center. PATIENTS: Thirty children with Cushing disease (13 females and 17 males with a mean age of 12 +/- 3 yr) were studied. INTERVENTIONS AND OUTCOME MEASURES: Imaging results were compared with surgical and pathological findings and the clinical outcome. RESULTS: Twenty-eight patients had microadenomas, and two had macroadenomas; the latter were identified by both MRI techniques. Precontrast SE and SPGR-MRI identified four and six of the microadenomas, respectively. Postcontrast SPGR-MRI identified the location of the tumor in 18 of 28 patients, whereas postcontrast SE-MRI identified the location and accurately estimated the size of the tumor in only five patients (P < 0.001). CONCLUSIONS: We conclude that conventional MRI, even with contrast enhancement, mostly failed to identify ACTH-secreting microadenomas in children and adolescents with Cushing disease. Postcontrast SPGR-MRI was superior to SE-MRI and should be used in addition to conventional SE-MRI in the pituitary evaluation of children and adolescents with suspected Cushing disease.     

Influence of ambient (outdoor) sources on residential indoor and personal PM2.5 concentrations: analyses of RIOPA data

The Relationship of Indoor, Outdoor and Personal Air (RIOPA) study was designed to investigate residential indoor, outdoor and personal exposures to several classes of air pollutants, including volatile organic compounds, carbonyls and fine particles (PM2.5). Samples were collected from summer, 1999 to spring, 2001 in Houston (TX), Los Angeles (CA) and Elizabeth (NJ). Indoor, outdoor and personal PM2.5 samples were collected at 212 nonsmoking residences, 162 of which were sampled twice. Some homes were chosen due to close proximity to ambient sources of one or more target analytes, while others were farther from sources. Median indoor, outdoor and personal PM2.5 mass concentrations for these three sites were 14.4, 15.5 and 31.4 microg/m3, respectively. The contributions of ambient (outdoor) and nonambient sources to indoor and personal concentrations were quantified using a single compartment box model with measured air exchange rate and a random component superposition (RCS) statistical model. The median contribution of ambient sources to indoor PM2.5 concentrations using the mass balance approach was estimated to be 56% for all study homes (63%, 52% and 33% for California, New Jersey and Texas study homes, respectively). Reasonable variations in model assumptions alter median ambient contributions by less than 20%. The mean of the distribution of ambient contributions across study homes agreed well for the mass balance and RCS models, but the distribution was somewhat broader when calculated using the mass balance model with measured air exchange rates.     

The antimicrobial activities of Psidium guajava and Juglans regia leaf extracts to acne-developing organisms

This study aims to present the in vitro inhibitory effect of Psidium guajava and Juglans regia leaf extracts on the main developer of acne lesions, Propionibacterium acnes (P. acnes), and other organisms that are isolated from acne lesions. Thirty-eight subjects (males and females) who had various types of acne were enrolled in the study. The contents of the acne lesions were cultured and the frequency of P. acnes (alone and with Staphylococci spp.) was 47%, whereas the frequencies for Staphylococcus aureus and Staphylococcus epidermidis were 13% and 24%, respectively. The antimicrobial activities of Psidium guajava and Juglans regia leaf extracts, determined by disk diffusion method (zone of inhibition), were compared to tea tree oil (TTO), doxycycline and clindamycin antibiotics. The zones of inhibition due to the Psidium guajava and Juglans regia leaf extracts ranged from 15.8-17.6 mm against P. acnes, 11.3-15.7 mm against S. aureus and 12.9-15.5 mm against S. epidermidis, respectively. These zones of inhibition were significantly higher than those of TTO and equivalent in case of Staphylococci spp., but less in case of P. acnes, to those obtained from doxycycline or clindamycin. It can be concluded that Psidium guajava and Juglans regia leaf extracts may be beneficial in treating acne especially when they are known to have anti-inflammatory activities.     

High-throughput protein expression analysis using tissue microarray technology of a large well-characterised series identifies biologically distinct classes of breast cancer confirming recent cDNA expression analyses

Recent studies on gene molecular profiling using cDNA microarray in a relatively small series of breast cancer have identified biologically distinct groups with apparent clinical and prognostic relevance. The validation of such new taxonomies should be confirmed on larger series of cases prior to acceptance in clinical practice. The development of tissue microarray (TMA) technology provides methodology for high-throughput concomitant analyses of multiple proteins on large numbers of archival tumour samples. In our study, we have used immunohistochemistry techniques applied to TMA preparations of 1,076 cases of invasive breast cancer to study the combined protein expression profiles of a large panel of well-characterized commercially available biomarkers related to epithelial cell lineage, differentiation, hormone and growth factor receptors and gene products known to be altered in some forms of breast cancer. Using hierarchical clustering methodology, 5 groups with distinct patterns of protein expression were identified. A sixth group of only 4 cases was also identified but deemed too small for further detailed assessment. Further analysis of these clusters was performed using multiple layer perceptron (MLP)-artificial neural network (ANN) with a back propagation algorithm to identify key biomarkers driving the membership of each group. We have identified 2 large groups by their expression of luminal epithelial cell phenotypic characteristics, hormone receptors positivity, absence of basal epithelial phenotype characteristics and lack of c-erbB-2 protein overexpression. Two additional groups were characterized by high c-erbB-2 positivity and negative or weak hormone receptors expression but showed differences in MUC1 and E-cadherin expression. The final group was characterized by strong basal epithelial characteristics, p53 positivity, absent hormone receptors and weak to low luminal epithelial cytokeratin expression. In addition, we have identified significant differences between clusters identified in this series with respect to established prognostic factors including tumour grade, size and histologic tumour type as well as differences in patient outcomes. The different protein expression profiles identified in our study confirm the biologic heterogeneity of breast cancer and demonstrate the clinical relevance of classification in this manner. These observations could form the basis of revision of existing traditional classification systems for breast cancer.