Endothelial nitric oxide gene polymorphisms,nitric oxide production and coronary artery disease risk in a South Indian population

Nitric oxide (NO) synthesized by vascular endothelial cells, is a vasodilator agent produced from endothelial NO synthase (eNOS). It has been reported that decreased bioavailability of NO plays an important role in the development and progression of atherosclerosis.Electrocardiographically proven 100 patients with acute myocardial infarction and 100 age and sex matched healthy individuals with normal coronary arteries were included for the study. The genotypes of a 27-bp insertion/deletion in intron 4 (eNOS 4b/4a) and G894T polymorphism in exon 7, were determined by PCR analysis based on the banding pattern on gel electrophoresis. The genotype frequencies were calculated following the Hardy–Weinberg law. Serum NO level was also estimated by the Griess method. NO levels in AMI patients were higher than those of the healthy subjects (median [interquartile range], (14.36[12.42–15.78]) μM compared with 11.28[10.32–11.89]) μM; p < 0.001; Mann–Whitney rank sum test, U = 285. Mutant “T” allele frequency of the eNOS-G894T polymorphism was found to be comparatively higher (0.29) in AMI patients than among the controls (0.17). The calculated Odds ratio showed that the occurrence of mutant allele “T” was 1.6 fold as frequent in cases than controls [OR = 1.6 (95%CI 0.898 to 2.833)].To conclude, in the present study, (i) NO levels were found to be increased in patients than in controls, (ii) the homozygous mutant (TT) genotype confers genetic susceptibility to coronary artery disease (iii) both the eNOS 4a/b and G894T polymorphisms were not associated with serum NO levels in a South Indian Tamil population.     

A primary current distribution model of a novel micro-electroporation channel configuration

Traditional macro and micro-electroporation devices utilize facing electrodes, which generate electric fields inversely proportional to their separation distance. Although the separation distances in micro-electroporation devices are significantly smaller than those in macro-electroporation devices, they are limited by cell size. Because of this, significant potential differences are required to induce electroporation. These potential differences are often large enough to cause water electrolysis, resulting in electrode depletion and bubble formation, both of which adversely affect the electroporation process. Here, we present a theoretical study of a novel micro-electroporation channel composed of an electrolyte flowing over a series of adjacent electrodes separated by infinitesimally small insulators. Application of a small, non-electrolysis inducing potential difference between the adjacent electrodes results in radially-varying electric fields that emanate from these insulators, causing cells flowing through the channel to experience a pulsed electric field. This eliminates the need for a pulse generator, making a minimal power source (such as a battery) the only electrical equipment that is needed. A non-dimensional primary current distribution model of the novel micro-electroporation channel shows that decreasing the channel height results in an exponential increase in the electric field magnitude, and that cells experience exponentially greater electric field magnitudes the closer they are to the channel walls. Finally, dimensional primary current distribution models of two potential applications, water sterilization and cell transfection, demonstrate the practical feasibility of the novel micro-electroporation channel.     

Hearing loss alters serotonergic modulation of intrinsic excitability in auditory cortex

Sensorineural hearing loss during early childhood alters auditory cortical evoked potentials in humans and profoundly changes auditory processing in hearing-impaired animals. Multiple mechanisms underlie the early postnatal establishment of cortical circuits, but one important set of developmental mechanisms relies on the neuromodulator serotonin (5-hydroxytryptamine [5-HT]). On the other hand, early sensory activity may also regulate the establishment of adultlike 5-HT receptor expression and function. We examined the role of 5-HT in auditory cortex by first investigating how 5-HT neurotransmission and 5-HT(2) receptors influence the intrinsic excitability of layer II/III pyramidal neurons in brain slices of primary auditory cortex (A1). A brief application of 5-HT (50 μM) transiently and reversibly decreased firing rates, input resistance, and spike rate adaptation in normal postnatal day 12 (P12) to P21 rats. Compared with sham-operated animals, cochlear ablation increased excitability at P12-P21, but all the effects of 5-HT, except for the decrease in adaptation, were eliminated in both sham-operated and cochlear-ablated rats. At P30-P35, cochlear ablation did not increase intrinsic excitability compared with shams, but it did prevent a pronounced decrease in excitability that appeared 10 min after 5-HT application. We also tested whether the effects on excitability were mediated by 5-HT(2) receptors. In the presence of the 5-HT(2)-receptor antagonist, ketanserin, 5-HT significantly decreased excitability compared with 5-HT or ketanserin alone in both sham-operated and cochlear-ablated P12-P21 rats. However, at P30-P35, ketanserin had no effect in sham-operated and only a modest effect cochlear-ablated animals. The 5-HT(2)-specific agonist 5-methoxy-N,N-dimethyltryptamine also had no effect at P12-P21. These results suggest that 5-HT likely regulates pyramidal cell excitability via multiple receptor subtypes with opposing effects. These data also show that early sensorineural hearing loss affects the ability of 5-HT receptor activation to modulate A1 pyramidal cell excitability.     

SELDI-TOF serum profiling for prognostic and diagnostic classification of breast cancers

Surface enhanced laser desorption/ionization (SELDI) time-of-flight mass spectrometry has emerged as a successful tool for serum based detection and differentiation of many cancer types, including breast cancers. In this study, we have applied the SELDI technology to evaluate three potential applications that could extend the effectiveness of established procedures and biomarkers used for prognostication of breast cancers. Paired serum samples obtained from women with breast cancers prior to surgery and post-surgery (6-9 mos.) were examined. In 14/16 post-treatment patients, serum protein profiles could be used to distinguish these samples from the pre-treatment cancer samples. When compared to serum samples from normal healthy women, 11 of these post-treatment samples retained global protein profiles not found in healthy women, including five low-mass proteins that remained elevated in both pre-treatment and post-treatment serum groups. In another pilot study, serum profiles were compared for a group of 30 women who were known BRCA-1 mutation carriers, half of whom subsequently developed breast cancer within three years of the sample procurement. SELDI protein profiling accurately classified 13/15 women with BRCA-1 breast cancers from the 15 non-cancer BRCA-1 carriers. Additionally, the ability of SELDI to distinguish between the serum profiles from sentinel lymph node positive and sentinel lymph node negative patients was evaluated. In sentinel lymph node positive samples, 22/27 samples were correctly classified, in comparison to the correct classification of 55/71 sentinel lymph node negative samples. These initial results indicate the utility of protein profiling approaches for developing new diagnostic and prognostic assays for breast cancers.     

Diagnosis of Ovarian Cancer Using Decision Tree Classification of Mass Spectral Data

Recent reports from our laboratory and others support the SELDI ProteinChip technology as a potential clinical diagnostic tool when combined with $n$ -dimensional analyses algorithms. The objective of this study was to determine if the commercially available classification algorithm biomarker patterns software (BPS), which is based on a classification and regression tree (CART), would be effective in discriminating ovarian cancer from benign diseases and healthy controls. Serum protein mass spectrum profiles from 139 patients with either ovarian cancer, benign pelvic diseases, or healthy women were analyzed using the BPS software. A decision tree, using five protein peaks resulted in an accuracy of 81.5% in the cross-validation analysis and 80%in a blinded set of samples in differentiating the ovarian cancer from the control groups. The potential, advantages, and drawbacks of the BPS system as a bioinformatic tool for the analysis of the SELDI high-dimensional proteomic data are discussed.     

Control of dendritic cell cross-presentation by the major histocompatibility complex class I cytoplasmic domain

Dendritic cells (DCs) can present extracellularly derived antigens in the context of major histocompatibility complex (MHC) class I molecules, a process called cross-presentation. Although recognized to be important for priming of T cell responses to many viral, bacterial and tumor antigens, the mechanistic details of this alternative antigen-presentation pathway are poorly understood. We demonstrate here the existence of an endolysosomal compartment in DCs where exogenously derived peptides can be acquired for presentation to T cells, and show that the MHC class I cytoplasmic domain contains a tyrosine-based targeting signal required for routing MHC class I molecules through these compartments. We also report that transgenic mice expressing H-2K(b) with a tyrosine mutation mount inferior H-2K(b)-restricted cytotoxic T lymphocyte responses against two immunodominant viral epitopes, providing evidence of a crucial function for cross-priming in antiviral immunity.     

PenBase, the shrimp antimicrobial peptide penaeidin database: sequence-based classification and recommended nomenclature

Antimicrobial peptides play a major role in innate immunity. The penaeidins, initially characterized from the shrimp Litopenaeus vannamei, are a family of antimicrobial peptides that appear to be expressed in all penaeid shrimps. As of recent, a large number of penaeid nucleotide sequences have been identified from a variety of penaeid shrimp species and these sequences currently reside in several databases under unique identifiers with no nomenclatural continuity. To facilitate research in this field and avoid potential confusion due to a diverse number of nomenclatural designations, we have made a systematic effort to collect, analyse, and classify all the penaeidin sequences available in every database. We have identified a common penaeidin signature and subsequently established a classification based on amino acid sequences. In order to clarify the naming process, we have introduced a 'penaeidin nomenclature' that can be applied to all extant and future penaeidins. A specialized database, PenBase, which is freely available at , has been developed for the penaeidin family of antimicrobial peptides, to provide comprehensive information about their properties, diversity and nomenclature.