Relatedness of Indian Flax Genotypes (<Emphasis Type="Italic">Linum usitatissimum</Emphasis> L.): An Inter-Simple Sequence Repeat (ISSR) Primer Assay

The objective of this study was to analyze the genetic relationships, using PCR-based ISSR markers, among 70 Indian flax (Linum usitatissimum L.) genotypes actively utilized in flax breeding programs. Twelve ISSR primers were used for the analysis yielding 136 loci, of which 87 were polymorphic. The average number of amplified loci and the average number of polymorphic loci per primer were 11.3 and 7.25, respectively, while the percent loci polymorphism ranged from 11.1 to 81.8 with an average of 63.9 across all the genotypes. The range of polymorphism information content scores was 0.03–0.49, with an average of 0.18. A dendrogram was generated based on the similarity matrix by the Unweighted Pair Group Method with Arithmetic Mean (UPGMA), wherein the flax genotypes were grouped in five clusters. The Jaccard’s similarity coefficient among the genotypes ranged from 0.60 to 0.97. When the omega-3 alpha linolenic acid (ALA) contents of the individual genotypes were correlated with the clusters in the dendrogram, the high ALA containing genotypes were grouped in two clusters. This study identified SLS 50, Ayogi, and Sheetal to be the most diverse genotypes and suggested their use in breeding programs and for developing mapping populations.     

Pervasive and cooperative deadenylation of 3'UTRs by embryonic microRNA families

To understand how miRNA-mediated silencing impacts on embryonic mRNAs, we conducted a functional survey of abundant maternal and zygotic miRNA families in the C. elegans embryo. We show that the miR-35-42 and the miR-51-56 miRNA families define maternal and zygotic miRNA-induced silencing complexes (miRISCs), respectively, that share a large number of components. Using a cell-free C. elegans embryonic extract, we demonstrate that the miRISC directs the rapid deadenylation of reporter mRNAs with natural 3'UTRs. The deadenylated targets are translationally suppressed and remarkably stable. Sampling of the predicted miR-35-42 targets reveals that roughly half are deadenylated in a miRNA-dependent manner, but with each target displaying a distinct efficiency and pattern of deadenylation. Finally, we demonstrate that functional cooperation between distinct miRISCs within 3'UTRs is required to potentiate deadenylation. With this report, we reveal the extensive and direct impact of miRNA-mediated deadenylation on embryonic mRNAs.     

Novel benzothiophene H1-antihistamines for the treatment of insomnia

SAR of lead benzothiophene H₁-antihistamine 2 was explored to identify backup candidates with suitable pharmacokinetic profiles for an insomnia program. Several potent and selective H₁-antihistamines with a range of projected half-lives in humans were identified. Compound 16d had a suitable human half-life as demonstrated in a human microdose study, but variability in pharmacokinetic profile, attributed to metabolic clearance, prevented further development of this compound. Compound 28b demonstrated lower predicted clearance in preclinical studies, and may represent a more suitable backup compound.     

Identification of a novel selective H1-antihistamine with optimized pharmacokinetic properties for clinical evaluation in the treatment of insomnia

Analogs of the known H₁-antihistamine R-dimethindene with suitable selectivity for key GPCRs, P450 enzymes and hERG channel were assessed for metabolism profile and in vivo properties. Several analogs were determined to exhibit diverse metabolism. One of these compounds, 10a, showed equivalent efficacy in a rat EEG/EMG model to a previously identified clinical candidate and a potentially superior pharmacokinetic profile as determined from a human microdose study.     

The habenula prevents helpless behavior in larval zebrafish

Animals quickly learn to avoid predictable danger. However, if pre-exposed to a strong stressor, they do not display avoidance even if this causes continued contact with painful stimuli [1, 2]. In rodents, lesioning the habenula, an epithalamic structure that regulates the monoaminergic system, has been reported to reduce avoidance deficits caused by inescapable shock [3]. This is consistent with findings that inability to overcome a stressor is accompanied by an increase in serotonin levels [4]. However, other studies conclude that habenula lesions cause avoidance deficits [5, 6]. These contradictory results may be caused by lesions affecting unintended regions [6]. To clarify the role of the habenula, we used larval zebrafish, whose transparency and amenability to genetic manipulation enables more precise disruption of cells. We show that larval zebrafish learn to avoid a light that has been paired with a mild shock but fail to do so when pre-exposed to inescapable shock. Photobleaching of habenula afferents expressing the photosensitizer KillerRed causes a similar failure in avoidance. Expression of tetanus toxin in dorsal habenula neurons is sufficient to prevent avoidance. We suggest that this region may signal the ability to control a stressor, and that its disruption could contribute to anxiety disorders.     

Flying Over an Infected Landscape: Distribution of Highly Pathogenic Avian Influenza H5N1 Risk in South Asia and Satellite Tracking of Wild Waterfowl

Highly pathogenic avian influenza (HPAI) H5N1 virus persists in Asia, posing a threat to poultry, wild birds, and humans. Previous work in Southeast Asia demonstrated that HPAI H5N1 risk is related to domestic ducks and people. Other studies discussed the role of migratory birds in the long distance spread of HPAI H5N1. However, the interplay between local persistence and long-distance dispersal has never been studied. We expand previous geospatial risk analysis to include South and Southeast Asia, and integrate the analysis with migration data of satellite-tracked wild waterfowl along the Central Asia flyway. We find that the population of domestic duck is the main factor delineating areas at risk of HPAI H5N1 spread in domestic poultry in South Asia, and that other risk factors, such as human population and chicken density, are associated with HPAI H5N1 risk within those areas. We also find that satellite tracked birds (Ruddy Shelduck and two Bar-headed Geese) reveal a direct spatio-temporal link between the HPAI H5N1 hot-spots identified in India and Bangladesh through our risk model, and the wild bird outbreaks in May-June-July 2009 in China (Qinghai Lake), Mongolia, and Russia. This suggests that the continental-scale dynamics of HPAI H5N1 are structured as a number of persistence areas delineated by domestic ducks, connected by rare transmission through migratory waterfowl.     

Growth kinetics and ginsenosides production in transformed hairy roots of American ginseng—Panax quinquefolium L

A thin, profusely branched, fast growing hairy root line of Panax quinquefolium (American ginseng) was established by co-culturing epicotyl explants with a wild type strain of Agrobacterium rhizogenes. The transformed roots grew by over 10-fold from the initial inoculum within 8 weeks. The crude ginsenosides content in the roots was about 0.2 g/g dry wt level up to the 10th week of culture. Ginsenosides Rb2, Rd, Re, Rf and Rg1 constituted 47–49% of the crude saponin fraction between 6 and 8 weeks of growth whereas, Rc ginsenoside was accumulated only after 9th weeks when the biomass started receding. PCR amplification analysis of the hairy roots confirmed their transgenic nature by showing the presence of Ri-TL DNA with rolA, rolB and rolC genes in their genome.     

PINK1 Is Selectively Stabilized on Impaired Mitochondria to Activate Parkin

Loss-of-function mutations in PINK1 and Parkin cause parkinsonism in humans and mitochondrial dysfunction in model organisms. Parkin is selectively recruited from the cytosol to damaged mitochondria to trigger their autophagy. How Parkin recognizes damaged mitochondria, however, is unknown. Here, we show that expression of PINK1 on individual mitochondria is regulated by voltage-dependent proteolysis to maintain low levels of PINK1 on healthy, polarized mitochondria, while facilitating the rapid accumulation of PINK1 on mitochondria that sustain damage. PINK1 accumulation on mitochondria is both necessary and sufficient for Parkin recruitment to mitochondria, and disease-causing mutations in PINK1 and Parkin disrupt Parkin recruitment and Parkin-induced mitophagy at distinct steps. These findings provide a biochemical explanation for the genetic epistasis between PINK1 and Parkin in Drosophila melanogaster. In addition, they support a novel model for the negative selection of damaged mitochondria, in which PINK1 signals mitochondrial dysfunction to Parkin, and Parkin promotes their elimination.     

Post-Stroke Inhibition of Induced NADPH Oxidase Type 4 Prevents Oxidative Stress and Neurodegeneration

Ischemic stroke is the second leading cause of death worldwide. Only one moderately effective therapy exists, albeit with contraindications that exclude 90% of the patients. This medical need contrasts with a high failure rate of more than 1,000 pre-clinical drug candidates for stroke therapies. Thus, there is a need for translatable mechanisms of neuroprotection and more rigid thresholds of relevance in pre-clinical stroke models. One such candidate mechanism is oxidative stress. However, antioxidant approaches have failed in clinical trials, and the significant sources of oxidative stress in stroke are unknown. We here identify NADPH oxidase type 4 (NOX4) as a major source of oxidative stress and an effective therapeutic target in acute stroke. Upon ischemia, NOX4 was induced in human and mouse brain. Mice deficient in NOX4 (Nox4 ^−/−) of either sex, but not those deficient for NOX1 or NOX2, were largely protected from oxidative stress, blood-brain-barrier leakage, and neuronal apoptosis, after both transient and permanent cerebral ischemia. This effect was independent of age, as elderly mice were equally protected. Restoration of oxidative stress reversed the stroke-protective phenotype in Nox4 ^−/− mice. Application of the only validated low-molecular-weight pharmacological NADPH oxidase inhibitor, VAS2870, several hours after ischemia was as protective as deleting NOX4. The extent of neuroprotection was exceptional, resulting in significantly improved long-term neurological functions and reduced mortality. NOX4 therefore represents a major source of oxidative stress and novel class of drug target for stroke therapy.     

Gender distribution in irritable bowel syndrome is proportional to the severity of constipation relative to diarrhea

Background: A significant gender disparity has been observed in individuals with irritable bowel syndrome (IBS), with females outnumbering males, especially in constipation-predominant IBS (C-IBS). However, this observation is based on Rome criteria categorization and does not take into account the severity of constipation or diarrhea.Objective: In a large prospective data set, gender differences across the severity of constipation and diarrhea were examined in patients with IBS.Methods: Consecutive adult patients with Rome I positive IBS who were referred to a tertiary care medical center (Cedars-Sinai Medical Center, Los Angeles, California) in 1999–2003 were given a questionnaire. The questionnaire asked subjects to rate their constipation and diarrhea according to perceived severity using a scale from 0 (none) to 5 (very severe). C-IBS was determined to be present if the severity of constipation was greater than the severity of diarrhea (sevC > sevD). If sevD > sevC, subjects were considered to have diarrhea-predominant IBS (D-IBS). To further categorize their symptoms, subjects were then grouped by the difference between the severity of constipation and diarrhea, creating a range of values from ?5 to +5. For each of these 11 constipation/diarrhea severity values, the female:male ratio was determined. The severity of constipation to diarrhea was compared by Spearman rank correlation.Results: A total of 429 subjects with IBS (325 women: mean [SD] age, 42.5 [0.8] years; 104 men: mean age, 42.2 [1.7] years) completed the questionnaire. Constipation occurred more frequently in women (79.7%) compared with men (61.5%) (odds ratio [OR] = 2.49; 95% CI, 1.55–4.02). The prevalence of diarrhea was similar between the sexes. Whereas C-IBS was more common in women (31.8%) than in men (26.0%) (OR = 2.03; 95% CI, 1.24–3.30), D-IBS was more prevalent in men (62.5%) than in women (36.3%) (OR = 2.39; 95% CI, 1.53–3.75). When the female:male ratio was evaluated across the 11 severity score categories of constipation and diarrhea, the greater the sevC — sevD score, the higher the proportion of women (R = 0.80; P = 0.003).Conclusion: In this study of patients with IBS, the observation of the association of constipation and gender in IBS is extended to indicate that the female:male ratio significantly increases according to the severity of constipation relative to the severity of diarrhea.