An efficient regeneration system via somatic embryogenesis in mango ginger (Curcuma amada Roxb.)

A reproducible protocol for somatic embryogenesis was established for mango ginger (Curcuma amada Roxb.)—an important horticultural aromatic rhizomatous plant. Embryogenic callus induction was obtained from leaf sheath explants of in vitro raised plants on Murashige and Skoog (MS) agar medium containing 2.0 mg/L 2,4-dichlorophenoxyacetic acid and 0.5 mg/L 6-benzyladenine (BA). Embryogenic callus proliferation, somatic embryo (SE) formation and subsequent plantlet conversion occurred under optimal culture conditions. The effects of MS medium strength, sucrose and BA on SE formation were also evaluated. Half strength MS liquid medium necessary for SE formation and optimal sucrose concentration was found to be 3.0 %. BA at 0.3 mg/L produced the highest number (84.71 %) of SEs from leaf sheath explants. Secondary somatic embryos originated from primary somatic embryos on the same medium supplemented with 0.4–0.6 mg/L BA. Stereo microscopic and scanning electron microscopic observation revealed that the globular and torpedo shaped somatic embryos resulted in suspension culture during development. Mature somatic embryos germinated readily and developed into normal plantlets after 3 weeks on half strength MS basal agar medium under dark condition. Well rooted plantlets were successfully acclimatized at the survival rate of 70 %.     

Vitamin E succinate inhibits survivin and induces apoptosis in pancreatic cancer cells

Pancreatic cancer is the fourth leading cause of cancer-related deaths in the United States. Identifying novel chemotherapeutic and chemopreventive approaches is critical in the prevention and treatment of cancers such as pancreatic cancer. Vitamin E succinate (VES) is a redox-silent analog of the fat-soluble vitamin alpha-tocopherol. In the present study, we explored the antiproliferative action of VES and its effects on inhibitor of apoptosis proteins in pancreatic cancer cells. We show that VES inhibits cell proliferation and induces apoptosis in pancreatic cancer cells. Further, we demonstrate that VES downregulates the expression of survivin and X-linked inhibitor of apoptosis proteins. The apoptosis induced by VES was augmented by siRNA-mediated inhibition of survivin in PANC-1 cells. In summary, our results suggest that VES targets survivin signaling and induces apoptosis in pancreatic cancer cells.     

Nitric oxide-dependent Src activation and resultant caveolin-1 phosphorylation promote eNOS/caveolin-1 binding and eNOS inhibition

Endothelial nitric oxide synthase (eNOS)-mediated NO production plays a critical role in the regulation of vascular function and pathophysiology. Caveolin-1 (Cav-1) binding to eNOS holds eNOS in an inactive conformation; however, the mechanism of Cav-1-mediated inhibition of activated eNOS is unclear. Here the role of Src-dependent Cav-1 phosphorylation in eNOS negative feedback regulation is investigated. Using fluorescence resonance energy transfer (FRET) and coimmunoprecipitation analyses, we observed increased interaction between eNOS and Cav-1 following stimulation of endothelial cells with thrombin, vascular endothelial growth factor, and Ca(2+) ionophore A23187, which is corroborated in isolated perfused mouse lung. The eNOS/Cav-1 interaction is blocked by eNOS inhibitor L-N(G)-nitroarginine methyl ester (hydrochloride) and Src kinase inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazolo [3, 4-d] pyrimidine. We also observe increased binding of phosphomimicking Y14D-Cav-1 mutant transduced in human embryonic kidney cells overexpressing eNOS and reduced Ca(2+)-induced NO production compared to cells expressing the phosphodefective Y14F-Cav-1 mutant. Finally, Src FRET biosensor, eNOS small interfering RNA, and NO donor studies demonstrate NO-induced Src activation and Cav-1 phosphorylation at Tyr-14, resulting in increased eNOS/Cav-1 interaction and inhibition of eNOS activity. Taken together, these data suggest that activation of eNOS promotes Src-dependent Cav-1-Tyr-14 phosphorylation and eNOS/Cav-1 binding, that is, eNOS feedback inhibition.     

Ecological Flexibility as Measured by the Use of Pioneer and Exotic Plants by Two Lemurids: <Emphasis Type="BoldItalic">Eulemur collaris</Emphasis> and <Emphasis Type="BoldItalic">Hapalemur meridionalis</Emphasis>

Primate responses to habitat alteration vary depending on the species’ dietary guild and forest type. Leaves from secondary vegetation can provide nutritious resources to folivorous primates, whereas frugivores, burdened with a scattered spatial and temporal distribution of fruiting resources, require larger home ranges, potentially limiting their ability to cope with altered landscapes. Within coastal southeastern Madagascar, we sought to determine whether two lemur species occupying contrasting ecological niches respond differently to the changing features of their degraded and fragmented habitat. We conducted behavioral observations between 2011 and 2013 on frugivorous collared brown lemurs (Eulemur collaris) and folivorous southern bamboo lemurs (Hapalemur meridionalis). To estimate the ability of lemurs to use pioneer species, we categorized all plants used for feeding and resting as fast growing, mid-growing, or slow growing. We fitted general linear mixed-effects models, one for each plant growth category with monthly proportional use rates as the dependent variable, and included species (E. collaris and H. meridionalis), activity (feeding and resting), and season (dry and wet) as fixed effects. Our results show that E. collaris used both slow- and mid-growing plant species most often, while H. meridionalis were more likely to use fast-growing plants, which indicated an ability to use secondary/disturbed vegetation. Frugivorous E. collaris appear more limited by climax plants, while folivorous H. meridionalis appear to be slightly more adaptable, a finding that is consistent with that for other primate folivores.     

Isolation and characterisation of Isaria farinosa and Purpureocillium lilacinum associated with emerald ash borer,Agrilus planipennis in Canada

Entomopathogenic fungi of the genera Isaria and Purpureocillium were recovered from infestation sites of emerald ash borer (EAB) in Southern Ontario, Canada. Isolates were identified using morphological characters and by sequencing the ITS1-5.8S-ITS2 ribosomal DNA gene and partial β-tubulin gene. Phylogenetic analysis and constructed trees based on the ITS and β-tubulin gene explicitly confirm isolates L66B, SY17-a and LHY46-a as Isaria farinosa and B3A, B59A and SY45B-a as Purpureocillium lilacinum. Pathogenicity was tested in the laboratory against adult EAB using a single concentration (2.0×10⁷ conidia/ml) applied topically to adults. Controls included three commercial isolates: Isaria fumosorosea LRC176, Metarhizium brunneum LRC187 and Beauveria bassiana strain GHA. The native isolates I. farinosa L66B and P. lilacinum SY45B-a killed 75 and 50% of the beetles 14 days post-inoculation. Although these indigenous entomogenous fungi were less virulent compared with the commercial isolates, yearly isolation from EAB populations suggests they are one of the natural mortality factors of EAB in Canada.     

Effects of sex steroids and their inhibitors on endocrine parameters and gender growth differences in Eurasian perch (Perca fluviatilis) juveniles

Sex steroid inhibitors were used to characterize the effects of 17beta-estradiol (E2) and testosterone (T) on the sexual growth dimorphism of Eurasian perch juveniles. In experiment 1, growth responses to different doses of either E2 (25, 50, 75, and 100 mgkg(diet)-1) or fadrozole (Fa; 50 and 100 mgkg(diet)-1) were compared in triplicate tanks of 30 fish each during 85 days. In experiment 2, five diets containing (50 mgkg(diet)-1) Tamoxifen (Ta), Flutamide (Flu), Fa, E2, and T were tested in triplicate tanks of 20 fish each during 90 days. Steroid supplementation or inhibition increased or decreased E2 and T plasma levels. Moreover, E2 treatment induced a higher plasma vitellogenin level but decreased triidothyronine levels. Brain aromatase activity (AA) was lower in Fa-treated fish than in other groups. In experiment 1, E2 supplementation did not promote growth, but high doses had negative effects as did Fa. In experiment 2, a greater growth response was observed only in E2-treated females in relation to higher food intake (FI) not feeding efficiency. Fa also promoted growth and FI both in females and males during the last month of the experiment. Other treatments did not affect growth, but T treatment decreased FI in males. In conclusion, the results did not provide clear evidence for E2 action on sexual growth dimorphism, but showed that testosterone may decrease growth in males by decreasing food intake in Eurasian perch. Therefore, the acceleration of male-to-female growth differences with age may not be a result of promotion of growth in females by estrogens, but a consequence of a reduction in growth by increased secretion of androgens in males.     

ETB receptor dependent alteration in aortic responses to ET-1 in the cardiomyopathic hamster

The aim of this study was to verify whether an alteration in the aortic endothelin-1 (ET-1) response takes place in UM-X7.1 cardiomyopathic hamsters. Our results showed that ET-1 (10(-12) - 10(-5) mol/L) induces dose-dependent sustained increases in tension in the intact and endothelium denuded aortas from both normal and cardiomyopathic hamsters. The EC50 values of ET-1 of both intact and endothelium denuded aortas of normal hamsters were similar (2.2 x 10(-9) mol/L and 1.8 x 10(-9) mol/L, respectively). However, in cardiomyopathic hamsters, the EC50 of ET-1 in intact aortas was higher (1.5 x 10(-8) mol/L) than that of the endothelium denuded preparations (2.7 x 10(-9) mol/L). The EC50 of ET-1 in normal and cardiomyopathic hamster denuded aortas were similar. However, the EC50 of ET-1 in intact aortas of cardiomyopathic hamster was higher (1.5 x 10(-8) mol/L) than that of normal hamsters (2.2 x 10(-9) mol/L). Pre-treatment with the ETA receptor antagonist ABT-627 (10(-5)mol/L) of intact and endothelium denuded aortas from both normal and cardiomyopathic hamsters significantly prevented ET-1 (10(-7) mol/L) from inducing an increase in tension. Pre-treatment with the ETB receptor antagonist A-192621 (10(-5) mol/L) had no effect on the ET-1-induced increase in tension in endothelium denuded aortas of both normal and cardiomyopathic hamsters, as well as in intact preparations of normal animals. However, blockade of the ETB receptors in intact aortas of cardiomyopathic hamsters significantly (p < 0.001) potentiated the ET-1-induced increase in tension. In summary, an attenuation of the contraction response to ET-1 was found in UM-X7.1 cardiomyopathic hamsters when compared with normal age-matched hamsters. This alteration of the ET-1 effect in the aortas of cardiomyopathic hamsters seems to be dependent on the presence of the endothelium and could be due, in part, to an increase in the contribution of endothelial ETB receptors to relaxation, which in turn acts as a physiological depressor of ET-1 vasoconstriction. Our results suggest that an increase in the endothelium ETB receptor density may play a role in the development of hypotension in UM-X7.1 cardiomyopathic hamsters.     

Mutations in MAP3K1 cause 46,XY disorders of sex development and implicate a common signal transduction pathway in human testis determination

Investigations of humans with disorders of sex development (DSDs) resulted in the discovery of many of the now-known mammalian sex-determining genes, including SRY, RSPO1, SOX9, NR5A1, WT1, NR0B1, and WNT4. Here, the locus for an autosomal sex-determining gene was mapped via linkage analysis in two families with 46,XY DSD to the long arm of chromosome 5 with a combined, multipoint parametric LOD score of 6.21. A splice-acceptor mutation (c.634-8T>A) in MAP3K1 segregated with the phenotype in the first family and disrupted RNA splicing. Mutations were demonstrated in the second family (p.Gly616Arg) and in two of 11 sporadic cases (p.Leu189Pro, p.Leu189Arg)-18% prevalence in this cohort of sporadic cases. In cultured primary lymphoblastoid cells from family 1 and the two sporadic cases, these mutations altered the phosphorylation of the downstream targets, p38 and ERK1/2, and enhanced binding of RHOA to the MAP3K1 complex. Map3k1 within the syntenic region was expressed in the embryonic mouse gonad prior to, and after, sex determination. Thus, mutations in MAP3K1 that result in 46,XY DSD with partial or complete gonadal dysgenesis implicate this pathway in normal human sex determination.