Insulin signaling, lifespan and stress resistance are modulated by metabotropic GABA receptors on insulin producing cells in the brain of Drosophila

Insulin-like peptides (ILPs) regulate growth, reproduction, metabolic homeostasis, life span and stress resistance in worms, flies and mammals. A set of insulin producing cells (IPCs) in the Drosophila brain that express three ILPs (DILP2, 3 and 5) have been the main focus of interest in hormonal DILP signaling. Little is, however, known about factors that regulate DILP production and release by these IPCs. Here we show that the IPCs express the metabotropic GABA(B) receptor (GBR), but not the ionotropic GABA(A) receptor subunit RDL. Diminishing the GBR expression on these cells by targeted RNA interference abbreviates life span, decreases metabolic stress resistance and alters carbohydrate and lipid metabolism at stress, but not growth in Drosophila. A direct effect of diminishing GBR on IPCs is an increase in DILP immunofluorescence in these cells, an effect that is accentuated at starvation. Knockdown of irk3, possibly part of a G protein-activated inwardly rectifying K(+) channel that may link to GBRs, phenocopies GBR knockdown in starvation experiments. Our experiments suggest that the GBR is involved in inhibitory control of DILP production and release in adult flies at metabolic stress and that this receptor mediates a GABA signal from brain interneurons that may convey nutritional signals. This is the first demonstration of a neurotransmitter that inhibits insulin signaling in its regulation of metabolism, stress and life span in an invertebrate brain.     

2′ Biaryl amides as novel and subtype selective M1 agonists. Part II: Further optimization and profiling

Further optimization of the biaryl amide series via extensively exploring structure–activity relationships resulted in potent and subtype selective M₁ agonists exemplified by compounds 9a and 9j with good rat PK properties including CNS penetration. Synthesis, structure–activity relationships, subtype selectivity for M₁ over M_2–5, and DMPK properties of these novel compounds are described.     

2′ Biaryl amides as novel and subtype selective M1 agonists. Part I: Identification,synthesis, and initial SAR

Biaryl amides were discovered as novel and subtype selective M₁ muscarinic acetylcholine receptor agonists. The identification, synthesis, and initial structure–activity relationships that led to compounds 3j and 4c, possessing good M₁ agonist potency and intrinsic activity, and subtype selectivity for M₁ over M_2–5, are described.     

Nutrient digestibility and prediction of metabolizable energy in total mixed rations for ruminants

The aim of the present study was to determine equations that predict ME in total mixed rations (TMR) based on routine methods. The ME content of 30 TMR for dairy cows was determined based on digestible crude nutrients obtained with wether sheep. Concentrations in the TMR (in g/kg DM) varied between 118 and 234 for crude protein, 26 and 48 for crude lipid, 131 and 250 for crude fibre, 281 and 488 for NDF, and 173 and 304 for ADF. Gas production ranged from 40.7 to 54.1 ml/200 mg DM, and enzymatically degraded organic matter from 652 to 800 g/kg DM. Digestibility [%] ranged from 68.6 to 84.0 for organic matter, from 55.6 to 84.3 for crude lipid, from 55.0 to 77.8 for crude fibre, from 57.6 to 77.0 for NDF and from 53.1 to 79.6 for ADF. ME ranged from 9.6 to 11.9 MJ/kg DM, and NEL from 5.7 to 7.4 MJ/kg DM. ME content was highly correlated with the concentration of both crude fibre and enzymatically degradable organic matter as well as with organic matter digestibility. A multiple regression equation based on crude fibre and crude lipid predicted ME with a reasonable goodness of fit (r² = 0.81; s_y.x = 2.4%). The inclusion of other nutrients, of neutral and acid detergent fibre, neither of gas production did improve the goodness of fit. The best prediction was achieved with inclusion of enzymatically degraded organic matter (r² = 0.90; s_y.x = 1.7%).     

Effects of Management Tactics on Meeting Conservation Objectives for Western North American Groundfish Fisheries

There is considerable variability in the status of fish populations around the world and a poor understanding of how specific management characteristics affect populations. Overfishing is a major problem in many fisheries, but in some regions the recent tendency has been to exploit stocks at levels below their maximum sustainable yield. In Western North American groundfish fisheries, the status of individual stocks and management systems among regions are highly variable. In this paper, we show the current status of groundfish stocks from Alaska, British Columbia, and the U.S. West Coast, and quantify the influence on stock status of six management tactics often hypothesized to affect groundfish. These tactics are: the use of harvest control rules with estimated biological reference points; seasonal closures; marine reserves; bycatch constraints; individual quotas (i.e., ‘catch shares’); and gear type. Despite the high commercial value of many groundfish and consequent incentives for maintaining stocks at their most productive levels, most stocks were managed extremely conservatively, with current exploitation rates at only 40% of management targets and biomass 33% above target biomass on average. Catches rarely exceeded TACs but on occasion were far below TACs (mean catch:TAC ratio of 57%); approximately $150 million of potential landed value was foregone annually by underutilizing TACs. The use of individual quotas, marine reserves, and harvest control rules with estimated limit reference points had little overall effect on stock status. More valuable fisheries were maintained closer to management targets and were less variable over time than stocks with lower catches or ex-vessel prices. Together these results suggest there is no single effective management measure for meeting conservation objectives; if scientifically established quotas are set and enforced, a variety of means can be used to ensure that exploitation rates and biomass levels are near to or more conservative than management targets.     

The Australian Multiple Sclerosis (MS) Immunotherapy Study: A Prospective,Multicentre Study of Drug Utilisation Using the MSBase Platform

Objective

To prospectively characterise treatment persistence and predictors of treatment discontinuation in an Australian relapsing-remitting multiple sclerosis (RRMS) population.

Methods

Tertiary MS treatment centres participating in the MSBase registry prospectively assessed treatment utilisation, persistence, predictors of treatment discontinuation and switch rates. Multivariable survival analyses were used to compare treatment persistence between drugs and to identify predictors of treatment discontinuation.

Results

1113 RRMS patients were studied. Patients persisted on their first disease-modifying therapy (DMT) for a median of 2.5 years. Treatment persistence on GA was shorter than on all IFNβ products (p<0.03). Younger age at treatment initiation and higher EDSS were predictive of DMT discontinuation. Patients persisted on subsequent DMTs, for 2.3 years. Patients receiving natalizumab (NAT) as a subsequent DMT persisted longer on treatment than those on IFNβ or GA (p<0.000). The primary reason for treatment discontinuation for any drug class was poor tolerability. Annualised switch or cessation rates were 9.5–12.5% for individual IFNβ products, 11.6% for GA and 4.4% for NAT.

Conclusion

This multicentre MS cohort study is the first to directly compare treatment persistence on IFNβ and GA to NAT. We report that treatment persistence in our Australian RRMS population is short, although patients receiving IFNβ as a first DMT persisted longer on treatment than those on GA. Additionally, patients receiving NAT as a subsequent DMT were more likely to persist on treatment than those switched to IFNβ or GA. EDSS and age at DMT initiation were predictive of DMT discontinuation. Treatment intolerance was the principal reason for treatment cessation.     

Pulmonary hypertension in the newborn GTP cyclohydrolase I-deficient mouse

Tetrahydrobiopterin (BH4) is a regulator of endothelial nitric oxide synthase (eNOS) activity. Deficient levels result in eNOS uncoupling, with a shift from nitric oxide to superoxide generation. The hph-1 mutant mouse has deficient GTP cyclohydrolase I (GTPCH1) activity, resulting in low BH4 tissue content. The adult hph-1 mouse has pulmonary hypertension, but whether such condition is present from birth is not known. Thus, we evaluated newborn animals’ pulmonary arterial medial thickness, biopterin content (BH4 + BH2), H₂O₂ and eNOS, right ventricle-to-left ventricle + septum (RV/LV + septum) ratio, near-resistance pulmonary artery agonist-induced force, and endothelium-dependent and -independent relaxation. The lung biopterin content was inversely related to age for both types, but significantly lower in hph-1 mice, compared to wild-type animals. As judged by the RV/LV + septum ratio, newborn hph-1 mice have pulmonary hypertension and, after a 2-week 13% oxygen exposure, the ratios were similar in both types. The pulmonary arterial agonist-induced force was reduced (P < 0.01) in hph-1 animals and no type-dependent difference in endothelium-dependent or -independent vasorelaxation was observed. Compared to wild-type mice, the lung H₂O₂ content was increased, whereas the eNOS expression was decreased (P < 0.01) in hph-1 animals. The pulmonary arterial medial thickness, a surrogate marker of vascular remodeling, was increased (P < 0.01) in hph-1 compared to wild-type mice. In conclusion, our data suggest that pulmonary hypertension is present from birth in the GTPCH1-deficient mice, not as a result of impaired vasodilation, but secondary to vascular remodeling.     

Molecular mechanism of the anti-cancer activity of cerivastatin,an inhibitor of HMG-CoA reductase,on aggressive human breast cancer cells

Statins are currently used for the treatment of hypercholesterolemia. Recently, we demonstrated that cerivastatin also reduces the proliferation and invasion of aggressive breast cancer cells, MDA-MB-231. In this report, a molecular mechanism to explain its anti-cancer action is proposed by combining the study of cerivastatin effect on both gene expression (microarray) and signal transduction pathways. Firstly, the expression of 13 genes was modified by cerivastatin and confirmed at protein level. They could contribute to the inhibition of both cell proliferation (down-regulation of cyclin D1, PCNA, c-myc and up-regulation p21(Waf1), p19(INK4d), integrin beta8) and cell invasion, either directly (decrease in u-PA, MMP-9, u-PAR, PAI-1 and increase in anti-oncogenes Wnt-5a and H-cadherin) or indirectly by stimulating an anti-angiogenic gene (thrombospondin-2). The anti-angiogenic activity was confirmed by in vivo experiments. Secondly, we demonstrated that the biochemical mechanism of its anti-cancer action could be mainly explained by the inhibition of RhoA-dependent cell signalling. This hypothesis was supported by the fact that a RhoA inhibitor (C3 exoenzyme) or a dominant negative mutant RhoA (N19RhoA) induced similar effects to those of cerivastatin. In conclusion, cerivastatin, by preventing RhoA prenylation, inhibits (i) the RhoA/ROCK pathway, leading to defective actin stress fibres formation responsible for the loss of traction forces required for cell motility and (ii) the RhoA/FAK/AKT signalling pathway that could explain the majority of cancer-related gene modifications described above. Thus, the inhibition of RhoA cell signalling could be a good strategy in therapy of aggressive forms of breast cancer.     

Post-translational regulation of the microtubule cytoskeleton: mechanisms and functions

Half a century of biochemical and biophysical experiments has provided attractive models that may explain the diverse functions of microtubules within cells and organisms. However, the notion of functionally distinct microtubule types has not been explored with similar intensity, mostly because mechanisms for generating divergent microtubule species were not yet known. Cells generate distinct microtubule subtypes through expression of different tubulin isotypes and through post-translational modifications, such as detyrosination and further cleavage to Δ2-tubulin, acetylation, polyglutamylation and polyglycylation. The recent discovery of enzymes responsible for many tubulin post-translational modifications has enabled functional studies demonstrating that these post-translational modifications may regulate microtubule functions through an amazing range of mechanisms.