Effect of type of intravaginal progestagen treatment of estrous response and reproductive performance of ewes

A comparison was made of the relative effectiveness of sponge pessaries impregnated with 40mg flourogestone acetate (FGA) or 60mg medroxyprogesterone acetate (MAP) to induce a synchronized estrus in ewes. Ewes were treated with sponge pessaries for 14 days and 500 IU pregnant mares' serum gonadotropin was injected i.m. at the time of sponge removal. The degree and pattern of mating response of ewes were similar, irrespective of the treatment used, approximately 92% of the ewes being marked by the ram by 72h after sponge removal. No significant differences in fertility or litter size were observed between the treatment groups. Ewes treated with FGA sponges had a fertility of 53% and litter size of 2.3 after mating at the synchronized estrus. The corresponding values for ewes treated with MAP sponges were 57% and 2.1. Use of MAP sponges was associated with a 17.8% sponge loss during treatment compared with 1% sponge loss in ewes treated with FGA sponges. Such losses could compromise the use of MAP sponges by reducing their overall efficacy.     

Pmch-Deficiency in Rats Is Associated with Normal Adipocyte Differentiation and Lower Sympathetic Adipose Drive

The orexigenic neuropeptide melanin-concentrating hormone (MCH), a product of Pmch, is an important mediator of energy homeostasis. Pmch-deficient rodents are lean and smaller, characterized by lower food intake, body-, and fat mass. Pmch is expressed in hypothalamic neurons that ultimately are components in the sympathetic nervous system (SNS) drive to white and interscapular brown adipose tissue (WAT, iBAT, respectively). MCH binds to MCH receptor 1 (MCH1R), which is present on adipocytes. Currently it is unknown if Pmch-ablation changes adipocyte differentiation or sympathetic adipose drive. Using Pmch-deficient and wild-type rats on a standard low-fat diet, we analyzed dorsal subcutaneous and perirenal WAT mass and adipocyte morphology (size and number) throughout development, and indices of sympathetic activation in WAT and iBAT during adulthood. Moreover, using an in vitro approach we investigated the ability of MCH to modulate 3T3-L1 adipocyte differentiation. Pmch-deficiency decreased dorsal subcutaneous and perirenal WAT mass by reducing adipocyte size, but not number. In line with this, in vitro 3T3-L1 adipocyte differentiation was unaffected by MCH. Finally, adult Pmch-deficient rats had lower norepinephrine turnover (an index of sympathetic adipose drive) in WAT and iBAT than wild-type rats. Collectively, our data indicate that MCH/MCH1R-pathway does not modify adipocyte differentiation, whereas Pmch-deficiency in laboratory rats lowers adiposity throughout development and sympathetic adipose drive during adulthood.     

Anti-CD70 Immunocytokines for Exploitation of Interferon-γ-Induced RIP1-Dependent Necrosis in Renal Cell Carcinoma

Metastatic renal cell carcinoma (RCC) is an incurable disease in clear need of new therapeutic interventions. In early-phase clinical trials, the cytokine IFN-γ showed promise as a biotherapeutic for advanced RCC, but subsequent trials were less promising. These trials, however, focused on the indirect immunomodulatory properties of IFN-γ, and its direct anti-tumor effects, including its ability to kill tumor cells, remains mostly unexploited. We have previously shown that IFN-γ induces RIP1 kinase-dependent necrosis in cells lacking NF-κB survival signaling. RCC cells display basally-elevated NF-κB activity, and inhibiting NF-κB in these cells, for example by using the small-molecule proteasome blocker bortezomib, sensitizes them to RIP1-dependent necrotic death following exposure to IFN-γ. While these observations suggest that IFN-γ-mediated direct tumoricidal activity will have therapeutic benefit in RCC, they cannot be effectively exploited unless IFN-γ is targeted to tumor cells in vivo. Here, we describe the generation and characterization of two novel ‘immunocytokine’ chimeric proteins, in which either human or murine IFN-γ is fused to an antibody targeting the putative metastatic RCC biomarker CD70. These immunocytokines display high levels of species-specific IFN-γ activity and selective binding to CD70 on human RCC cells. Importantly, the IFN-γ immunocytokines function as well as native IFN-γ in inducing RIP1-dependent necrosis in RCC cells, when deployed in the presence of bortezomib. These results provide a foundation for the in vivo exploitation of IFN-γ-driven tumoricidal activity in RCC.     

Nitrogen‐Doped CNx/CNTs Heteroelectrocatalysts for Highly Efficient Dye‐Sensitized Solar Cells

The use of polydopamine as a nitrogen containing precursor to generate catalytically active nitrogen‐doped carbon (CN_x) materials on carbon nanotubes (CNTs) is reported. These N‐doped CN_x/CNT materials display excellent electrocatalytic activity toward the reduction of triiodide electrolyte in dye‐sensitized solar cells (DSSCs). Further, the influence of various synthesis parameters on the catalytic performance of CN_x/CNTs is investigated in detail. The best performing device fabricated with the CN_x/CNTs material delivers power conversion efficiency of 7.3%, which is comparable or slightly higher than that of Pt (7.1%) counter electrode‐based DSSC. These CN_x/CNTs materials show great potential to address the issues associated with the Pt electrocatalyst including the high cost and scarcity.     

Fitness consequences of maternal and grandmaternal effects

Transgenerational effects are broader than only parental relationships. Despite mounting evidence that multigenerational effects alter phenotypic and life‐history traits, our understanding of how they combine to determine fitness is not well developed because of the added complexity necessary to study them. Here, we derive a quantitative genetic model of adaptation to an extraordinary new environment by an additive genetic component, phenotypic plasticity, maternal and grandmaternal effects. We show how, at equilibrium, negative maternal and negative grandmaternal effects maximize expected population mean fitness. We define negative transgenerational effects as those that have a negative effect on trait expression in the subsequent generation, that is, they slow, or potentially reverse, the expected evolutionary dynamic. When maternal effects are positive, negative grandmaternal effects are preferred. As expected under Mendelian inheritance, the grandmaternal effects have a lower impact on fitness than the maternal effects, but this dual inheritance model predicts a more complex relationship between maternal and grandmaternal effects to constrain phenotypic variance and so maximize expected population mean fitness in the offspring.     

Analysis of conditions affecting auto-phosphorylation of human kinases during expression in bacteria

Bacterial over-expression of kinases is often associated with high levels of auto-phosphorylation resulting in heterogeneous recombinant protein preparations or sometimes in insoluble protein. Here we present expression systems for nine kinases in Escherichia coli and, for the most heavily phosphorylated, the characterisation of factors affecting auto-phosphorylation. Experiments showed that the level of auto-phosphorylation was proportional to the rate of expression. Comparison of phosphorylation states following in vitro phosphorylation with phosphorylation states following expression in E. coli showed that the non-physiological 'hyper-phosphorylation' was occurring at sites that would require local unfolding to be accessible to a kinase active site. In contrast, auto-phosphorylation on unphosphorylated kinases that had been expressed in bacteria overexpressing λ-phosphatase was only observed on distinct exposed sites. Remarkably, the Ser/Thr kinase PLK4 auto-phosphorylated on a tyrosine residue (Tyr177) located in the activation segment. The results give support to a mechanism in which auto-phosphorylation occurs before or during protein folding. In addition, the expression systems and protocols presented will be a valuable resource to the research community.     

Structural Basis of Differential Neutralization of DENV-1 Genotypes by an Antibody that Recognizes a Cryptic Epitope

We previously developed a panel of neutralizing monoclonal antibodies against Dengue virus (DENV)-1, of which few exhibited inhibitory activity against all DENV-1 genotypes. This finding is consistent with reports observing variable neutralization of different DENV strains and genotypes using serum from individuals that experienced natural infection or immunization. Herein, we describe the crystal structures of DENV1-E111 bound to a novel CC′ loop epitope on domain III (DIII) of the E protein from two different DENV-1 genotypes. Docking of our structure onto the available cryo-electron microscopy models of DENV virions revealed that the DENV1-E111 epitope was inaccessible, suggesting that this antibody recognizes an uncharacterized virus conformation. While the affinity of binding between DENV1-E111 and DIII varied by genotype, we observed limited correlation with inhibitory activity. Instead, our results support the conclusion that potent neutralization depends on genotype-dependent exposure of the CC′ loop epitope. These findings establish new structural complexity of the DENV virion, which may be relevant for the choice of DENV strain for induction or analysis of neutralizing antibodies in the context of vaccine development.     

CXCL-16, IL-17, and bone morphogenetic protein 2 (BMP-2) are associated with overweight and obesity conditions in middle-aged and elderly women

Background

The current concept of overweight/obesity is most likely related to a combination of increased caloric intake and decreased energy expenditure. Widespread inflammation, associated with both conditions, appears to contribute to the development of some obesity-related comorbidities. Interventions that directly or indirectly target individuals at high risk of developing obesity have been largely proposed because of the increasing number of overweight/obese cases worldwide. The aim of the present study was to assess CXCL16, IL-17, and BMP-2 plasma factors in middle-aged and elderly women and relate them to an overweight or obese status. In total, 117 women were selected and grouped as eutrophic, overweight, and obese, according to anthropometric parameters. Analyses of anthropometric and circulating biochemical parameters were followed by plasma immunoassays for CXCL-16, IL-17, and BMP-2.

Results

Plasma mediators increased in all overweight and obese individuals, with the exception of BMP-2 in the elderly group, whereas CXCL16 levels were shown to differentiate overweight and obese individuals. Overweight and/or obese middle-aged and elderly individuals presented with high LDL, triglycerides, and glycemia levels. Anthropometric parameters indicating increased-cardiovascular risk were positively correlated with CXCL-16, BMP-2, and IL-17 levels in overweight and obese middle-aged and elderly individuals.

Conclusion

This study provides evidence that CXCL-16, IL-17, and BMP-2 are potential plasma indicators of inflammatory status in middle-aged and elderly women; therefore, further investigation of obesity-related comorbidities is recommended. CXCL16, in particular, could be a potential marker for middle-aged and elderly individuals transitioning from eutrophic to overweight body types, which represents an asymptomatic and dangerous condition.

     

Synthesis and biological evaluation of water-soluble progesterone-conjugated probes for magnetic resonance imaging of hormone related cancers

Progesterone receptor (PR) is strongly associated with disease prognosis and therapeutic efficacy in hormone-related diseases such as endometriosis and breast, ovarian, and uterine cancers. Receptor status is currently determined by immunohistochemistry assays. However, noninvasive PR imaging agents could improve disease detection and help elucidate pathological molecular pathways, leading to new therapies and animal disease models. A series of water-soluble PR-targeted magnetic resonance imaging (MRI) probes were synthesized using Cu(I)-catalyzed click chemistry and evaluated in vitro and in vivo. These agents demonstrated activation of PR in vitro and preferential accumulation in PR(+) compared to PR(-) human breast cancer cells with low toxicity. In xenograft tumor models, the agents demonstrated enhanced signal intensity in PR(+) tumors compared to PR(-) tumors. The results suggest that these agents may be promising MRI probes for PR(+) diseases.