Actions and interactions of growth hormone and insulin-like growth factor-II: body and organ growth of transgenic mice

To characterize long-term actions and interactions of growth hormone (GH) and insulin-like growth factor-II (IGF-II) on postnatal body and organ growth, hemizygous phosphoenolpyruvate carboxykinase (PEPCK)-human IGF-II transgenic mice were crossed with hemizygous PEPCK-bovine GH transgenic mice. The latter are characterized by two-fold increased serum levels of IGF-I and exhibit markedly increased body, skeletal and organ growth. Four different genetic groups were obtained: mice harbouring the IGF-II transgene (I), the bGH transgene (B), or both transgenes (IB), and non- transgenic controls (C). These groups of mice have previously been studied for circulating IGF-I levels (Wolf et al., 1995a), whereas the present study deals with body and organ growth. Growth curves (week 3 to 12) were estimated by regression with linear and quadratic components of age on body weight and exhibited significantly (p < 0.001) greater linear coefficients in B and IB than in I and C mice. The linear coefficients of male I and C mice were significantly (p < 0.001) greater than those of their female counterparts, whereas this sex-related difference was absent in the bGH transgenic groups. The weights of internal organs as well as the weights of abdominal fat, skin and carcass were recorded from 3.5- to 8- month-old mice. In addition, organ weight-to-body weight-ratios (relative organ weights) were calculated. Except for the weight of abdominal fat, absolute organ weights were as a rule significantly greater in B and IB than in I and C mice. IGF-II overproduction as a tendency increased the weights of kidneys, adrenal glands, pancreas and uterus both in the absence and presence of the bGH transgene. Analysis of relative organ weights demonstrated significant (p < 0.05) effects of elevated IGF- II on the relative growth of kidneys (males and females) and adrenal glands (females), confirming our previous report on organ growth of PEPCK-IGF-II transgenic mice. In females, IGF-II and GH overproduction were additive in stimulating the growth of spleen and uterus, providing evidence for tissue-specific postnatal growth promoting effects by IGF-II in the presence of elevated IGF-I     

The connecting cilium inner scaffold provides a structural foundation that protects against retinal degeneration

Inherited retinal degeneration due to loss of photoreceptor cells is a leading cause of human blindness. These cells possess a photosensitive outer segment linked to the cell body through the connecting cilium (CC). While structural defects of the CC have been associated with retinal degeneration, its nanoscale molecular composition, assembly, and function are barely known. Here, using expansion microscopy and electron microscopy, we reveal the molecular architecture of the CC and demonstrate that microtubules are linked together by a CC inner scaffold containing POC5, CENTRIN, and FAM161A. Dissecting CC inner scaffold assembly during photoreceptor development in mouse revealed that it acts as a structural zipper, progressively bridging microtubule doublets and straightening the CC. Furthermore, we show that Fam161a disruption in mouse leads to specific CC inner scaffold loss and triggers microtubule doublet spreading, prior to outer segment collapse and photoreceptor degeneration, suggesting a molecular mechanism for a subtype of retinitis pigmentosa.

Inherited retinal degeneration due to loss of photoreceptor cells is a leading cause of human blindness. Ultrastructure expansion microscopy on mouse retina reveals the presence of a novel structure inside the photoreceptor connecting cilium, the inner scaffold, that protects the outer segment against degeneration.     

Simulation of direct shear tests on rooted and non-rooted soil using finite element analysis

The finite element (FE) method has been used in recent years to simulate overturning processes in trees and to better comprehend plant anchorage mechanics. We aimed at understanding the fundamental mechanisms of root-soil reinforcement by simulating direct shear of rooted and non-rooted soil. Two- (2D) and three-dimensional (3D) FE simulations of direct shear box tests were carried out using readily available software for routine strength assessment of the root-soil composite. Both rooted and non-rooted blocks of soil were modelled using a simplified model of root distribution and root material properties representative of real roots. Linear elastic behaviour was assumed for roots and the soil was modelled as an ideally plastic medium. FE analysis showed that direct shear tests were dependent on the material properties specified for both the soil and roots. 2D and 3D simulations of direct shear of non-rooted soil produced similar results and any differences between 2D and 3D simulations could be explained with regard to the spatial complexity of roots used in the root distribution model. The application of FE methods was verified through direct shear tests on soil with analogue roots and the results compared to in situ tests on rooted soil in field conditions.     

Shear Viscosity Investigation on Mango Juice with High Frequency Longitudinal Ultrasonic Waves and Rotational Viscosimetry

Ultrasonic velocity and attenuation measurements were performed on mango juices at 25 MHz in order to estimate longitudinal viscosity. Juices were extracted from fruits, removed periodically from fruit batches undergoing ripening for 3 weeks under controlled conditions. The correlation between longitudinal viscosity and apparent dynamic shear viscosity, obtained from flow tests, showed that up to 12–13 wt.% of Soluble Solids Content (SSC), the juices presented a Newtonian behavior. In this case the relation between longitudinal viscosity measured by ultrasound and shear viscosity measured by flow tests was very simple leading to the conclusion that ultrasound could replace rotating viscosimeters for specific applications. Over this limit, the results were also clearly correlated but the correlation depended on the shear rate because of the shear thinning behavior of the juices certainly due to soluble pectins. The use of longitudinal ultrasonic waves as a tool for viscosity determination on large batches of samples is discussed at the end of this communication.     

Estimating Ecosystem Metabolism to Entire River Networks

Ecosystems - River ecosystem metabolism (REM) is a promising cost-effective measure of ecosystem functioning, as it integrates many different ecosystem processes and is affected by both rapid...     

Distinct role of subcomplexes of the COPI coat in the regulation of ArfGAP2 activity

COPI vesicles serve for transport of proteins and membrane lipids in the early secretory pathway. Their coat protein (coatomer) is a heptameric complex that is recruited to the Golgi by the small GTPase Arf1. Although recruited en bloc, coatomer can be viewed as a stable assembly of an adaptin‐like tetrameric subcomplex (CM4) and a trimeric ‘cage’ subcomplex (CM3). Following recruitment, coatomer stimulates ArfGAP‐dependent GTP hydrolysis on Arf1. Here, we employed recombinant coatomer subcomplexes to study the role of coatomer components in the regulation of ArfGAP2, an ArfGAP whose activity is strictly coatomer‐dependent. Within CM4, we define a novel hydrophobic pocket for ArfGAP2 interaction on the appendage domain of γ₁‐COP. The CM4 subcomplex (but not CM3) is recruited to membranes through Arf1 and can subsequently recruit ArfGAP2. Neither CM3 nor CM4 in itself is effective in stimulating ArfGAP2 activity, but stimulation is regained when both subcomplexes are present. Our findings point to a distinct role of each of the two coatomer subcomplexes in the regulation of ArfGAP2‐dependent GTP hydrolysis on Arf1, where the CM4 subcomplex functions in GAP recruitment, while, similarly to the COPII system, the cage‐like CM3 subcomplex stimulates the catalytic reaction.     

Genome-Wide Analysis of Cold Adaptation in Indigenous Siberian Populations

Following the dispersal out of Africa, where hominins evolved in warm environments for millions of years, our species has colonised different climate zones of the world, including high latitudes and cold environments. The extent to which human habitation in (sub-)Arctic regions has been enabled by cultural buffering, short-term acclimatization and genetic adaptations is not clearly understood. Present day indigenous populations of Siberia show a number of phenotypic features, such as increased basal metabolic rate, low serum lipid levels and increased blood pressure that have been attributed to adaptation to the extreme cold climate. In this study we introduce a dataset of 200 individuals from ten indigenous Siberian populations that were genotyped for 730,525 SNPs across the genome to identify genes and non-coding regions that have undergone unusually rapid allele frequency and long-range haplotype homozygosity change in the recent past. At least three distinct population clusters could be identified among the Siberians, each of which showed a number of unique signals of selection. A region on chromosome 11 (chr11:66–69 Mb) contained the largest amount of clustering of significant signals and also the strongest signals in all the different selection tests performed. We present a list of candidate cold adaption genes that showed significant signals of positive selection with our strongest signals associated with genes involved in energy regulation and metabolism (CPT1A, LRP5, THADA) and vascular smooth muscle contraction (PRKG1). By employing a new method that paints phased chromosome chunks by their ancestry we distinguish local Siberian-specific long-range haplotype signals from those introduced by admixture.     

An Allergic Lung Microenvironment Suppresses Carbon Nanotube-Induced Inflammasome Activation via STAT6-Dependent Inhibition of Caspase-1

Background

Multi-walled carbon nanotubes (MWCNTs) represent a human health risk as mice exposed by inhalation display pulmonary fibrosis. Production of IL-1β via inflammasome activation is a mechanism of MWCNT-induced acute inflammation and has been implicated in chronic fibrogenesis. Mice sensitized to allergens have elevated T-helper 2 (Th2) cytokines, IL-4 and IL-13, and are susceptible to MWCNT-induced airway fibrosis. We postulated that Th2 cytokines would modulate MWCNT-induced inflammasome activation and IL-1β release in vitro and in vivo during allergic inflammation.

Methods

THP-1 macrophages were primed with LPS, exposed to MWCNTs and/or IL-4 or IL-13 for 24 hours, and analyzed for indicators of inflammasome activation. C57BL6 mice were sensitized to house dust mite (HDM) allergen and MWCNTs were delivered to the lungs by oropharyngeal aspiration. Mice were euthanized 1 or 21 days post-MWCNT exposure and evaluated for lung inflammasome components and allergic inflammatory responses.

Results

Priming of THP-1 macrophages with LPS increased pro-IL-1β and subsequent exposure to MWCNTs induced IL-1β secretion. IL-4 or IL-13 decreased MWCNT-induced IL-1β secretion by THP-1 cells and reduced pro-caspase-1 but not pro-IL-1β. Treatment of THP-1 cells with STAT6 inhibitors, either Leflunomide or JAK I inhibitor, blocked suppression of caspase activity by IL-4 and IL-13. In vivo, MWCNTs alone caused neutrophilic infiltration into the lungs of mice 1 day post-exposure and increased IL-1β in bronchoalveolar lavage fluid (BALF) and pro-caspase-1 immuno-staining in macrophages and airway epithelium. HDM sensitization alone caused eosinophilic inflammation with increased IL-13. MWCNT exposure after HDM sensitization increased total cell numbers in BALF, but decreased numbers of neutrophils and IL-1β in BALF as well as reduced pro-caspase-1 in lung tissue. Despite reduced IL-1β mice exposed to MWCNTs after HDM developed more severe airway fibrosis by 21 days and had increased pro-fibrogenic cytokine mRNAs.

Conclusions

These data indicate that Th2 cytokines suppress MWCNT-induced inflammasome activation via STAT6-dependent down-regulation of pro-caspase-1 and suggest that suppression of inflammasome activation and IL-1β by an allergic lung microenvironment is a mechanism through which MWCNTs exacerbate allergen-induced airway fibrosis.     

External validation of Adjuvant! Online breast cancer prognosis tool. Prioritising recommendations for improvement

Background

Adjuvant! Online is a web-based application designed to provide 10 years survival probability of patients with breast cancer. Several predictors have not been assessed in the original Adjuvant! Online study. We provide the validation of Adjuvant! Online algorithm on two breast cancer datasets, and we determined whether the accuracy of Adjuvant! Online is improved with other well-known prognostic factors.

Patients and Methods

The French data set is composed of 456 women with early breast cancer. The Dutch data set is composed of 295 women less than 52 years of age. Agreement between observation and Adjuvant! Online prediction was checked, and logistic models were performed to estimate the prognostic information added by risk factors to Adjuvant! Online prediction.

Results

Adjuvant! Online prediction was overall well-calibrated in the French data set but failed in some subgroups of such high grade and HER2 positive patients. HER2 status, Mitotic Index and Ki67 added significant information to Adjuvant! Online prediction. In the Dutch data set, the overall 10-year survival was overestimated by Adjuvant! Online, particularly in patients less than 40 years old.

Conclusion

Adjuvant! Online needs to be updated to adjust overoptimistic results in young and high grade patients, and should consider new predictors such as Ki67, HER2 and Mitotic Index.