VACTERL/caudal regression/Currarino syndrome-like malformations in mice with mutation in the proprotein convertase Pcsk5

We have identified an ethylnitrosourea (ENU)-induced recessive mouse mutation (Vcc) with a pleiotropic phenotype that includes cardiac, tracheoesophageal, anorectal, anteroposterior patterning defects, exomphalos, hindlimb hypoplasia, a presacral mass, renal and palatal agenesis, and pulmonary hypoplasia. It results from a C470R mutation in the proprotein convertase PCSK5 (PC5/6). Compound mutants (Pcsk5(Vcc/null)) completely recapitulate the Pcsk5(Vcc/Vcc) phenotype, as does an epiblast-specific conditional deletion of Pcsk5. The C470R mutation ablates a disulfide bond in the P domain, and blocks export from the endoplasmic reticulum and proprotein convertase activity. We show that GDF11 is cleaved and activated by PCSK5A, but not by PCSK5A-C470R, and that Gdf11-deficient embryos, in addition to having anteroposterior patterning defects and renal and palatal agenesis, also have a presacral mass, anorectal malformation, and exomphalos. Pcsk5 mutation results in abnormal expression of several paralogous Hox genes (Hoxa, Hoxc, and Hoxd), and of Mnx1 (Hlxb9). These include known Gdf11 targets, and are necessary for caudal embryo development. We identified nonsynonymous mutations in PCSK5 in patients with VACTERL (vertebral, anorectal, cardiac, tracheoesophageal, renal, limb malformation OMIM 192350) and caudal regression syndrome, the phenotypic features of which resemble the mouse mutation. We propose that Pcsk5, at least in part via GDF11, coordinately regulates caudal Hox paralogs, to control anteroposterior patterning, nephrogenesis, skeletal, and anorectal development.     

A long-term study of bone mineral density in patients with phenylketonuria under diet therapy

Introduction

Dietary control of classic phenylketonuria (PKU) needs restriction of natural proteins; adequate protein intake is achieved by adding low phenylalanine (phe) formulae. The adequacy of this diet for normal bone mineralization had not been sufficiently evaluated. Our aim was to evaluate and follow up bone mineral density (BMD) in children and adolescents with PKU within a 2-year time interval to assess the adequacy of a phenylalanine restricted diet for bone mineralization and to search for a possible relationship between BMD, dietary control and blood phenylalanine (phe) concentrations.

Material and methods

Thirty-two patients with classic PKU (3-19 years) were evaluated for their bone mineral status using dual energy X-ray absorptiometry (DEXA) both at the beginning (baseline) and the end (follow-up) of the study.

Results

Low BMD was detected in 31.25% at the start and in 6.25% of patients after 2 years follows-up. No relationship was found between BMD and the duration of diet compliance and phe level as well.

Conclusions

In this study the low BMD detected in our patients was both at baseline and follow-up independent of diet restriction. A yearly DEXA would be highly beneficial for early detection and treatment, thus preventing osteoporosis and decreasing the risk of fractures. We also suggest the importance of searching for new emerging therapies such as enzyme substitution or gene therapy as low protein diet compliance was not enough to maintain normal bone mineral density.     

Thymocyte expression of cathepsin L is essential for NKT cell development

CD1d antigen presentation to natural killer T (NKT) cells expressing the semi-invariant T cell receptor V(alpha)14J(alpha)18 requires CD1d trafficking through endosomal compartments; however, the endosomal events remain undefined. We show that mice lacking the endosomal protease cathepsin L (catL) have greatly reduced numbers of V(alpha)14(+)NK1.1(+) T cells. In addition, catL expression in thymocytes is critical not only for selection of these cells in vivo but also for stimulation of V(alpha)14(+)NK1.1(+) T cells in vitro. CD1d cell-surface expression and intracellular localization appear normal in catL-deficient thymocytes, as does the lysosomal morphology; this implies a specific role for catL in regulating presentation of natural CD1d ligands mediating V(alpha)14(+)NK1.1(+) T cell selection. These data implicate lysosomal proteases as key regulators of not only classical major histocompatibility complex class II antigen presentation but also nonclassical CD1d presentation.     

α7 Nicotinic Acetylcholine Receptor Regulates Airway Epithelium Differentiation by Controlling Basal Cell Proliferation

Airway epithelial basal cells are known to be critical for regenerating injured epithelium and maintaining tissue homeostasis. Recent evidence suggests that the α7 nicotinic acetylcholine receptor (nAChR), which is highly permeable to Ca²⁺, is involved in lung morphogenesis. Here, we have investigated the potential role of the α7 nAChR in the regulation of airway epithelial basal cell proliferation and the differentiation of the human airway epithelium. In vivo during fetal development and in vitro during the regeneration of the human airway epithelium, α7 nAChR expression coincides with epithelium differentiation. Inactivating α7 nAChR function in vitro increases cell proliferation during the initial steps of the epithelium regeneration, leading to epithelial alterations such as basal cell hyperplasia and squamous metaplasia, remodeling observed in many bronchopulmonary diseases. The regeneration of the airway epithelium after injury in α7^−/− mice is delayed and characterized by a transient hyperplasia of basal cells. Moreover, 1-year-old α7^−/− mice more frequently present basal cells hyperplasia. Modulating nAChR function or expression shows that only α7 nAChR, as opposed to heteropentameric α_xβ_y nAChRs, controls the proliferation of human airway epithelial basal cells. These findings suggest that α7 nAChR is a key regulator of the plasticity of the human airway epithelium by controlling basal cell proliferation and differentiation pathway and is involved in airway remodeling during bronchopulmonary diseases.The respiratory epithelium, which is constantly exposed to airborne pollutants, is frequently injured, which results in altered epithelial functions. To restore these functions, the respiratory epithelium must undergo rapid repair via epithelial cell spreading and migration and regenerate its structure via basal cell proliferation and differentiation.¹ These processes are tightly controlled to restore the pseudostratified architecture of the normal mucociliary epithelium. However, in most respiratory diseases, alterations of the regeneration processes induce epithelial remodeling such as hyperplasia, metaplasia, and fibrosis. Understanding the sequence of processes involved in cell proliferation and differentiation is therefore of crucial importance. Both in vivo and in vitro, human airway basal cells are able to proliferate and reconstitute a fully differentiated and functional epithelium.² These cells are such considered as progenitors of the human airway epithelium and important actors of the airway epithelium regeneration.The nonneuronal cholinergic system is thought to be involved in the regulation of cell functions such as cell-cell interaction, apoptosis, and proliferation.³ It is now established that human bronchial epithelial cells contain all of the machinery for the production, storage, secretion, and degradation of acetylcholine, which acts as an autocrine or paracrine hormone.^4,5 Acetylcholine exerts its effects through muscarinic and nicotinic acetylcholine receptors. Nicotinic acetylcholine receptors (nAChRs) are composed of five subunits, arranged as α/β heteromeric or α homomeric nAChRs, and assembled around a central ion channel, mediating the influx of Ca²⁺.⁶ The airway epithelium expresses α3, α4, α5, α7, α9, β2, and β4 subunits for nAChRs.^7,8,9α7 nAChR is characterized by an elevated Ca²⁺ permeability¹⁰ and has been involved in several important biological processes such as cell proliferation, apoptosis, and angiogenesis in cancer.^11,12 Prenatal nicotine exposure significantly increases pulmonary α7 nAChR expression and alters fetal lung development¹³ and subsequently pulmonary function in newborn.¹⁴ In particular, alteration of lung branching morphogenesis induced by nicotine is mediated by α7 nAChR.¹⁵ Altogether, these observations led us to investigate whether the α7 nAChR could be involved in the differentiation of the respiratory epithelium. In the human airway epithelium, we observed α7 nAChR expression in basal cells, which play a critical role in the epithelial regeneration. Both in vivo and in vitro, the α7 nAChR expression is associated with the airway epithelium differentiation. Moreover, in vitro inactivating α7 nAChR or in vivo disrupting genetic α7 nAChR expression induces airway epithelium remodeling by modulating basal cell proliferation. This study thus provides several lines of evidence that α7 nAChR is significant for airway epithelial differentiation and suggests that α7 nAChR is a key regulator of the plasticity of the airway epithelium.     

High-fat diet induced an oxidative stress in white adipose tissue and disturbed plasma transition metals in rat: prevention by grape seed and skin extract

Obesity is a public health problem characterized by increased accumulation of fat into adipose tissues leading to oxidative stress, dyslipidemia, and chronic inflammatory status. We used an experimental model of high-fat diet-induced obesity to analyze the link between dyslipidemia, oxidative stress, and fat accumulation into adipose tissue of rats, as well as the involvement of intracellular mediators such as transition metals on signal transduction. We also looked at the ability of a grape seed and skin extract (GSSE) from a Tunisian cultivar to prevent fat-induced disturbances. Data showed that a high-fat diet (HFD) provoked dyslipidemia into plasma which is linked to an oxidative stress, an accumulation of transition metals such as manganese, copper, and zinc and a depletion of iron. GSSE prevented dyslipidemia by modulating lipase activity, together with increased antioxidant capacity and depletion of transition metals as well as of free radicals such as O₂ ^? and OH. These data indicated that GSSE has important preventive effects on HFD-induced obesity and oxidative stress whose transduction seems to involve transition metals. GSSE should be used as a safe anti-obesity agent that could find potential applications in metabolic disorders involving transition metals dyshomeostasis.     

A brain‐potential correlate of task‐set conflict

Brain‐potential correlates of response conflict are well documented, but those of task conflict are not. Task‐switching studies have suggested a plausible correlate of task conflict—a poststimulus posterior negativity—however, in such paradigms the negativity may also reflect poststimulus task‐set reconfiguration postulated in some models. Here, participants alternated between single‐task blocks of classifying letters and digits; hence, no within‐block task‐set reconfiguration was required. Presenting letters alongside digits slowed responses to the digits and elicited an ERP negativity from ～350 ms, relative to task‐neutral symbols presented alongside digits, consistent with task conflict. The negativity was also present for congruent digit‐letter stimuli; this and the lack of behavioral response congruency effects indicate conflict at the level of task‐set rather than response selection.     

RyR1 Deficiency in Congenital Myopathies Disrupts Excitation–Contraction Coupling

In skeletal muscle, excitation–contraction (EC) coupling is the process whereby the voltage‐gated dihydropyridine receptor (DHPR) located on the transverse tubules activates calcium release from the sarcoplasmic reticulum by activating ryanodine receptor (RyR1) Ca²⁺ channels located on the terminal cisternae. This subcellular membrane specialization is necessary for proper intracellular signaling and any alterations in its architecture may lead to neuromuscular disorders. In this study, we present evidence that patients with recessive RYR1‐related congenital myopathies due to primary RyR1 deficiency also exhibit downregulation of the alfa 1 subunit of the DHPR and show disruption of the spatial organization of the EC coupling machinery. We created a cellular RyR1 knockdown model using immortalized human myoblasts transfected with RyR1 siRNA and confirm that knocking down RyR1 concomitantly downregulates not only the DHPR but also the expression of other proteins involved in EC coupling. Unexpectedly, this was paralleled by the upregulation of inositol‐1,4,5‐triphosphate receptors; functionally however, upregulation of the latter Ca²⁺ channels did not compensate for the lack of RyR1‐mediated Ca²⁺ release. These results indicate that in some patients, RyR1 deficiency concomitantly alters the expression pattern of several proteins involved in calcium homeostasis and that this may influence the manifestation of these diseases.