Rapamycin impairs trabecular bone acquisition from high‐dose but not low‐dose intermittent parathyroid hormone treatment

The osteo‐anabolic effects of intermittent parathyroid hormone (PTH) treatment require insulin‐like growth factor (IGF) signaling through the IGF‐I receptor. A major downstream target of the IGF‐I receptor (via Akt) is the mammalian target of rapamycin (mTOR), a kinase involved in protein synthesis. We investigated whether the bone‐building effects of intermittent PTH require functional mTOR signaling. Mice were treated with daily PTH 1–34 (0, 10, 30, or 90 µg/kg) for 6 weeks in the presence or absence of rapamycin, a selective inhibitor of mTOR. We found that all PTH doses were effective in enhancing bone mass, whether rapamycin was present or not. Rapamycin had little to no effect on the anabolic response at low (10 µg) PTH doses, small effects in a minority of anabolic measures at moderate doses (30 µg), but the anabolic effects of high‐dose PTH (90 µg) were consistently and significantly suppressed by rapamycin (～4–36% reduction). Serum levels of Trap5b, a marker of resorption, were significantly enhanced by rapamycin, but these effects were observed whether PTH was absent or present. Our data suggest that intermittent PTH, particularly at lower doses, is effective in building bone mass in the presence of rapamycin. However, the full anabolic effects of higher doses of PTH are significantly suppressed by rapamycin, suggesting that PTH might normally activate additional pathways (including mTOR) for its enhanced high‐dose anabolic effects. Clinical doses of intermittent PTH could be an effective treatment for maintaining or increasing bone mass among patients taking rapamycin analogs for unrelated health issues. J. Cell. Physiol. 221: 579–585, 2009. © 2009 Wiley‐Liss, Inc.     

Parental relatedness and major histocompatibility effects on early embryo survivorship in Atlantic salmon

Salmon have provided key insights into the relative influence of natural and sexual selection on major histocompatibility complex (MHC) variation. Natural selection on salmon MHC genes has been demonstrated in pathogen studies, and there is evidence of MHC-based mate choice (sexual selection). We tested whether parental MHC genes affect survivorship of juvenile Atlantic salmon (Salmo salar) by quantifying the influence of parental genome-wide relatedness and MHC genotype on survivorship to the swim-up stage. Thirteen microsatellite loci were used to estimate the influence of genome-wide relatedness between parents on offspring survivorship and MHC genotypes were determined by sequencing part of the class IIβ gene. Our results revealed no significant relationship between early offspring survivorship and genome-wide relatedness, predicted MHC heterozygosity, or MHC allelic similarity. Overall, our data are consistent with the contention that excess MHC heterozygosity in Atlantic salmon juveniles is due to sexual selection as well as differential survival of offspring due to MHC genotype.     

Soft tissue reconstruction of Homotherium latidens (Mammalia, Carnivora, Felidae). Implications for the possibility of representations in Palaeolithic art

We reconstruct the life appearance of the head and body of the sabretoothed felid Homotherium latidens through the study of osteological correlates of soft tissue attributes, incorporating data from the dissection of several large felids and using the Extant Phylogenetic Bracket to infer the condition of unpreserved attributes where morphological evidence is inconclusive. Our reconstruction shows that Homotherium would have differed from modern felids in aspects of the general proportions of the head, having a straighter dorsal outline and a long, square muzzle with an angular “chin”, although large pantherines may mimic to some degree the angular shape of the machairodontine mandibular symphysis with the growth of long hair in the chin area. The tips of the sabres of Homotherium would have been visible in life, protruding beyond the lips. Our reconstructed body proportions of Homotherium imply a sloping back. These conclusions contradict a previous proposal that postulated a different soft tissue anatomy for Homotherium, and which led to interpretation of a Palaeolithic figurine as a rendering of the sabretooth. Our results suggest that the figurine in question is a depiction of a cave lion, and that there is no single known representation of a machairodont in prehistoric art. The implications for our current understanding of the Late Pleistocene large-carnivore guild are discussed.     

Wolbachia Lipoprotein Stimulates Innate and Adaptive Immunity through Toll-like Receptors 2 and 6 to Induce Disease Manifestations of Filariasis

Wolbachia endosymbiotic bacteria have been implicated in the inflammatory pathogenesis of filariasis. Inflammation induced by Brugia malayi female worm extract (BMFE) is dependent on Toll-like receptors 2 and 6 (TLR2/6) with only a partial requirement for TLR1. Removal of Wolbachia, lipids, or proteins eliminates all inflammatory activity. Wolbachia bacteria contain the lipoprotein biosynthesis genes Ltg and LspA but not Lnt, suggesting Wolbachia proteins cannot be triacylated, accounting for recognition by TLR2/6. Lipoprotein databases revealed 3–11 potential lipoproteins from Wolbachia. Peptidoglycan-associated lipoprotein (PAL) and Type IV secretion system-VirB6 were consistently predicted, and B. malayi Wolbachia PAL (wBmPAL) was selected for functional characterization. Diacylated 20-mer peptides of wBmPAL (Diacyl Wolbachia lipopeptide (Diacyl WoLP)) showed a near identical TLR2/6 and TLR2/1 usage compared with BMFE and bound directly to TLR2. Diacyl WoLP induced systemic tumor necrosis factor-α and neutrophil-mediated keratitis in mice. Diacyl WoLP activated monocytes induce up-regulation of gp38 on human lymphatic endothelial cells and induced dendritic cell maturation and activation. Dendritic cells primed with BMFE generated a non-polarized Th1/Th2 CD4⁺ T cell profile, whereas priming with Wolbachia depleted extracts (following tetracycline treatment; BMFEtet) polarized to a Th2 profile that could be reversed by reconstitution with Diacyl WoLP. BMFE generated IgG1 and IgG2c antibody responses, whereas BMFEtet or inoculation of TLR2 or MyD88^−/− mice produced defective IgG2c responses. Thus, in addition to innate inflammatory activation, Wolbachia lipoproteins drive interferon-γ-dependent CD4⁺ T cell polarization and antibody switching.Human filariasis is a major neglected tropical disease. More than 150 million individuals are infected with the filarial worms responsible for lymphatic filariasis (LF)⁴ (Wuchereria bancrofti and Brugia malayi) and onchocerciasis (Onchocerca volvulus). Over 40 million suffer from disfiguring and incapacitating disease with an estimated 1.5 billion people at risk of infection, ranking filariasis as one of the major causes of global morbidity (1).A feature of filarial pathogenesis is a host inflammatory response provoked by the death of larvae and adult stages within parasitized tissues (2). All causative agents of LF and O. volvulus harbor an intracellular symbiotic bacterium, Wolbachia, and are reliant on this endosymbiont for embryogenesis, growth, and survival (3). Previous studies have determined that the inflammatory potential of B. malayi and O. volvulus is dependent on the presence of Wolbachia. For example, Wolbachia-containing filarial extracts induce activation and tolerance in murine macrophages (4, 5), activate human monocytes (6), and activate human and murine neutrophils (7, 8). In addition, O. volvulus and B. malayi extracts containing Wolbachia stimulate neutrophil recruitment to the corneal stroma and development of corneal haze in a murine model of ocular onchocerciasis, in contrast with an aposymbiotic filaria (9). Moreover, isolated Wolbachia from filaria or from insect cells can replicate these effects (8, 10). The activation of neutrophils results in further neutrophil recruitment leading to the disruption of normal corneal clarity and development of stromal haze (11).Activation and subsequent desensitization of macrophages by Wolbachia molecules has been shown to be dependent on TLR2 and the adaptor molecule MyD88 (5, 10). Further studies have established that Wolbachia-induced inflammation is dependent on TLR2 and TLR6 recognition and signaling through the MyD88/Mal pathway and are independent of TRIF and TRAM (12). However, Wolbachia ligands for TLR2/TLR6 have not been characterized. To address this, we used the TLR receptor recognition profile to identify TLR2/6 ligands in the Wolbachia genome. In this study, we demonstrate that Wolbachia-derived diacyl-lipoproteins are candidate stimulatory molecules required for TLR2/6 ligation and production of pro-inflammatory cytokine and chemokine responses. Furthermore, we show that a synthetic Wolbachia lipopeptide (Diacyl WoLP) induces TLR2/6-dependent corneal inflammation, and TLR2-dependent TNFα responses in filarial disease models and up-regulates surface markers of human lymphatic endothelium. Diacyl WoLP also induced activation and maturation of dendritic cells and generated type 1 CD4⁺ T cell and antibody responses to filarial antigens.     

Neprilysin gene expression requires binding of the amyloid precursor protein intracellular domain to its promoter: implications for Alzheimer disease

Amyloid β‐peptide (Aβ) accumulation leads to neurodegeneration and Alzheimer disease; however, amyloid metabolism is a dynamic process and enzymic mechanisms exist for Aβ removal. Considerable controversy surrounds whether the intracellular domain of the amyloid precursor protein (AICD) regulates expression of the Aβ‐degrading metalloprotease, neprilysin (NEP). By comparing two neuroblastoma cell lines differing substantially in NEP expression, we show by chromatin immunoprecipitation (ChIP) that AICD is bound directly to the NEP promoter in high NEP‐expresser (NB7) cells but not in low‐expresser (SH‐SY5Y) cells. The methylation status of the NEP promoter does not regulate expression in these cells, whereas the histone deacetylase inhibitors trichostatin A and valproate partly restore NEP expression and activity in SH‐SY5Y cells. ChIP analysis also reveals AICD binding to the NEP promoter in rat primary neurons but not in HUVEC cells. Chromatin remodelling of crucial Alzheimer disease‐related genes by valproate could provide a new therapeutic strategy.     

The Composite Task Reveals Stronger Holistic Processing in Children than Adults for Child Faces

Background

While own-age faces have been reported to be better recognized than other-age faces, the underlying cause of this phenomenon remains unclear. One potential cause is holistic face processing, a special kind of perceptual and cognitive processing reserved for perceiving upright faces. Previous studies have indeed found that adults show stronger holistic processing when looking at adult faces compared to child faces, but whether a similar own-age bias exists in children remains to be shown.

Methodology/Principal Findings

Here we used the composite face task – a standard test of holistic face processing – to investigate if, for child faces, holistic processing is stronger for children than adults. Results showed child participants (8–13 years) had a larger composite effect than adult participants (22–65 years).

Conclusions/Significance

Our finding suggests that differences in strength of holistic processing may underlie the own-age bias on recognition memory. We discuss the origin of own-age biases in terms of relative experience, face-space tuning, and social categorization.     

Disruption of Mitochondrial DNA Replication in Drosophila Increases Mitochondrial Fast Axonal Transport In Vivo

Mutations in mitochondrial DNA polymerase (pol γ) cause several progressive human diseases including Parkinson''s disease, Alper''s syndrome, and progressive external ophthalmoplegia. At the cellular level, disruption of pol γ leads to depletion of mtDNA, disrupts the mitochondrial respiratory chain, and increases susceptibility to oxidative stress. Although recent studies have intensified focus on the role of mtDNA in neuronal diseases, the changes that take place in mitochondrial biogenesis and mitochondrial axonal transport when mtDNA replication is disrupted are unknown. Using high-speed confocal microscopy, electron microscopy and biochemical approaches, we report that mutations in pol γ deplete mtDNA levels and lead to an increase in mitochondrial density in Drosophila proximal nerves and muscles, without a noticeable increase in mitochondrial fragmentation. Furthermore, there is a rise in flux of bidirectional mitochondrial axonal transport, albeit with slower kinesin-based anterograde transport. In contrast, flux of synaptic vesicle precursors was modestly decreased in pol γ−α mutants. Our data indicate that disruption of mtDNA replication does not hinder mitochondrial biogenesis, increases mitochondrial axonal transport, and raises the question of whether high levels of circulating mtDNA-deficient mitochondria are beneficial or deleterious in mtDNA diseases.     

A Structured Model of Video Reproduces Primary Visual Cortical Organisation

The visual system must learn to infer the presence of objects and features in the world from the images it encounters, and as such it must, either implicitly or explicitly, model the way these elements interact to create the image. Do the response properties of cells in the mammalian visual system reflect this constraint? To address this question, we constructed a probabilistic model in which the identity and attributes of simple visual elements were represented explicitly and learnt the parameters of this model from unparsed, natural video sequences. After learning, the behaviour and grouping of variables in the probabilistic model corresponded closely to functional and anatomical properties of simple and complex cells in the primary visual cortex (V1). In particular, feature identity variables were activated in a way that resembled the activity of complex cells, while feature attribute variables responded much like simple cells. Furthermore, the grouping of the attributes within the model closely parallelled the reported anatomical grouping of simple cells in cat V1. Thus, this generative model makes explicit an interpretation of complex and simple cells as elements in the segmentation of a visual scene into basic independent features, along with a parametrisation of their moment-by-moment appearances. We speculate that such a segmentation may form the initial stage of a hierarchical system that progressively separates the identity and appearance of more articulated visual elements, culminating in view-invariant object recognition.     

Functional Substitution by TAT-Utrophin in Dystrophin-Deficient Mice

Background

The loss of dystrophin compromises muscle cell membrane stability and causes Duchenne muscular dystrophy and/or various forms of cardiomyopathy. Increased expression of the dystrophin homolog utrophin by gene delivery or pharmacologic up-regulation has been demonstrated to restore membrane integrity and improve the phenotype in the dystrophin-deficient mdx mouse. However, the lack of a viable therapy in humans predicates the need to explore alternative methods to combat dystrophin deficiency. We investigated whether systemic administration of recombinant full-length utrophin (Utr) or ΔR4-21 “micro” utrophin (μUtr) protein modified with the cell-penetrating TAT protein transduction domain could attenuate the phenotype of mdx mice.

Methods and Findings

Recombinant TAT-Utr and TAT-μUtr proteins were expressed using the baculovirus system and purified using FLAG-affinity chromatography. Age-matched mdx mice received six twice-weekly intraperitoneal injections of either recombinant protein or PBS. Three days after the final injection, mice were analyzed for several phenotypic parameters of dystrophin deficiency. Injected TAT-μUtr transduced all tissues examined, integrated with members of the dystrophin complex, reduced serum levels of creatine kinase (11,290±920 U versus 5,950±1,120 U; PBS versus TAT), the prevalence of muscle degeneration/regeneration (54%±5% versus 37%±4% of centrally nucleated fibers; PBS versus TAT), the susceptibility to eccentric contraction-induced force drop (72%±5% versus 40%±8% drop; PBS versus TAT), and increased specific force production (9.7±1.1 N/cm² versus 12.8±0.9 N/cm²; PBS versus TAT).

Conclusions

These results are, to our knowledge, the first to establish the efficacy and feasibility of TAT-utrophin-based constructs as a novel direct protein-replacement therapy for the treatment of skeletal and cardiac muscle diseases caused by loss of dystrophin.