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Hair cells, the mechanosensory receptors of the inner ear, are responsible for hearing and balance. Hair cell death and consequent hearing loss are common results of treatment with ototoxic drugs, including the widely used aminoglycoside antibiotics. Induction of heat shock proteins (HSPs) confers protection against aminoglycoside-induced hair cell death via paracrine signaling that requires extracellular heat shock 70-kDa protein (HSP70). We investigated the mechanisms underlying this non-cell-autonomous protective signaling in the inner ear. In response to heat stress, inner ear tissue releases exosomes that carry HSP70 in addition to canonical exosome markers and other proteins. Isolated exosomes from heat-shocked utricles were sufficient to improve survival of hair cells exposed to the aminoglycoside antibiotic neomycin, whereas inhibition or depletion of exosomes from the extracellular environment abolished the protective effect of heat shock. Hair cell-specific expression of the known HSP70 receptor TLR4 was required for the protective effect of exosomes, and exosomal HSP70 interacted with TLR4 on hair cells. Our results indicate that exosomes are a previously undescribed mechanism of intercellular communication in the inner ear that can mediate nonautonomous hair cell survival. Exosomes may hold potential as nanocarriers for delivery of therapeutics against hearing loss., Introduction Disabling hearing loss affects approximately 6.1% of the global population (1). Most hearing loss is due to death of sensory hair cells of the inner ear. These mechanosensitive receptor [...]

Antisense oligonucleotides (ASOs) targeting pathologic RNAs have shown promising therapeutic corrections for many genetic diseases including myotonic dystrophy (DM1). Thus, ASO strategies for DM1 can abolish the toxic RNA gain-of-function mechanism caused by nucleus-retained mutant DMPK (DM1 protein kinase) transcripts containing CUG expansions (CUGexps). However, systemic use of ASOs for this muscular disease remains challenging due to poor drug distribution to skeletal muscle. To overcome this limitation, we test an arginine-rich Pip6a cell-penetrating peptide and show that Pip6aconjugated morpholino phosphorodiamidate oligomer (PMO) dramatically enhanced ASO delivery into striated muscles of DM1 mice following systemic administration in comparison with unconjugated PMO and other ASO strategies. Thus, low-dose treatment with Pip6a-PMO-CAG targeting pathologic expansions is sufficient to reverse both splicing defects and myotonia in DM1 mice and normalizes the overall disease transcriptome. Moreover, treated DM1 patient-derived muscle cells showed that Pip6a-PMO-CAG specifically targets mutant CUGexp-DMPK transcripts to abrogate the detrimental sequestration of MBNL1 splicing factor by nuclear RNA foci and consequently MBNL1 functional loss, responsible for splicing defects and muscle dysfunction. Our results demonstrate that Pip6a-PMO-CAG induces long- lasting correction with high efficacy of DM1-associated phenotypes at both molecular and functional levels, and strongly support the use of advanced peptide conjugates for systemic corrective therapy in DM1., Introduction Myotonic dystrophy (DM1), one of the most common muscular dystrophies in adults (1), is an RNA-dominant disorder caused by the expression of expanded microsatellite repeats located in the 3' [...]

Antisense oligonucleotides (AONs) hold promise for therapeutic correction of many genetic diseases via exon skipping, and the first AON-based drugs have entered clinical trials for neuromuscular disorders (1,2). However, despite advances in AON chemistry and design, systemic use of AONs is limited because of poor tissue uptake, and recent clinical reports confirm that sufficient therapeutic efficacy has not yet been achieved. Here we present a new class of AONs made of tricyclo-DNA (tcDNA), which displays unique pharmacological properties and unprecedented uptake by many tissues after systemic administration. We demonstrate these properties in two mouse models of Duchenne muscular dystrophy (DMD), a neurogenetic disease typically caused by frame-shifting deletions or nonsense mutations in the gene encoding dystrophin (3,4) and characterized by progressive muscle weakness, cardiomyopathy, respiratory failure (5) and neurocognitive impairment (6). Although current naked AONs do not enter the heart or cross the blood-brain barrier to any substantial extent, we show that systemic delivery of tcDNA-AONs promotes a high degree of rescue of dystrophin expression in skeletal muscles, the heart and, to a lesser extent, the brain. Our results demonstrate for the first time a physiological improvement of cardio-respiratory functions and a correction of behavioral features in DMD model mice. This makes tcDNA-AON chemistry particularly attractive as a potential future therapy for patients with DMD and other neuromuscular disorders or with other diseases that are eligible for exon-skipping approaches requiring whole-body treatment., Exon skipping using AONs to restore an open reading frame, thereby establishing expression of a truncated but functional dystrophin, is currently one of the most attractive approaches for the treatment [...]

X-linked agammaglobulinemia (XLA) is an inherited immunodeficiency that results from mutations within the gene encoding Bruton's tyrosine kinase (BTK). Many XLA-associated mutations affect splicing of BTK pre-mRNA and severely impair B cell development. Here, we assessed the potential of antisense, splice-correcting oligonucleotides (SCOs) targeting mutated BTK transcripts for treating XLA. Both the SCO structural design and chemical properties were optimized using 2'-0-methyl, locked nucleic acid, or phosphorodiamidate morpholino backbones. In order to have access to an animal model of XLA, we engineered a transgenic mouse that harbors a BAC with an authentic, mutated, splice-defective human BTK gene. BTK transgenic mice were bred onto a Btk knockout background to avoid interference of the orthologous mouse protein. Using this model, we determined that BTK-specific SCOs are able to correct aberrantly spliced BTK in B lymphocytes, including pro-B cells. Correction of BTK mRNA restored expression of functional protein, as shown both by enhanced lymphocyte survival and reestablished BTK activation upon B cell receptor stimulation. Furthermore, SCO treatment corrected splicing and restored BTK expression in primary cells from patients with XLA. Together, our data demonstrate that SCOs can restore BTK function and that BTK-targeting SCOs have potential as personalized medicine in patients with XLA., Introduction X-linked agammaglobulinemia (XLA) is the most common form of inherited agammaglobulinemia, accounting for 85% of all cases (1, 2). In this disease, a B cell lineage developmental block, manifested [...]

Drawing on agency and configurations theories, this study examines how perceived level of control over the entrepreneur influences venture capitalist (VC) decision making. We model the direct effects of perceived [...]

Given the increasing attention to traditionally less 'rational' entrepreneurial behaviors, such as bricolage, we used grounded theory techniques to study 23 diverse entrepreneurs. From this, we developed a five-category typology [...]

Previous attempts to identify neuroprotective targets by studying the ischemic cascade and devising ways to suppress it have failed to translate to efficacious therapies for acute ischemic stroke (1). We hypothesized that studying the molecular determinants of endogenous neuroprotection in two well-established paradigms, the resistance of CA3 hippocampal neurons to global ischemia (2) and the tolerance conferred by ischemic preconditioning (IPC) (3), would reveal new neuroprotective targets. We found that the product of the tuberous sclerosis complex 1 gene (TSC1), hamartin, is selectively induced by ischemia in hippocampal CA3 neurons. In CA1 neurons, hamartin was unaffected by ischemia but was upregulated by IPC preceding ischemia, which protects the otherwise vulnerable CA1 cells. Suppression of hamartin expression with TSC1 shRNA viral vectors both in vitro and in vivo increased the vulnerability of neurons to cell death following oxygen glucose deprivation (OGD) and ischemia. In vivo, suppression of TSC1 expression increased locomotor activity and decreased habituation in a hippocampal-dependent task. Overexpression of hamartin increased resistance to OGD by inducing productive autophagy through an mTORC1-dependent mechanism., Transient global ischemia can be caused by cardiac arrest or open heart surgery, resulting in selective and delayed cell death of CA1 hippocampal neurons (4). Notably, neighboring CA3 neurons are [...]

Silvicultural treatments that aim to improve tree growth rates also have the potential to alter physical characteristics of the tree stem and thus affect the recovery of solid-wood products. We tested the hypothesis that manifest crown asymmetry in thinned Eucalyptus nitens (Deane & Maiden) Maiden plantations was affecting the development of stem shape. The crown and stem characteristics of 15 E. nitens trees from a 22-year-old thinning trial in northeastern Tasmania were examined. The trial had been thinned 16 years previously. Lowering the intensity of local intraspecific competition through thinning increased the crown dry mass in the north-facing aspect. No direct link was found between crown dry mass distribution and stem eccentricity. The direction of pith eccentricity at 3.0 m height was confined to the northwest and southeast sectors and averaged 11%; the degree of noncircularity in stems at 3.0 m height was strongly related to the ratio of stem diameter to total height squared. These results suggest that the dynamic loading from wind exposure plays a greater role in determining the extent and direction of pith eccentricity and stem cross-sectional circularity in E. nitens than does the static load from asymmetrical crown dry mass distribution. Les traitements sylvicoles qui visent a ameliorer le taux de croissance des arbres ont aussi la capacite de modifier les caracteristiques physiques du tronc des arbres et par consequent d'avoir un effet sur la recuperation des produits en bois solide. Nous avons teste l'hypothese que l'asymetrie evidente de la cime dans les plantations eclaircies d'Eucalyptus nitens (Deane & Maiden) Maiden avait un effet sur le developpement de la forme du tronc. Les caracteristiques de la cime et du tronc de 15 tiges d'E. nitens dans un essai d'eclaircie de 22 ans situe dans le nordest de la Tasmanie ont ete etudiees. L'eclaircie avait ete effectuee 16 ans auparavant. La reduction de l'intensite de la competition intraspecifique locale due a l'eclaircie a augmente la masse anhydre de la cime du cote expose au nord. Aucun lien direct n'a ete observe entre la distribution de la masse anhydre de la cime et l'excentricite du tronc. La direction de l'excentricite de la moelle a une hauteur de 3,0 m etait confinee aux secteurs nord-ouest et sud-est et elle atteignait en moyenne 11 %; le degre de non-circularite du tronc a une hauteur de 3,0 m etait etroitement relie au rapport entre le diametre du tronc et la hauteur totale au carre. Ces resultats indiquent que la charge dynamique due a l'exposition au vent a un impact plus important sur l'ampleur et la direction de l'excentricite de la moelle et sur la circularite de la section transversale du tronc chez E. nitens que la charge statique due a la distribution asymetrique de la masse anhydre de la cime. [Traduit par la Redaction], Introduction Stem shape is a crucial variable that determines the recovery of high-value products from eucalypt logs (Shi and Walker 2006). Automated sawing and rotary peeling systems require circular logs [...]

This article develops a theoretical model that suggests that differential levels of uncertainty, knowledge relatedness, and richness of information will have a substantial impact on the decision to engage in [...]

This paper offers an integrative theory, through the use of transaction cost theory principles, that attempts to match the attributes of university-held innovations with the specific organizational form that best [...]

Within the sociology of religion there has emerged a discourse on spirituality that views contemporary developments as involving the assertion of individuals' self-authority. This perspective's theoretical roots have been persistently criticized for their conceptualization of agency; in contrast, this article draws on Bourdieu's concept of strategy to examine action in an English religious network of the sort often classified as 'New Age'. In particular; one informant is discussed in order to provide focus for an understanding of what Lahire calls sociology at the level of the individual. Her actions, better explained as strategic improvisations than as choices made on the basis of self-authority, help to illuminate the peculiarities of this religious setting, which is characterized in terms of 'nonformativeness'. By emphasizing social contextualization, this approach addresses people's meaningful actions in a way that may be applied not only more widely within the religious field but also in other fields of action. KEY WORDS authority / Bourdieu / choice / New Age movement / spirituality / secularization

The OxyR transcription factor is a key regulator of the Escherichia coli response to oxidative stress. Previous studies showed that OxyR binding to a target promoter enhances RNA polymerase binding and vice versa, suggesting a direct interaction between OxyR and RNA polymerase. To identify the region of OxyR that might contact RNA polymerase, we carried out alanine scanning and random mutagenesis of oxyR. The combination of these approaches led to the identification of several mutants defective in the activation of an OxyR target gene. A subset of the mutations map to the DNA-binding domain, other mutations appear to affect dimerization of the regulatory domain, while another group is suggested to affect disulfide bond formation. The two mutations, D142A and R273H, giving the most dramatic phenotype are located in a patch on the surface of the oxidized OxyR protein and possibly define an activating region on OxyR.

Author(s): Stuart P. Sharp (corresponding author) [1]; Andrew McGowan [2]; Matthew J. Wood [3]; Ben J. Hatchwell [1] In many cooperatively breeding birds, kin selection has an important role in [...]

Author(s): Matthew J. Wood [1]; Gisela Storz (corresponding author) [1]; Nico Tjandra (corresponding author) [2] The ability of organisms to alter their gene expression patterns in response to environmental changes [...]

Author(s): A. Graham Fieggen, Kirsten A. Donald, Dan J. Stein, Matthew J. A. Wood Author Affiliations: Africa -- first institute for neuroscience A new institute for neuroscience, the first of [...]

Results point out that the conformational dimorphism of the prion protein fragment of residues 110-136 is due to the charge state of a single histidine residue. The helical conformation of prion spans the bilayer.

The photolysis reaction of CO-bound species was investigated via time resolved absorption and time-resolved circular dichroism (TRCD) methods to determine protein folding intermediates of cytochrome c. Results of TRCD studies revealed four intermediates with two, 50, 225 and 880 microsecond lifetimes. The findings further showed that folding after two microseconds may be controlled by litigation of His26/His33 using Fe(II).

Non-replicating adenovirus vectors are being developed as vehicles for gene transfer into cells of the nervous system. An important requirement for successful gene transfer is the absence of deleterious cytotoxic or inflammatory side effects of the delivery system. Despite offering relatively stable reporter gene expression, currently available adenovirus vectors also elicit immune responses in the brain, both at the site of vector delivery and at synaptically linked distant sites. However, although an anti-viral T-lymphocyte response eliminates the vector and damages local tissue in many peripheral organs, the immune response to adenovirus in the brain is less effective and enables the vector to persist. Nevertheless, in this persistent state the adenovirus vector remains a potential target for a destructive immune response that can also cause local demyelination. The development of strategies to minimize this damaging immune response, through either vector modification or immunomodulation, will be crucial for the future success of genetic therapies in the brain.

Author(s): Matthew J.A. Wood [1]; Angela Vincent [2] Stroke is one of the most common neurological disorders, and much of the damage that follows the primary insult (hemorrhage or thrombosis) [...]

A study was conducted to examine the acute effect of caffeine on the exercise-associated increases in testosterone and cortisol. Results conclude that the anabolic effects of the increase in testosterone concentration caffeine has some potential to benefit training outcomes but can be counteracted by increase in cortisol and resulting decline in testosterone cortisol ratio.

A redox-regulated domain from the Yap1 transcription factor is utilized to trap the proteins that form cysteine sulfenic acid (Cys-SOH) in vivo. The Yap1 probe is found to be extremely efficient for monitoring Cys-SOH in any organism.

Author(s): Ed S. Lein (corresponding author) [1, 4]; Michael J. Hawrylycz [1, 4]; Nancy Ao [2]; Mikael Ayres [1]; Amy Bensinger [1]; Amy Bernard [1]; Andrew F. Boe [1]; Mark [...]

The thermodynamic basis of the disulfide/dithiol couples are examined in the pathway of the Yap1 oxidative stress signal transduction found in Saccharomyces cerevisiae. It is observed that the oxidized basis form of the Yap1 redox domain fragment, derived from the Yap1 transcription factor contains two disulfide bonds, one between Cys303 and Cys598 and one between Cys310 and Cys629.

The NMR backbone dynamics experiments are carried out on the individual fourth and fifth Epidermal Growth Factor (EGF)-like domains as well as on the wild type, the Met 388 Leu mutant, and the variant in which Met 388 is oxidized. The results presented show that changes at Met 388 cause significant changes in backbone dynamics in both the fourth and fifth EGF-like domains of thrombomodulin (TM).