From morphological heterogeneity at alveolar level to the overall mechanical lung behavior: an in vivo microscopic imaging study.

In six male anesthetized, tracheotomized, and mechanically ventilated rabbits, we imaged subpleural alveoli under microscopic view (60×) through a "pleural window" obtained by stripping the endothoracic fascia and leaving the parietal pleura intact. Three different imaging scale levels were identified for the analysis on increasing stepwise local distending pressure (P ld) up to 16.5 cmH2O: alveoli, alveolar cluster, and whole image field. Alveolar profiles were manually traced, clusters of alveoli of similar size were identified through a contiguity-constrained hierarchical agglomerative clustering analysis and alveolar surface density (ASD) was estimated as the percentage of air on the whole image field. Alveolar area distributions were remarkably right-skewed and showed an increase in median value with a large topology-independent heterogeneity on increasing P ld. Modeling of alveolar area distributions on increasing P ld led to hypothesize that absolute alveolar compliance (change in surface area over change in P ld) increases fairly linearly with increasing initial alveolar size, the corollary of this assumption being a constant specific compliance. Clusters were reciprocally interweaved due to their highly variable complex shapes. ASD was found to increase with a small coefficient of variation (CV <25\%) with increasing P ld. The CV of lung volume at each transpulmonary pressure was further decreased (about 6\%). The results of the study suggest that the considerable heterogeneity of alveolar size and of the corresponding alveolar mechanical behavior are homogenously distributed, resulting in a substantially homogenous mechanical behavior of lung units and whole organ

Probing the mechanism for hydrogel-based stasis induction in human pluripotent stem cells : is the chemical functionality of the hydrogel important?

It is well-known that pluripotent human embryonic stem cells (hPSC) can differentiate into any cell type. Recently, we reported that hPSC colonies enter stasis when immersed in an extremely soft hydrogel comprising hydroxyl-functional block copolymer worms (I. Canton, N. J. Warren, A. Chahal, K. Amps, A. Wood, R. Weightman, E. Wang, H. Moore and S. P. Armes, ACS Centr. Sci., 2016, 2, 65–74). The gel modulus and chemical structure of this synthetic hydrogel are similar to that of natural mucins, which are implicated in the mechanism of diapause for mammalian embryos. Does stasis induction occur merely because of the very soft nature of such hydrogels or does chemical functionality also play a role? Herein, we address this key question by designing a new hydrogel of comparable softness in which the PGMA stabilizer chains are replaced with non-hydroxylated poly(ethylene glycol) [PEG]. Immunolabeling studies confirm that hPSC colonies immersed in such PEG-based hydrogels do not enter stasis but instead proliferate (and differentiate if no adhesion substrate is present). However, pluripotency is retained if an appropriate adhesion substrate is provided. Thus, the chemical functionality of the hydrogel clearly plays a decisive role in the stasis induction mechanism

Terneza, grasa intramuscular y de cobertura en carne de novillos faenados en Corrientes (Argentina)

La terneza de la carne es el atributo más apreciado por los consumidores, encontrándose condicionada por muchos factores. El objetivo de este trabajo fue generar información sobre terneza objetiva post maduración de la carne bovina y determinar valores de grasa intramuscular y de cobertura en animales de diferentes biotipos y edades faenados en Corrientes. El trabajo se realizó en un frigorífico tipo A y en la Facultad de Ciencias Veterinarias de la UNNE. Se evaluó el músculo longisimus dorsi de novillos tipo Brangus y Braford, de 4, 6 y 8 dientes. Se registró el peso individual de res caliente, así como la conformación y terminación. Las muestras divididas en dos fueron maduradas durante 7 y 14 días envasadas al vacío. La terneza se evaluó por la cizalla de Warner-Bratzler, la grasa total por el método de Soxhlet y la grasa de cobertura con un escalímetro. Se utilizó el análisis de la covarianza a tres vías incluyendo el peso de la res como covariable. El periodo de maduración afectó la terneza. Se registraron diferencias estadísticas entre tratamientos a los 14 días, no así a los 7 días. Durante este periodo, la diferencia de peso lograda se atribuyó al número de dientes y la covariable. El espesor de grasa dorsal se vio afectado por el número de dientes al igual que en la grasa intramuscular, donde además afectó la covariable. La maduración al vacío mejoró la terneza de la carne, siendo este efecto manifiesto en individuos más jóvenes, incrementándose la grasa de cobertura con la edad, no así la grasa intramuscular. La carne producto de animales faenados en la Provincia de Corrientes debe ser considerada de buena calidad, ya que según características de terneza y porcentajes de grasa intramuscular y de cobertura encontradas, responde a las más altas exigencias del mercado.The tenderness of meat is the most valued attribute for the market, being influenced by many factors. The aims of this study were to generate information on post objective tenderness values of beef maturation and to estimate both intramuscular and cover fat values in animals of different ages and biotypes slaughtered in Corrientes, Argentina. Assay was performed in a type A slaughterhouse and at the Faculty of Veterinary Science UNNE. Longisimus dorsi muscle of 4, 6 and 8 teeth Brangus and Braford steers was evaluated. Individual weight, conformation and termination on the hot carcass were registered. The samples were divided in two groups vacuum packed, maturated for 7 and 14 days. Tenderness was assessed by Warner-Bratzler shear, total fat by the Soxhlet method, and fat coverage with a scaler. We used analysis of covariance for three tracks including the carcass weight as covariable. The period of maturation affected tenderness. There were statistical differences between treatments at 14 days, but not at 7. During this period, the difference of weight achieved was attributable to number of teeth and the covariable. The back fat thickness was affected by the number of teeth as in intramuscular fat which also affected the covariable. Vacuum maturation accentuated the tenderness of beef, being more evident in younger individuals, with cover fat increasing according age; on the contrary, this effect was not observed for intramuscular fat. Meat products from animals slaughtered in Corrientes must be consid-ered of very good quality, according to the characteristics of tenderness and intramuscular as well as cover fat percentages that meet high market demands.EEA MercedesFil: Ynsaurralde Rivolta, Amanda Eugenia. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Mercedes; ArgentinaFil: Rebak, Gladis I. Universidad Nacional del Nordeste. Facultad de Ciencias Veterinarias; ArgentinaFil: Sanchez, S. Universidad Nacional del Nordeste. Facultad de Ciencias Veterinarias; ArgentinaFil: Capellari, Adriana. Universidad Nacional del Nordeste. Facultad de Ciencias Veterinarias; Argentin

Genomic analysis of the function of the transcription factor gata3 during development of the Mammalian inner ear

We have studied the function of the zinc finger transcription factor gata3 in auditory system development by analysing temporal profiles of gene expression during differentiation of conditionally immortal cell lines derived to model specific auditory cell types and developmental stages. We tested and applied a novel probabilistic method called the gamma Model for Oligonucleotide Signals to analyse hybridization signals from Affymetrix oligonucleotide arrays. Expression levels estimated by this method correlated closely (p<0.0001) across a 10-fold range with those measured by quantitative RT-PCR for a sample of 61 different genes. In an unbiased list of 26 genes whose temporal profiles clustered most closely with that of gata3 in all cell lines, 10 were linked to Insulin-like Growth Factor signalling, including the serine/threonine kinase Akt/PKB. Knock-down of gata3 in vitro was associated with a decrease in expression of genes linked to IGF-signalling, including IGF1, IGF2 and several IGF-binding proteins. It also led to a small decrease in protein levels of the serine-threonine kinase Akt2/PKB beta, a dramatic increase in Akt1/PKB alpha protein and relocation of Akt1/PKB alpha from the nucleus to the cytoplasm. The cyclin-dependent kinase inhibitor p27(kip1), a known target of PKB/Akt, simultaneously decreased. In heterozygous gata3 null mice the expression of gata3 correlated with high levels of activated Akt/PKB. This functional relationship could explain the diverse function of gata3 during development, the hearing loss associated with gata3 heterozygous null mice and the broader symptoms of human patients with Hearing-Deafness-Renal anomaly syndrome

Preliminary Evidence of “Other-Race Effect”-Like Behavior Induced by Cathodal-tDCS over the Right Occipital Cortex, in the Absence of Overall Effects on Face/Object Processing

Neuromodulation techniques such as tDCS have provided important insight into the neurophysiological mechanisms that mediate cognition. Albeit anodal tDCS (a-tDCS) often enhances cognitive skills, the role of cathodal tDCS (c-tDCS) in visual cognition is largely unexplored and inconclusive. Here, in a single-blind, sham-controlled study, we investigated the offline effects of 1.5 mA c-tDCS over the right occipital cortex of 86 participants on four tasks assessing perception and memory of both faces and objects. Results demonstrated that c-tDCS does not overall affect performance on the four tasks. However, post-hoc exploratory analysis on participants' race (Caucasian vs. non-Caucasians), showed a “face-specific” performance decrease (≈10%) in non-Caucasian participants only. This preliminary evidence suggests that c-tDCS can induce “other-race effect (ORE)-like” behavior in non-Caucasian participants that did not show any ORE before stimulation (and in case of sham stimulation). Our results add relevant information about the breadth of cognitive processes and visual stimuli that can be modulated by c-tDCS, about the design of effective neuromodulation protocols, and have important implications for the potential neurophysiological bases of ORE

Genomic and transcriptomic landscape of conjunctival melanoma.

Conjunctival melanoma (CJM) is a rare but potentially lethal and highly-recurrent cancer of the eye. Similar to cutaneous melanoma (CM), it originates from melanocytes. Unlike CM, however, CJM is relatively poorly characterized from a genomic point of view. To fill this knowledge gap and gain insight into the genomic nature of CJM, we performed whole-exome (WES) or whole-genome sequencing (WGS) of tumor-normal tissue pairs in 14 affected individuals, as well as RNA sequencing in a subset of 11 tumor tissues. Our results show that, similarly to CM, CJM is also characterized by a very high mutation load, composed of approximately 500 somatic mutations in exonic regions. This, as well as the presence of a UV light-induced mutational signature, are clear signs of the role of sunlight in CJM tumorigenesis. In addition, the genomic classification of CM proposed by TCGA seems to be well-applicable to CJM, with the presence of four typical subclasses defined on the basis of the most frequently mutated genes: BRAF, NF1, RAS, and triple wild-type. In line with these results, transcriptomic analyses revealed similarities with CM as well, namely the presence of a transcriptomic subtype enriched for immune genes and a subtype enriched for genes associated with keratins and epithelial functions. Finally, in seven tumors we detected somatic mutations in ACSS3, a possible new candidate oncogene. Transfected conjunctival melanoma cells overexpressing mutant ACSS3 showed higher proliferative activity, supporting the direct involvement of this gene in the tumorigenesis of CJM. Altogether, our results provide the first unbiased and complete genomic and transcriptomic classification of CJM

Modulation of the intrinsic neuronal excitability by multifunctional liposomes tailored for the treatment of alzheimer&#8217;s disease

Purpose: Nanotechnologies turned out to be promising in the development of diagnostic and therapeutic approaches toward neurodegenerative disorders. However, only a very scant number of nanodevices until now proved to be effective on preclinical animal models. Although specific tests in vivo are available to assess the potential toxicity of these nanodevices on cognitive functions, those to evaluate their biosafety in vitro on neurons are still to be improved. Materials and methods: We utilized the patch-clamp technique on primary cultures of cortical neural cells isolated from neonatal rats, aiming to evaluate their electrical properties after the incubation with liposomes (mApoE-PA-LIPs), previously proved able to cross the blood\u2013brain barrier and to be effective on mouse models of Alzheimer\u2019s disease (AD), both in the absence and in the presence of \u3b2-amyloid peptide oligomers. Results: Data show a high degree of biocompatibility, evaluated by lactate dehydrogenase (LDH) release and MTT assay, and the lack of cellular internalization. After the incubation with mApoE-PA-LIPs, neuronal membranes show an increase in the input resistance (from 724.14\ub176 M\u3a9 in untreated population to 886.06\ub186 M\u3a9 in the treated one), a reduction in the rheobase current (from 29.6\ub13 to 24.2\ub13 pA in untreated and treated, respectively), and an increase of the firing frequency, consistent with an ultimate increase in intrinsic excitability. Data obtained after co-incubation of mApoE-PA-LIPs with \u3b2-amyloid peptide oligomers suggest a retention of liposome efficacy. Conclusion: These data suggest the ability of liposomes to modulate neuronal electrical properties and are compatible with the previously demonstrated amelioration of cognitive functions induced by treatment of AD mice with liposomes. We conclude that this electrophysiological approach could represent a useful tool for nanomedicine to evaluate the effect of nanoparticles on intrinsic neuronal excitability

TMS-EEG Signatures of GABAergic Neurotransmission in the Human Cortex

Combining transcranial magnetic stimulation (TMS) and electroencephalography (EEG) constitutes a powerful tool to directly assess human cortical excitability and connectivity. TMS of the primary motor cortex elicits a sequence of TMS-evoked EEG potentials (TEPs). It is thought that inhibitory neurotransmission through GABA-A receptors (GABAAR) modulates early TEPs (�50 ms after TMS), whereas GABA-B receptors (GABABR) play a role for later TEPs (at�100 ms after TMS). However, the physiological underpinnings of TEPs have not been clearly elucidated yet. Here, we studied the role of GABAA/B-ergic neurotransmission for TEPs in healthy subjects using a pharmaco-TMS-EEG approach. In Experiment 1, we tested the effects of a single oral dose of alprazolam (a classical benzodiazepine acting as allosteric-positive modulator at �1, �2, �3, and �5 subunit-containing GABAARs) and zolpidem (a positive modulator mainly at the�1 GABAAR) in a double-blind, placebo-controlled, crossover study. In Experiment 2, we tested the influence of baclofen (a GABABRagonist) and diazepam (a classical benzodiazepine) versus placebo on TEPs. Alprazolam and diazepam increased the amplitude of the negative potential at 45 ms after stimulation (N45) and decreased the negative component at 100 ms (N100), whereas zolpidem increased the N45 only. In contrast, baclofen specifically increased the N100 amplitude. These results provide strong evidence that the N45 represents activity of �1-subunit-containing GABAARs, whereas the N100 represents activity of GABABRs. Findings open a novel window of opportunity to study alteration of GABAA-/GABAB-related inhibition in disorders, such as epilepsy or schizophrenia

A detailed clinical and molecular survey of subjects with nonsyndromic USH2A retinopathy reveals an allelic hierarchy of disease-causing variants.

Defects in USH2A cause both isolated retinal disease and Usher syndrome (ie, retinal disease and deafness). To gain insights into isolated/nonsyndromic USH2A retinopathy, we screened USH2A in 186 probands with recessive retinal disease and no hearing complaint in childhood (discovery cohort) and in 84 probands with recessive retinal disease (replication cohort). Detailed phenotyping, including retinal imaging and audiological assessment, was performed in individuals with two likely disease-causing USH2A variants. Further genetic testing, including screening for a deep-intronic disease-causing variant and large deletions/duplications, was performed in those with one likely disease-causing change. Overall, 23 of 186 probands (discovery cohort) were found to harbour two likely disease-causing variants in USH2A. Some of these variants were predominantly associated with nonsyndromic retinal degeneration ('retinal disease-specific'); these included the common c.2276 G>T, p.(Cys759Phe) mutation and five additional variants: c.2802 T>G, p.(Cys934Trp); c.10073 G>A, p.(Cys3358Tyr); c.11156 G>A, p.(Arg3719His); c.12295-3 T>A; and c.12575 G>A, p.(Arg4192His). An allelic hierarchy was observed in the discovery cohort and confirmed in the replication cohort. In nonsyndromic USH2A disease, retinopathy was consistent with retinitis pigmentosa and the audiological phenotype was variable. USH2A retinopathy is a common cause of nonsyndromic recessive retinal degeneration and has a different mutational spectrum to that observed in Usher syndrome. The following model is proposed: the presence of at least one 'retinal disease-specific' USH2A allele in a patient with USH2A-related disease results in the preservation of normal hearing. Careful genotype-phenotype studies such as this will become increasingly important, especially now that high-throughput sequencing is widely used in the clinical setting.European Journal of Human Genetics advance online publication, 4 February 2015; doi:10.1038/ejhg.2014.283