Assembly of the Auditory Circuitry by a Hox Genetic Network in the Mouse Brainstem

Rhombomeres (r) contribute to brainstem auditory nuclei during development. Hox genes are determinants of rhombomere-derived fate and neuronal connectivity. Little is known about the contribution of individual rhombomeres and their associated Hox codes to auditory sensorimotor circuitry. Here, we show that r4 contributes to functionally linked sensory and motor components, including the ventral nucleus of lateral lemniscus, posterior ventral cochlear nuclei (VCN), and motor olivocochlear neurons. Assembly of the r4-derived auditory components is involved in sound perception and depends on regulatory interactions between Hoxb1 and Hoxb2. Indeed, in Hoxb1 and Hoxb2 mutant mice the transmission of low-level auditory stimuli is lost, resulting in hearing impairments. On the other hand, Hoxa2 regulates the Rig1 axon guidance receptor and controls contralateral projections from the anterior VCN to the medial nucleus of the trapezoid body, a circuit involved in sound localization. Thus, individual rhombomeres and their associated Hox codes control the assembly of distinct functionally segregated sub-circuits in the developing auditory brainstem

Design and development of topical liposomal formulations in a regulatory perspective

The skin is the absorption site for drug substances intended to treat loco-regional diseases, although its barrier properties limit the permeation of drug molecules. The growing knowledge of the skin structure and its physiology have supported the design of innovative nanosystems (e.g. liposomal systems) to improve the absorption of poorly skin-permeable drugs. However, despite the dozens of clinical trials started, few topically applied liposomal systems have been authorized both in the EU and the USA. Indeed, the intrinsic complexity of the topically applied liposomal systems, the higher production costs, the lack of standardized methods and the more stringent guidelines for assessing their benefit/risk balance can be seen as causes of such inefficient translation. The present work aimed to provide an overview of the physicochemical and biopharmaceutical characterization methods that can be applied to topical liposomal systems intended to be marketed as medicinal products, and the current regulatory provisions. The discussion highlights how such methodologies can be relevant for defining the critical quality attributes of the final product, and they can be usefully applied based on the phase of the life cycle of a liposomal product: to guide the formulation studies in the early stages of development, to rationally design preclinical and clinical trials, to support the pharmaceutical quality control system and to sustain post-marketing variations. The provided information can help define harmonized quality standards able to overcome the case-by-case approach currently applied by regulatory agencies in assessing the benefit/risk of the topically applied liposomal systems. Graphical abstract: [Figure not available: see fulltext.

Cortical cell stiffness is independent of substrate mechanics.

Cortical stiffness is an important cellular property that changes during migration, adhesion and growth. Previous atomic force microscopy (AFM) indentation measurements of cells cultured on deformable substrates have suggested that cells adapt their stiffness to that of their surroundings. Here we show that the force applied by AFM to a cell results in a significant deformation of the underlying substrate if this substrate is softer than the cell. This 'soft substrate effect' leads to an underestimation of a cell's elastic modulus when analysing data using a standard Hertz model, as confirmed by finite element modelling and AFM measurements of calibrated polyacrylamide beads, microglial cells and fibroblasts. To account for this substrate deformation, we developed a 'composite cell-substrate model'. Correcting for the substrate indentation revealed that cortical cell stiffness is largely independent of substrate mechanics, which has major implications for our interpretation of many physiological and pathological processes

Reciprocal regulation between Smad7 and Sirt1 in the gut

In inflammatory bowel disease (IBD) mucosa, there is over-expression of Smad7, an intracellular inhibitor of the suppressive cytokine transforming growth factor-β1, due to post-transcriptional mechanisms that enhance Smad7 acetylation status thus preventing ubiquitination-mediated proteosomal degradation of the protein. IBD-related inflammation is also marked by defective expression of Sirt1, a class III NAD+-dependent deacetylase, which promotes ubiquitination-mediated proteosomal degradation of various intracellular proteins and triggers anti-inflammatory signals. The aim of our study was to determine whether, in IBD, there is a reciprocal regulation between Smad7 and Sirt1. Smad7 and Sirt1 were examined in mucosal samples of IBD patients and normal controls by Western blotting and immunohistochemistry, and Sirt1 activity was assessed by a fluorimetric assay. To determine whether Smad7 is regulated by Sirt1, normal or IBD lamina propria mononuclear cells (LPMC) were cultured with either Sirt1 inhibitor (Ex527) or activator (Cay10591), respectively. To determine whether Smad7 controls Sirt1 expression, ex vivo organ cultures of IBD mucosal explants were treated with Smad7 sense or antisense oligonucleotide. Moreover, Sirt1 expression was evaluated in LPMC isolated from Smad7-transgenic mice given dextran sulfate sodium (DSS). Upregulation of Smad7 was seen in both the epithelial and lamina propria compartments of IBD patients and this associated with reduced expression and activity of Sirt1. Activation of Sirt1 in IBD LPMC with Cay10591 reduced acetylation and enhanced ubiquitination-driven proteasomal-mediated degradation of Smad7, while inhibition of Sirt1 activation in normal LPMC with Ex527 increased Smad7 expression. Knockdown of Smad7 in IBD mucosal explants enhanced Sirt1 expression, thus suggesting a negative effect of Smad7 on Sirt1 induction. Consistently, mucosal T cells of Smad7-transgenic mice contained reduced levels of Sirt1, a defect that was amplified by induction of DSS colitis. The data suggest the existence of a reciprocal regulatory mechanism between Smad7 and Sirt1, which could contribute to amplify inflammatory signals in the gut

Mechanosensing is critical for axon growth in the developing brain.

During nervous system development, neurons extend axons along well-defined pathways. The current understanding of axon pathfinding is based mainly on chemical signaling. However, growing neurons interact not only chemically but also mechanically with their environment. Here we identify mechanical signals as important regulators of axon pathfinding. In vitro, substrate stiffness determined growth patterns of Xenopus retinal ganglion cell axons. In vivo atomic force microscopy revealed a noticeable pattern of stiffness gradients in the embryonic brain. Retinal ganglion cell axons grew toward softer tissue, which was reproduced in vitro in the absence of chemical gradients. To test the importance of mechanical signals for axon growth in vivo, we altered brain stiffness, blocked mechanotransduction pharmacologically and knocked down the mechanosensitive ion channel piezo1. All treatments resulted in aberrant axonal growth and pathfinding errors, suggesting that local tissue stiffness, read out by mechanosensitive ion channels, is critically involved in instructing neuronal growth in vivo.This work was supported by the German National Academic Foundation (scholarship to D.E.K.), Wellcome Trust and Cambridge Trusts (scholarships to A.J.T.), Winston Churchill Foundation of the United States (scholarship to S.K.F.), Herchel Smith Foundation (Research Studentship to S.K.F.), CNPq 307333/2013-2 (L.d.F.C.), NAP-PRP-USP and FAPESP 11/50761-2 (L.d.F.C.), UK EPSRC BT grant (J.G.), Wellcome Trust WT085314 and the European Research Council 322817 grants (C.E.H.); an Alexander von Humboldt Foundation Feodor Lynen Fellowship (K.F.), UK BBSRC grant BB/M021394/1 (K.F.), the Human Frontier Science Program Young Investigator Grant RGY0074/2013 (K.F.), the UK Medical Research Council Career Development Award G1100312/1 (K.F.) and the Eunice Kennedy Shriver National Institute Of Child Health & Human Development of the National Institutes of Health under Award Number R21HD080585 (K.F.).This is the author accepted manuscript. The final version is available from Nature Publishing Group via https://doi.org/10.1038/nn.439

Innovation in Phytotherapy : is a New Regulation the Feasible Perspective in Europe?

Classical multicomponent preparations mostly derived from traditional usages in Western and Eastern phytotherapy have been under-evaluated for a long time as potential new pharmaceutical products. The regulatory scenario, in particular at the European level, has only recently considered these aspects proposing harmonized guidelines for the pharmaceutical registration of traditional herbal products. Nevertheless, a specific regulation for innovative products based on the combination of precious knowledge arising from traditional usages and modern scientific advancements is still missing. In this paper, we propose a critical review of the current situation with the specific aim of contributing to create a more favorable regulatory environment for the pharmaceutical registration of new and innovative herbal medicinal products

Tbet Expression in Regulatory T Cells Is Required to Initiate Th1-Mediated Colitis

In normal conditions gut homeostasis is maintained by the suppressive activity of regulatory T cells (Tregs), characterized by the expression of the transcription factor FoxP3. In human inflammatory bowel disease, which is believed to be the consequence of the loss of tolerance toward antigens normally contained in the gut lumen, Tregs have been found to be increased and functionally active, thus pointing against their possible role in the pathogenesis of this immune-mediated disease. Though, in inflammatory conditions, Tregs have been shown to upregulate the T helper (Th) type 1-related transcription factor Tbet and to express the pro-inflammatory cytokine IFN\u3b3, thus suggesting that at a certain point of the inflammatory process, Tregs might contribute to inflammation rather than suppress it. Starting from the observation that Tregs isolated from the lamina propria of active but not inactive IBD patients or uninflamed controls express Tbet and IFN\u3b3, we investigated the functional role of Th1-like Tregs in the dextran sulfate model of colitis. As observed in human IBD, Th1-like Tregs were upregulated in the inflamed lamina propria of treated mice and the expression of Tbet and IFN\u3b3 in Tregs preceded the accumulation of conventional Th1 cells. By using a Treg-specific Tbet conditional knockout, we demonstrated that Tbet expression in Tregs is required for the development of colitis. Indeed, Tbet knockout mice developed milder colitis and showed an impaired Th1 immune response. In these mice not only the Tbet deficient Tregs but also the Tbet proficient conventional T cells showed reduced IFN\u3b3 expression. However, Tbet deficiency did not affect the Tregs suppressive capacity in vitro and in vivo in the adoptive transfer model of colitis. In conclusion here we show that Tbet expression by Tregs sustains the early phase of the Th1-mediated inflammatory response in the gut

Performance and characteristics of the Newborn Hearing Screening Program in Campania region (Italy) between 2013 and 2019

Purpose: Universal newborn hearing screening (UNHS) in the first month of life is crucial for facilitating both early hearing detection and intervention (EHDI) of significant permanent hearing impairment (PHI). In Campania region, UNHS has been introduced in 2003 by the Regional Council Resolution and started on January 2007. The aim of this paper is to update a previous article describing the performance of the program since its implementation in the period between 2013 and 2019. Methods: A longitudinal retrospective study was carried at the Regional Reference Center III on 350,178 babies born in the analysis period. The paper reports the main results of overall coverage, referral rate, lost-to-follow-up rate,yield for PHI and shall determine various risk factor associations with hearing impairment Results: In Campania region, 318,878 newborns were enrolled at I level, with a coverage rate of 91.06%, 301,818 (86.18%) Well Infant Nurseries (WIN) and 17,060 (5.35%) Neonatal Intensive Care Unit (NICU) babies. PHI was identified in 413 children, 288 (69.73%) bilaterally and 125 (30.26%) unilaterally. The overall cumulative incidence rate of PHI was 1.29 per 1000 live-born infants (95% CI 1.17–1.42) with a quite steady tendency during the whole study period. Conclusions: This study confirms the feasibility and effectiveness of UNHS in Campania region also in a setting with major socioeconomic and health organization restrictions.The program meets quality benchmarks to evaluate the progress of UNHS. Nowadays, it is possible to achieve an early diagnosis of all types of HL avoiding the consequences of hearing deprivation