Still Left in the Dark : The Confrontation Clause and Child Abuse Cases After Davis v. Washington

In his concurring opinion in Crawford v. Washington, Chief Justice Rehnquist criticized the majority for holding that the Confrontation Clause applies to “testimonial” statements but leaving for “another day” any effort to define sufficiently what “testimonial” means. Prosecutors and defendants, he said, “should not be left in the dark in this manner.” Over the next two years, both sides grappled with the meaning of testimonial, each gleaning import from sections of Crawford that seemingly proved their test was the right one. When the Court granted certiorari in Davis v. Washington and Hammon v. Indiana (hereinafter Davis), hopes were high that the Court would provide the answers—the definitive test for testimonial. Not surprisingly, it did not

Still Left in the Dark : The Confrontation Clause and Child Abuse Cases After Davis v. Washington

In his concurring opinion in Crawford v. Washington, Chief Justice Rehnquist criticized the majority for holding that the Confrontation Clause applies to “testimonial” statements but leaving for “another day” any effort to define sufficiently what “testimonial” means. Prosecutors and defendants, he said, “should not be left in the dark in this manner.” Over the next two years, both sides grappled with the meaning of testimonial, each gleaning import from sections of Crawford that seemingly proved their test was the right one. When the Court granted certiorari in Davis v. Washington and Hammon v. Indiana (hereinafter Davis), hopes were high that the Court would provide the answers—the definitive test for testimonial. Not surprisingly, it did not

Interleukin-34 promotes tumorigenic signals for colon cancer cells

Colorectal carcinoma (CRC) is one of the most common forms of malignancy in the Western world. Accumulating evidence indicates that colon carcinogenesis is tightly controlled by tumour-associated immune cells and stromal cells, which can either stimulate or suppress CRC cell growth and survival, mainly via the production of cytokines. Interleukin-34 (IL-34), a cytokine known to regulate mainly monocyte/macrophage survival and function, is highly produced within the CRC microenvironment by several cell types, including cancer cells, tumour-associated macrophages (TAMs) and cancer-associated fibroblasts (CAFs), and regulates the pro-tumoural functions of such cells. In this article, we summarize the available data supporting the multiple effects of IL-34 in human CRC

CNS cell distribution and axon orientation determine local spinal cord mechanical properties.

Mechanical signaling plays an important role in cell physiology and pathology. Many cell types, including neurons and glial cells, respond to the mechanical properties of their environment. Yet, for spinal cord tissue, data on tissue stiffness are sparse. To investigate the regional and direction-dependent mechanical properties of spinal cord tissue at a spatial resolution relevant to individual cells, we conducted atomic force microscopy (AFM) indentation and tensile measurements on acutely isolated mouse spinal cord tissue sectioned along the three major anatomical planes, and correlated local mechanical properties with the underlying cellular structures. Stiffness maps revealed that gray matter is significantly stiffer than white matter irrespective of directionality (transverse, coronal, and sagittal planes) and force direction (compression or tension) (K(g) = ∼ 130 P(a) vs. K(w) = ∼ 70 Pa); both matters stiffened with increasing strain. When all data were pooled for each plane, gray matter behaved like an isotropic material under compression; however, subregions of the gray matter were rather heterogeneous and anisotropic. For example, in sagittal sections the dorsal horn was significantly stiffer than the ventral horn. In contrast, white matter behaved transversely isotropic, with the elastic stiffness along the craniocaudal (i.e., longitudinal) axis being lower than perpendicular to it. The stiffness distributions we found under compression strongly correlated with the orientation of axons, the areas of cell nuclei, and cellular in plane proximity. Based on these morphological parameters, we developed a phenomenological model to estimate local mechanical properties of central nervous system (CNS) tissue. Our study may thus ultimately help predicting local tissue stiffness, and hence cell behavior in response to mechanical signaling under physiological and pathological conditions, purely based on histological data.The authors thank the CECAD Imaging Facility (and their staff members), Andreas Christ, Jochen Guck, Jolanta Kozlowski, Ryan MacDonald, Graham Sheridan, and Alex Winkel for helpful discussions and/or technical support. This work was supported by Köln Fortune Program/Faculty of Medicine, University of Cologne (Fellowship to D.E.K.), German National Academic Foundation (Scholarship to D.E.K.), Herchel Smith Foundation (Fellowship to E.M.), DAAD-PROMOS-Program (Scholarship to J.H.), Deutsche Forschungsgemeinschaft (grant KU2760/2-1 to S.K.), UK Medical Research Council (Career Development Award to K.F.), and the Human Frontier Science Program (Young Investigator Grant to K.F.).This is the final version of the article. It first appeared from Cell Press via http://dx.doi.org/10.1016/j.bpj.2015.03.03

Spatial heterogeneity of cell-matrix adhesive forces predicts human glioblastoma migration.

BACKGROUND: Glioblastoma (GBM) is a highly aggressive incurable brain tumor. The main cause of mortality in GBM patients is the invasive rim of cells migrating away from the main tumor mass and invading healthy parts of the brain. Although the motion is driven by forces, our current understanding of the physical factors involved in glioma infiltration remains limited. This study aims to investigate the adhesion properties within and between patients' tumors on a cellular level and test whether these properties correlate with cell migration. METHODS: Six tissue samples were taken from spatially separated sections during 5-aminolevulinic acid (5-ALA) fluorescence-guided surgery. Navigated biopsy samples were collected from strongly fluorescent tumor cores, a weak fluorescent tumor rim, and nonfluorescent tumor margins. A microfluidics device was built to induce controlled shear forces to detach cells from monolayer cultures. Cells were cultured on low modulus polydimethylsiloxane representative of the stiffness of brain tissue. Cell migration and morphology were then obtained using time-lapse microscopy. RESULTS: GBM cell populations from different tumor fractions of the same patient exhibited different migratory and adhesive behaviors. These differences were associated with sampling location and amount of 5-ALA fluorescence. Cells derived from weak- and nonfluorescent tumor tissue were smaller, adhered less well, and migrated quicker than cells derived from strongly fluorescent tumor mass. CONCLUSIONS: GBM tumors are biomechanically heterogeneous. Selecting multiple populations and broad location sampling are therefore important to consider for drug testing

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

Lyophilization of Liposomal Formulations: Still Necessary, Still Challenging

Nowadays, the freeze-drying of liposome dispersions is still necessary to provide a solid dosage form intended for different routes of administration (i.e., parenteral, oral, nasal and/or pulmonary). However, after decades of studies the optimization of process conditions remains still challenging since the freezing and the dehydration destabilize the vesicle organization with the concomitant drug leakage. Starting from the thermal properties of phospholipids, this work reviews the main formulation and process parameters which can guarantee a product with suitable characteristics and increase the efficiency of the manufacturing process. In particular, an overview of the cryo- and/or lyo-protective mechanisms of several excipients and the possible use of co-solvent mixtures is provided. Attention is also focused on the imaging methods recently proposed to characterize the appearance of freeze-dried products and liposome dispersions upon reconstitution. The combination of such data would allow a better knowledge of the factors causing inter-vials variability in the attempt to improve the quality of the final medicinal product

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

GAS1 is required for NOTCH-dependent facilitation of SHH signaling in the ventral forebrain neuroepithelium

Growth arrest-specific 1 (GAS1) acts as a co-receptor to patched 1, promoting sonic hedgehog (SHH) signaling in the developing nervous system. GAS1 mutations in humans and animal models result in forebrain and craniofacial malformations, defects ascribed to a function for GAS1 in SHH signaling during early neurulation. Here, we confirm loss of SHH activity in the forebrain neuroepithelium in GAS1-deficient mice and in induced pluripotent stem cell-derived cell models of human neuroepithelial differentiation. However, our studies document that this defect can be attributed, at least in part, to a novel role for GAS1 in facilitating NOTCH signaling, which is essential to sustain a persistent SHH activity domain in the forebrain neuroepithelium. GAS1 directly binds NOTCH1, enhancing ligand-induced processing of the NOTCH1 intracellular domain, which drives NOTCH pathway activity in the developing forebrain. Our findings identify a unique role for GAS1 in integrating NOTCH and SHH signal reception in neuroepithelial cells, and they suggest that loss of GAS1-dependent NOTCH1 activation contributes to forebrain malformations in individuals carrying GAS1 mutations

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