Usp22 deficiency impairs intestinal epithelial lineage specification in vivo.

Epigenetic regulatory mechanisms play a central role in controlling gene expression during development, cell differentiation and tumorigenesis. Monoubiquitination of histone H2B is one epigenetic modification which is dynamically regulated by the opposing activities of specific ubiquitin ligases and deubiquitinating enzymes (DUBs). The Ubiquitin-specific Protease 22 (USP22) is the ubiquitin hydrolase component of the human SAGA complex which deubiquitinates histone H2B during transcription. Recently, many studies have investigated an oncogenic potential of USP22 overexpression. However, its physiological function in organ maintenance, development and its cellular function remain largely unknown. A previous study reported embryonic lethality in Usp22 knockout mice. Here we describe a mouse model with a global reduction of USP22 levels which expresses the LacZ gene under the control of the endogenous Usp22 promoter. Using this reporter we found Usp22 to be ubiquitously expressed in murine embryos. Notably, adult Usp22(lacZ/lacZ) displayed low residual Usp22 expression levels coupled with a reduced body size and weight. Interestingly, the reduction of Usp22 significantly influenced the frequency of differentiated cells in the small intestine and the brain while H2B and H2Bub1 levels remained constant. Taken together, we provide evidence for a physiological role for USP22 in controlling cell differentiation and lineage specification

Direct targeting of hippocampal neurons for apoptosis by glucocorticoids is reversible by mineralocorticoid receptor activation

Prova tipográfica (In Press)An important question arising from previous observations in vivo is whether glucocorticoids can directly influence neuronal survival in the hippocampus. To this end, a primary postnatal hippocampal culture system containing mature neurons and expressing both glucocorticoid (GR) and mineralocorticoid (MR) receptors was developed. Results show that the GR agonist dexamethasone (DEX) targets neurons (microtubule-associated protein 2-positive cells) for death through apoptosis. GR-mediated cell death was counteracted by the MR agonist aldosterone (ALDO). Antagonism of MR with spironolactone ([7a-(acetylthio)-3-oxo-17a-pregn- 4-ene,21 carbolactone] (SPIRO)) causes a dose-dependent increase in neuronal apoptosis in the absence of DEX, indicating that nanomolar levels of corticosterone present in the culture medium, which are sufficient to activate MR, can mask the apoptotic response to DEX. Indeed, both SPIRO and another MR antagonist, oxprenoate potassium ((7a,17a)-17-Hydroxy-3-oxo-7- propylpregn-4-ene-21-carboxylic acid, potassium salt (RU28318)), accentuated DEX-induced apoptosis. These results demonstrate that GRs can act directly to induce hippocampal neuronal death and that demonstration of their full apoptotic potency depends on abolition of survival-promoting actions mediated by MR

In-vivo X-ray Dark-Field Chest Radiography of a Pig

X-ray chest radiography is an inexpensive and broadly available tool for initial assessment of the lung in clinical routine, but typically lacks diagnostic sensitivity for detection of pulmonary diseases in their early stages. Recent X-ray dark-field (XDF) imaging studies on mice have shown significant improvements in imaging-based lung diagnostics. Especially in the case of early diagnosis of chronic obstructive pulmonary disease (COPD), XDF imaging clearly outperforms conventional radiography. However, a translation of this technique towards the investigation of larger mammals and finally humans has not yet been achieved. In this letter, we present the first in-vivo XDF full-field chest radiographs (32 × 35 cm²) of a living pig, acquired with clinically compatible parameters (40s scan time, approx. 80 μSv dose). For imaging, we developed a novel high-energy XDF system that overcomes the limitations of currently established setups. Our XDF radiographs yield sufficiently high image quality to enable radiographic evaluation of the lungs. We consider this a milestone in the bench-to-bedside translation of XDF imaging and expect XDF imaging to become an invaluable tool in clinical practice, both as a general chest X-ray modality and as a dedicated tool for high-risk patients affected by smoking, industrial work and indoor cooking

DOT1L Activity Promotes Proliferation and Protects Cortical Neural Stem Cells from Activation of ATF4-DDIT3-Mediated ER Stress In Vitro

Growing evidence suggests that the lysine methyltransferase DOT1L/KMT4 has important roles in proliferation, survival, and differentiation of stem cells in development and in disease. We investigated the function of DOT1L in neural stem cells (NSCs) of the cerebral cortex. The pharmacological inhibition and shRNA-mediated knockdown of DOT1L impaired proliferation and survival of NSCs. DOT1L inhibition specifically induced genes that are activated during the unfolded protein response (UPR) in the endoplasmic reticulum (ER). Chromatin-immunoprecipitation analyses revealed that two genes encoding for central molecules involved in the ER stress response, Atf4 and Ddit3 (Chop), are marked with H3K79 methylation. Interference with DOT1L activity resulted in transcriptional activation of both genes accompanied by decreased levels of H3K79 dimethylation. Although downstream effectors of the UPR, such as Ppp1r15a/Gadd34, Atf3, and Tnfrsf10b/Dr5 were also transcriptionally activated, this most likely occurred in response to increased ATF4 expression rather than as a direct consequence of altered H3K79 methylation. While stem cells are particularly vulnerable to stress, the UPR and ER stress have not been extensively studied in these cells yet. Since activation of the ER stress program is also implicated in directing stem cells into differentiation or to maintain a proliferative status, the UPR must be tightly regulated. Our and published data suggest that histone modifications, including H3K4me3, H3K14ac, and H3K79me2, are implicated in the control of transcriptional activation of ER stress genes. In this context, the loss of H3K79me2 at the Atf4- and Ddit3-promoters appears to mark a point-of-no-return that activates the death program in NSCs

Facilitated diagnosis of pneumothoraces in newborn mice using x-ray dark-field radiography.

OBJECTIVE: The aim of this study was to evaluate the diagnostic value of x-ray dark-field imaging in projection radiography-based depiction of pneumothoraces in the neonatal murine lung, a potentially life-threatening medical condition that requires a timely and correct diagnosis. MATERIALS AND METHODS: By the use of a unique preclinical model, 7-day-old C57Bl/6N mice received mechanical ventilation for 2 or 8 hours with oxygen-rich gas (FIO2 = 0.4; n = 24). Unventilated mice either spontaneously breathed oxygen-rich gas (FIO2 = 0.4) for 2 or 8 hours or room air (n = 22). At the end of the experiment, lungs were inflated with a standardized volume of air after a lethal dose of pentobarbital was administered to the pups. All lungs were imaged with a prototype grating-based small-animal scanner to acquire x-ray transmission and dark-field radiographs. Image contrast between the air-filled pleural space and lung tissue was quantified for both transmission and dark-field radiograms. After the independent expert's assessment, 2 blinded readers evaluated all dark-field and transmission images for the presence or absence of pneumothoraces. Contrast ratios, diagnostic accuracy, as well as reader's confidence and interreader agreement were recorded for both imaging modalities. RESULTS: Evaluation of both x-ray transmission and dark-field radiographs by independent experts revealed the development of a total of 10 pneumothoraces in 8 mice. Here, the contrast ratio between the air-filled pleural space of the pneumothoraces and the lung tissue was significantly higher in the dark field (8.4 ± 3.5) when compared with the transmission images (5.1 ± 2.8; P < 0.05). Accordingly, the readers' diagnostic confidence for the diagnosis of pneumothoraces was significantly higher for dark-field compared with transmission images (P = 0.001). Interreader agreement improved from moderate for the analysis of transmission images alone (κ = 0.41) to very good when analyzing dark-field images alone (κ = 0.90) or in combination with transmission images (κ = 0.88). Diagnostic accuracy significantly improved for the analysis of dark-field images alone (P = 0.04) or in combination with transmission images (P = 0.02), compared with the analysis of transmission radiographs only. CONCLUSIONS: The significant improvement in contrast ratios between lung parenchyma and free air in the dark-field images allows the facilitated detection of pneumothoraces in the newborn mouse. These preclinical experiments indicate the potential of the technique for future clinical applications

Visualization of neonatal lung injury associated with mechanical ventilation using X-ray dark-field radiography.

Mechanical ventilation (MV) and supplementation of oxygen-enriched gas, often needed in postnatal resuscitation procedures, are known to be main risk factors for impaired pulmonary development in the preterm and term neonates. Unfortunately, current imaging modalities lack in sensitivity for the detection of early stage lung injury. The present study reports a new imaging approach for diagnosis and staging of early lung injury induced by MV and hyperoxia in neonatal mice. The imaging method is based on the Talbot-Lau x-ray grating interferometry that makes it possible to quantify the x-ray small-angle scattering on the air-tissue interfaces. This so-called dark-field signal revealed increasing loss of x-ray small-angle scattering when comparing images of neonatal mice undergoing hyperoxia and MV-O2 with animals kept at room air. The changes in the dark field correlated well with histologic findings and provided superior differentiation than conventional x-ray imaging and lung function testing. The results suggest that x-ray dark-field radiography is a sensitive tool for assessing structural changes in the developing lung. In the future, with further technical developments x-ray dark-field imaging could be an important tool for earlier diagnosis and sensitive monitoring of lung injury in neonates requiring postnatal oxygen or ventilator therapy

Learning the Combinatorial Structure of Demonstrated Behaviors with Inverse Feedback Control

International audienceIn many applications, such as virtual agents or humanoid robots, it is difficult to represent complex human behaviors and the full range of skills necessary to achieve them. Real life human behaviors are often the combination of several parts and never reproduced in the exact same way. In this work we introduce a new algorithm that is able to learn behaviors by assuming that the observed complex motions can be represented in a smaller dictionary of concurrent tasks. We present an optimization formalism and show how we can learn simultaneously the dictionary and the mixture coefficients that represent each demonstration. We present results on a idealized model where a set of potential functions represents human objectives or preferences for achieving a task

Longitudinal characterization of biomarkers for spinal muscular atrophy

Objective Recent advances in understanding Spinal Muscular Atrophy (SMA) etiopathogenesis prompted development of potent intervention strategies and raised need for sensitive outcome measures capable of assessing disease progression and response to treatment. Several biomarkers have been proposed; nevertheless, no general consensus has been reached on the most feasible ones. We observed a wide range of measures over 1 year to assess their ability to monitor the disease status and progression. Methods 18 SMA patients and 19 healthy volunteers (HV) were followed in this 52‐weeks observational study. Quantitative‐MRI (qMRI) of both thighs and clinical evaluation of motor function was performed at baseline, 6, 9 and 12 months follow‐up. Blood samples were taken in patients for molecular characterization at screening, 9 and 12 month follow‐up. Progression, responsiveness and reliability of collected indices were quantified. Correlation analysis was performed to test for potential associations. Results QMRI indices, clinical scales and molecular measures showed high to excellent reliability. Significant differences were found between qMRI of SMA patients and HV. Significant associations were revealed between multiple qMRI measures and functional clinical scales. None of the qMRI, clinical, or molecular measures was able to detect significant disease progression over 1 year. Interpretation We probed a variety of quantitative measures for SMA in a slowly‐progressing disease population over 1 year. The presented measures demonstrated potential to provide a closer link to underlying disease biology as compared to conventional functional scales. The proposed biomarker framework can guide implementation of more sensitive endpoints in future clinical trials and prove their utility in search for novel disease‐modifying therapies