Histone deacetylase 4 is crucial for proper skeletal muscle development and disease

Epigenetics plays a pivotal role in modulating gene response to physiological or pathological stimuli. Histone Deacetylase inhibitors (HDACi) have been used in the treatment of various cancers1, are ef-fective in several animal models of neurodegenerative diseases, including amyotrophic lateral scle-rosis (ALS), and are currently in clinical trial to promote muscle repair in muscular dystrophies2. However, long-term use of pan-HDAC inhibitors is not tolerated3. The assignment of distinct biologi-cal functions to individual HDACs in skeletal muscle is a prerequisite to improve the efficacy of pharmacological treatments based on HDACi. HDAC4 is a member of class II HDACs that mediates many cellular responses. Clinical reports suggest that inhibition of HDAC4 can be beneficial to cancer cachexia, dystrophic or ALS patients. All the above conditions are characterized by progressive mus-cle wasting and up-regulation of HDAC4 expression in skeletal muscle, suggesting a potential role for this protein in regulating these diseases. To study the role of HDAC4 with a genetic approach, we generated several models of muscle disease in mice lacking HDAC4 in skeletal muscle: cancer ca-chexia, by implanting Lewis lung carcinoma (LLC), muscular dystrophy, by using mdx mice, or ALS, by using SODG93A mice. Lack of HDAC4 worsens skeletal muscle atrophy induced by both LLC and ALS, demonstrated by a reduction in muscle mass and myofibers size. Conversely, dystrophic mice lacking HDAC4 in skeletal muscle show an increased number of necrotic myofibers and run less efficiently than mdx mice. The aggravation of the dystrophic phenotype may be partially due to the impairment in skeletal muscle regeneration observed in mice lacking HDAC4 in skeletal muscle. Our results indi-cate that HDAC4 is necessary for maintaining skeletal muscle homeostasis and function. Current studies aim to investigate the molecular mechanisms underlying the role of HDAC4 in skeletal mus-cle maintenance in response to cancer cachexia, ALS or muscular dystrophy

mRNA-mediated delivery of gene editing tools to human primary muscle stem cells

Muscular dystrophies are about 50 devastating untreatable monogenic diseases leading to progressive muscle degeneration and atrophy. Gene correction of transplantable cells using CRISPR/Cas9-based tools is a realistic scenario for autologous cell replacement therapies to restore organ function in many genetic disorders. However, muscle stem cells have so far lagged behind due to the absence of methods to isolate and propagate them and their susceptibility to extensive ex vivo manipulations. Here, we show that mRNA-based delivery of SpCas9 and an adenine base editor results in up to >90% efficient genome editing in human muscle stem cells from many donors regardless of age and gender, and without any enrichment step. Using NCAM1 as an endogenous reporter locus expressed by all muscle stem cells and whose knock-out does not affect cell fitness, we show that cells edited with mRNA fully retain their myogenic marker signature, proliferation capacity and functional attributes. Moreover, mRNA-based delivery of a base editor led to highly efficient repair of a muscular dystrophy-causing SGCA mutation in a single selection-free step. In sum, our work establishes mRNA-mediated delivery of CRISPR/Cas9-based tools as a promising and universal approach for taking gene edited muscle stem cells into clinical application to treat muscle disease

Implications of Clock Distribution Faults and Issues with Screening Them During Manufacturing Testing

Based on real process data of a reference microprocessor, fault models are derived for the manufacturing defects most likely to affect signals of the clock distribution network. Their probability is estimated with Inductive Fault Analysis performed on the actual layout of the reference microprocessor. The effects of the most likely faults have been evaluated by electrical level simulations. We have found that, contrary to common assumptions, only a small percentage of such faults result in catastrophic failures easily detected during manufacturing testing. On the contrary, the majority of such faults lead to local failures not likely to be detected during manufacturing testing, despite their possibly compromising the microprocessor operation and reliability. In particular, we have found that the clock faults can be detected during manufacturing testing in only 12 percent of cases. Even more surprisingly, we have also found that, in 10 percent of cases, the undetected clock faults also invalidate the testing procedure itself

Scan flip-flops with on-line testing ability with respect to input delay and crosstalk faults

We propose a possible modification to the internal structure of scan flip-flops, which allows the online detection of delay and crosstalk faults affecting their input. Our solution allows to obtain, together with the flip-flop output datum, an indication denoting whether or not the provided datum is incorrect, because of an input crosstalk or delay fault. The proposed solution features self-checking ability with respect to a wide set of possible internal faults, including node stuck-ats, transistor stuck-ons and stuck-opens, resistive bridgings, delays, transient and crosstalk faults

On-Chip Clock Faults' Detector

This paper proposes an on-chip detector for the on-line testing of faults affecting clock signals and making them change with incorrect duty-cycle. Our scheme is particularly suitable to be integrated within Systems-On-a-Chip (SOCs), in order to avoid their possible incorrect operation because of faults affecting clock signals, thus solving their extreme criticality in clock faults' testing. In particular, our detector is suitable to be applied to clock signals within each SOC digital core, to the clock signals at the interface between the diverse cores, as well as to those driving the DFT and BIST structures used to perform the SOC test. Our scheme features self-checking ability with respect to its possible internal faults belonging to a realistic set including stuck-ats, transistor stuck-ons, stuck-opens and resistive bridgings