Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant muscle disorder caused by complex genetic and epigenetic mechanisms. Previous studies showed that transcription de- repression of double homeobox protein 4 (DUX4) due to epigenetic changes in the D4Z4 region causes FSHD. The epigenetic changes are caused by either contraction of the D4Z4 array from 11-150 repeat units in unaffected individuals to 1-10 repeat units in roughly 95% of patients (FSHD1), or mutations in epigenetic regulators of the D4Z4 region (FSHD2). The expression of DUX4 leads to downstream molecular and cellular changes, which contribute to disease progression. However the cellular mechanisms cause FSHD are not clear. In our preliminary studies we identified cell membrane repair deficit in FSHD myoblasts in comparison to myoblasts from unaffected siblings. Moderate reduction of DUX4 in the FSHD cells partially improved the repair capacity. This repair deficit was also observed in skeletal muscle from an FSHD-like mouse model (FLExDUX4). It is known that FSHD myoblasts are more susceptible to oxidative stress. While reactive oxygen species (ROS) regulation plays an important role in sarcolemmal membrane repair, excessive or prolonged oxidative stress in cells lead to membrane repair deficits. Our novel finding provides a plausible link between the molecular pathways mis-regulated by DUX4 (e.g. oxidative stress) and the FSHD phenotype secondary to reduced membrane repair capacity. The goal of the study is to further investigate the observed membrane repair deficit and test the hypothesis that the membrane repair capacity is modulated by DUX4 levels in myofibers.
In aim 1, we will determine if DUX4 expression has a dose-dependent effect on sarcolemmal repair deficit using skeletal muscle-specific inducible mouse model and an antisense oligonucleotide that can modulate DUX4 expression in FLExDXU4 mice. Molecular mechanisms of repair deficits will be investigated.
In aim 2, we will evaluate the effect of DUX4 on sarcolemmal repair in FSHD patient myofibers using a xenograft mouse model of FSHD. We will produce human myofibers and test their sarcolemmal repair ability and determine whether AON-mediated knockdown of DUX4 affects the membrane repair ability. The study will identify a novel mechanism, which links previously reported molecular deficits to the disease phenotypes. In addition, the proposed study will determine whether membrane repair can be an appropriate acute readout for therapeutic approaches that aim to reduce DUX4, and its related pathways, in FSHD skeletal muscle.

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Facioscapulohumeral muscular dystrophy (FSHD) is caused by aberrant expression of double homeobox 4 (DUX4). The goal of the study is to investigate a novel cellular defect, membrane repair deficit, in FSHD using mouse and human xenograft models. In addition, we will evaluate whether the membrane repair capacity can be used as an acute readout for DUX4 reduction in muscles of these models.

National Institute of Health (NIH)
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Exploratory/Developmental Grants (R21)
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Therapeutic Approaches to Genetic Diseases Study Section (TAG)
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King, Tracy
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Children's Research Institute
United States
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