Abstract Details

Title Single-cell Transcriptomes in Facioscapulohumeral Muscular Dystrophy

Topic Neuromuscular and Clinical Neurophysiology (EMG)

Presentation(s) P9 - Poster Session 9 (12:00 PM-1:00 PM)

Poster/Presentation
Number 1-012

Objective We performed single-cell transcriptome analyses of primary cells from facioscapulohumeral muscular dystrophy (FSHD) muscle biopsies and iPSC-derived secondary myoblasts. We aim to gain mechanistic insights regarding FSHD and to identify novel biomarkers of disease progression.

Background FSHD causes progressive muscle wasting triggered by aberrant de-repression of DUX4, a pro-apoptotic transcription factor, in <1:200 muscle cells. We adopted a single-cell approach to understand 1) cellular heterogeneity and DUX4-regulated changes in rare expressing cells), 2) properties of distinct cell populations undergoing myogenic reprogramming, and 3) immune cell phenotypes that may be relevant to FSHD pathophysiology.

Design/Methods Starting with iPS cells from 8 individuals with early-onset or late-onset FSHD or controls, we induced PAX3 myogenic lineages using a gene-free growth factor protocol. This included a skeletal muscle progenitor induction stage (S1 cells), a primary myoblast stage (S2 cells), a myocyte differentiation stage (S3 cells), and induced secondary myoblasts (iSM cells) isolated and expanded from S3 myocytes. Reads were aligned to the human reference genome GRCh38 using STAR, a digital expression matrix was extracted, and clustering and expression analyses were performed using Seurat v3.

Results Proliferating myogenic cells (S1 and iSM) showed distinct single-cell gene expression patterns that also differed from those of low-passage biopsy-derived proliferating cells. Muscle progenitor cells (S1 cells) with 2-4 D4Z4 repeats (74I and 85I, early-onset phenotype) showed a higher frequency of DUX4 biomarker expression compared to S1 cells harboring 5-8 D4Z4 repeats (15A and 17A, adult-onset phenotype). Proliferating secondary myoblasts (iSM cells) and biopsy-derived cells did not express detectable DUX4 biomarkers. Further studies are in progress to characterize phenotypes of immune cells isolated from FSHD tissues.

Conclusions These results may provide insight into mechanisms contributing to the wide variance in disease severity among early-onset, adult-onset, and non-manifesting FSHD muscles and among corresponding iPSC myogenic lineages.

Authors/Disclosures
Lawrence J. Hayward, MD, PhD (UMass Chan Medical School) PRESENTER	Dr. Hayward has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Myocea, Inc. Dr. Hayward has received personal compensation in the range of $0-$499 for serving as a Consultant for HC Wainwright. Dr. Hayward has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Fulcrum Therapeutics. The institution of Dr. Hayward has received research support from Fulcrum Therapeutics. The institution of Dr. Hayward has received research support from Solve FSHD. The institution of Dr. Hayward has received research support from Myscular Dystrophy Association. The institution of Dr. Hayward has received research support from Swan Bio Therapeutics. The institution of Dr. Hayward has received research support from NIH.
	No disclosure on file
Kathryn R. Wagner, MD, PhD (The Kennedy Krieger Institute)	Dr. Wagner has received personal compensation for serving as an employee of F. Hoffmann-La Roche Ltd. Dr. Wagner has stock in F. Hoffmann-La Roche.
	No disclosure on file
	No disclosure on file

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Abstract Details