A human immune/muscle xenograft model of FSHD muscle pathology.

Background: Facioscapulohumeral muscular dystrophy (FSHD) disease progression is associated with muscle inflammation, although its role in FSHD muscle pathology is unknown. Methods: We have developed a novel humanized mouse strain, NSG-SGM3-W41, that supports the co- engraftment of human hematopoietic stem cells (HSCs) and muscle myoblasts as an experimental model to investigate the role of innate immunity in FSHD muscle pathology. Results: The NSG-SGM3-W41 mouse supports the selective expansion of human innate immune cell lineages following engraftment of human HSCs and the co-engraftment and differentiation of patient-derived FSHD or control muscle myoblasts. Immunohistological and NanoString RNA expression assays establish that muscle xenografts from three FSHD subjects were immunogenic compared to those from unaffected first-degree relatives. FSHD muscle xenografts preferentially accumulated human macrophages and B cells and expressed early complement genes of the classical and alternative pathways including complement factor C3 protein, which is a mediator of early complement function through opsonization to mark damaged cells for macrophage engulfment. FSHD muscle xenografts also underwent immune donor dependent muscle turnover as assayed by human spectrin ß1 immunostaining of muscle fibers and by NanoString RNA expression assays of muscle differentiation genes. Conclusions: The NSG-SGM3-W41 mouse provides an experimental model to investigate the role of innate immunity and complement in FSHD muscle pathology and to develop FSHD therapeutics targeting DUX4 and the innate immunity inflammatory responses.

Generation and molecular characterization of human pluripotent stem cell-derived pharyngeal foregut endoderm.

Approaches to study human pharyngeal foregut endoderm-a developmental intermediate that is linked to various human syndromes involving pharynx development and organogenesis of tissues such as thymus, parathyroid, and thyroid-have been hampered by scarcity of tissue access and cellular models. We present an efficient stepwise differentiation method to generate human pharyngeal foregut endoderm from pluripotent stem cells. We determine dose and temporal requirements of signaling pathway engagement for optimized differentiation and characterize the differentiation products on cellular and integrated molecular level. We present a computational classification tool, "CellMatch," and transcriptomic classification of differentiation products on an integrated mouse scRNA-seq developmental roadmap confirms cellular maturation. Integrated transcriptomic and chromatin analyses infer differentiation stage-specific gene regulatory networks. Our work provides the method and integrated multiomic resource for the investigation of disease-relevant loci and gene regulatory networks and their role in developmental defects affecting the pharyngeal endoderm and its derivatives.

Human 'brite/beige' adipocytes develop from capillary networks, and their implantation improves metabolic homeostasis in mice.

Uncoupling protein 1 (UCP1) is highly expressed in brown adipose tissue, where it generates heat by uncoupling electron transport from ATP production. UCP1 is also found outside classical brown adipose tissue depots, in adipocytes that are termed 'brite' (brown-in-white) or 'beige'. In humans, the presence of brite or beige (brite/beige) adipocytes is correlated with a lean, metabolically healthy phenotype, but whether a causal relationship exists is not clear. Here we report that human brite/beige adipocyte progenitors proliferate in response to pro-angiogenic factors, in association with expanding capillary networks. Adipocytes formed from these progenitors transform in response to adenylate cyclase activation from being UCP1 negative to being UCP1 positive, which is a defining feature of the beige/brite phenotype, while displaying uncoupled respiration. When implanted into normal chow-fed, or into high-fat diet (HFD)-fed, glucose-intolerant NOD-scid IL2rg(null) (NSG) mice, brite/beige adipocytes activated in vitro enhance systemic glucose tolerance. These adipocytes express neuroendocrine and secreted factors, including the pro-protein convertase PCSK1, which is strongly associated with human obesity. Pro-angiogenic conditions therefore drive the proliferation of human beige/brite adipocyte progenitors, and activated beige/brite adipocytes can affect systemic glucose homeostasis, potentially through a neuroendocrine mechanism.