Heterotopic ossification (HO), the formation of bone in skeletal muscle and associated soft tissues, can result from traumatic injury or disease. The most extreme form of HO is manifested in the rare, autosomal- dominant genetic disorder, Fibrodysplasia ossificans progressiva (FOP), in which HO continues progressively throughout life, resulting in devastating effects on health and life expectancy. We developed a new genetic model of FOP based on conditional expression of the disease-causing BMP receptor, Acvr1(R206H). Using this model, we identified fibro-adipogenic progenitors (FAPs), a multipotent mesenchymal progenitor in muscle tissue, as the disease-causing cell population. Notably, we have shown that intramuscular injection of BMP2 into wild type mice also leads to FAP-derived HO, suggesting mechanistic similarities of HO pathogenesis in FOP patients and the general population. Studies of HO have focused almost entirely on mechanisms of bone growth, yet severe muscle loss can be a significant contributing factor to patient morbidity. The overarching objectives of this research are to 1) understand how FAP programming becomes subverted for pathological bone formation; 2) understand how regeneration is inhibited in skeletal muscle susceptible to HO; 3) determine whether HO of skeletal muscle is entirely dependent on FAPs. By targeting Acvr1R206H expression to FAPs using PdgrfaCreER and Tie2-Cre drivers, Aim 1 will quantify regeneration impairment, and will determine whether muscle stem cell (satellite cell; SC) dysfunction is responsible for regeneration deficits. These studies will quantify SC proliferation and apoptosis after muscle injury and will define the SC mRNA transcriptome.
Aim 1 will also determine whether Acvr1(R206H) expression in FAPs disrupts FAP-SC interactions.
Aim 2 will use RNA-Seq to define the FAP transcriptome at early, critical, times after injury to identify gene targets of Acvr1(R206H) signaling. Gro-Seq (Genome-wide Run-On) analyses will quantify changes in active gene transcription, will identify candidate genes regulated by promoter-proximal polymerase pausing, and will identify potential enhancer targets of Acvr1(R206H) signaling. Lineage tracing will determine whether the abnormal muscle environment differentially affects the fate of normal and mutant FAPs.
In Aim 3, DTA ablation approaches will determine whether FAPs are the sole source of osteogenic cells in muscle, information that is essential for evaluating the possible efficacy of cell-specific therapies. Activin inhibition has recently emerged as a powerful potential therapy for FOP.
Aim 4 will use lineage tracing to address how Activin blockade affects proliferation, survival and developmental capacity of FAPs and SCs. RNA-Seq will define the extent to which Activin inhibition ?normalizes? FAP and SC transcriptomes. The proposed research will contribute significantly to an understanding of the cells and cellular interactions responsible for HO and associated inhibition of muscle regeneration, and may lead to the development of strategies for cell-based therapies.
The proposed research will reveal underlying molecular and cellular mechanisms responsible for the ossification of skeletal muscle (heterotopic ossification), a debilitating condition caused by injury or disease. The project focuses on understanding how the offending cell population (fibro-adipogenic progenitors) become programmed for pathological bone formation, and determining how this programming negatively impacts the capacity of skeletal muscle to regenerate. These studies may lead to cell-specific therapeutic approaches to prevent heterotopic ossification.
|Lees-Shepard, John B; Goldhamer, David J (2018) Stem cells and heterotopic ossification: Lessons from animal models. Bone 109:178-186|
|Lees-Shepard, John B; Yamamoto, Masakazu; Biswas, Arpita A et al. (2018) Activin-dependent signaling in fibro/adipogenic progenitors causes fibrodysplasia ossificans progressiva. Nat Commun 9:471|