Abstract
Heterotopic ossification (HO), the formation of bone outside of the skeleton, occurs in response to severe trauma and in rare genetic diseases such as progressive osseous heteroplasia (POH). In POH, which is caused by inactivation of GNAS, a gene that encodes the alpha stimulatory subunit of G proteins (Gsα), HO typically initiates within subcutaneous soft tissues before progressing to deeper connective tissues. To mimic POH, we used conditional Gnas-null mice which form HO in subcutaneous tissues upon Gnas inactivation. In response to Gnas inactivation, we determined that prior to detection of heterotopic bone, dermal adipose tissue changed dramatically, with progressively decreased adipose tissue volume and increased density of extracellular matrix over time. Upon depletion of the adipose tissue, heterotopic bone progressively formed in those locations. To investigate the potential relevance of the tissue microenvironment for HO formation, we implanted Gnas-null or control mesenchymal progenitor cells into Gnas-null or control host subcutaneous tissues. We found that mutant cells in a Gnas-null tissue environment induced a robust HO response while little/no HO was detected in control hosts. Additionally, a Gnas-null tissue environment appeared to support the recruitment of control cells to heterotopic bone, although control cell implants were associated with less HO formation compared to mutant cells. Our data support that Gnas inactivation alters the tissue microenvironment to influence mutant and wild-type progenitor cells to contribute to HO formation.
Highlights
The formation of extra-skeletal bone within soft tissues, known as heterotopic ossification (HO), is a frequent patho-physiological response to severe tissue trauma such as combat blast injuries, high impact trauma, and hip replacements (Meyers et al, 2019)
To facilitate in vivo investigations of Heterotopic ossification (HO) initiation, we developed an approach using a mouse model with inducible postnatal homozygous deletion of Gnas that reliably forms heterotopic bone over a shorter time period
While many cells lining the surface of the ectopic bone were Ai9 positive, both Ai9-positive and Ai9-negative cells were within the ectopic bone (Figures 1D–D ), indicating the presence of unrecombined and recombined Gnas-null cells within the HO and suggesting that cells without Gnas inactivation can contribute to the heterotopic bone
Summary
The formation of extra-skeletal bone within soft tissues, known as heterotopic ossification (HO), is a frequent patho-physiological response to severe tissue trauma such as combat blast injuries, high impact trauma, and hip replacements (Meyers et al, 2019). Rare genetic disorders caused by heterozygous inactivating mutations of the GNAS locus, including progressive osseous heteroplasia (POH), are associated with HO that forms within the skin (Shore et al, 2002; Adegbite et al, 2008; Turan and Bastepe, 2015). In POH, this subcutaneous HO progresses into deeper connective tissues over time (Kaplan et al, 1994; Aynaci and Mu, 2002; Pignolo et al, 2015) Both endochondral and intramembranous mechanisms of ossification have been observed in POH patient lesions (Adegbite et al, 2008; Pignolo et al, 2015; Ware et al, 2019). Activating mutations of the GNAS locus are associated with fibrous dysplasia, a disorder of weakened skeletal structure (Zhao et al, 2018), further highlighting the critical role of the GNAS locus in the formation and maintenance of bone tissues
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