The HIF-1a/PLOD2 axis integrates extracellular matrix organization and cell metabolism leading to aberrant musculoskeletal repair.

While hypoxic signaling has been shown to play a role in many cellular processes, its role in metabolism linked extracellular matrix (ECM) organization and downstream processes of cell fate after musculoskeletal injury remains to be determined. Heterotopic ossification (HO) is a debilitating condition where abnormal bone formation occurs within extra-skeletal tissues. Hypoxia and hypoxia-inducible factor 1a (HIF-1a) activation have been shown to promote HO. However, the underlying molecular mechanisms by which the HIF-1a pathway in mesenchymal progenitor cells (MPCs) contributes to pathologic bone formation remain to be elucidated. Here we used a proven mouse injury-induced HO model to investigate the role of HIF-1a on aberrant cell fate. Using single-cell RNA-sequencing (scRNA-Seq), we found that collagen ECM organization is the most highly up-regulated biological process in MPCs. Zeugopod mesenchymal cell-specific deletion of Hif1a (Hoxa11-CreERT2; Hif1afl/fl) significantly mitigated HO in vivo. ScRNA-Seq analysis of these Hoxa11-CreERT2; Hif1afl/fl mice identified the PLOD2/LOX pathway for collagen cross-linking as downstream of the HIF-1a regulation of HO. Importantly, our scRNA-seq data and mechanistic studies further uncovered that glucose metabolism in MPCs is most highly impacted by HIF-1a deletion. Overall design: To investigate the effects of Hoxa11(+) lineage specific deletion of Hif1a on burn/tenotomy injury-induced heterotopic ossification, cells from tenotomy inury site were isolated at 7 days post-injury for scRNA-seq. Sample A1,B1: 10 week old male C57BL/6 mice were subjected to the burn/tenotomy procedure. Samples were collected 7 days post-injury