title: Inhibition of Hif1α prevents both trauma-induced and genetic heterotopic ossification.
abstract: Pathologic extraskeletal bone formation, or heterotopic ossification (HO), occurs following mechanical trauma, burns, orthopedic operations, and in patients with hyperactivating mutations of the type I bone morphogenetic protein receptor ACVR1 (Activin type 1 receptor). Extraskeletal bone forms through an endochondral process with a cartilage intermediary prompting the hypothesis that hypoxic signaling present during cartilage formation drives HO development and that HO precursor cells derive from a mesenchymal lineage as defined by Paired related homeobox 1 (Prx). Here we demonstrate that Hypoxia inducible factor-1α (Hif1α), a key mediator of cellular adaptation to hypoxia, is highly expressed and active in three separate mouse models: trauma-induced, genetic, and a hybrid model of genetic and trauma-induced HO. In each of these models, Hif1α expression coincides with the expression of master transcription factor of cartilage, Sox9 [(sex determining region Y)-box 9]. Pharmacologic inhibition of Hif1α using PX-478 or rapamycin significantly decreased or inhibited extraskeletal bone formation. Importantly, de novo soft-tissue HO was eliminated or significantly diminished in treated mice. Lineage-tracing mice demonstrate that cells forming HO belong to the Prx lineage. Burn/tenotomy performed in lineage-specific Hif1α knockout mice (Prx-Cre/Hif1α(fl:fl)) resulted in substantially decreased HO, and again lack of de novo soft-tissue HO. Genetic loss of Hif1α in mesenchymal cells marked by Prx-cre prevents the formation of the mesenchymal condensations as shown by routine histology and immunostaining for Sox9 and PDGFRα. Pharmacologic inhibition of Hif1α had a similar effect on mesenchymal condensation development. Our findings indicate that Hif1α represents a promising target to prevent and treat pathologic extraskeletal bone.
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Shailesh
authors.initials: S
authors.lastname: Agarwal
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Shawn
authors.initials: S
authors.lastname: Loder
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Cameron
authors.initials: C
authors.lastname: Brownley
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: David
authors.initials: D
authors.lastname: Cholok
authors.affiliation: Department of Orthopedic Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Laura
authors.initials: L
authors.lastname: Mangiavini
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: John
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Christopher
authors.initials: C
authors.lastname: Breuler
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Hsiao H
authors.initials: HH
authors.lastname: Sung
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Shuli
authors.initials: S
authors.lastname: Li
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Kavitha
authors.initials: K
authors.lastname: Ranganathan
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109;
authors.forename: Joshua
authors.initials: J
authors.lastname: Peterson
authors.affiliation: Department of Surgery, Massachusetts General Hospital, Boston, MA 02114;
authors.forename: Ronald
authors.initials: R
authors.lastname: Tompkins
authors.affiliation: Department of Surgery, Shriners Hospital for Children and University of Texas Medical Branch, Galveston, TX 77555;
authors.forename: David
authors.initials: D
authors.lastname: Herndon
authors.affiliation: Department of Surgery, Genome Technology Center, Stanford University, Palo Alto, CA 94305;
authors.forename: Wenzhong
authors.initials: W
authors.lastname: Xiao
authors.affiliation: Department of Developmental Biology, Harvard Dental School, Boston, MA 02115;
authors.forename: Dolrudee
authors.initials: D
authors.lastname: Jumlongras
authors.affiliation: Department of Developmental Biology, Harvard Dental School, Boston, MA 02115;
authors.forename: Bjorn R
authors.initials: BR
authors.lastname: Olsen
authors.affiliation: Regenerative Medicine Department, Naval Medical Research Center, Silver Spring, MD 20910;
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Davis
authors.affiliation: Department of Biologic and Materials Sciences, University of Michigan, Ann Arbor, MI 48109.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Mishina
authors.affiliation: Department of Orthopedic Surgery, University of Michigan, Ann Arbor, MI 48109; eschipan@med.umich.edu blevi@umich.edu.
authors.forename: Ernestina
authors.initials: E
authors.lastname: Schipani
authors.affiliation: Department of Surgery, University of Michigan, Ann Arbor, MI 48109; eschipan@med.umich.edu blevi@umich.edu.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Levi
mesh_terms: D030201:Activin Receptors, Type I / Q000378:metabolism; D000273:Adipose Tissue / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D002056:Burns / Q000150:complications / Q000235:genetics; D020219:Chondrogenesis / Q000187:drug effects / Q000235:genetics; D004195:Disease Models, Animal; D053263:Gene Regulatory Networks / Q000187:drug effects; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D019426:Integrases / Q000378:metabolism; D008163:Luminescent Measurements; D059630:Mesenchymal Stem Cells / Q000187:drug effects; D018345:Mice, Knockout; D008954:Models, Biological; D009150:Mustard Compounds / Q000494:pharmacology; D009999:Ossification, Heterotopic / Q000000981:diagnostic imaging / Q000188:drug therapy / Q000235:genetics* / Q000517:prevention & control*; D010666:Phenylpropionates / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020796:Receptor, Platelet-Derived Growth Factor alpha / Q000378:metabolism; D055758:SOX9 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000494:pharmacology; D013710:Tendons / Q000187:drug effects / Q000473:pathology / Q000601:surgery; D058971:Tenotomy; D015854:Up-Regulation / Q000187:drug effects; D014945:Wound Healing / Q000187:drug effects; D014947:Wounds and Injuries / Q000150:complications* / Q000473:pathology; D055114:X-Ray Microtomography
chemical_list: C492908:2-amino-3-(4'-N,N-bis(2-chloroethyl)amino)phenylpropionic acid N-oxide; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009150:Mustard Compounds; D010666:Phenylpropionates; D012333:RNA, Messenger; D055758:SOX9 Transcription Factor; D020796:Receptor, Platelet-Derived Growth Factor alpha; D030201:Activin Receptors, Type I; C045073:Cre recombinase; D019426:Integrases; D020123:Sirolimus
keywords: HIF1α; Prx; cartilage; heterotopic ossification; mesenchymal condensation