Hemin and iron increase synthesis and trigger export of xanthine oxidoreductase from hepatocytes to the circulation
Evan R. DeVallance,
Heidi M. Schmidt,
Madison Seman,
Sara E. Lewis,
Katherine C. Wood,
Schuyler D. Vickers,
Scott A. Hahn,
Murugesan Velayutham,
Emily A. Hileman,
Dario A. Vitturi,
Roberta Leonardi,
Adam C. Straub,
Eric E. Kelley
Affiliations
Evan R. DeVallance
Center for Inhalation Toxicology, West Virginia University School of Medicine, Morgantown, WV, USA; Department of Physiology and Pharmacology, Health Sciences Center, West Virginia University, Morgantown, WV, USA
Heidi M. Schmidt
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA
Madison Seman
Department of Physiology and Pharmacology, Health Sciences Center, West Virginia University, Morgantown, WV, USA
Sara E. Lewis
Department of Physiology and Pharmacology, Health Sciences Center, West Virginia University, Morgantown, WV, USA
Katherine C. Wood
Heart, Lung, Blood and Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, PA, USA
Schuyler D. Vickers
Department of Biochemistry, West Virginia University, Morgantown, WV, 26505, USA
Scott A. Hahn
Heart, Lung, Blood and Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, PA, USA
Murugesan Velayutham
Department of Biochemistry, West Virginia University, Morgantown, WV, 26505, USA
Emily A. Hileman
Department of Physiology and Pharmacology, Health Sciences Center, West Virginia University, Morgantown, WV, USA
Dario A. Vitturi
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA; Heart, Lung, Blood and Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, PA, USA
Roberta Leonardi
Department of Biochemistry, West Virginia University, Morgantown, WV, 26505, USA
Adam C. Straub
Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA; Heart, Lung, Blood and Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, PA, USA; Corresponding author. University of Pittsburgh School of Medicine, Department of Pharmacology and Chemical Biology, Heart, Lung, Blood and Vascular Medicine Institute, E1254 Biomedical Science Tower, 200 Lothrop St., Pittsburgh, PA, 15216, USA.
Eric E. Kelley
Department of Physiology and Pharmacology, Health Sciences Center, West Virginia University, Morgantown, WV, USA; Corresponding author. West Virginia University School of Medicine, Department of Physiology and Pharmacology, 3074 Health Sciences Center North, 1 Medical Center Dr., Morgantown, WV, 15902, USA.
We recently reported a previously unknown salutary role for xanthine oxidoreductase (XOR) in intravascular heme overload whereby hepatocellular export of XOR to the circulation was identified as a seminal step in affording protection. However, the cellular signaling and export mechanisms underpinning this process were not identified. Here, we present novel data showing hepatocytes upregulate XOR expression/protein abundance and actively release it to the extracellular compartment following exposure to hemopexin-bound hemin, hemin or free iron. For example, murine (AML-12 cells) hepatocytes treated with hemin (10 μM) exported XOR to the medium in the absence of cell death or loss of membrane integrity (2.0 ± 1.0 vs 16 ± 9 μU/mL p controls for both, p < 0.05) and that silencing either or TLR4 with siRNA prevented hemin-induced XOR upregulation (p < 0.01). Finally, to confirm direct action of these transcription factors on the Xdh gene, chromatin immunoprecipitation was performed indicating that hemin significantly enriched (∼5-fold) both Sp1 and NF-kB near the transcription start site. In summary, our study identified a previously unknown pathway by which XOR is upregulated via SP1/NF-kB and subsequently exported to the extracellular environment. This is, to our knowledge, the very first study to demonstrate mechanistically that XOR can be specifically targeted for export as the seminal step in a compensatory response to heme/Fe overload.
