title: High mobility group Box 1 inhibits human pulmonary artery endothelial cell migration via a Toll-like receptor 4- and interferon response factor 3-dependent mechanism(s).
abstract: In pulmonary hypertension the loss of precapillary arterioles results from vascular injury causing endothelial dysfunction. Endothelial cell migration and proliferation are critical for vascular regeneration. This study focused on the effect of high mobility group box 1 protein (HMGB1) on these critical processes. HMGB1 had no effect on human pulmonary artery endothelial cell (HPAEC) proliferation. In contrast, treatment of HPAECs with HMGB1 dose-dependently inhibited VEGF-stimulated HPAEC migration. The effect of HMGB1 on HPAEC migration was TLR4-dependent because it was reversed by TLR4 siRNA or TLR4-neutralizing antibody. Exposure of HPAECs to hypoxia caused translocation and release of HMGB1 and inhibition of HPAEC migration. The effect of hypoxia on HPAEC migration was mediated by HMGB1 because HMGB1-neutralizing antibody but not control IgG restored HPAEC migration. Likewise, TLR4 siRNA but not control siRNA reversed the inhibitory effect of hypoxia in HPAECs. The canonical TLR4 signaling pathway requires the adaptor protein MyD88 and leads to downstream NFκB activation. Interestingly, HMGB1 failed to stimulate NFκB translocation to the nucleus, but instead activated an alternative pathway characterized by activation of interferon response factor 3 (IRF3). This was in contrast to human umbilical vein endothelial cells in which HMGB1 stimulated nuclear translocation of NFκB but not IRF3. IRF3 siRNA, but not MyD88 siRNA, reversed the inhibitory effect of HMGB1 on HPAEC migration. These data demonstrate that HMGB1 inhibits HPAEC migration, a critical process for vascular regeneration, via TLR4- and IRF3-dependent mechanisms.
authors.affiliation: Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213, USA.
authors.forename: Eileen M
authors.initials: EM
authors.lastname: Bauer
authors.forename: Richard
authors.initials: R
authors.lastname: Shapiro
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Billiar
authors.forename: Philip M
authors.initials: PM
authors.lastname: Bauer
mesh_terms: D015153:Blotting, Western; D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D005455:Fluorescent Antibody Technique; D024243:HMGB1 Protein / Q000502:physiology*; D006801:Humans; D050838:Interferon Regulatory Factor-3 / Q000235:genetics / Q000502:physiology*; D053594:Myeloid Differentiation Factor 88 / Q000502:physiology; D011651:Pulmonary Artery / Q000166:cytology*; D034741:RNA, Small Interfering; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000502:physiology*; D042461:Vascular Endothelial Growth Factor A / Q000502:physiology
chemical_list: D024243:HMGB1 Protein; C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; C507779:MYD88 protein, human; D053594:Myeloid Differentiation Factor 88; D034741:RNA, Small Interfering; D051197:Toll-Like Receptor 4; D042461:Vascular Endothelial Growth Factor A