Anti-inflammatory properties of histone deacetylase inhibitors: a mechanistic study.
BACKGROUND
We have demonstrated that postshock administration of suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, can significantly improve early survival in a highly lethal model of hemorrhagic shock. As the primary insult in hemorrhagic shock is cellular hypoxia, and transcription factor hypoxia-inducible factor-1α (HIF-1α) controls proinflammatory gene expression in macrophages, we hypothesized that SAHA would attenuate the HIF-1α associated proinflammatory pathway in a hypoxic macrophage model.


METHODS
Mouse macrophages were exposed to hypoxic conditions (0.5% O2, 10% CO2, and 89.5% N2) at 37°C in the presence or absence of SAHA (10 μmol/L). The cells and culture medium were harvested at 1 hour, 4 hours, and 8 hours. Sham (no hypoxia, no SAHA) served as a control. Western blots were performed to assess protein levels of prolyl hydroxylase 2 (PHD2), HIF-1α, and inducible nitric oxide synthase (iNOS) in the cells. Colorimetric biochemical assay and enzyme-linked immunosorbent assay were used to analyze the release of nitric oxide (NO) and secretion of tumor necrosis factor α (TNF-α), respectively, in the cell culture medium.


RESULTS
Hypoxia significantly increased cellular level of HIF-1α (1 hour and 4 hours), gene transcription of iNOS (4 hours and 8 hours), iNOS protein (8 hours), NO production (8 hours), and TNF-α secretion (4 hours and 8 hours). SAHA treatment attenuated all of the above hypoxia-induced alterations in the macrophages. In addition, SAHA treatment significantly increased cellular level of PHD2, one of the upstream negative regulators of HIF-1α, at 1 hour.


CONCLUSIONS
Treatment with SAHA attenuates hypoxia-HIF-1α-inflammatory pathway in macrophages and suppresses hypoxia-induced release of proinflammatory NO and TNF-α. SAHA also causes an early increase in cellular PHD2, which provides, at least in part, a new explanation for the decrease in the HIF-1α protein levels.
Division of Trauma, Emergency Surgery and Surgical Critical Care, Department of Surgery, Massachusetts General Hospital, 165 Cambridge St., Boston, MA 02114, USA.
Wei
W
Chong
Yongqing
Y
Li
Baoling
B
Liu
Zhengcai
Z
Liu
Ting
T
Zhao
Diane R
DR
Wonsey
Changmin
C
Chen
George C
GC
Velmahos
Marc A
MA
deMoya
David R
DR
King
Andrew L
AL
Kung
Hasan B
HB
Alam
D000818:Animals; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D015153:Blotting, Western; D015687:Cell Hypoxia; D002478:Cells, Cultured; D003124:Colorimetry; D004797:Enzyme-Linked Immunosorbent Assay; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology*; D006877:Hydroxamic Acids / Q000494:pharmacology*; D008264:Macrophages / Q000187:drug effects* / Q000378:metabolism*; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D012313:RNA / Q000032:analysis; D060888:Real-Time Polymerase Chain Reaction; D012771:Shock, Hemorrhagic / Q000188:drug therapy*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D000077337:Vorinostat
D000893:Anti-Inflammatory Agents; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide; D000077337:Vorinostat; D012313:RNA; D052247:Nitric Oxide Synthase Type II