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Dan-Ning Hu, Richard B Rosen, Codrin Eugen Iacob; Hypoxia induces VEGF secretion in uveal melanocytes through increased protein levels of hypoxia-inducible factors-1α. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):223. doi: https://doi.org/.
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Hypoxia leads to the accumulation of hypoxia-inducible factor-1α (HIF-1α) protein, which in turn causes the increase in VEGF secretion in various cell types; however, the effects of hypoxia on the expression of VEGF and HIF-1α in uveal melanocytes (UM) have never been reported. We hypothesize that hypoxia may stimulate the secretion of VEGF in cultured human UM via the accumulation of HIF-1α protein.
An in vitro UM hypoxia model was established by cultured cells in a sealed chamber at 37°C for 24 h in a controlled environment of 1% O2, 5% CO2 and 94% N2 using a PROOX 100 System (BioSherix, Redfield, NT). Cells cultured under standard conditions (21% O2, 5% CO2, and 74% N2) served as controls. Conditioned media and cells were then collected from cultures under normoxic and hypoxic conditions. VEGF and HIF-1α protein levels and RNA were measured using specific ELISA kits and RT-PCR analysis, respectively. One-way ANIVA was used for statistical analysis.
UM demonstrated a relatively low expression and secretion of VEGF under normoxic conditions. Hypoxia significantly increased the VEGF RNA expression (P <0.05). VEGF protein levels in the conditioned medium of UM cultured under hypoxic condition increased to 3.12-fold over that of normoxic conditions (P<0.05). Hypoxia also significantly increased HIF-1α protein levels in the protein extracted from UM (P<0.05), but not the expression of HIF-1α RNA (P > 0.05), suggesting that the increase of HIF-1α protein was due to hypoxia induced-stabilization and accumulation of HIF-1α protein rather than by an increase in the expression of HIF-1α RNA.
Hypoxia appears to induce VEGF secretion in UM through increased protein levels of hypoxia-inducible factor-1α. This mechanism may play a role in the pathogenesis of choroid neovascular membrane in the age-related macular degeneration.
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