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Bongsu Kim, Rania Kusibati, Elaine Binkley, Mohamed H Abdel-Rahman, Colleen M Cebulla; Up-Regulation of VEGF by Hyaluronic Acid in Microglia. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2261.
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© ARVO (1962-2015); The Authors (2016-present)
Vascular endothelial growth factor (VEGF) expression is critical to many diseases at the vitreoretinal interface. Inflammatory cells, particularly macrophages and microglia, play an important role in disease modulation and may be a source of VEGF. We investigated the hypotheses that vitreous component hyaluronic acid (HA) and sheared HA (HAS - low molecular weight) induce VEGF expression in murine microglia and that AZD 6244, a MAPK/ERK kinase (MEK) inhibitor down-regulates this VEGF expression.
Murine microglial BV-2 cells or primary microglia isolated from CNS were treated with HA and HAS in serum free media (SFM), with or without AZD6244 or DMSO vehicle. Total RNA, cell lysate and culture supernatant were collected at 6 and 12 hours. Real time q-PCR was performed to evaluate gene expression. Western blot and ELISA assays were performed to evaluate the protein expression.
At 12 hours, BALBC primary microglia show Vegf expression increased by 1.56-fold in HAS but not HA treatment. HA and HAS induced Vegf gene expression in BV-2 microglia by 1.27- and 1.44-fold, respectively. VEGF protein on Western blot was increased 3.09-fold in HA and 2.22-fold in HAS compared to SFM. HAS, but not HA, also increased VEGF expression 1.48-fold at 6hr. The ELISA assay result shows that HA and HAS with DMSO induce VEGF expression in cell culture supernatant by 4.01-fold and 3.97-fold at 6hours and 1.51-fold and 3.13-fold at 12hours, respectively. AZD6244 decreased VEGF supernatant concentrations in HA and HAS to that in SFM with DMSO in all time points.
HA and HAS induced VEGF expression in murine microglia. MEK inhibitor AZD6244 decreased VEGF supernatant concentrations in BV-2 cells treated by HA and HAS.
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