Effects of Cytokine Activation and Oxidative Stress on the Function of the Human Embryonic Stem Cell–Derived Retinal Pigment Epithelial Cells

Kati Juuti-Uusitalo, Monica Nieminen, Felix Treumer, Minna Ampuja, Anne Kallioniemi, Alexa Klettner, Heli Skottman
2015 Investigative Ophthalmology and Visual Science  
Citation: Juuti-Uusitalo K, Nieminen M, Treumer F, et al. Effects of the cytokine activation and oxidative stress to the function of the human embryonic stem cell derived retinal pigment epithelial cells. Invest Ophthalmol Vis Sci. 2015;56:6265-6274. PURPOSE. In several retinal complications, such as age-dependent macular degeneration (AMD), oxidative stress is increased and cytokine level is elevated. These are shown to alter the activation and expression of matrix metalloproteinase (MMP) both
more » ... oteinase (MMP) both in human primary and immortalized retinal pigment epithelial (RPE) cells. However, the effects on human embryonic stem cell (hESC)-derived RPE cells remain to be elucidated. METHODS. The mature hESC-RPE cells were exposed to inflammatory cytokines (IFN-c or TNF-a) for 24 hours or oxidative stress (H 2 O 2 ) for 1 hour. Effects on barrier properties were analyzed with transepithelial electrical resistance (TEER), the expression of MMP-1, MMP-2, MMP-3, MMP-9, collagen I, and collagen IV genes with quantitative RT-PCR, and the expression of MMP-1 and MMP-3 proteins with Western blot or ELISA, respectively. Also, activation and secretion of MMP-2 and -9 proteins were analyzed with zymography. RESULTS. In normal state, mature hESC-RPE cells expressed MMP-1, -2, -3, and -9 genes in low levels, respectively. Tumor necrosis factor-a increased MMP-1 and -2 gene expression, and H 2 O 2 increased MMP-3 and -9 gene expression. Zymography revealed IFN-c-and TNF-ainduced secretion of MMP-2 and high-molecular-weight species of MMP (HMW MMP), but H 2 O 2 decreased their secretion. Furthermore, TNF-a and H 2 O 2 significantly decreased barrier properties. CONCLUSIONS. Here, cytokines induced the MMP-1 and -2 gene and protein expression. Also, H 2 O 2 induced MMP-3 and -9 gene expression, but not their protein secretion. These data propose that under oxidative stress and cytokine stimuli, mature hESC-RPE cells resemble their native counterpart in the human eye in regard to MMP secretion and expression and could be used to model retinal disorders involving alterations in MMP activity such as AMD, diabetic retinopathy, or proliferative vitreoretinopathy in vitro.
doi:10.1167/iovs.15-17333 pmid:26431480 fatcat:6vazbw2gvveithbzxxpetkxoya