Monocyte-induced cytokine expression in cultured human retinal pigment epithelial cells

Abstract

Monocytes and retinal pigment epithelial cells are intimately associated in membranes of eyes with proliferative vitreoretinopathy and in certain types of uveitis. The goal of this study was to determine whether monocytes modulate cytokine expression in retinal pigment epithelial cells, and if so, to identify the monocyte products responsible for this effect. Cultured human retinal pigment epithelial cells were exposed to varying concentrations of monocyte-conditioned medium from unstimulated human monocytes for 1-48 hr, or from monocytes prestimulated with lipopolysaccharide. mRNA expression of interleukin-1 beta, interleukin-6, interleukin-8, melanoma growth stimulating activity/gro alpha and gamma, macrophage colony stimulating factor, transforming growth factor-beta 2, basic fibroblast growth factor and activin beta A chain was determined by reverse transcription polymerase chain reaction. Protein secretion of selected cytokines, interleukin-1 beta, interleukin-6, interleukin-8, macrophage colony stimulating factor and transforming growth factor-beta 2 was measured in RPE-conditioned medium by ELISA. Retinal pigment epithelial cells constitutively expressed mRNA for interleukin-6, macrophage colony stimulating factor, transforming growth factor-beta 2, basic fibroblast growth factor and activin beta A chain. Interleukin-1 beta, melanoma growth stimulating activity/gro alpha and gamma and interleukin-8 were not expressed under basal conditions. Stimulated monocyte-conditioned medium markedly induced mRNA of all cytokines except basic fibroblast growth factor and transforming growth factor-beta 2 in a dose- and time-dependent manner. Unstimulated monocyte-conditioned medium was a less potent inducing agent, but still enhanced mRNA expression of interleukin-6, interleukin-8 and melanoma growth stimulating activity/gro alpha. Stimulated monocyte-conditioned medium also induced a time-dependent increase in interleukin-6, Interleukin-8, macrophage colony stimulation factor and transforming growth factor-beta 2, but not interleukin-1 beta protein secretion (p < 0.05 for all time points). Neutralizing antibodies to interleukin-1 beta, or tumour necrosis factor alpha, but not interleukin-1 alpha, significantly reduced cytokine mRNA expression induced by stimulated monocyte-conditioned medium. The combination of all three neutralizing antibodies almost entirely eliminated monocyte-induced mRNA expression and protein production of all cytokines studied. Activated monocytes secrete a heterogeneous mixture of products that together strongly induce expression of multiple cytokines in human retinal pigment epithelial cells. Most if not all of the inducing effect can be accounted for by interleukin-1 beta and tumour necrosis factor alpha. Because cytokines have been implicated in proliferative vitreoretinopathy and uveitis, monocyte-mediated cytokine expression by RPE cells may serve to initiate and perpetuate these diseases.