Abstract
Inflammation and immune-mediated processes are pivotal to the pathogenic progression of age-related macular degeneration (AMD). Although plasma levels of C-reactive protein (CRP) have been shown to be associated with an increased risk for AMD, the pathophysiological importance of the prototypical acute-phase reactant in the etiology of the disease is unknown, and data regarding the exact role of CRP in ocular inflammation are limited. In this study, we provide mechanistic insight into how CRP contributes to the development of AMD. In particular, we show that monomeric CRP (mCRP) but not the pentameric form (pCRP) upregulates IL-8 and CCL2 levels in retinal pigment epithelial cells. Further, we show that complement factor H (FH) binds mCRP to dampen its proinflammatory activity. FH from AMD patients carrying the “risk” His402 polymorphism displays impaired binding to mCRP, and therefore proinflammatory effects of mCRP remain unrestrained.
Highlights
In plasma, CRP exists as a cyclic non-covalent pentamer of 125-kDa composed of five identical Ca2+-stabilized subunits, which are placed in the corners of a regular pentagon19. (See Supplemental Figure 1A for a representation of the three-dimensional (3D) crystal structure of pentameric CRP; notice that a phosphocholine molecule is bound to each of the five independent binding sites)[20]
The data supports our hypothesis that mCRP contributes to the development of Age-related macular degeneration (AMD) through direct proinflammatory effects on retinal pigment epithelial cells, and that this proinflammatory activity is unchecked in subjects with the “at risk” Tyr402His polymorphism in CFH, due to an impaired interaction with mCRP
To determine the effect of CRP isoforms on the expression of inflammatory mediators, ARPE-19 cells were exposed for 24 h to different concentrations of either mCRP or pCRP. mRNA levels of IL8 and CCL2 were determined by quantitative RT-polymerase chain reaction (PCR), and protein concentrations of both cytokines in the supernatants were quantified by Enzyme-linked immunosorbent assay (ELISA)
Summary
CRP exists as a cyclic non-covalent pentamer of 125-kDa composed of five identical Ca2+-stabilized subunits, which are placed in the corners of a regular pentagon19. (See Supplemental Figure 1A for a representation of the three-dimensional (3D) crystal structure of pentameric CRP (pCRP); notice that a phosphocholine molecule is bound to each of the five independent binding sites)[20]. To determine the effect of CRP isoforms on the expression of inflammatory mediators, ARPE-19 cells were exposed for 24 h to different concentrations of either mCRP or pCRP. Similar to the effect on IL-8, only mCRP significantly increased CCL2 mRNA expression levels in a dose-dependent manner, reaching significance at 5 μg/mL (Fig. 1C).
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