Clearance of dying ARPE-19 cells by professional and nonprofessional phagocytesin vitro- implications for age-related macular degeneration (AMD)

Abstract

ABSTRACT.Purpose: Failure of retinal pigment epithelial (RPE) cells and macrophages to engulf different dying cells in the retina may result in accumulation of debris and development of age-related macular degeneration (AMD). The dynamics and influence of different treatments on this clearance process can be studied in vitro using human ARPE-19 cells and macrophages as phagocytes modelling dry and wet type of AMD, respectively.Methods: Death through extracellular matrix detachment using polyHEMA-coated surfaces (anoikis) and UV irradiation (apoptosis) was induced in ARPE-19 cells. Twocoloured phagocytic assays were performed to quantify the amount of dying cells phagocytes engulfed (flow cytometry) and for visualization (fluorescent and scanning electron microscopy). The effect of phosphatidylserine inhibition with recombinant annexin-V and glucocorticoid (triamcinolone) treatment on the phagocytic process was tested.Results: The clearance of anoikic and apoptotic cells by nondying ARPE-19 cells over 8 hr of co-incubation increased over time (at 8 hr, over 53% and 35% of the phagocytes contained engulfed dying cells, respectively). The human macrophages engulfed the anoikic and apoptotic ARPE-19 cells with seven and four times lower capacity, respectively. Phosphatidylserine appearance on the dying cells did not affect, but triamcinolone treatment enhanced the phagocytosis of the dying cells by macrophages.Conclusions: ARPE-19 cells are more efficient in clearing anoikic than UV-induced apoptotic cells. Macrophages are less efficient in the clearance process than ARPE-19 cells. The present model can be used for studying both dry and wet type of AMD in vitro and for testing different pharmacological aspects affecting this disease.

Differential effects of bevacizumab, ranibizumab and aflibercept on cell viability, phagocytosis and mitochondrial bioenergetics of retinal pigment epithelial cell

“…) and the development of AMD (Petrovski et al. ). In our study, exposure to bevacizumab, but not aflibercept or ranibizumab, suppressed the phagocytotic activity of ARPE‐19 cells.…”

“…) and AMD pathology (Petrovski et al. ). Both phagocytosis and autophagy are dynamic, multistep processes that can be modulated at several points (Valapala et al.…”

Aging is not a disease: Distinguishing age-related macular degeneration from aging

“…At the same time, with age macrophages are increasingly prevalent in the choroid and express complement receptor CRIg (Xu et al, 2009), which is involved in complement-mediated phagocytosis (Gorgani et al, 2008; Helmy et al, 2006). The finding that ARPE-19 cells are better-able to phagocytose apoptotic cells than are macrophages in vitro (Petrovski et al, 2011) allows for the possibility that age-related alterations in macrophages together with reduced viability of RPE lends to chronic para-inflammation in the retinal tissues.…”

NETs: the missing link between cell death and systemic autoimmune diseases?

“…Apoptotic cells also expose “eat-me” signals on their surface (e.g., phosphatidylserine, PS) that are recognized by phagocytes through specific engulfment receptors. Interestingly, other forms of death including anoikis (death induced by detachment of anchorage-dependent cells), autophagic cell death, caspase-independent apoptosis, and necroptosis also expose PS as a mechanism for efficient non-inflammatory clearance by phagocytes (Hirt and Leist, 2003; Brouckaert et al, 2004; Petrovski et al, 2007a, 2011). In addition, cells dying by anoikis and necroptosis are efficiently engulfed by PS-independent process (Hirt et al, 2000; Petrovski et al, 2007a, 2011).…”