Abstract
Retinal pigment epithelial (RPE) cells increase in size and multinucleate during aging. We have shown using human and mouse cell lines that oxidised photoreceptor outer segments (oxPOS)-induced cytokinesis failure is related to RPE cell multinucleation, although the underlying mechanism remains unknown. This study investigated the role of the PKC pathway in oxPOS-induced RPE multinucleation using ARPE19 cells. oxPOS treatment promoted PKC activity and upregulated the mRNA expression of PKC α, δ, ζ, ι and μ. Inhibition of PKCα with Gö6976 resulted in a 33% reduction of multinucleate ARPE19 cells, whereas inhibition of PKCζ with Gö6983 led to a 50% reduction in multinucleate ARPE19 cells. Furthermore, oxPOS treatment induced a PKCζ-dependent upregulation of the Cdk inhibitor p27kip1, its inhibition using A2CE reduced oxPOS-induced ARPE19 multinucleation. Our results suggest that oxPOS-induced ARPE19 cytokinesis failure is, at least in part, due to the upregulation of p27kip1 through activating the PKC, particularly PKCζ pathway. Targeting the PKCζ-p27kip1 signalling axis may be a novel approach to restore RPE repair capacity during aging.
Highlights
Multinucleate cells, i.e., multiple nuclei share one common cytoplasm, are frequently observed in various patho-physiological conditions, including development, aging, inflammation and malignant tumor
PMA, a Protein Kinase C (PKC) activator, and photoreceptor outer segments (POS) or oxidised photoreceptor outer segments (oxPOS) did not affect the expression of total PKC protein (Fig. 1A-1C), the expression was suppressed by a cocktail of PKC isoform inhibitors (Fig. 1A, 1B)
We have shown that POS and oxPOS increased reactive oxygen species (ROS) production and DNA oxidation in Retinal pigment epithelial (RPE) cells, and we postulate that oxPOS-induced DNA oxidation may contribute to RPE multinucleation [8]
Summary
Multinucleate cells, i.e., multiple nuclei share one common cytoplasm, are frequently observed in various patho-physiological conditions, including development, aging, inflammation and malignant tumor. Multinucleate cells can be induced by the fusion of multiple cells or formed by nuclear division that is not followed by cytokinesis [1]. Ts’o and Friedman’s landmark studies in the late 1960’s noted a variable increase in RPE cell size as well as multinucleation with age in humans [5]. Al-Hussaini et al further reported multinucleate RPE cells in proximity to drusen and they are greater in number in AMD compared to age matched healthy controls [6]. We reported an age-dependent increase in the number and size of multinucleate RPE cells in mice [8]. The underlying mechanism related to age-induced multinucleation of RPE cells remains poorly defined. Del Priore [9] showed that there is little evidence for overall cell loss in the human RPE with age [9] and Al Hussaini [10] observed few dividing RPE in rats with BrdU
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