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Mariacarmela Allocca, Aprotim Mazumder, Leona D Samson; Necroptosis mediates alkylation-induced retinal degeneration in mouse. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2744.
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Alkylating agents represent one class of commonly-utilized chemotherapeutic agents that generate numerous types of alkylated DNA base lesions. Although the base excision repair (BER) pathway can repair DNA alkylation damage, under certain conditions, the initiation of BER produces toxic repair intermediates that damage healthy tissues. We have shown that the alkyladenine DNA glycosylase Aag (a.k.a. Mpg), an enzyme that initiates BER, mediates alkylation-induced retinal degeneration (RD). RD is wholly prevented by Aag deficiency. The purpose of this work was to study the involvement of necroptosis in alkylation-induced RD.
RD was induced by intraperitoneal injections of the alkylating agent, methyl-methanesulfonate (MMS). Morphological and biochemical assessment of degeneration, necroptosis and inflammation was performed by electron microscopy, histology, immunofluorescence and gene expression analyses.
MMS-injected male mice showed a reduced number of photoreceptor (PR) nuclei, active gliosis, macrophages infiltration into the outer retina and vacuolated retinal pigment epithelial cells (RPE) as early as 3 days post-treatment. Ultra-structural analysis of retinas revealed that a fraction of dying PR exhibited necrotic morphology. The expression of RIP1 and RIP3, key regulators of necroptosis, was elevated in the neural retina but not RPE cells. Moreover, poly(ADP-ribose) polymerase (PARP) activity was localized to PR cells and associated with release of high-mobility group-1 (HMGB1) from PR nuclei. High levels of the pro-inflammatory cytokine TNFα and the chemokine CCl2 as well as increased levels of anti-inflammatory cytokines such, as IL-10, were also observed in the neural retina. Moreover, deficiency of IL-10 resulted in more severe RD. Female mice were partially protected from MMS-induced RD and showed reduced evidence of necroptosis and inflammation when compared to males. Aag-/- mice did not show any sign of necroptosis or inflammation.
Our findings provide in vivo evidence that alkylating agents induce sex-dependent RD mainly by induction of necroptosis in PR cells and activation of the inflammatory response. Given the extensive use of alkylating agents as chemotherapeutics and the wide range of Aag activity in the human population, understanding the molecular mechanisms underlying alkylation-induced PR cell death may be crucial to prevent potential side-effects.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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