UNC569-induced Morphological Changes in Pigment Epithelia and Photoreceptor Cells in the Retina through MerTK Inhibition in Mice

Abstract

Mer proto-oncogene tyrosine kinase (MerTK), which is expressed in the retinal pigment epithelium (RPE), regulates phagocytosis of shed photoreceptor outer segments (POS). To investigate the effects of drug-induced MerTK inhibition on the retina, UNC569, a specific MerTK inhibitor, was orally administered to male mice at a concentration of 60, 100, or 150 mg/kg for up to 14 days. Furthermore, MerTK inhibition in the retinal tissue sample was examined using a phosphorylation assay following a single dose of UNC569 at 100 mg/kg. In electron microscopic examination, UNC569 at 100 mg/kg or more increased phagosomes and phagolysosomes in the RPE. In addition, UNC569 at 150 mg/kg increased chromatin-condensed nuclei in the outer nuclear layer, indicating the early phase of apoptosis of photoreceptor cells. MiR-183, miR-96, and miR-124, which are enriched in photoreceptor cells, were elevated in the plasma of mice following treatment of 150-mg/kg UNC569, in conjunction with the photoreceptor lesion. Additionally, 100-mg/kg UNC569 inhibited MerTK phosphorylation in the retina. These results suggest that MerTK inhibition impaired phagocytic function of the retina, leading to accumulation of shed POS within the POS layer and increasing phagosomes and phagolysosomes in the RPE to delay POS renewal, resulting in apoptosis of photoreceptor cells.

Targeting Tyro3, Axl and MerTK (TAM receptors): implications for macrophages in the tumor microenvironment

“…UNC569 induces apoptosis, improves sensitivity to cytotoxic chemotherapy and reduces colony formation in ALL cell lines [146]. UNC569 has also been utilized in vivo to study phagocytosis by retinal pigment epithelium cells [151]. UNC1062, as well as UNC569, induces apoptosis and decreases cell growth in AML cell lines [147].…”

The Impact of the Timing of Dosing on the Severity of UNC569-Induced Ultrastructural Changes in the Mouse Retina

“…The previous study demonstrated that the physiological MerTK phosphorylation continued from ZT2 to ZT5, and UNC569 inhibited the phosphorylation for 17 h after 100 mg/kg of dosing. 14 Because of this 17 h inhibition period, the inhibitory activity in the ZT22 UNC569 group was thought to have persisted up to ZT15 (ZT22 + 17 h = ZT39 = ZT15) and those in the ZT5.5 UNC569 groups was up to ZT22.5 (ZT5.5 + 17 h = ZT22.5). The former period included the duration from ZT2 to ZT5, the duration of physiological MerTK phosphorylation, whereas the latter period did not (Figure 5).…”

“…Ultrastructurally, membrane-bound multilamellar bodies in the RPE increased in both the ZT5.5 and ZT22 UNC569 groups after 14- and 28-day administration similar to our previous study. 14 These structures in the RPE were considered to be phagosomes and phagolysosomes, and a variety of appearances including electron-lucent areas and electron-dense cores were suggested to have reflected the different stages of the intracellular digestion process. Interestingly, dilatation of the ER in the RPE, indicative of the ER stress, and chromatin-condensed nuclei of the photoreceptor cells, and indicative of the early phase of apoptosis were observed only in the ZT22 UNC569 group after the 28-day administration.…”

“…The dose levels were set based on our previous results, in which ultrastructural changes in the RPE were observed without death or moribundity in animals during the 14-day administration period. 14 The first day of dosing was defined as day 1. The animals were euthanized by exsanguinations under isoflurane anesthesia and necropsied at ZT2 on day 15 or 29.…”

“…Recently, we reported that UNC569, a specific MerTK inhibitor, 13 induces ultrastructural changes both in the RPE and the photoreceptor cells of mice. 14 Since physiological MerTK phosphorylation and RPE phagocytosis are synchronized with circadian rhythms, we hypothesized that mice demonstrate diurnal variation in MerTK-induced retinal toxicity, that is, chronotoxicity. In the present study, ultrastructural changes in the retina were investigated in mice orally administered UNC569 at 2 different Zeitgeber times (ZT) to evaluate the effects of dosing time on the retinal toxicity caused by MerTK inhibition.…”