Minocycline Prevents Hyperglycemia-Induced Caspase Activation in Retina of Diabetic Mice and in Retinal Müller Cells

Abstract

Abstract: : Purpose: Caspase-1 has been associated with the production of pro-inflammatory cytokines and the induction of apoptosis. We have previously shown that hyperglycemia induces caspase activation, especially the activation of caspase-1 family, in the retina of diabetic mice and humans, and retinal in Müller cells. Therefore, this study is focused on the effect of minocycline, a second-generation tetracycline and proposed caspase-1 inhibitor, on hyperglycemia-induced caspase-1 activation in retinal Müller cells and retinas of diabetic mice. Methods: Mice made diabetic using streptozotocin were injected intraperitoneal with minocycline (5mg/kg body weight) 3 or 7 times a week for 10 weeks. Transformed rat retinal Müller cells (rMC-1) were treated with 25 mM glucose in the presence or absence of minocycline (50-100 µM) for up to 5 days. Cells treated with 5 mM glucose served as controls. Caspase activities were measured by incubating whole cell lysates or retinal lysates with fluorogenic substrates specific for the individual caspases. Mitochondria mediated oxidative stress was detected using MitoTrackerRed (CM-H₂XRos) followed by confocal microscopy analysis. Apoptosis was measured using AnnexinV staining. Results: Minocycline prevented hyperglycemia-induced activation of caspase-1 and subsequent interleukin-1ß production, as well as the activation of caspase-2, 6, and 8 in retinas of diabetic mice at 2 months of diabetes in vivo. Minocycline also inhibited high glucose induced caspase activation (cas-1, 2, 3, 6, 8, and 9), production of interleukin-1ß, formation of mitochondrial superoxide, and apoptosis in retinal Müller cells. Conclusions: The results indicate that minocycline is a potent caspases inhibitor for hyperglycemia-induced caspase activation in retinal cells in vivo and in vitro. Therefore, minocycline might represent a potential new drug for investigating the role of caspases and inflammation in the pathogenesis of diabetic retinopathy.

Keywords: diabetic retinopathy • apoptosis/cell death • Muller cells