Abstract
Purpose: :
Ischemic injury of the retina and optic nerve is typically followed by neuroinflammation, a major contributor to neurological diseases in the retina. Activation of the multi-protein complex termed the inflammasome (IFSM), a key regulator of neuroinflammation in professional inflammatory cells, glial cells and neurons facilitates caspase1-mediated processing of the interleukin (IL)-1 cytokines IL-1β and IL-18. In this study, we aimed to characterize the role of pannexin-1 (Panx-1) on IFSM activation in the retina after ischemia-reperfusion (IR) and tested the hypothesis that the IFSM contributes to the pathogenesis of ischemic injury in a Panx-1-dependent pathway.
Methods: :
We examined the expression of IFSM components at gene and protein levels and utilized Panx1 knockout (KO) mice to test our hypothesis in a mouse model of retinal IR. Retinal ganglion cell (RGC) tolerance to IR was tested by counting the class-III β-Tubulin-labeled neurons in the ganglion cell layer in flat mounted retinas. Cell death in cultured cells was assayed using Annexin V/propidium iodide labeling.
Results: :
Our analysis revealed expression of major IFSM proteins ASC, NALP1, Caspase1 in mouse RGCs, showed IL-1β induction, processing and release after ischemia in vitro and in vivo. The analysis of ischemic wild type (WT) retinas showed robust caspase-1 activation and IL-1β production in the inner retina, particularly in RGCs. Immunohistochemistry and western blot analysis suggests that the IFSM is rapidly activated in retinal neurons and glia after IR. Morevoer, we show that this effect is dependent on the presence of functional Panx1 channels in the membrane. This activation was blocked in the KO mice. Panx1 deficiency correlated with 98% survival of RGCs, a 29% decrease relative to wild type (WT) controls. Importantly, the percentage of apoptotic cells decreased and necrotic cells were virtually eliminated in post-ischemic KO vs. WT retinas.
Conclusions: :
Panx1-dependent activation of the IFMS complex in post-ischemic retinas is required for IL-1β production and represent a novel pathway facilitating RGCs loss after ischema. Panx1 is known to integrate signaling from several neurotoxic pathways, suggesting it as a potential new target for therapy of ischemic damage to the retina
Keywords: inflammation • ganglion cells • ischemia