Abstract Title

C1Q EXPRESSION AND GLIAL ACTIVITY IN THE MOUSE RETINA FOLLOWING ISCHEMIA/REPERFUSION INJURY

Presenter Name

Sean Silverman

Abstract

We are using a mouse model whereby blood flow to the eye is blocked by raising the pressure in the eye in order to mimic damage caused by glaucoma. Our interest is to see how levels of C1q, a protein typically associated with the immune system as well as injury responsive cells of the eye are changed.

Purpose (a):

The complement cascade has become of increasing interest in several neurodegenerative diseases, including glaucoma, a leading cause of blindness. C1q has been observed as one of the earliest upregulated genes in the optic nerve head, the initial site of glaucoma injury preceding pathological changes. Here we use a glaucoma-like model of retinal ischemia/reperfusion (I/R) to mimic clinical changes in visual function and cellular loss.

Methods (b):

Deeply anesthetized C57BL/6J received a cannula to the anterior chamber of their left eye, through which their intraocular pressure (IOP) was raised to 120mmHg for 60 minutes leading to complete retinal ischemia. The cannula was then removed and blood flow was naturally reperfused. The right eye was uninjured as a contralateral control. Mice were sacrificed and enucleated at 3, 7, 14, 21, and 28 days. Eyes were fixed in 4% PFA and frozen for immunofluorescence or in situ hybridization studies. Microglia and astrocytes were identified using Iba1 and GFAP, respectively. Quantifications were performed using ImageJ Analysis software(NIH).

Results (c):

Initial changes in C1q expression were observed as early as 72 hours following injury, with a nearly two-fold increase compared to uninjured controls. Upregulated C1q was observed only in the ganglion cell (GCL) and inner plexiform (IPL) layers. Maximum intensity of C1q expression was observed 14 days post injury. Fluorescent in situ hybridization (FISH) studies reveal primarily microglia, not astrocytes, colocalized with expression of C1q in the retina.

Conclusions (d):

Following retinal I/R injury, C1q expression is actively upregulated, which appears to spatio-temporally correlate with changes in microglial, astrocyte, and Mueller cell homeostasis. Our FISH studies identify microglial cells as the primary producers of C1q following I/R injury. This suggests the elevated levels of C1q may stimulate astrocyte activation. There appears to be an interplay between microglia and astrocytes, both of which have been directly implicated in neurodegenerative diseases, including loss of RGCs in glaucoma. We propose C1q is an integral part of this mechanism, and by removing C1q we hope to preserve visual function and prevent degeneration in the visual system following injury.

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C1Q EXPRESSION AND GLIAL ACTIVITY IN THE MOUSE RETINA FOLLOWING ISCHEMIA/REPERFUSION INJURY

We are using a mouse model whereby blood flow to the eye is blocked by raising the pressure in the eye in order to mimic damage caused by glaucoma. Our interest is to see how levels of C1q, a protein typically associated with the immune system as well as injury responsive cells of the eye are changed.