Preventive Effects of Rapamycin on Inflammation and Capillary Degeneration in a Rat Model of NMDA-Induced Retinal Injury

Abstract

Inhibitors of the mammalian target of rapamycin (mTOR) have been shown to protect against neuronal injury, but the mechanisms underlying this effect are not fully understood. The present study aimed to examine the effects of rapamycin, an inhibitor of the mTOR pathway, on inflammation and capillary degeneration in a rat model of N-methyl-D-aspartate (NMDA)-induced retinal neurotoxicity. Inflammation and capillary degeneration were evaluated by counting the numbers of CD45-positive leukocytes and Iba1-positive microglia, and by measuring the length of empty basement membrane sleeves, respectively. Marked increases in the numbers of leukocytes and microglia were observed 1 d after intravitreal injection of NMDA (200 nmol), and significant capillary degeneration was observed after 7 d. These NMDA-induced changes were significantly reduced by the simultaneous injection of rapamycin (20 nmol) with NMDA. These results suggest that rapamycin has preventive effects on inflammation and capillary degeneration during retinal injury.Key words endothelial cell; excitotoxicity; leukocyte; mammalian target of rapamycin; microglia Glutamate is a major excitatory neurotransmitter, and excessive extracellular levels of glutamate cause neuronal cell death. The neurotoxicity induced by elevated glutamate levels is implicated in some ocular diseases, including diabetic retinopathy and glaucoma. 1,2) In many cases, glutamate-induced neurotoxicity has been predominantly attributed to overstimulation of N-methyl-D-aspartate (NMDA) receptors. 3,4) In addition to direct effects on neurons, indirect effects, such as upregulation of pro-inflammatory cytokines and recruitment of leukocytes into the retina, are involved in NMDA-induced retinal neuronal damage. 5,6) Recent studies have shown that rapamycin, an inhibitor of the mammalian target of rapamycin (mTOR), exerts neuroprotective and axon growth-promoting effects during neuronal injury. 7,8) The mTOR is a serine/threonine kinase that regulates a wide array of cellular functions, including cell proliferation and cell-cycle control. 9) There are two distinct functional mTOR complexes, mTOR complex 1 and 2 (mTORC1 and mTORC2). Rapamycin inhibits mTORC1, which prevents phosphorylation of at least 2 well-characterized effectors: the p70S6 kinases (S6K1 and S6K2) and eIF4E-binding proteins (4E-BP1). 10) Inactivation of mTORC1 can cause autophagy, which has an important role in cellular homeostasis. 11) A role for autophagy has been suggested as a neuroprotective mechanism, whereby it enhances the clearance of harmful protein aggregates. 12,13) In addition, rapamycin could prevent inflammatory responses during brain injury, including activation of microglia. 14) Thus, multiple mechanisms might be involved in the neuroprotective effects of rapamycin.In the present study, we examined the effects of rapamycin on inflammation and capillary degeneration in a rat model of retinal neurotoxicity induced by overstimulation of NMDA receptors. MATERIALS AND METHODSAnimals Male Sprague-Dawley rats weighi...

Protective Effects of Everolimus against <i>N</i>-Methyl-D-aspartic Acid-Induced Retinal Damage in Rats

“…34) Previous studies showed that the suppressive effect of rapamycin on inflammatory responses, such as infiltration of leukocytes and activation of microglia, might prevent secondary retinal damage. 13) Similarly, in the present study, we found that co-injection of everolimus with NMDA prevented increases in CD45-positive cells and Iba1-positive cells and the morphologic transformation of Iba1-positive cells from the ramified form to an amoeboid shape 24 h after NMDA injection. The reduction of inflammatory responses by everolimus may contribute to the prevention of additional retinal damage.…”

“…13,22) The numbers of CD45-positive cells and Iba1-positve cells were counted 24 h after intravitreal injection of vehicle (n=8), NMDA (n=4) or NMDA+everolimus (n=5).…”

“…12,13,22) Briefly, systemic vascular perfusion was performed 6 or 24 h after the intravitreal injection. Following perfusion, the eyes were removed and stored in the fixative for 1 h at 4°C.…”

“…[9][10][11] Our recent studies have shown that rapamycin, an inhibitor of the mammalian target of rapamycin (mTOR), exerts protective effects on NMDA-induced retinal damage in rats. 12,13) mTOR is a serine/threonine kinase that signals through two distinct functional mTOR complexes, mTOR complexes 1 and 2 (mTORC1 and mTORC2). 14) Rapamycin inhibits mTORC1, which prevents phosphorylation of at least 2 well-characterized effectors: the p70S6 kinases (S6K1 and S6K2) and eIF4E-binding proteins (4E-BP1).…”

“…[17][18][19][20] In addition, rapamycin could prevent neuroinflammatory responses, including activation of microglia. 13,21) Multiple mechanisms are likely involved in the neuroprotective effects of rapamycin.…”