Purpose: We demonstrated previously that pro-survival insulin receptor, PI3K-Akt, and p70 S6K signaling is diminished in models of diabetic retinopathy. As mammalian target of rapamycin (mTOR), an upstream activator of p70 S6Kinase is, in part, regulated by lipid-derived second messengers, such as phosphatidic acid (PA), we sought to determine if diminished mTOR/p70 S6Kinase signaling in diabetic retinas may reflect diminished PA levels.
Methods: Alterations in PA mass from retinas of control and streptozotocin-induced diabetic rats were determined by mass spectrometry. The biochemical and biophysical mechanisms underlying the actions of PA on insulin-activated mTOR/p70 S6Kinase signaling were determined using R28 retinal neuronal cells.
Results: We demonstrate a significant decrease in PA in R28 retinal neuronal cells exposed to hyperglycemia as well as in streptozotocin-induced diabetic rat retinas. Exogenous PA augmented insulin-induced protection from interleukin-1β-induced apoptosis. Moreover, exogenous PA and insulin cooperatively activated mTOR survival pathways in R28 neuronal cultures. Exogenous PA colocalized with activated mTOR/p70 S6kinase signaling elements within lipid microdomains. The biochemical consequences of this biophysical mechanism is reflected by differential phosphorylation of tuberin at threonine 1462 and serine 1798, respectively, by PA and insulin, which reduce this suppressor of mTOR/S6Kinase signaling within lipid microdomains.
Conclusions: These results identify PA-enriched microdomains as a putative lipid-based signaling element responsible for mTOR-dependent retinal neuronal survival. Moreover, diabetic retinal neuronal apoptosis may reflect diminished PA mass. Elevating PA concentrations and restoring mTOR signaling may be an effective therapeutic modality to reduce neuronal cell death in diabetic retinopathy.