Department of Pharmacology, Penn State College of Medicine, Hershey, PA 17033, USA.
Invest Ophthalmol Vis Sci. 2012 Oct 19;53(11):7257-67. doi: 10.1167/iovs.11-7626.
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.
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.
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.
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.
我们之前已经证明，在糖尿病性视网膜病变模型中，生存促进型胰岛素受体、PI3K-Akt 和 p70 S6K 信号转导会减弱。由于哺乳动物雷帕霉素靶蛋白（mTOR）是 p70 S6K 的上游激活物，部分受到脂质衍生的第二信使（如磷脂酸（PA））的调节，我们试图确定糖尿病视网膜中 mTOR/p70 S6K 信号转导减弱是否反映了 PA 水平降低。
通过质谱法测定对照和链脲佐菌素诱导的糖尿病大鼠视网膜中 PA 的质量变化。使用 R28 视网膜神经元细胞确定 PA 对胰岛素激活的 mTOR/p70 S6K 信号转导作用的生化和生物物理机制。
我们证明，暴露于高血糖以及链脲佐菌素诱导的糖尿病大鼠视网膜中的 R28 视网膜神经元细胞中的 PA 显著减少。外源性 PA 增强了胰岛素诱导的对白细胞介素-1β诱导的细胞凋亡的保护作用。此外，外源性 PA 和胰岛素在 R28 神经元培养物中协同激活 mTOR 存活途径。外源性 PA 与在脂质微域中激活的 mTOR/p70 S6kinase 信号转导元件共定位。这种生物物理机制的生化后果反映在 PA 和胰岛素分别通过 Thr1462 和 Ser1798 对 tuberin 的不同磷酸化上，这会降低脂质微域中 mTOR/S6Kinase 信号的抑制作用。
这些结果表明，富含 PA 的微域是负责 mTOR 依赖性视网膜神经元存活的假定脂质基信号元件。此外，糖尿病性视网膜神经元凋亡可能反映了 PA 质量的降低。升高 PA 浓度并恢复 mTOR 信号可能是减少糖尿病性视网膜病变中神经元细胞死亡的有效治疗方式。
