Early Intervention of Tyrosine Nitration Prevents Vaso-Obliteration and Neovascularization in Ischemic Retinopathy
Early Intervention of Tyrosine Nitration Prevents Vaso-Obliteration and Neovascularization in Ischemic Retinopathy
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DOI：
10.1124/jpet.109.157941
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发表时间：
2010-01-01
影响因子：
3.5
通讯作者：
El-Remessy, Azza B.
中科院分区：
文献类型：
--
作者：
Abdelsaid, Mohammed A.;Pillai, Bindu A.;El-Remessy, Azza B.
糖尿病视网膜病变和早产儿视网膜病变是一种致盲性疾病，其病理模式为缺血性视网膜病变，影响早产儿和工作年龄的成人。然而，治疗选择仅限于激光光凝。本研究的目的是阐明分子机制，并检查在缺血性视网膜病变模型中抑制酪氨酸硝化对保护早期视网膜血管细胞死亡和晚期新生血管的治疗效果。通过将新生小鼠暴露于75%的氧气[出生后第7天（p）-p12]，然后暴露于常氧（21%的氧气）（p12-p17），形成缺血性视网膜病变。给予过氧亚硝酸盐分解催化剂5，10，15，20-四（4-磺酸基苯基）卟啉合氯化铁III（FeTPPS）（1 mg/kg）、硝化抑制剂表儿茶素（10 mg/kg）或巯基供体N-乙酰半胱氨酸（NAC，150 mg/kg）（p7-p12）或（p7-p17）。将血管内皮细胞在高氧（40%氧）或常氧（21%氧）下孵育48 h。在用异凝集素B4标记的视网膜平坦载片中测定血管密度。Western blot分析血管内皮生长因子、caspase-3和聚（ADP核糖）聚合酶（PARP）的表达，Akt和p38丝裂原活化蛋白激酶（MAPK）的活化，以及磷脂酰肌醇（PI）3-激酶p85亚基的酪氨酸硝化。高氧诱导的过氧亚硝酸盐引起内皮细胞凋亡，如通过表达裂解的caspase-3和PARP导致血管闭塞所示。这些作用与PI 3-激酶的p85亚基的显著酪氨酸硝化、Akt活化降低和p38 MAPK活化增强有关。用表儿茶素或NAC阻断PI 3-激酶的酪氨酸硝化恢复Akt磷酸化，并抑制p12的血管闭塞和p17的新生血管形成，与FeTPPS相当。早期使用表儿茶素或NAC抑制酪氨酸硝化可以代表缺血性视网膜病变中安全有效的血管保护剂。
Diabetic retinopathy and retinopathy of prematurity are blinding disorders that follow a pathological pattern of ischemic retinopathy and affect premature infants and working-age adults. Yet, the treatment options are limited to laser photocoagulation. The goal of this study is to elucidate the molecular mechanism and examine the therapeutic effects of inhibiting tyrosine nitration on protecting early retinal vascular cell death and late neovascularization in the ischemic retinopathy model. Ischemic retinopathy was developed by exposing neonatal mice to 75% oxygen [postnatal day (p) 7-p12] followed by normoxia (21% oxygen) (p12-p17). Peroxynitrite decomposition catalyst 5,10,15,20-tetrakis(4-sulfonatophenyl) porphyrinato iron III chloride (FeTPPS) (1 mg/kg), the nitration inhibitor epicatechin (10 mg/kg) or the thiol donor N-acetylcysteine (NAC, 150 mg/kg) were administered (p7-p12) or (p7-p17). Vascular endothelial cells were incubated at hyperoxia (40% oxygen) or normoxia (21% oxygen) for 48 h. Vascular density was determined in retinal flat mounts labeled with isolectin B4. Expression of vascular endothelial growth factor, caspase-3, and poly(ADP ribose) polymerase (PARP), activation of Akt and p38 mitogen-activated protein kinase (MAPK), and tyrosine nitration of the phosphatidylinositol (PI) 3-kinase p85 subunit were analyzed by Western blot. Hyperoxia-induced peroxynitrite caused endothelial cell apoptosis as indicated by expression of cleaved caspase-3 and PARP leading to vaso-obliteration. These effects were associated with significant tyrosine nitration of the p85 subunit of PI 3-kinase, decreased Akt activation, and enhanced p38 MAPK activation. Blocking tyrosine nitration of PI 3-kinase with epicatechin or NAC restored Akt phosphorylation, and inhibited vaso-obliteration at p12 and neovascularization at p17 comparable with FeTPPS. Early inhibition of tyrosine nitration with use of epicatechin or NAC can represent safe and effective vascular-protective agents in ischemic retinopathy.