Novel atypical PKC inhibitors prevent vascular endothelial growth factor-induced blood-retinal barrier dysfunction.
Novel atypical PKC inhibitors prevent vascular endothelial growth factor-induced blood-retinal barrier dysfunction.
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DOI：
10.1042/bj20111961
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发表时间：
2012-09-15
期刊：
影响因子：
--
通讯作者：
Antonetti DA
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文献类型：
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作者：
Titchenell PM;Lin CM;Keil JM;Sundstrom JM;Smith CD;Antonetti DA
促炎性细胞因子和生长因子如血管内皮生长因子（VEGF）在各种视网膜病变中导致血-视网膜屏障（BRB）的丧失和随后的黄斑水肿。VEGF信号传导需要常规的PKC（PKCβ）活性;然而，PKCβ抑制仅部分阻止VEGF诱导的内皮渗透性，不影响促炎性精氨酸诱导的渗透性，表明参与了替代信号传导途径。在这里，我们提供的证据参与非典型蛋白激酶C（aPKC）信号在VEGF诱导的内皮细胞通透性，并确定了一类新的抑制剂的aPKC，防止BRB在体内的崩溃。使用aPKC亚型的遗传和药理学操作来评估它们对培养物中内皮渗透性的贡献。使用体外激酶测定筛选化学文库以鉴定新的小分子抑制剂，并进行进一步的药物化学以描绘新的药效团。我们证明aPKC亚型是VEGF诱导的内皮通透性的充分和必需的。此外，这些特异性的、有效的、非竞争性的小分子抑制剂在原代培养物和啮齿类动物视网膜中均阻止VEGF诱导的紧密连接内化和视网膜内皮通透性对VEGF的响应。这些数据表明，可以开发用2-氨基-4-苯基-噻吩衍生物抑制aPKC，以在视网膜疾病如糖尿病性视网膜病变或葡萄膜炎和脑肿瘤存在下的血脑屏障（BBB）中保护BRB。
Pro-inflammatory cytokines and growth factors such as vascular endothelial growth factor (VEGF) contribute to the loss of the blood-retinal barrier (BRB) and subsequent macular edema in various retinal pathologies. VEGF signaling requires conventional PKC (PKCβ) activity; however, PKCβ inhibition only partially prevents VEGF-induced endothelial permeability and does not affect pro-inflammatory cytokine-induced permeability suggesting the involvement of alternative signaling pathways. Here, we provide evidence for the involvement of atypical protein kinase C (aPKC) signaling in VEGF-induced endothelial permeability and identify a novel class of inhibitors of aPKC that prevent BRB breakdown in vivo. Genetic and pharmacological manipulations of aPKC isoforms were used to assess their contribution to endothelial permeability in culture. A chemical library was screened using an in vitro kinase assay to identify novel small molecule inhibitors and further medicinal chemistry was performed to delineate a novel pharmacophore. We demonstrate that aPKC isoforms are both sufficient and required for VEGF-induced endothelial permeability. Furthermore, these specific, potent, non-competitive, small molecule inhibitors prevented VEGF-induced tight junction internalization and retinal endothelial permeability in response to VEGF in both primary culture and in rodent retina. These data suggest that aPKC inhibition with 2-amino-4-phenyl-thiophene derivatives may be developed to preserve the BRB in retinal diseases such as diabetic retinopathy or uveitis and the blood-brain barrier (BBB) in the presence of brain tumors.