Notch Signaling Functions in Retinal Pericyte Survival
Notch Signaling Functions in Retinal Pericyte Survival
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DOI：
10.1167/iovs.14-14046
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发表时间：
2014-08-01
影响因子：
4.4
通讯作者：
D'Amore, Patricia A.
中科院分区：
文献类型：
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作者：
Arboleda-Velasquez, Joseph F.;Primo, Vincent;D'Amore, Patricia A.
目的.周细胞是构成毛细血管外层的血管细胞，已被证明在血管发育和稳定性中具有至关重要的作用。在小血管疾病（包括糖尿病视网膜病变）中，周细胞的损失先于内皮细胞功能障碍和血管变性。尽管它们的临床意义，控制视网膜周细胞的存活的细胞通路仍然在很大程度上未被表征。因此，我们研究了Notch信号传导（细胞命运决定的主要调节因子）在视网膜周细胞存活中的作用。使用原代培养的视网膜周细胞和来源于表达Delta样1 Notch配体的诱导型小鼠模型的间充质细胞系开发配体依赖性Notch信号传导的共培养系统。该模型用于使用定量PCR（qPCR）和光诱导细胞死亡测定来检查Notch活性对周细胞存活的影响。使用抗体阵列在视网膜周细胞的单培养物中分析Notch功能获得和丧失的影响，以询问视网膜变性相关蛋白的表达。原代培养的视网膜周细胞差异表达Notch途径的关键分子，并显示经典Notch/RBPJK（J-kappa重组信号结合蛋白1）下游靶点的强表达。使用功能获得和功能丧失方法的基因表达筛选将与细胞存活相关的基因鉴定为视网膜周细胞中Notch活性的下游靶标。配体介导的Notch活性保护视网膜周细胞免受光诱导的细胞死亡。我们的研究结果已经确定了视网膜周细胞中Notch活性下游的标记基因，并表明Notch信号传导的严格调节对周细胞存活至关重要。
PURPOSE. Pericytes, the vascular cells that constitute the outer layer of capillaries, have been shown to have a crucial role in vascular development and stability. Loss of pericytes precedes endothelial cell dysfunction and vascular degeneration in small-vessel diseases, including diabetic retinopathy. Despite their clinical relevance, the cellular pathways controlling survival of retinal pericytes remain largely uncharacterized. Therefore, we investigated the role of Notch signaling, a master regulator of cell fate decisions, in retinal pericyte survival.METHODS. A coculture system of ligand-dependent Notch signaling was developed using primary cultured retinal pericytes and a mesenchymal cell line derived from an inducible mouse model expressing the Delta-like 1 Notch ligand. This model was used to examine the effect of Notch activity on pericyte survival using quantitative PCR (qPCR) and a light-induced cell death assay. The effect of Notch gain- and loss-of-function was analyzed in monocultures of retinal pericytes using antibody arrays to interrogate the expression of apoptosis-related proteins.RESULTS. Primary cultured retinal pericytes differentially expressed key molecules of the Notch pathway and displayed strong expression of canonical Notch/RBPJK (recombination signal-binding protein 1 for J-kappa) downstream targets. A gene expression screen using gain- and loss-of-function approaches identified genes relevant to cell survival as downstream targets of Notch activity in retinal pericytes. Ligand-mediated Notch activity protected retinal pericytes from light-induced cell death.CONCLUSIONS. Our results have identified signature genes downstream of Notch activity in retinal pericytes and suggest that tight regulation of Notch signaling is crucial for pericyte survival.