Zou Jing, Chen Zhaoyu, Wei Xiaobo, Chen Zhigang, Fu Yongmei, Yang Xiaoyan, Chen Dan, Wang Rui, Jenner Peter, Lu Jia-Hong, Li Min, Zhang Zhuohua, Tang Beisha, Jin Kunlin, Wang Qing
Department of Neurology, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, Guangdong 510630, China.
Department of Emergency, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, Guangdong 510630, China.
Cell Death Dis. 2017 Jun 1;8(6):e2854. doi: 10.1038/cddis.2017.240.
Cystatin C (CYS C, Cst3) is an endogenous cysteine protease inhibitor that plays neuroprotective roles in neurodegenerative diseases. We aimed to explore the association of CYS C with Parkinson's disease (PD) models and investigate its involvement in the role of neurovascular units (NVUs) in PD neuro-pathogenesis. We used A53T α-synuclein (SNCA) transgenic mice and 6-hydroxydopamine-lesioned DAergic PC12 cells as experimental PD models to investigate the mechanisms behind this association. The injections of CYS C were administered to the right substantia nigra (SN) of A53T SNCA transgenic mice to measure the effects of CYS C in transgenic A53T SNCA mice. To explore the angiogenesis in vivo and in vitro, we used the chick embryo chorioallantoic membrane (CAM) assay and tube formation (TF) assay. We found that CYS C has a neuroprotective effect in this in vivo PD model. We observed increased VEGF, NURR1 and autophagy markers LC3B and decreased SNCA and apoptosis marker cleaved CASP3 in different brain regions of CYS C-treated A53T SNCA transgenic mice. In vitro, we observed that CYS C-induced VEGF, a secreted protein, attenuated 6-OHDA-lesioned DAergic PC12 cell degeneration by regulating p-PKC-α/p-ERK1/2-Nurr1 signaling and inducing autophagy. VEGF-mediated angiogenesis was markedly enhanced in the conditioned media of 6-OHDA-lesioned PC12 cells with CYS C-overexpression, whereas blockage of autophagy in CYS C-overexpressing PC12 cells significantly downregulated VEGF expression and the associated angiogenesis. Our data indicate that CYS C displays dual neuronal-vascular functions, promoting PC12 cell survival and angiogenesis via regulating the level of secreted VEGF in NVUs. Our study provides evidence that may aid in the development of an alternative approach for the treatment of PD through modulation of CYS C-mediated neuronal-vascular pathways.
胱抑素C（CYS C，Cst3）是一种内源性半胱氨酸蛋白酶抑制剂，在神经退行性疾病中发挥神经保护作用。我们旨在探讨CYS C与帕金森病（PD）模型的关联，并研究其在PD神经发病机制中神经血管单元（NVU）作用的参与情况。我们使用A53T α-突触核蛋白（SNCA）转基因小鼠和6-羟基多巴胺损伤的多巴胺能PC12细胞作为实验性PD模型，以研究这种关联背后的机制。将CYS C注射到A53T SNCA转基因小鼠的右侧黑质（SN）中，以测量CYS C在转基因A53T SNCA小鼠中的作用。为了探究体内和体外的血管生成情况，我们使用了鸡胚绒毛尿囊膜（CAM）试验和管形成（TF）试验。我们发现CYS C在这种体内PD模型中具有神经保护作用。我们观察到，在经CYS C处理的A53T SNCA转基因小鼠的不同脑区中，血管内皮生长因子（VEGF）、核受体相关因子1（NURR1）和自噬标志物微管相关蛋白1轻链3β（LC3B）增加，而SNCA和凋亡标志物裂解的半胱天冬酶3（cleaved CASP3）减少。在体外，我们观察到CYS C诱导分泌的蛋白VEGF，通过调节p-蛋白激酶C-α/p-细胞外信号调节激酶1/2-Nurr1信号通路并诱导自噬，减轻6-羟基多巴胺损伤的多巴胺能PC12细胞变性。在CYS C过表达的6-羟基多巴胺损伤的PC12细胞的条件培养基中，VEGF介导的血管生成明显增强，而在CYS C过表达的PC12细胞中阻断自噬显著下调VEGF表达及相关的血管生成。我们的数据表明，CYS C具有双重神经血管功能，通过调节NVU中分泌的VEGF水平促进PC12细胞存活和血管生成。我们的研究提供了证据，可能有助于开发一种通过调节CYS C介导的神经血管途径来治疗PD的替代方法。
