Department of Ophthalmology & Visual Science, Eye & ENT Hospital, Shanghai Medical College, Fudan University, Shanghai, China.
Bioinformatics, School of Life Sciences and Biotechnology, Tongji University.
Mol Vis. 2020 Feb 25;26:76-90. eCollection 2020.
This study investigates the impact of aging on the miRNA expression profile in porcine angular aqueous plexus (AAP) cells, which are the porcine equivalent of human Schlemm's canal endothelial cells.
AAP endothelial cells were isolated and cultured in physiologic (5% O) or hyperoxic condition (40% O) for 14 days to induce cell senescence. miRNA and protein expression profiles of control and senescent cells were analyzed with miRNA microarray and isobaric tags for relative and absolute quantification (iTRAQ), respectively.
The miRNA microarray identified 33 differentially expressed miRNAs in senescent cells compared with controls (p<0.05), and quantitative real-time PCR (qRT-PCR) confirmed 12 of them (p<0.05). iTRAQ analysis identified 148 upregulated and 222 downregulated proteins (p<0.05, fold change>1.2). Bioinformatics analysis of miRNA microarray and proteomics data predicted that six out of seven miRNAs are associated with aqueous humor outflow by targeting integrin and the downstream pathways (Src/Rho kinase, focal adhesion kinase (FAK)/NO-cGMP), and one miRNA might influence gap junction by targeting the Inositol trisphosphate receptor (IP3R) /Protein kinase C (PKC) pathway.
This study identified miRNAs in senescent AAP cells that might regulate aqueous humor outflow by targeting proteins involved in focal adhesion, cytoskeleton, NO-cGMP signaling, and gap junction.
本研究旨在探讨衰老对猪角状房水丛（AAP）细胞中 microRNA（miRNA）表达谱的影响，AAP 内皮细胞是猪 Schlemm 管内皮细胞的等效物。
分离并培养 AAP 内皮细胞，使其在生理（5%O2）或高氧（40%O2）条件下培养 14 天以诱导细胞衰老。用 miRNA 微阵列和等重同位素标记相对和绝对定量（iTRAQ）分别分析对照组和衰老组细胞的 miRNA 和蛋白质表达谱。
miRNA 微阵列鉴定出衰老细胞中与对照组相比有 33 个差异表达的 miRNA（p<0.05），实时定量 PCR（qRT-PCR）验证了其中的 12 个（p<0.05）。iTRAQ 分析鉴定出 148 个上调和 222 个下调的蛋白质（p<0.05，倍数变化>1.2）。miRNA 微阵列和蛋白质组学数据的生物信息学分析预测，七个 miRNA 中有六个通过靶向整合素及其下游途径（Src/Rho 激酶、粘着斑激酶（FAK）/NO-cGMP）与房水流出有关，一个 miRNA 可能通过靶向三磷酸肌醇受体（IP3R）/蛋白激酶 C（PKC）途径影响间隙连接。
本研究鉴定出衰老的 AAP 细胞中的 miRNA，这些 miRNA 可能通过靶向参与粘着斑、细胞骨架、NO-cGMP 信号和间隙连接的蛋白质来调节房水流出。
