Bit1-a potential positive regulator of epithelial-mesenchymal transition in lens epithelial cells
Bit1-a potential positive regulator of epithelial-mesenchymal transition in lens epithelial cells
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DOI：
10.1007/s00417-016-3357-3
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发表时间：
2016-07-01
影响因子：
2.7
通讯作者：
Luo, Min
中科院分区：
文献类型：
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作者：
Wu, Xinhua;Ruan, Jing;Luo, Min
后囊混浊（PCO）是白内障手术常见的术后并发症。晶状体上皮细胞 (LEC) 的上皮间质转化 (EMT) 是 PCO 发病机制的重要初始步骤。我们之前已经证明 Bit1 在大鼠 LEC 中表达。本研究旨在探讨Bit1在人LECs EMT中的作用。首先通过实时PCR和免疫荧光染色在人PCO附着的LECs和人晶状体细胞系SRA01/04中检测到Bit1的表达。通过细胞计数、伤口愈合实验和Transwell迁移实验评估Bit1敲低的SRA01/04细胞的增殖和迁移。 TGF-β2触发LECs的EMT，然后通过siRNA敲低实验分析Bit1对EMT关键生物标志物α-平滑肌肌动蛋白（α-SMA）的影响，并通过实时PCR和蛋白质印迹验证EMT的逆转。Bit1在PCO组织来源的LECs中明显增强，并且Bit1在培养的SRA01/04细胞的细胞质中高水平表达。在 Bit1 敲低的 SRA01/04 细胞中，细胞增殖、侵袭和迁移以及 α-SMA 表达显着降低。虽然 TGF-β 2 以剂量依赖性方式升高 Bit1 和 α-SMA 表达水平（在 10 ng/ml TGF-β 2 处理时达到峰值水平），但 SRA01/04 细胞中 Bit1 的抑制逆转了 EMT 过程。 TGF-β2诱导的α-SMA表达水平升高以及迁移和侵袭能力均被Bit1缺陷抑制。这些发现表明，Bit1促进TGF-β2诱导的α-SMA表达，并作为EMT的正调节因子。抑制 Bit1 可抑制 LEC 的增殖、迁移和 EMT。 Bit1可能是预防和治疗PCO的潜在新治疗靶点。
Posterior capsule opacification (PCO) is a common postoperative complication of cataract surgery. Epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs) is an important initial step of PCO pathogenesis. We have previously shown that Bit1 expresses in rat LECs. In this study, we aim to investigate the role of Bit1 in the EMT of human LECs.The expression of Bit1 was firstly detected in human PCO-attached LECs and human lens cell line SRA01/04 by real-time PCR and immunofluorescence staining. The proliferation and migration of Bit1 knockdown SRA01/04 cells were evaluated by cell counting, wound-healing assay, and transwell migration assay. The EMT of LECs was triggered by TGF-beta 2, and then the effect of Bit1 on EMT with a key biomarker of alpha-smooth muscle actin (alpha-SMA) was analyzed by siRNA knockdown assay, and the reversal of EMT was validated by real-time PCR and western blot.Bit1 was obviously augmented in LECs derived from PCO tissues and Bit1 expressed with high levels in the cytoplasm of cultured SRA01/04 cells. Cell proliferation, invasion, and migration, as well as alpha-SMA expression, were significantly decreased in Bit1 knockdown SRA01/04 cells. While TGF-beta 2 elevated Bit1 and alpha-SMA expression levels in a dose-dependent manner, with peak levels at 10 ng/ml of TGF-beta 2 treatment, suppression of Bit1 in SRA01/04 cells reversed the EMT process. TGF-beta 2 induced elevation of alpha-SMA expression level, as well as migration, and invasion abilities were all suppressed by Bit1 deficiency.These findings reveal that Bit1 promotes TGF-beta 2 induced alpha-SMA expression and acts as an positive regulator of EMT. Suppressing Bit1 inhibits the proliferation, migration, and EMT of LECs. Bit1 may be a potential novel therapeutic target for the prevention and treatment of PCO.