Reduced NGF Level Promotes Epithelial–Mesenchymal Transition in Human Lens Epithelial Cells Exposed to High Dexamethasone Concentrations

<b>Purpose</b>: To investigate the protective effects of nerve growth factor (NGF) against steroid-induced cataract formation in dexamethasone (Dex)-treated human lens epithelial B-3 (HLE-B3) cells and the possible molecular mechanisms underlying this protection. <b>Materials and Methods</b>: HLE-B3 cells were treated with Dex, and cell viability was assessed using the Cell Counting Kit-8 (CCK-8) assay. The levels of expression of NGF, fibronectin, α-smooth muscle actin (α-SMA), and E-cadherin mRNAs were measured by real-time quantitative polymerase chain reaction (qPCR), and the levels of NGF, fibronectin, α-SMA, E-cadherin, tropomyosin receptor kinase A (TrkA), and Akt proteins were measured by Western blot analysis. Gene expression profiles of growth factors in Dex-treated HLE-B3 cells were determined by PCR arrays. In addition, anterior capsule tissue was obtained during cataract surgery, and the specimens were also examined expressions of NGF. <b>Results</b>: NGF was expressed in HLE-B3 cells and also in lens epithelial cells of anterior lens capsules. Dex treatment of HLE-B3 cells increased their expression of epithelial–mesenchymal transition (EMT) markers and migration activity, while markedly downregulating the expression of NGF. NGF treatment significantly reduced the expression of α-SMA and fibronectin, as well as cell proliferation. The decreased phosphorylation of p38 MAPK and Akt induced by Dex treatment was significantly reversed by treatment with NGF. <b>Conclusion</b>: NGF/TrkA may repress EMT by targeting the p38 MAPK and pAkt pathways in Dex-treated HLE-B3 cells. NGF may be a novel therapeutic target for patients with steroid-induced cataract.

**研究目的**：探讨神经生长因子（nerve growth factor, NGF）对地塞米松（dexamethasone, Dex）处理的人晶状体上皮B-3（human lens epithelial B-3, HLE-B3）细胞中类固醇诱导性白内障形成的保护作用，及其潜在的分子机制。**材料与方法**：将HLE-B3细胞以地塞米松处理，采用细胞计数试剂盒-8（Cell Counting Kit-8, CCK-8）法检测细胞活力；通过实时定量聚合酶链反应（real-time quantitative polymerase chain reaction, qPCR）检测NGF、纤连蛋白、α-平滑肌肌动蛋白（α-smooth muscle actin, α-SMA）及E-钙粘蛋白的mRNA表达水平；通过蛋白质印迹分析检测NGF、纤连蛋白、α-SMA、E-钙粘蛋白、原肌球蛋白受体激酶A（tropomyosin receptor kinase A, TrkA）及Akt的蛋白表达水平。采用PCR基因芯片分析经地塞米松处理的HLE-B3细胞的生长因子基因表达谱。此外，在白内障手术中获取前囊膜组织，检测标本中NGF的表达情况。**结果**：NGF在HLE-B3细胞及前囊膜的晶状体上皮细胞中均有表达。地塞米松处理HLE-B3细胞后，上皮间质转化（epithelial–mesenchymal transition, EMT）标志物的表达及细胞迁移活性均升高，同时NGF的表达显著下调。外源性NGF处理可显著降低α-SMA与纤连蛋白的表达水平，并抑制细胞增殖。地塞米松诱导的p38丝裂原活化蛋白激酶（p38 MAPK）及Akt磷酸化水平降低，可被NGF处理显著逆转。**结论**：在经地塞米松处理的HLE-B3细胞中，NGF/TrkA可能通过靶向p38 MAPK及pAkt通路抑制上皮间质转化。NGF或可成为类固醇诱导性白内障患者的新型治疗靶点。