Zhang Kaiye, Zhang Yu, Wan Xiaojie, Mou Yujie, Huang Xiaodan
Eye Center, The Second Affiliated Hospital, School of Medicine, Zhejiang University, Zhejiang Provincial Key Laboratory of Ophthalmology, Zhejiang Provincial Clinical Research Center for Eye Diseases, Zhejiang Provincial Engineering Institute on Eye Diseases, Hangzhou, China.
Department of Ophthalmology, Beijing Key Laboratory of Restoration of Damaged Ocular Nerve, Peking University Third Hospital, Beijing, China.
Invest Ophthalmol Vis Sci. 2025 Aug 1;66(11):76. doi: 10.1167/iovs.66.11.76.
To investigate glycolytic and inflammatory changes on the ocular surface caused by dry eye disease (DED) and the regulatory effect of 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3)-dependent glycolysis on the nuclear factor kappa B (NF-κB) pathway.
Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and a lactate assay were used to evaluate the expression of glycolytic genes, lactate secretion, and inflammatory factors in human corneal epithelial cells (HCECs) under hyperosmotic conditions, which served as an in vitro DED model. Transcriptome sequencing identified key regulatory genes in HCECs under hyperosmotic stimulation. PFKFB3 overexpression plasmids and the small molecule inhibitor 3-(3-pyridinyl)-1-(4-pyridinyl)-2-propen-1-one or small interfering RNA (siRNA) were used to validate the role of PFKFB3 in glycolytic reprogramming and NF-κB pathway activation.
Hyperosmotic stress significantly upregulated glycolytic metabolic enzymes, increased lactate production, and induced inflammatory cytokine secretion in HCECs. Transcriptomics revealed a marked upregulation of the glycolytic regulator PFKFB3 and NF-κB-related genes. Overexpression of PFKFB3 further enhanced NF-κB pathway activation. Inhibition of PFKFB3 reversed hyperosmotic-induced glycolytic activation, suppressed NF-κB phosphorylation, and reduced tumor necrosis factor alpha (TNF-α) secretion.
Hyperosmotic stress activated the NF-κB pathway through PFKFB3-dependent glycolytic reprogramming, forming a vicious metabolic-inflammatory cycle. Targeting PFKFB3 may block this interaction and provide a novel therapeutic strategy for DED.
研究干眼疾病（DED）引起的眼表糖酵解和炎症变化，以及6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶3（PFKFB3）依赖性糖酵解对核因子κB（NF-κB）通路的调节作用。
采用逆转录定量聚合酶链反应（RT-qPCR）和乳酸测定法，评估在高渗条件下作为体外DED模型的人角膜上皮细胞（HCECs）中糖酵解基因的表达、乳酸分泌和炎症因子。转录组测序确定了高渗刺激下HCECs中的关键调控基因。使用PFKFB3过表达质粒和小分子抑制剂3-(3-吡啶基)-1-(4-吡啶基)-2-丙烯-1-酮或小干扰RNA（siRNA）来验证PFKFB3在糖酵解重编程和NF-κB通路激活中的作用。
高渗应激显著上调HCECs中的糖酵解代谢酶，增加乳酸生成，并诱导炎症细胞因子分泌。转录组学显示糖酵解调节因子PFKFB3和NF-κB相关基因显著上调。PFKFB3的过表达进一步增强了NF-κB通路的激活。抑制PFKFB3可逆转高渗诱导的糖酵解激活，抑制NF-κB磷酸化，并减少肿瘤坏死因子α（TNF-α）分泌。
高渗应激通过PFKFB3依赖性糖酵解重编程激活NF-κB通路，形成恶性循环的代谢-炎症循环。靶向PFKFB3可能阻断这种相互作用，并为DED提供一种新的治疗策略。
