PDK2-enhanced glycolysis promotes fibroblast proliferation in thyroid-associated ophthalmopathy
PDK2-enhanced glycolysis promotes fibroblast proliferation in thyroid-associated ophthalmopathy
复制标题
PDK2增强的糖酵解促进甲状腺相关眼病中成纤维细胞的增殖
DOI：
10.1530/jme-20-0143
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发表时间：
2020-11-01
影响因子：
3.5
通讯作者：
Qian, Jiang
中科院分区：
文献类型：
--
作者：
Ma, Ruiqi;Gan, Lu;Qian, Jiang
该研究旨在探讨丙酮酸脱氢酶激酶（PDK）在甲状腺相关眼病（TAO）发病过程中调节肌周眶成纤维细胞（pOF）糖酵解和增殖中的作用。进行 EdU 和 BrdU 掺入测定以检查细胞增殖。进行乳酸产生和耗氧量测定以评估糖酵解。采用实时 PCR 来量化 PDK mRNA 水平。采用毛细管蛋白质免疫分析法对蛋白质样品中的 PDK2、Akt、pAkt(308) 和 GAPDH 进行定量。与对照 pOF 相比，TAO pOF 表现出更强的增殖活性、更高的细胞内乳酸浓度和更低的耗氧率。 PDK 抑制剂二氯乙酸 (DCA) 剂量依赖性地抑制 TAO 和对照 pOF 的增殖。 DCA 减少了 TAO pOF 中的乳酸生成并促进了氧消耗，但对对照 pOF 中的糖酵解没有显着影响。在四种 PDK 同种型中，PDK2 在 TAO pOF 中过表达。潜在的 PDK 信号传导介质细胞质 Akt 在 TAO pOF 中比对照 pOF 更丰富。 PDK2 的敲除导致 TAO pOF 中乳酸产量降低、耗氧量增强、增殖活性减弱以及细胞质 Akt 减少，但在对照 pOF 中没有显示出显着影响。 Akt 抑制剂 MK2206 抑制 TAO 和对照 pOF 中的增殖，MK2206 抑制 TAO OF 中的乳酸产生，但不抑制对照 pOF 中的乳酸产生。总之，PDK2 过表达可通过 TAO pOF 中的 Akt 信号传导增强糖酵解并促进增殖。这些发现表明 PDK2 是 TAO 治疗的潜在治疗靶点。
The study aimed to investigate the role of pyruvate dehydrogenase kinase (PDK) in regulating glycolysis and proliferation of perimysial orbital fibroblasts (pOFs) during the pathogenesis of thyroid-associated ophthalmopathy (TAO). EdU and BrdU incorporation assays were performed to examine cell proliferation. Lactate production and oxygen consumption assays were conducted to evaluate glycolysis. Real-time PCR was adapted to quantify PDK mRNA levels. Capillary Western immunoassay was adapted to quantify PDK2, Akt, pAkt(308) and GAPDH in protein samples. The TAO pOFs exhibited stronger proliferation activity, higher intracellular lactate concentration, and lower oxygen consumption rate than the control pOFs. The PDK inhibitor dichloroacetic acid (DCA) dose-dependently suppressed the proliferation of both TAO and control pOFs. DCA reduced lactate production and promoted oxygen consumption in the TAO pOFs but showed no significant effects on glycolysis in the control pOFs. Among four PDK isotypes, PDK2 was overexpressed in the TAO pOFs. The potential PDK signaling mediator, cytoplasmic Akt, was more abundant in TAO pOFs than control pOFs. Knockdown of PDK2 resulted in lower lactate production, stronger oxygen consumption, weaker proliferation activity, and less cytoplasmic Akt in the TAO pOFs but showed no significant effects in the control pOFs. The Akt inhibitor MK2206 suppressed proliferation in both TAO and control pOFs, and lactate production was inhibited by MK2206 in the TAO OFs but not the control pOFs. To conclude, PDK2 overexpression enhances glycolysis and promotes proliferation via Akt signaling in the TAO pOFs. These findings yield insights that PDK2 is a potential therapeutic target for TAO treatment.