Wei Xiang, Jin Xiao-Hong, Meng Xiao-Wen, Hua Jie, Ji Fu-Hai, Wang Li-Na, Yang Jian-Ping
Department of Anesthesiology and Pain Management, the First Affiliated Hospital of Soochow University, Suzhou, China.
Ann Transl Med. 2020 Nov;8(21):1456. doi: 10.21037/atm-20-6502.
Astrocytes are highly glycolytic cells that play a crucial role in chronic pain. Recently it has been found that inflammation and metabolism are related to the inflammatory stimuli closely that cause cellular metabolic changes. Pyruvate kinase M2 (PKM2) is a critical metabolic kinase in aerobic glycolysis or the Warburg effect. Besides, it also plays a crucial role in cell proliferation and signal transduction, but its role in astrocytes is still unclear.
The chronic inflammatory pain model was set up by intraplantar injection of complete Freund's adjuvant (CFA) in Sprague Dawley (SD) rats as well as the cell model was constructed by lipopolysaccharide-treated primary astrocytes. Von Frey filament stimulation was used to continuously observe the changes of pain behavior in rats after modeling. Then, immunofluorescence staining and Western blot tests were used to observe the expression levels of glial fibrillary acidic protein (GFAP), pyruvate kinase (PKM2), signal transducers and activators of transcription 3 (STAT3) and high mobility group box-1 protein (HMGB1). After that, specific kits measured lactate contents. Finally, we observed the platelet-rich plasma's (PRP) effect on mechanical hyperalgesia in rats with inflammatory pain induced by CFA and its effect on related signal molecules.
We found that in the CFA-induced inflammatory pain model, astrocytes were significantly activated, GFAP was increased, PKM2 was significantly up-regulated, and the glycolytic product lactate was increased. Also, intrathecal injection of PRP increased the pain threshold, inhibited the activation of astrocytes, and decreased the expression of PKM2 and aerobic glycolysis; in LPS-activated primary astrocytes as an model, we found PKM2 translocation activationSTAT3 signaling resulted in sustained activation of astrocyte marker GFAP, and the expression level and localization of p-STAT3 were correlated with PKM2. PRP could inhibit the activation of astrocytes, reduce the expression of PKM2 and the expression levels of glycolysis and GFAP, GLUT1, and p-STAT3 in astrocytes.
Our findings suggest PKM2 not only plays a glycolytic role in astrocytes, but also plays a crucial role in astrocyte-activated signaling pathways, and PRP attenuates CFA induced inflammatory pain by inhibiting aerobic glycolysis in astrocytes, providing a new therapeutic target for the treatment of inflammatory pain.
星形胶质细胞是高度糖酵解的细胞，在慢性疼痛中起关键作用。最近发现炎症与新陈代谢密切相关，炎症刺激会导致细胞代谢变化。丙酮酸激酶M2（PKM2）是有氧糖酵解或瓦伯格效应中的关键代谢激酶。此外，它在细胞增殖和信号转导中也起关键作用，但其在星形胶质细胞中的作用仍不清楚。
通过在Sprague Dawley（SD）大鼠足底注射完全弗氏佐剂（CFA）建立慢性炎性疼痛模型，并通过脂多糖处理的原代星形胶质细胞构建细胞模型。采用von Frey细丝刺激法持续观察建模后大鼠疼痛行为的变化。然后，通过免疫荧光染色和蛋白质印迹试验观察胶质纤维酸性蛋白（GFAP）、丙酮酸激酶（PKM2）、信号转导子和转录激活子3（STAT3）及高迁移率族蛋白B1（HMGB1）的表达水平。之后，使用特定试剂盒检测乳酸含量。最后，我们观察了富血小板血浆（PRP）对CFA诱导的炎性疼痛大鼠机械性痛觉过敏的影响及其对相关信号分子的影响。
我们发现，在CFA诱导的炎性疼痛模型中，星形胶质细胞显著激活，GFAP增加，PKM2显著上调，糖酵解产物乳酸增加。此外，鞘内注射PRP提高了疼痛阈值，抑制了星形胶质细胞的激活，并降低了PKM2的表达及有氧糖酵解；在以脂多糖激活的原代星形胶质细胞为模型中，我们发现PKM2易位激活STAT3信号通路，导致星形胶质细胞标志物GFAP持续激活，且p-STAT3的表达水平和定位与PKM2相关。PRP可抑制星形胶质细胞的激活，降低PKM2的表达以及星形胶质细胞中糖酵解、GFAP、葡萄糖转运蛋白1（GLUT1）和p-STAT3的表达水平。
我们的研究结果表明，PKM2不仅在星形胶质细胞中起糖酵解作用，还在星形胶质细胞激活信号通路中起关键作用，且PRP通过抑制星形胶质细胞的有氧糖酵解减轻CFA诱导的炎性疼痛，为炎性疼痛的治疗提供了新的治疗靶点。
