PDF(Pubmed)
Abstract：
BACKGROUND: Lactate has emerged as a critical regulator within the tumor microenvironment, including glioma. However, the precise mechanisms underlying how lactate influences the communication between tumor cells and tumor-associated macrophages (TAMs), the most abundant immune cells in glioma, remain poorly understood. This study aims to elucidate the impact of tumor-derived lactate on TAMs and investigate the regulatory pathways governing TAM-mediated tumor-promotion in glioma.
METHODS: Bioinformatic analysis was conducted using datasets from TCGA and CGGA. Single-cell RNA-seq datasets were analyzed by using UCSC Cell Browser and Single Cell Portal. Cell proliferation and mobility were evaluated through CCK8, colony formation, wound healing, and transwell assays. Western blot and immunofluorescence staining were applied to assess protein expression and cell distribution. RT-PCR and ELISA were employed to identify the potential secretory factors. Mechanistic pathways were explored by western blotting, ELISA, shRNA knockdown, and specific inhibitors and activators. The effects of pathway blockades were further assessed using subcutaneous and intracranial xenograft tumor models in vivo.
RESULTS: Elevated expressions of LDHA and MCT1 were observed in glioma and exhibited a positive correlation with M2-type TAM infiltration. Lactate derived from glioma cells induced TAMs towards M2-subtype polarization, subsequently promoting glioma cells proliferation, migration, invasion, and mesenchymal transition. GPR65, highly expressed on TAMs, sensed lactate-stimulation in the TME, fueling glioma cells malignant progression through the secretion of HMGB1. GPR65 on TAMs triggered HMGB1 release in response to lactate stimulation via the cAMP/PKA/CREB signaling pathway. Disrupting this feedback loop by GPR65-knockdown or HMGB1 inhibition mitigated glioma progression in vivo.
CONCLUSIONS: These findings unveil the intricate interplay between TAMs and tumor cells mediated by lactate and HMGB1, driving tumor progression in glioma. GPR65, selectively highly expressed on TAMs in glioma, sensed lactate stimulation and fostered HMGB1 secretion via the cAMP/PKA/CREB signaling pathway. Blocking this feedback loop presents a promising therapeutic strategy for GBM.
METHODS: Bioinformatic analysis was conducted using datasets from TCGA and CGGA. Single-cell RNA-seq datasets were analyzed by using UCSC Cell Browser and Single Cell Portal. Cell proliferation and mobility were evaluated through CCK8, colony formation, wound healing, and transwell assays. Western blot and immunofluorescence staining were applied to assess protein expression and cell distribution. RT-PCR and ELISA were employed to identify the potential secretory factors. Mechanistic pathways were explored by western blotting, ELISA, shRNA knockdown, and specific inhibitors and activators. The effects of pathway blockades were further assessed using subcutaneous and intracranial xenograft tumor models in vivo.
RESULTS: Elevated expressions of LDHA and MCT1 were observed in glioma and exhibited a positive correlation with M2-type TAM infiltration. Lactate derived from glioma cells induced TAMs towards M2-subtype polarization, subsequently promoting glioma cells proliferation, migration, invasion, and mesenchymal transition. GPR65, highly expressed on TAMs, sensed lactate-stimulation in the TME, fueling glioma cells malignant progression through the secretion of HMGB1. GPR65 on TAMs triggered HMGB1 release in response to lactate stimulation via the cAMP/PKA/CREB signaling pathway. Disrupting this feedback loop by GPR65-knockdown or HMGB1 inhibition mitigated glioma progression in vivo.
CONCLUSIONS: These findings unveil the intricate interplay between TAMs and tumor cells mediated by lactate and HMGB1, driving tumor progression in glioma. GPR65, selectively highly expressed on TAMs in glioma, sensed lactate stimulation and fostered HMGB1 secretion via the cAMP/PKA/CREB signaling pathway. Blocking this feedback loop presents a promising therapeutic strategy for GBM.
摘要：
背景：乳酸盐已成为肿瘤微环境中的关键调节因子，包括神经胶质瘤.然而,乳酸如何影响肿瘤细胞和肿瘤相关巨噬细胞（TAMs）之间的通讯的确切机制，神经胶质瘤中最丰富的免疫细胞，仍然知之甚少。本研究旨在阐明肿瘤源性乳酸对TAM的影响，并研究控制TAM介导的神经胶质瘤肿瘤促进的调节途径。
方法：使用来自TCGA和CGGA的数据集进行生物信息学分析。通过使用UCSC细胞浏览器和单细胞门户分析单细胞RNA-seq数据集。通过CCK8，集落形成评估细胞增殖和迁移率，伤口愈合，和transwell分析。应用蛋白质印迹和免疫荧光染色评估蛋白质表达和细胞分布。RT-PCR和ELISA用于鉴定潜在的分泌因子。通过蛋白质印迹法探索了机制途径，ELISA,shRNA敲除，和特异性抑制剂和活化剂。在体内使用皮下和颅内异种移植肿瘤模型进一步评估途径阻断的效果。
结果：胶质瘤中LDHA和MCT1的表达升高，并与M2型TAM浸润呈正相关。来自神经胶质瘤细胞的乳酸诱导TAMs向M2亚型极化，随后促进神经胶质瘤细胞增殖，迁移,入侵,和间质转化。GPR65在TAMs上高表达，感觉到TME中的乳酸刺激，通过分泌HMGB1促进胶质瘤细胞恶性进展。TAM上的GPR65通过cAMP/PKA/CREB信号通路响应乳酸刺激而触发HMGB1释放。通过GPR65敲低或HMGB1抑制破坏该反馈回路减轻了体内神经胶质瘤的进展。
结论：这些发现揭示了TAMs和由乳酸和HMGB1介导的肿瘤细胞之间复杂的相互作用，驱动神经胶质瘤的肿瘤进展。GPR65在胶质瘤中的TAMs上选择性高表达，通过cAMP/PKA/CREB信号通路感知乳酸刺激并促进HMGB1分泌。阻断该反馈回路为GBM提供了有希望的治疗策略。
方法：使用来自TCGA和CGGA的数据集进行生物信息学分析。通过使用UCSC细胞浏览器和单细胞门户分析单细胞RNA-seq数据集。通过CCK8，集落形成评估细胞增殖和迁移率，伤口愈合，和transwell分析。应用蛋白质印迹和免疫荧光染色评估蛋白质表达和细胞分布。RT-PCR和ELISA用于鉴定潜在的分泌因子。通过蛋白质印迹法探索了机制途径，ELISA,shRNA敲除，和特异性抑制剂和活化剂。在体内使用皮下和颅内异种移植肿瘤模型进一步评估途径阻断的效果。
结果：胶质瘤中LDHA和MCT1的表达升高，并与M2型TAM浸润呈正相关。来自神经胶质瘤细胞的乳酸诱导TAMs向M2亚型极化，随后促进神经胶质瘤细胞增殖，迁移,入侵,和间质转化。GPR65在TAMs上高表达，感觉到TME中的乳酸刺激，通过分泌HMGB1促进胶质瘤细胞恶性进展。TAM上的GPR65通过cAMP/PKA/CREB信号通路响应乳酸刺激而触发HMGB1释放。通过GPR65敲低或HMGB1抑制破坏该反馈回路减轻了体内神经胶质瘤的进展。
结论：这些发现揭示了TAMs和由乳酸和HMGB1介导的肿瘤细胞之间复杂的相互作用，驱动神经胶质瘤的肿瘤进展。GPR65在胶质瘤中的TAMs上选择性高表达，通过cAMP/PKA/CREB信号通路感知乳酸刺激并促进HMGB1分泌。阻断该反馈回路为GBM提供了有希望的治疗策略。
