由神经胶质瘤细胞释放的细胞外体高迁移率族蛋白 B3 赋予肿瘤相关巨噬细胞中 NLRP3 炎性体的激活和细胞焦亡。

Exosomal HMGB3 released by glioma cells confers the activation of NLRP3 inflammasome and pyroptosis in tumor-associated macrophages.

机构信息

Department of Neurosurgery, the 2nd affiliated hospital, Jiangxi Medical College, Nanchang University, Nanchang, Jiangxi Province 330006, PR China.

出版信息

Tissue Cell. 2024 Jun;88:102406. doi: 10.1016/j.tice.2024.102406. Epub 2024 May 12.

DOI:10.1016/j.tice.2024.102406

PMID:38761792

Abstract

BACKGROUND

Previous evidences has highlighted the pivotal role of NOD-like receptor family pyrin domain-containing 3 (NLRP3)-mediated inflammasomes and pyroptosis activation in driving tumor malignancy and shaping the tumor microenvironment. Herein, we aimed to elucidate the impact of high-mobility group box 3 (HMGB3) released in glioma-derived exosomes on macrophage infiltration in gliomas, NLRP3 inflammasome activation and polarization.

METHODS

Transcripts and protein levels of HMGB3, and cytokines associated with macrophage phenotypes and pyroptosis were assessed in glioma tissues and cell lines (U251, LN229, T98G, A172) using qRT-PCR and/or Western blot analysis. Exosomes secreted from LN229 and NHA cells were isolated via differential ultracentrifugation and characterized by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and analysis of exosome-related markers. PKH67 staining was employed to examine exosomes uptake by THP-1 differentiated macrophages. Flow cytometry was utilized to assess macrophage pyroptotic rates and polarization-related markers.

RESULTS

HMGB3 expression was elevated in glioma tissues, serum samples and tumor cell lines. Kaplan-Meier curves revealed a positive correlation between higher HMGB3 expression and poor overall survival and recurrence-free survival. Moreover, glioma tissues with increased HMGB3 expression exhibited significant upregulation of M2 macrophages markers (CD68, CD206, Arg1) and NLRP3 inflammasome components (NLRP3, IL-1β, ASC), suggesting that HMGB3 was closely associated with macrophage infiltration and NLRP3 inflammasome activation. Notably, HMGB3 was found to be enriched in glioma cell- secreted exosomes and could be internalized by macrophages. Knockdown of HMGB3 in glioma cell exosomes could restrain M2 macrophage polarization, NLRP3 inflammasome activation and pyroptosis.

CONCLUSION

These findings suggested that glioma cells secreted exosomal HMGB3 could facilitate macrophage M2 polarization, pyroptosis and inflammatory infiltration, indicating HMGB3 might be a poor prognosis factor for glioma.

摘要

背景

先前的研究已经强调了 NOD 样受体家族吡啶结构域包含 3（NLRP3）介导的炎性小体和细胞焦亡激活在驱动肿瘤恶性和塑造肿瘤微环境中的关键作用。在此，我们旨在阐明胶质瘤衍生外泌体中高迁移率族蛋白 B3（HMGB3）的释放对胶质瘤中巨噬细胞浸润、NLRP3 炎性小体激活和极化的影响。

方法

使用 qRT-PCR 和/或 Western blot 分析评估胶质瘤组织和细胞系（U251、LN229、T98G、A172）中 HMGB3 的转录物和蛋白水平，以及与巨噬细胞表型和细胞焦亡相关的细胞因子。通过差速超速离心分离 LN229 和 NHA 细胞分泌的外泌体，并通过透射电子显微镜（TEM）、纳米颗粒跟踪分析（NTA）和外泌体相关标志物分析进行表征。PKH67 染色用于检测 THP-1 分化的巨噬细胞摄取外泌体。流式细胞术用于评估巨噬细胞细胞焦亡率和极化相关标志物。

结果

HMGB3 在胶质瘤组织、血清样本和肿瘤细胞系中表达上调。Kaplan-Meier 曲线显示，HMGB3 表达较高与总生存期和无复发生存期较差呈正相关。此外，HMGB3 表达增加的胶质瘤组织中 M2 巨噬细胞标志物（CD68、CD206、Arg1）和 NLRP3 炎性小体成分（NLRP3、IL-1β、ASC）显著上调，表明 HMGB3 与巨噬细胞浸润和 NLRP3 炎性小体激活密切相关。值得注意的是，HMGB3 在胶质瘤细胞分泌的外泌体中富集，并可被巨噬细胞内化。在胶质瘤细胞外泌体中敲低 HMGB3 可抑制 M2 巨噬细胞极化、NLRP3 炎性小体激活和细胞焦亡。