人脐带间充质干细胞通过外泌体递送外源性 miR-26a-5p 抑制核因子κB 炎性小体通路抑制髓核细胞焦亡
Human umbilical cord mesenchymal stem cells deliver exogenous miR-26a-5p via exosomes to inhibit nucleus pulposus cell pyroptosis through METTL14/NLRP3.
机构信息
Spine Center, Department of Orthopaedics, Changzheng Hospital, Naval Medical University, No 415 Fengyang Road, Shanghai, 200003, China.
出版信息
Mol Med. 2021 Aug 19;27(1):91. doi: 10.1186/s10020-021-00355-7.
BACKGROUND
Intervertebral disc degeneration (IVDD) is the breakdown of the discs supporting the vertebrae. It is one of the most frequent causes of back pain worldwide. Currently, the clinical interventions for IVDD are mainly focused on symptom releases. Thus, new therapeutic options are needed.
METHODS
Nucleus pulposus (NP) samples were obtained from 20 patients experiencing IVDD and 10 healthy volunteers compared for mRNA N-methyladenosine (mA) mRNA modification as well as methyltransferase (METT) like METTL3, METTL14, and Wilms' tumor 1-associated protein mRNA and protein abundance following exosomes exposure from mesenchymal stem cells. In addition, microRNA expressions were also compared. The correlation between the NLR family pyrin domain containing 3 (NLRP3) and METTL14 was measured by luciferase reporter assay. Cytokines were evaluated using an enzyme-linked immunosorbent assay. METTL14, NLRP3, and insulin-like growth factor 2 mRNA-binding protein 2 mRNAs were measured via a quantitative reverse transcription-polymerase chain reaction. Protein was assayed using western blots. Cell death was assessed by propidium iodide staining, lactate dehydrogenase release, gasdermin-N domain abundance, and caspase-1 activation.
RESULTS
The human umbilical cord mesenchymal stem cell (hucMSC) exosomes were found to effectively improve the viability of NP cells and protect them from pyroptosis through targeting METTL14, with a methyltransferase catalyzing mA modification. METTL14 was highly present in NP cells from IVDD patients, which stabilize NLRP3 mRNA in an IGFBP2-dependent manner. The elevated NLRP3 levels result in the increase of interleukin 1β (IL-1β) and IL-18 levels and trigger pyroptotic NP cell death. Such pathogenic axis could be blocked by hucMSC exosomes, which directly degrade METTL14 through exosomal miR-26a-5p.
CONCLUSIONS
The results of the current study revealed the beneficial effects of hucMSC exosomes on NP cells and determined a potential mechanism inducing IVDD.
背景
椎间盘退行性变（IVDD）是椎骨支撑椎间盘的破裂。它是全球最常见的腰痛原因之一。目前，IVDD 的临床干预主要集中在症状缓解上。因此，需要新的治疗选择。
方法
从 20 名患有 IVDD 的患者和 10 名健康志愿者中获取髓核（NP）样本，比较它们的 mRNA N-甲基腺苷（mA）mRNA 修饰以及甲基转移酶（METT）样 METTL3、METTL14 和Wilms 肿瘤 1 相关蛋白 mRNA 和蛋白质丰度，然后将间充质干细胞的外泌体暴露于这些样本中。此外，还比较了 microRNA 的表达。通过荧光素酶报告基因测定测量 NLR 家族吡啶结构域包含 3（NLRP3）和 METTL14 之间的相关性。使用酶联免疫吸附测定法评估细胞因子。通过定量逆转录-聚合酶链反应测量 METTL14、NLRP3 和胰岛素样生长因子 2 mRNA 结合蛋白 2 mRNA。使用蛋白质印迹法测定蛋白质。通过碘化丙啶染色、乳酸脱氢酶释放、gasdermin-N 结构域丰度和半胱天冬酶-1 激活评估细胞死亡。
结果
发现人脐带间充质干细胞（hucMSC）外泌体可通过靶向 METTL14 有效提高 NP 细胞的活力并保护其免受细胞焦亡，而 METTL14 是一种催化 mA 修饰的甲基转移酶。METTL14 在 IVDD 患者的 NP 细胞中高度存在，它以 IGFBP2 依赖的方式稳定 NLRP3 mRNA。升高的 NLRP3 水平导致白细胞介素 1β（IL-1β）和白细胞介素 18（IL-18）水平升高，并引发 NP 细胞焦亡。这种致病轴可以被 hucMSC 外泌体阻断，hucMSC 外泌体通过外泌体 miR-26a-5p 直接降解 METTL14。
结论
本研究结果揭示了 hucMSC 外泌体对 NP 细胞的有益作用，并确定了诱导 IVDD 的潜在机制。
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