Department of Pediatrics, University of Michigan Medical School, Ann Arbor, MI, United States.
Front Immunol. 2023 Sep 8;14:1156842. doi: 10.3389/fimmu.2023.1156842. eCollection 2023.
Bronchopulmonary dysplasia (BPD) is a chronic lung disease in preterm birth survivors characterized by inflammation, impaired alveolarization and dysmorphic vasculature. Activated IL-17A+ lymphocytes are key drivers of inflammation in preterm infants. We have shown that in immature mice chronic airway exposure to lipopolysaccharide (LPS) induces pulmonary inflammation, increased IL-17a expression, and hypoalveolarization, a BPD-like phenotype. The source of IL-17a and contribution to lung pathology is unknown. The natural-killer group 2, member D (NKG2D) receptor mediates activation and IL-17a production in γδ T cells by binding to stress molecules. LPS induces NKG2D ligand expression, including Rae-1 and MULT1. We hypothesized that IL-17a+ γδ T cells and NKG2D signaling mediate neonatal LPS-induced lung injury. Immature C57BL/6J (wild type), Nkg2d-/- or Tcrd-/- (lacking γδ T cells) mice were inoculated with 3ug/10ul of LPS from E. coli O26:B6 or 10ul of PBS intranasally on day of life 3, 5, 7, and 10. Selected mice were treated with neutralizing antibodies against IL-17a, or NKG2D intraperitoneally. Lung immune cells were assessed by flow cytometry and gene expression was analyzed by qPCR. Alveolar growth was assessed by lung morphometry. We established that anti-IL-17a antibody treatment attenuated LPS-induced hypoalveolarization. We found that LPS induced the fraction of IL-17a+NKG2D+ γδ T cells, a major source of IL-17a in the neonatal lung. LPS also induced lung mRNA expression of NKG2D, Rae-1, MULT1, and the DNA damage regulator p53. Anti-NKG2D treatment attenuated the effect of LPS on γδ T cell IL-17a expression, immune cell infiltration and hypoalveolarization. LPS-induced hypoalveolarization was also attenuated in Nkg2d-/- and Tcrd-/- mice. In tracheal aspirates of preterm infants IL-17A and its upstream regulator IL-23 were higher in infants who later developed BPD. Also, human ligands of NKG2D, MICA and MICB were present in the aspirates and MICA correlated with median FiO2. Our novel findings demonstrate a central role for activated IL-17a+ γδ T cells and NKG2D signaling in neonatal LPS-induced lung injury. Future studies will determine the role of NKG2D ligands and effectors, other NKG2D+ cells in early-life endotoxin-induced lung injury and inflammation with a long-term goal to understand how inflammation contributes to BPD pathogenesis.
支气管肺发育不良（BPD）是一种早产儿幸存者的慢性肺部疾病，其特征为炎症、肺泡化受损和血管形态异常。活化的白细胞介素-17A（IL-17A）+淋巴细胞是早产儿炎症的关键驱动因素。我们已经表明，在不成熟的小鼠中，慢性气道暴露于脂多糖（LPS）会诱导肺部炎症、增加 IL-17a 表达和肺泡化不足，形成类似 BPD 的表型。IL-17a 的来源及其对肺部病理学的贡献尚不清楚。自然杀伤细胞组 2，成员 D（NKG2D）受体通过与应激分子结合来介导 γδ T 细胞的激活和 IL-17a 产生。LPS 诱导 NKG2D 配体表达，包括 Rae-1 和 MULT1。我们假设 IL-17a+γδ T 细胞和 NKG2D 信号转导介导新生儿 LPS 诱导的肺损伤。将不成熟的 C57BL/6J（野生型）、Nkg2d-/-或 Tcrd-/-（缺乏 γδ T 细胞）小鼠在生后第 3、5、7 和 10 天用 3μg/10μl 的大肠杆菌 O26:B6 LPS 或 10μl 的 PBS 经鼻腔接种。选择的小鼠用抗 IL-17a 或 NKG2D 的中和抗体经腹腔内给药。通过流式细胞术评估肺免疫细胞，通过 qPCR 分析基因表达。通过肺形态计量学评估肺泡生长。我们证实抗 IL-17a 抗体治疗可减轻 LPS 诱导的肺泡化不足。我们发现 LPS 诱导了 IL-17a+NKG2D+γδ T 细胞的分数，这是新生儿肺部 IL-17a 的主要来源。LPS 还诱导了肺中 NKG2D、Rae-1、MULT1 和 DNA 损伤调节剂 p53 的 mRNA 表达。抗 NKG2D 治疗减轻了 LPS 对 γδ T 细胞 IL-17a 表达、免疫细胞浸润和肺泡化不足的影响。在 Nkg2d-/-和 Tcrd-/-小鼠中，LPS 诱导的肺泡化不足也减轻了。在早产儿的气管抽吸物中，后来发展为 BPD 的婴儿的 IL-17A 及其上游调节剂 IL-23 较高。此外，NKG2D 的人配体 MICA 和 MICB 也存在于抽吸物中，MICA 与中位数 FiO2 相关。我们的新发现表明，活化的 IL-17a+γδ T 细胞和 NKG2D 信号在新生儿 LPS 诱导的肺损伤中起核心作用。未来的研究将确定 NKG2D 配体和效应物以及其他 NKG2D+细胞在早期内毒素诱导的肺损伤和炎症中的作用，其长期目标是了解炎症如何导致 BPD 发病机制。
