Interferon regulatory factor 4 (IRF4), a transcription factor recognized as a key regulator of lymphoid and myeloid cell differentiation, has recently been recognized as a critical mediator of macrophage activation. Previously we have reported that IRF4 signaling is closely correlated with anti-inflammatory polarization of microglia in adult mice after stroke. However, IRF4's role in the inflammatory response in the immature brain is unknown. Using a model of neonatal hypoxic ischemic encephalopathy (HIE) we investigated the regulatory action of IRF4 signaling in the activation of microglia and monocytes after HIE. IRF4 myeloid cell conditional knockout (CKO) postnatal day 10 (P10) male pups were subjected to a 60-min hypoxic-ischemic insult by the Rice-Vanucci model (RVM). IRF4 gene floxed mice (IRF4fl/fl) were used as controls. Brain atrophy and behavioral deficits were measured 7 days after HIE. Flow cytometry (FC) was performed to examine central (microglial activation) and peripheral immune cell responses by both cell membrane and intracellular marker staining. Serum levels of cytokines were determined by ELISA. The results showed that IRF4 CKO pups had increased tissue loss and worse behavioral deficits than IRF4fl/fl mice seven days after HIE. FC demonstrated significantly more infiltration of monocytes and neutrophils in the ischemic brains of IRF4 CKO vs IRF4fl/fl pups. IRF4 CKO ischemic microglia were more pro-inflammatory as evidenced by higher expression of the pro-inflammatory marker CD68, and increased intracellular TNFα and IL-1β levels compared to controls. In addition, IRF4 deletion from myeloid cells resulted in increased levels of circulating pro-inflammatory cytokines and higher endothelial MMP9 expression after HIE. These data indicate that IRF4 signaling in myeloid cells plays a regulatory role in neuroinflammation and that deletion of myeloid IRF4 is detrimental to HIE injury, suggesting that IRF4 could serve as a potential therapeutic target for neonatal ischemic brain injury.

中文翻译：

---

骨髓细胞IRF4信号传导可保护新生儿大脑免受缺氧缺血性脑病的侵害。

干扰素调节因子4（IRF4）是公认的淋巴和髓样细胞分化关键调节因子，是一种转录因子，最近被认为是巨噬细胞激活的关键介体。以前我们已经报道过IRF4信号传导与中风后成年小鼠的小胶质细胞的抗炎极化密切相关。但是，IRF4在未成熟大脑炎症反应中的作用尚不清楚。使用新生儿缺氧缺血性脑病（HIE）模型，我们调查了IRF4信号在HIE后激活小胶质细胞和单核细胞中的调节作用。通过Rice-Vanucci模型（RVM）对IRF4骨髓细胞条件性敲除（CKO）产后第10天（P10）的雄性幼犬进行60分钟的缺氧缺血性损伤。使用IRF4基因多态小鼠（IRF4f1 / f1）作为对照。HIE 7天后测量脑萎缩和行为缺陷。进行流式细胞术（FC），以通过细胞膜和细胞内标记物染色检查中枢（小胶质细胞激活）和外周免疫细胞反应。通过ELISA测定血清细胞因子水平。结果显示，HIE后7天，IRF4 CKO幼犬比IRF4fl / fl小鼠具有更多的组织损失和更严重的行为缺陷。与IRF4fl / fl幼犬相比，FC证实IRF4 CKO缺血性脑中单核细胞和中性粒细胞的浸润明显增加。与对照组相比，IRF4 CKO缺血性小胶质细胞的促炎性更强，这由促炎性标记CD68的更高表达以及细胞内TNFα和IL-1β水平升高所证明。此外，从HIE后，髓样细胞的IRF4缺失导致循环促炎细胞因子水平升高和内皮MMP9表达更高。这些数据表明，髓样细胞中的IRF4信号传导在神经炎症中起调节作用，而髓样IRF4的缺失对HIE损伤有害，这表明IRF4可以作为新生儿缺血性脑损伤的潜在治疗靶标。