State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, the First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310003, China.
Center for Stem Cell and Regenerative Medicine, Zhejiang University School of Medicine, Hangzhou 310058, China.
J Zhejiang Univ Sci B. 2020;21(9):727-739. doi: 10.1631/jzus.B2000249.
Acute liver failure (ALF) is a type of disease with high mortality and rapid progression with no specific treatment methods currently available. Glucocorticoids exert beneficial clinical effects on therapy for ALF. However, the mechanism of this effect remains unclear and when to use glucocorticoids in patients with ALF is difficult to determine. The purpose of this study was to investigate the specific immunological mechanism of dexamethasone (Dex) on treatment of ALF induced by lipopolysaccharide (LPS)/D-galactosamine (D-GaIN) in mice.
Male C57BL/6 mice were given LPS and D-GaIN by intraperitoneal injection to establish an animal model of ALF. Dex was administrated to these mice and its therapeutic effect was observed. Hematoxylin and eosin (H&E) staining was used to determine liver pathology. Multicolor flow cytometry, cytometric bead array (CBA) method, and next-generation sequencing were performed to detect changes of messenger RNA (mRNA) in immune cells, cytokines, and Kupffer cells, respectively.
A mouse model of ALF can be constructed successfully using LPS/D-GaIN, which causes a cytokine storm in early disease progression. Innate immune cells change markedly with progression of liver failure. Earlier use of Dex, at 0 h rather than 1 h, could significantly improve the progression of ALF induced by LPS/D-GaIN in mice. Numbers of innate immune cells, especially Kupffer cells and neutrophils, increased significantly in the Dex-treated group. In vivo experiments indicated that the therapeutic effect of Dex is exerted mainly via the glucocorticoid receptor (Gr). Sequencing of Kupffer cells revealed that Dex could increase mRNA transcription level of nuclear receptor subfamily 4 group A member 1 (Nr4a1), and that this effect disappeared after Gr inhibition.
In LPS/D-GaIN-induced ALF mice, early administration of Dex improved ALF by increasing the numbers of innate immune cells, especially Kupffer cells and neutrophils. Gr-dependent Nr4a1 upregulation in Kupffer cells may be an important ALF effect regulated by Dex in this process.
急性肝衰竭（ALF）是一种死亡率高、进展迅速的疾病，目前尚无特效治疗方法。糖皮质激素对 ALF 的治疗具有良好的临床效果。然而，其作用机制尚不清楚，在 ALF 患者中何时使用糖皮质激素也难以确定。本研究旨在探讨地塞米松（Dex）治疗脂多糖（LPS）/D-半乳糖胺（D-GaIN）诱导的小鼠 ALF 的具体免疫学机制。
雄性 C57BL/6 小鼠腹腔注射 LPS 和 D-GaIN 建立 ALF 动物模型，给予 Dex 并观察其治疗效果。苏木精-伊红（H&E）染色观察肝组织病理学变化。多色流式细胞术、细胞因子珠阵列（CBA）法和下一代测序分别检测免疫细胞、细胞因子和库普弗细胞中信使 RNA（mRNA）的变化。
采用 LPS/D-GaIN 成功构建了小鼠 ALF 模型，该模型在疾病早期进展过程中会引发细胞因子风暴。固有免疫细胞在肝衰竭进展过程中发生明显变化。早期（0 h 而非 1 h）使用 Dex 可显著改善 LPS/D-GaIN 诱导的小鼠 ALF 进展。Dex 治疗组固有免疫细胞数量，尤其是库普弗细胞和中性粒细胞数量明显增加。体内实验表明，Dex 的治疗作用主要通过糖皮质激素受体（Gr）发挥。库普弗细胞测序显示，Dex 可增加核受体亚家族 4 组 A 成员 1（Nr4a1）的 mRNA 转录水平，而 Gr 抑制后该作用消失。
在 LPS/D-GaIN 诱导的 ALF 小鼠中，早期给予 Dex 通过增加固有免疫细胞，尤其是库普弗细胞和中性粒细胞的数量，改善 ALF。Gr 依赖性库普弗细胞中 Nr4a1 的上调可能是 Dex 在此过程中调节 ALF 的一个重要作用机制。
