Objective

Autoimmune hepatitis (AIH) is a chronic liver disease associated with impaired regulatory T cell (Treg) number and function. Erythropoietin (EPO) is a kidney-produced erythropoietic hormone that has known immune-modulating effects, including Treg induction. Whether EPO administration increases Treg in patients with AIH is unknown.

Methods

We treated six stable AIH patients with a single 1000 IU dose of EPO and comprehensively characterized changes in Treg overall and in Treg subsets before and at 4 and 12 weeks after treatment using mass cytometry (CyTOF) combined with an unbiased clustering approach (Phenograph) based on 22 Treg-relevant cell-surface markers.

Results

EPO was well-tolerated and no patients showed significant changes in hematological parameters, liver enzymes, or IgG levels from baseline to 12 weeks following EPO administration. Total Treg and Treg/CD8⁺ T cell ratios significantly increased at 4 weeks and returned to baseline levels at 12 weeks after EPO injection. We identified 17 Treg subsets of which CD4⁺CD25^HICD127^NEG HLADR⁺ Treg had the highest increase and the most favorable Treg/CD8⁺ ratio upon EPO treatment. At 12 weeks after EPO administration, the HLADR⁺ Treg subset also returned to values comparable to those at baseline. Ex vivo assays documented that Treg were functional and the ones isolated at 12 weeks after EPO injection were significantly more suppressive than the ones isolated at baseline. In Treg-depleted assays, EPO did not show a significant effect on IFN-γ+, IL-2⁺, and IL-17⁺ CD4⁺ T cells.

Conclusion

In stable AIH patients, EPO increases overall Treg and particularly those expressing the high function marker HLA-DR. These results provide the rationale for future studies testing the hypothesis that EPO or EPO analogues improve outcomes of AIH patients by increasing Treg.

中文翻译：

---

促红细胞生成素可扩大自身免疫性肝炎患者的调节性 T 细胞

客观的

自身免疫性肝炎 (AIH) 是一种与调节性 T 细胞 (Treg) 数量和功能受损相关的慢性肝病。促红细胞生成素 (EPO) 是一种肾脏产生的促红细胞生成激素，具有已知的免疫调节作用，包括 Treg 诱导作用。尚不清楚 EPO 是否会增加 AIH 患者的 Treg。

方法

我们使用单次 1000 IU 剂量的 EPO 治疗了 6 名稳定的 AIH 患者，并使用质量流式细胞术 (CyTOF) 结合基于无偏聚类方法 (Phenograph) 的治疗前后 4 周和 12 周的 Treg 总体和 Treg 亚群的变化进行了综合表征在 22 个 Treg 相关的细胞表面标志物上。

结果

EPO 耐受性良好，从基线到 EPO 给药后 12 周，没有患者出现血液学参数、肝酶或 IgG 水平的显着变化。总 Treg 和 Treg/CD8 ⁺ T 细胞比率在 4 周时显着增加，并在 EPO 注射后 12 周恢复到基线水平。我们确定了 17 个 Treg 亚群，其中 CD4 ⁺ CD25 ^HI CD127 ^NEG HLADR ⁺ Treg在 EPO 处理后具有最高的增加和最有利的 Treg/CD8 ^+比率。在 EPO 给药后 12 周，HLADR ⁺Treg 子集也返回到与基线相当的值。离体试验证明 Treg 是有功能的，在 EPO 注射后 12 周分离的 Treg 比基线时分离的 Treg 具有明显更强的抑制作用。在 Treg 耗竭试验中，EPO 对 IFN-γ+、IL-2 ⁺和 IL-17 ⁺ CD4 ⁺ T 细胞没有显着影响。

结论

在稳定的 AIH 患者中，EPO 增加整体 Treg，尤其是那些表达高功能标志物 HLA-DR 的患者。这些结果为未来研究检验 EPO 或 EPO 类似物通过增加 Treg 改善 AIH 患者预后的假设提供了依据。