外泌体递送的血红素氧合酶-1 修饰的骨髓间充质干细胞来源的 miR-124-3p 抑制铁死亡从而减轻肝脂肪变性供体肝再灌注损伤。

miR-124-3p delivered by exosomes from heme oxygenase-1 modified bone marrow mesenchymal stem cells inhibits ferroptosis to attenuate ischemia-reperfusion injury in steatotic grafts.

机构信息

School of Medicine, Nankai University, Tianjin, People's Republic of China.

Tianjin First Central Hospital Clinic Institute, Tianjin Medical University, Tianjin, 300070, People's Republic of China.

出版信息

J Nanobiotechnology. 2022 Apr 22;20(1):196. doi: 10.1186/s12951-022-01407-8.

BACKGROUND

Steatotic livers tolerate ischemia-reperfusion injury (IRI) poorly, increasing the risk of organ dysfunction. Ferroptosis is considered the initiating factor of organ IRI. Heme oxygenase oxygen-1 (HO-1)-modified bone marrow mesenchymal stem cells (BMMSCs) (HO-1/BMMSCs) can reduce hepatic IRI; however, the role of ferroptosis in IRI of steatotic grafts and the effect of HO-1/BMMSCs-derived exosomes (HM-exos) on ferroptosis remain unknown.

METHODS

A model of rat liver transplantation (LT) with a severe steatotic donor liver and a model of hypoxia and reoxygenation (H/R) of steatotic hepatocytes were established. Exosomes were obtained by differential centrifugation, and the differentially expressed genes (DEGs) in liver after HM-exo treatment were detected using RNA sequencing. The expression of ferroptosis markers was analyzed. microRNA (miRNA) sequencing was used to analyze the miRNA profiles in HM-exos.

RESULTS

We verified the effect of a candidate miRNA on ferroptosis of H/R treated hepatocytes, and observed the effect of exosomes knockout of the candidate miRNA on hepatocytes ferroptosis. In vitro, HM-exo treatment reduced the IRI in steatotic grafts, and enrichment analysis of DEGs suggested that HM-exos were involved in the regulation of the ferroptosis pathway. In vitro, inhibition of ferroptosis by HM-exos reduced hepatocyte injury. HM-exos contained more abundant miR-124-3p, which reduced ferroptosis of H/R-treated cells by inhibiting prostate six transmembrane epithelial antigen 3 (STEAP3), while overexpression of Steap3 reversed the effect of mir-124-3p. In addition, HM-exos from cell knocked out for miR-124-3p showed a weakened inhibitory effect on ferroptosis. Similarly, HM-exo treatment increased the content of miR-124-3p in grafts, while decreasing the level of STEAP3 and reducing the degree of hepatic ferroptosis.

CONCLUSION

Ferroptosis is involved in the IRI during LT with a severe steatotic donor liver. miR-124-3p in HM-exos downregulates Steap3 expression to inhibit ferroptosis, thereby attenuating graft IRI, which might be a promising strategy to treat IRI in steatotic grafts.

背景

脂肪肝肝脏对缺血再灌注损伤（IRI）的耐受性差，增加了器官功能障碍的风险。铁死亡被认为是器官 IRI 的起始因素。血红素加氧酶-1（HO-1）修饰的骨髓间充质干细胞（BMMSCs）（HO-1/BMMSCs）可减少肝 IRI；然而，铁死亡在脂肪肝供体肝移植（LT）中的作用以及 HO-1/BMMSCs 衍生的外泌体（HM-exos）对铁死亡的影响尚不清楚。

方法

建立大鼠严重脂肪肝供肝 LT 模型和脂肪肝细胞缺氧复氧（H/R）模型。采用差速离心法获取外泌体，采用 RNA 测序检测 HM-exo 处理后肝组织中差异表达基因（DEGs）。分析铁死亡标志物的表达。采用 microRNA（miRNA）测序分析 HM-exos 中的 miRNA 谱。

结果

验证了候选 miRNA 对 H/R 处理的肝细胞铁死亡的影响，并观察了外泌体敲除候选 miRNA 对肝细胞铁死亡的影响。体外，HM-exo 处理减轻了脂肪肝供体肝的 IRI，DEGs 富集分析提示 HM-exos 参与了铁死亡途径的调节。体外，HM-exos 通过抑制前列腺六跨膜上皮抗原 3（STEAP3）抑制铁死亡，减少 H/R 处理细胞的肝细胞损伤。HM-exos 含有更丰富的 miR-124-3p，通过抑制 STEAP3 减少 H/R 处理细胞的铁死亡，而过表达 Steap3 则逆转了 miR-124-3p 的作用。此外，miR-124-3p 敲除细胞的 HM-exos 显示出对铁死亡的抑制作用减弱。同样，HM-exo 处理增加了移植物中 miR-124-3p 的含量，降低了 STEAP3 的水平，减轻了肝铁死亡的程度。