Hexa Histidine-Tagged Recombinant Human Cytoglobin Deactivates Hepatic Stellate Cells and Inhibits Liver Fibrosis by Scavenging Reactive Oxygen Species

Hepatology. 2021 Jun;73(6):2527-2545. doi: 10.1002/hep.31752. Epub 2021 May 22.


Background and aims: Antifibrotic therapy remains an unmet medical need in human chronic liver disease. We report the antifibrotic properties of cytoglobin (CYGB), a respiratory protein expressed in hepatic stellate cells (HSCs), the main cell type involved in liver fibrosis.

Approach and results: Cygb-deficient mice that had bile duct ligation-induced liver cholestasis or choline-deficient amino acid-defined diet-induced steatohepatitis significantly exacerbated liver damage, fibrosis, and reactive oxygen species (ROS) formation. All of these manifestations were attenuated in Cygb-overexpressing mice. We produced hexa histidine-tagged recombinant human CYGB (His-CYGB), traced its biodistribution, and assessed its function in HSCs or in mice with advanced liver cirrhosis using thioacetamide (TAA) or 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). In cultured HSCs, extracellular His-CYGB was endocytosed and accumulated in endosomes through a clathrin-mediated pathway. His-CYGB significantly impeded ROS formation spontaneously or in the presence of ROS inducers in HSCs, thus leading to the attenuation of collagen type 1 alpha 1 production and α-smooth muscle actin expression. Replacement the iron center of the heme group with cobalt nullified the effect of His-CYGB. In addition, His-CYGB induced interferon-β secretion by HSCs that partly contributed to its antifibrotic function. Momelotinib incompletely reversed the effect of His-CYGB. Intravenously injected His-CYGB markedly suppressed liver inflammation, fibrosis, and oxidative cell damage in mice administered TAA or DDC mice without adverse effects. RNA-sequencing analysis revealed the down-regulation of inflammation- and fibrosis-related genes and the up-regulation of antioxidant genes in both cell culture and liver tissues. The injected His-CYGB predominantly localized to HSCs but not to macrophages, suggesting specific targeting effects. His-CYGB exhibited no toxicity in chimeric mice with humanized livers.

Conclusions: His-CYGB could have antifibrotic clinical applications for human chronic liver diseases.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Anti-Inflammatory Agents / pharmacology
  • Antioxidants / pharmacology
  • Cholestasis / drug therapy
  • Cholestasis / metabolism
  • Cytoglobin / metabolism*
  • Drug Discovery
  • Fatty Liver* / drug therapy
  • Fatty Liver* / metabolism
  • Hepatic Stellate Cells* / drug effects
  • Hepatic Stellate Cells* / metabolism
  • Liver Cirrhosis* / metabolism
  • Liver Cirrhosis* / pathology
  • Liver Cirrhosis* / prevention & control
  • Mice
  • Mice, Knockout
  • Protective Agents / pharmacology
  • Recombinant Proteins / pharmacology
  • Treatment Outcome


  • Anti-Inflammatory Agents
  • Antioxidants
  • CYGB protein, human
  • Cygb protein, mouse
  • Cytoglobin
  • Protective Agents
  • Recombinant Proteins