Hepatic stellate cells are critically involved in modulation of inflammatory microenvironment during acute liver injury
Authors: Sharma Akanksha , Singh Atul K. , Kudira Ramesh , Miethke Alexander , Wang Jiang , Gandhi Chandrashekhar R. 
Published: April 2020

Background and Aims

Depletion of hepatic stellate cells (HSCs) was shown to protect the liver from acute injury due to ischemia/reperfusion and concanavalin A. Depletion protocol involves 3 administrations of carbon tetrachloride (CCl4) (3 days apart) to mice transgenic for thymidine kinase under the glial fibrillary acidic protein (GFAP (expressed exclusively by HSCs in the liver) promoter (GFAP‐TK/TG). This causes activation of HSCs that become vulnerable to ganciclovir (GCV)‐induced death. After 10‐days GCV treatment, 70–75% HSCs are depleted and the liver completely recovers from prior CCl4‐induced injury. We aimed to determine CCl4‐induced acute liver injury in HSC‐depleted mice and the role of HSCs in regulating hepatic inflammatory microenvironment.

Methods

GFAP‐TK/TG or wild type (WT) B6 mice (Control) were treated with 0.16 ml/kg CCl4 (3 injections) then 40 μg/g/d GCV for 10 days. Other controls were GFAP‐TK/TG mice treated with CCl4 but not GCV (does not deplete HSCs) or naïve B6 mice. Mice then received 0.16 ml/kg CCl4 and mechanisms of liver injury were determined at 24h.

Results

HSC‐depleted mice showed similar CCl4‐induced necrotic liver damage and oxidative stress as HSC‐sufficient mice. CCl4 administration to HSC‐sufficient mice and not HSC‐depleted mice caused reduction in F4/80+ macrophages, but CD68+ cells and inflammation as measured by expression of TNFα, IL6 and IFNβ showed greater increase in HSC‐depleted mice than HSC‐sufficient mice. Surprisingly, CCl4 administration to HSC‐sufficient mice caused rapid activation of HSCs and significant fibrosis accompanied by increased expression of collagen 1a1, TGFβ1 and TGFβR1. As expected, α‐SMA+ activated fibrogenic HSCs were found in HSC‐sufficient and not HSC‐depleted mice. CCl4‐treated HSC‐sufficient mice also increased hepatic IL17‐producing γδT cells and Ly6c+ monocytes. In contrast, MMP13 and MMP9 (collagenases) expression increased in CCl4‐treated HSC‐depleted mice indicating increase in restorative macrophages (found during fibrosis resolution). Finally, in co‐culture, HSCs were found to affect the expression of various inflammatory cytokines/cytokines by CD4+ T cells, regulatory T cells and Kupffer cells variably.

Conclusion

These data suggest that CCl4 has direct damaging effect on hepatocytes but HSCs regulate inflammatory response of the resident and infiltrating cells. Rapid activation of HSCs and fibrosis after CCl4 administration to CCl4/GCV‐pre‐treated and recovered WT mice indicate that even transiently activated HSCs that revert to the quiescent physiological phenotype upon removal of injury stimulus remain primed to become quickly re‐activated to cause fibrosis. This should be an important consideration when targeting HSCs to treat fibrosis clinically.

Support or Funding Information

Support: VA Merit Review Award 1IO1BX001174 and DoD W81XWH‐14‐PRMRP‐IIRA to CRG and NIH P30 DK078392 to the Digestive Diseases Health Center at Cincinnati Children’s Hospital Medical Center.