Differential recruitment of monocyte‐derived macrophages in control and stellate cell‐depleted mice during recurrent carbon tetrachloride‐induced acute liver injury

Abstract

Liver depleted of hepatic stellate cells (HSCs) is resistant to ischemia/reperfusion-, concanavalin A-, and acetaminophen-induced acute injury. Whether HSCs regulate carbon tetrachloride (CCl 4 )-induced acute liver injury is not known. CCl 4 treatment damages pericentral hepatocytes that express CCl 4 -metabolizing Cyp2E1 and activates HSCs. We investigated whether HSC-depletion in mice transgenic for thymidine kinase under the glial fibrillary acidic protein promoter (GFAP-TK-Tg) confers resistance to injury and inflammation due to CCl 4 rechallenge. GFAP-TK-Tg or wild type (WT) mice were administered 0.16 ml/kg CCl 4 (3× at 3 days intervals), then 40 μg/g/day ganciclovir for 10 days. The treatment depletes ~70%-75% HSCs from GFAP-TK-Tg but not WT mice while the liver recovers from earlier CCl 4induced injury. Mice were then administered CCl 4 , and liver injury and inflammation were determined at 24 h. HSC-depleted and HSC-sufficient mice showed similar CCl 4 -induced hepatocyte necrosis and oxidative stress. However, increase in F4/80 + macrophages, but not CD68 + cells, was greater in CCl 4 rechallenged HSC-depleted compared to HSC-sufficient mice. Expression of tumor necrosis factor-α (TNF-α), CCL2, and CXCL1 increased similarly, whereas increase in interleukin-6 (IL6), IL1β, and IL10 expression was higher in CCl 4 rechallenged HSC-depleted compared to HSC-sufficient mice. CCl 4 rechallenge of HSC-sufficient mice rapidly activated HSCs causing significant fibrosis with increased expression of Col1a1, transforming growth factor β1 (TGFβ1), tissue inhibitors of metalloproteinases 1 (TIMP1); increase in TIPM1 was much lower and metalloproteinases 13 (MMP13) greater in CCl 4 rechallenged HSC-depleted mice. Interestingly, hepatic recruitment of both profibrogenic (Ly6C hi ) and antifibrogenic restorative (Ly6C lo ) macrophages, and neutrophils was significantly greater in CCl 4 rechallenged HSC-depleted mice. These data suggest that CCl 4 directly damages hepatocytes but HSCs regulate inflammation. Rapid fibrogenesis in CCl 4 rechallenged HSC-sufficient mice recovered from earlier injury indicates that even transiently activated HSCs that had reverted to the quiescent phenotype remain primed to become reactivated.

Assessing the combined impact of fatty liver-induced TGF-β1 and LPS-activated macrophages in fibrosis through a novel 3D serial section methodology

“…As recent studies have shown that macrophages characterized by high F4/80 and Ly6C expression, which are activated in response to hepatocyte necrosis and inflammation, produce inflammatory cytokines such as TNF-α and IL-1β 33 , 34 , which exacerbates inflammation. In this study, the proportions of F4/80 + and Ly6C + macrophages in the liver were analyzed.…”

“…Once activated, macrophages secrete factors that exacerbate inflammation. Specifically, in the liver, F4/80 + Ly6C + macrophages are known to produce inflammatory cytokines such as TNF-α and IL-1β 33 , 34 . These cytokines activate HSCs, leading to collagen fiber production, which is foundational to fibrosis.…”

Stellate cell in hepatic inflammation and acute injury

“…Since expression of cytoglobin, which regulates CYP2E1‐induced APAP metabolism in hepatocytes and is expressed primarily by HSCs (Teranishi et al, 2015), is reduced in HSC‐depleted mice, it is plausible that the metabolism of APAP to toxic metabolites is inhibited in vivo in HSC‐depleted mice. However, when HSC‐depleted mice were challenged with CCl4, which is also metabolized by CYP2E1 to the toxic trichloromethyl (CCl3˙) and trichloromethyl peroxyl (CCl3COO˙) radicals, hepatic injury was similar to that in HSC‐sufficient mice (Sharma et al, 2022). This suggests that despite lower activity of CYP2E1 in vivo due to cytoglobin deficiency, the liver cells in HSC‐depleted mice are able to metabolize CCl4 with similar efficiency as HSC‐sufficient mice.…”