In addition, the hepatic mRNA

levels of fibrosis-related genes, including Z-IETD-FMK purchase collagen α1(I), α-SMA, and TGF-β1, were low in NOX1KO and NOX2KO mice compared with WT mice after CCl4 treatment (Fig. 3D) or BDL (Fig. 3E). There was no difference in hepatic expression of M1 or M2 macrophage markers between WT, NOX1KO, or NOX2KO mice (Supporting information Fig. 3A,B). These results suggest that both NOX1 and NOX2 may be directly involved in the activation of HSCs. We measured the lipid peroxidation products 4-hydroxynonenal and malondialdehyde as indicators of oxidative stress in the liver in NOX1KO, NOX2KO, and WT mice after CCl4 or BDL treatment. Immunofluorescence staining showed lower hepatic 4-hydroxynonenal Selleckchem Trametinib levels in NOX1KO and NOX2KO mice compared with WT mice after CCl4 or BDL treatment (Fig. 4A). Measurement of malondialdehyde using thiobarbituric acid–reactive

substances showed that NOX1KO and NOX2KO mice have lower levels of lipid peroxidation compared with WT mice after CCl4 or BDL treatment (Fig. 4B,C), suggesting that both NOX1 and NOX2 play an important role in the generation of hepatic oxidative stress in response to CCl4 or BDL in mice. To characterize the contributory roles of NOX1 and NOX2 in hepatic fibrosis in different liver cell populations, we generated NOX1 and NOX2 BM chimeric mice using a combination of lethal irradiation, KC depletion by way of clodronate injection, and BMT. This combination generates complete substitution of KCs and other BM-derived cells, but not of resident hepatic cell populations, including HSCs and SECs.18, 19 Eight weeks after BMT, hepatic fibrosis was induced by way of CCl4 treatment for 1 month. Serum ALT levels were lower in NOX1 chimeric mice with NOX1-deficient endogenous liver cells (WT BMNOX1KO and NOX1KO BMNOX1KO) compared with WT mice transplanted with WT BM (Fig. 5B). As expected, NOX1KO mice transplanted with NOX1KO BM had reduced hepatic fibrosis

compared with WT mice transplanted with WT BM. NOX1 chimeric mice that express NOX1 in BM-derived selleck products cells but not endogenous liver cells (WT BMNOX1KO) showed the similar reduction of hepatic fibrosis as mice with complete NOX1 deficiency. However, NOX1 chimeric mice that expressed NOX1 in endogenous liver cells but not BM-derived cells (NOX1KO BMWT) showed the same levels of fibrosis as WT mice (Fig. 5A,C). Serum ALT levels were reduced in NOX2 chimeric mice with NOX2-deficient endogenous liver cells (WT BMNOX2KO and NOX2KO BMNOX2KO) compared with WT mice transplanted with WT BM (Fig. 5E). NOX2KO mice transplanted with NOX2-deficient BM had reduced hepatic fibrosis compared with WT mice transplanted with WT BM. NOX2 chimeric mice that expressed NOX2 in BM-derived cells but not endogenous liver cells (WT BMNOX2KO) showed a reduction in hepatic fibrosis similar to those with complete NOX2 deficiency.