Furthermore, apoptotic tumor cells were more frequently observed in tumors from TLR4−/− mice than in tumors from wt mice (Fig. 2E). Moreover, the serum ALT was modestly reduced Selleckchem Palbociclib in tumor-bearing TLR4−/− mice compared with tumor-bearing wt mice, indicating a lower tumor load indirectly (Supporting Information Fig. 2A). Because TLR4 activation of innate immune cells resulted in the production of several inflammatory cytokines that stimulated tumor growth,

we thus assessed whether the absence of TLR4 influences cancer-linked inflammatory responses. Indeed, in addition to the smaller number and size of tumors in TLR4−/− mice, these lesions were consistently associated with reduced infiltration of macrophages (F4/80 staining) compared to wt mice (Supporting Information Fig. 2B). Concordantly, the expression levels of hepatomitogens (TNFα and IL-6) were evidently reduced in TLR4−/− HCCs relative to controls (Fig. 2F). However, unlike the DEN-induced rat HCC model, no evident liver fibrosis CHIR-99021 order was found in this model (Supporting Information Fig. 2C). Thus, the loss of TLR4 protects the liver from chemically induced carcinogenesis, possibly because

of less pronounced inflammation, reduced proliferation, and enhanced apoptosis in tumor cells. The finding that loss of TLR4 reduced the susceptibility of mice to chemical hepatocarcinogenesis prompted us to examine the early effects Morin Hydrate of DEN on cell behavior and signal transduction. At 24 or 48 hours after DEN injection, TLR4−/−

males displayed a considerable elevation of ALT in serum and an increased number of TUNEL-positive cells in liver, indicating the presence of exacerbated hepatocyte damage (Fig. 3A,B,D). The histological evidence of damage was likewise increased in TLR4−/− mice compared to wt mice (Supporting Information Fig. 3). DEN administration led to a rapid increase in expression of the p53 target genes p21 and Mdm2, but the response was similar in wt and TLR4−/− mice, excluding the possibility that TLR4 affects DEN metabolism (Supporting Information Fig. 4). These data suggest that deletion of TLR4 may result in more DEN-induced cell death. The mammalian liver possesses an extraordinary capacity for compensatory growth and thereby maintains liver mass after liver loss or injury.17 We analyzed 5-ethynyl-2′-deoxyuridine (EdU) incorporation 72 and 96 hours after DEN administration.18 As compared with wt mice, loss of TLR4 resulted in a substantial decrease in proliferating hepatocytes (Fig. 3C,D). Deletion of TLR4 significantly reduced the magnitude and duration of Jnk and Erk mitogenic signals after DEN exposure compared to wt mice (Fig. 3E). Therefore, both the enhanced cell apoptosis and reduced proliferative response likely account for the observed lower susceptibility of TLR4−/− mice to chemical hepatocarcinogenesis.