Inhibiting the toll-like receptor 4 Toll/interleukin-1 receptor domain protects against hepatic warm ischemia and reperfusion injury in mice.

OBJECTIVE: During Toll-like receptor 4 signaling, the Toll/interleukin-1 receptor domain is essential for interactions with downstream Toll/interleukin-1 receptor domain-containing adaptor proteins. The aim of this study is to investigate the role of the Toll/interleukin-1 receptor domain in the Toll-like receptor 4 signaling pathway during hepatic ischemia and reperfusion injury. DESIGN: We genetically blocked the function of Toll/interleukin-1 receptor domain in mice and examined the effect on Toll-like receptor 4 signaling and the response to hepatic ischemia and reperfusion. SETTING: University research laboratory. SUBJECTS: Male BALB/c mice. INTERVENTIONS: Male BALB/c mice were hydrodynamically administrated the target gene plasmid pTIR-IRES2-EGFP, empty vector containing enhanced green fluorescent protein, or normal saline. Animals underwent 90 minutes of partial hepatic ischemia, followed by 6 or 24 hours of reperfusion. Hepatic injury was assessed by measuring serum alanine transaminase, hepatic histology, and malondialdehyde. The expression of inflammatory cytokines and nuclear factor-κB phosphorylation was examined in liver tissues. Hepatic apoptosis was evaluated by caspase-3 assays, terminal deoxynucleotidyl transferase-mediated deoxyuridinetriphosphate nick-end labeling staining, and the activation of Jun N-terminal kinase and p38 mitogen-activated protein kinase. MEASUREMENTS AND MAIN RESULTS: Blocking the Toll/interleukin-1 receptor domain resulted in markedly lower serum alanine transaminase levels, reduced histologic injury, and lower malondialdehyde levels following 6 or 24 hours of hepatic reperfusion than mice receiving the vector alone or normal saline. Anti-Toll/interleukin-1 receptor treatment also reduced Toll-like receptor 4 expression and disrupted Toll/interleukin-1 receptor-Toll/interleukin-1 receptor interactions in Toll-like receptor 4 signaling pathways. Blocking the Toll/interleukin-1 receptor domain also prevented Toll-like receptor 4-mediated mitogen-activated protein kinase activation (via Jun N-terminal kinase and p38 mitogen-activated protein kinase), an activation that mediated liver ischemia and reperfusion injury via caspase-3 activation, resulting in increased hepatocellular apoptosis. Lastly, blocking the Toll/interleukin-1 receptor domain decreased inflammatory cytokine production by inhibiting nuclear factor-κB activation. CONCLUSIONS: Toll/interleukin-1 receptor domain inhibition disrupts the interaction of Toll-like receptor 4 with its adaptor proteins, which abrogates downstream signaling pathways and prevents the activation of nuclear factor-κB and Jun N-terminal kinase/p38. This reduction in signaling consequently reduces hepatic inflammation, cell apoptosis, and hepatic damage. Toll/interleukin-1 receptor domain-targeted therapy thus represents a new approach to inhibit the intracellular Toll-like receptor 4 signaling pathway and reveals novel therapeutic target sites, which will facilitate the development of specific therapeutic agents.

The association between Toll-like receptor 2 single-nucleotide polymorphisms and hepatocellular carcinoma susceptibility.

BACKGROUND: Toll-like receptors (TLR) are key innate immunity receptors participating in an immune response. Growing evidence suggests that mutations of TLR2/TLR9 gene are associated with the progress of cancers. The present study aimed to investigate the temporal relationship of single nucleotide polymorphisms (SNP) of TLR2/TLR9 and the risk of hepatocellular carcinoma (HCC). METHODS: In this single center-based case-control study, SNaPshot method was used to genotype sequence variants of TLR2 and TLR9 in 211 patients with HCC and 232 subjects as controls. RESULTS: Two synonymous SNPs in the exon of TLR2 were closely associated with risk of HCC. Compared with those carrying wild-type homozygous genotypes (T/T), risk of HCC decreased significantly in individuals carrying the heterozygous genotypes (C/T) of the rs3804099 (adjusted odds ratio (OR), 0.493, 95% CI 0.331 - 0.736, P < 0.01) and rs3804100 (adjusted OR, 0.509, 95% CI 0.342 - 0.759, P < 0.01). There was no significant association found in two TLR9 SNPs concerning the risk of HCC. The haplotype TT for TLR2 was associated significantly with the decreased risk of HCC (OR 0.524, 95% CI 0.394 - 0.697, P = 0.000). Inversely, the risk of HCC increased significantly in patients with the haplotype CC (OR 2.743, 95% CI 1.915 - 3.930, P = 0.000). CONCLUSIONS: These results suggested that TLR2 rs3804099 C/T and rs3804100 C/T polymorphisms were closely associated with HCC. In addition, the haplotypes composed of these two TLR2 synonymous SNPs have stronger effects on the susceptibility of HCC.

Single nucleotide polymorphisms of Toll-like receptor 4 decrease the risk of development of hepatocellular carcinoma.

BACKGROUND: Toll-like receptor 4 (TLR4) is a key innate immunity receptor that initiates an inflammatory response. Growing evidence suggests that mutation of TLR4 gene may play a role in the development of cancers. This study aimed to investigate the temporal relationship of single nucleotide polymorphisms of TLR4 and the risk of hepatocellular carcinoma, a single center-based case-control study was conducted. METHODS: A systematic genetic analysis of sequence variants of TLR4 by evaluating ten single-nucleotide polymorphisms was performed from 216 hepatocellular carcinoma cases and 228 controls. RESULTS: Six single nucleotide polymorphisms of the TLR4 in the 5'-untranslated region and intron were associated with risk of hepatocellular carcinoma. Individuals carrying the heterozygous genotypes for the rs10759930, rs2737190, rs10116253, rs1927914, rs12377632 and rs1927911 had significantly decreased risk of hepatocellular carcinoma (adjusted odds ratio [OR], from 0.527 to 0.578, P<0.01) comparing with those carrying wild-type homozygous genotypes. In haplotype analysis, one haplotype (GCCCTTAG) of TLR4 was associated significantly with decrease of the occurrence of hepatocellular carcinoma (OR, 0.556, 95% confidence interval [CI], 0.407-0.758, P = 0.000). CONCLUSIONS: Collectively, these results suggested that the risk of hepatocellular carcinoma was associated with TLR4 sequence variation. TLR4 single nucleotide polymorphisms may play an important protective role in the development of hepatocellular carcinoma.