Download files
Abstract
Gastric cancer (GC) is the fifth most common cancer and the third most common cause of cancer-related death worldwide. While Helicobacter pylori infection has been established as the most important risk factor for GC, previous evidence suggests that host intrinsic factors such as the immune response also contribute to the development of GC. Initial host recognition of H. pylori is known to involve pattern recognition receptors (PRRs) (mainly Toll-like receptors (TLRs) and NOD-like receptors (NLRs)), protease activated receptors (PARs) (mainly PAR-1 and PAR-2) and autophagy. Activation of these signalling pathways leads to the production of a large array of pro-inflammatory and anti-microbial molecules that are critical for the elimination of the pathogen, but under chronic conditions, can become detrimental to the host. Thus, the overall aim of the current PhD thesis was to provide a major advance in our understanding of innate immune recognition of H. pylori, and to investigate the association of this immune response with the development of GC.
As there is limited evidence on the association between polymorphisms in genes encoding TLRs, NLRs, PARs and molecules involved in autophagy, and risk of GC, and given that the incidence of GC in Chinese individuals represents 42% of the cases worldwide, we conducted case-control studies comprising 310 ethnic Chinese individuals (87 non-cardia GC cases and 223 controls with functional dyspepsia), in which 81 polymorphisms (25 in the TLR signalling pathway, 51 in the NLR signalling pathway, 3 in autophagy and 2 in PAR-1) were investigated in relation to GC. Further, given the limited understanding of the role of the NLR and autophagy signalling pathways during H. pylori infection, we investigated the gene expression of 168 molecules involved in these signalling pathways in mammalian cells (gastric epithelial and/or monocytic cell lines) challenged with different H. pylori strains (H. pylori GC026, a highly virulent strain isolated from a GC patient, and H. pylori 26695, a reference strain isolated from a gastritis patient).
In Publications I, III, IV and V, novel polymorphisms in genes encoding molecules involved in the TLR, NLR, PARs and autophagy signalling pathways, were associated with GC in an ethnic Chinese population. In multivariate analyses, five polymorphisms significantly modified the risk of GC in ethnic Chinese individuals (TLR4 rs11536889, CARD8 rs11672725, ATG16L1 rs2241880, IRGM rs4958847 and PAR-1 IVSn -14 A>T). Further, TLR4 rs10759932, NLRP12 rs2866112 and ATG16L1 rs2241880, were found to modulate the risk of H. pylori infection in this population, the main risk factor for GC. Strikingly, statistical analyses assessing the joint effect of H. pylori and the selected polymorphisms revealed that H. pylori-infected individuals harbouring 19 polymorphisms involved in the mentioned signalling pathways (TLR2 rs3804100, TLR2 -196 to -174del, TLR4 rs11536889, MD-2 rs11465996, MD-2 rs16938755, LBP rs2232578, TIRAP rs7932766, CARD8 rs10405717, NLRP3 rs12079994, NLRP3 rs3806265, NLRP3 rs4612666, NLRP12 rs2866112, NLRP12 rs4419163, NLRX1 rs10790286, CASP1 rs2282659, CASP1 rs530537, CASP1 rs61751523, ATG16L1 rs2241880 and IRGM rs13361189), were at most risk of developing GC.
In Publication II, we performed the first global meta-analysis to assess the role of TLR2, TLR4 and CD14 polymorphisms in gastric carcinogenesis, in an attempt to clarify the limited and current conflicting evidence, and to establish the true impact of the TLR signalling pathway in this pathology. This meta-analysis, which included 18 case-control studies conducted in Caucasian, Asian and Latin American populations, showed that TLR4 Asp299Gly was a definitive risk factor for GC in Western populations (pooled OR: 1.87, 95% CI: 1.31â 2.65). In addition, there was a potential association between TLR2 -196 to -174 and GC in Japanese (pooled OR: 1.18, 95% CI: 0.96â 1.45).
Further, in Publications III and IV, H. pylori was shown to significantly regulate the expression of a number of molecules involved in the NLR and autophagy signalling pathways. In Publication III, NLRP12 and NLRX1, two known NF-κB negative regulators, were markedly down-regulated, and NFKB1 and several NF-κB target genes encoding pro-inflammatory cytokines, chemokines and molecules involved in carcinogenesis were markedly up-regulated in THP-1-derived macrophages infected with a highly virulent H. pylori ¬strain isolated from a GC patient. These novel findings highlight the relevance of the NLR signalling pathways in gastric carcinogenesis and their close interaction with NF-κB. Publication IV is the first study to provide evidence that H. pylori modulates the expression of core autophagy proteins and autophagy regulators in both non-immune (gastric epithelial) and immune (monocytic) host cells. In addition, a number of molecules involved in autophagy that are also known tumour suppressors (Atg4c, Atg5, Beclin-1, UVRAG and PTEN) and enhancers (p62/SQSTM1) were significantly down-regulated and up-regulated in H. pylori GC026-challenged AGS cells, respectively.
In conclusion, the findings reported in this thesis not only provide further evidence that host genetic factors play an important role in GC pathogenesis but also improve our understanding of the host innate immune response to H. pylori infection. In addition, these advances add evidence to the notion that GC is a multifactorial process that is initiated by H. pylori and perpetuated by host immunogenetics.