Opisthorchis viverrini excretory/secretory products induce toll-like receptor 4 upregulation and production of interleukin 6 and 8 in cholangiocyte

Abstract

Biliary tract infection with the Group I carcinogenic liver fluke Opisthorchis viverrini is associated with severe inflammation leading to cholangiocarcinoma – a major biliary cancer in Southeast Asia. However, mechanism(s) by which the liver fluke induces host mucosal immune/inflammatory responses are unclear. In the present study we address whether a normal immortalized human cholangiocyte cell line (H69 cells) recognizes and responds to O. viverrini excretory/secretory products (OVES). Expression of multiple TLRs, activation of NF-κB, and expression of proinflammatory cytokines were monitored in the presence and absence of OVES. Our results showed that OVES induced increased cholangiocyte TLR4 mRNA expression, induced IκB-α degradation in a MyD88-dependent manner, and activated NF-κB nuclear translocation. Moreover, OVES induced expression and secretion of the strong chemoattractant chemokine interleukin 8 (IL-8) and pro-inflammatory cytokine IL-6. These results demonstrate that secreted/excreted products of O. viverrini are recognized by human cholangiocytes and initiate innate mucosal immunity/inflammatory cascades, a primary event in the pathogenesis of opisthorchiasis and cholangiocarcinoma.

Excretory/secretory products of the carcinogenic liver fluke are endocytosed by human cholangiocytes and drive cell proliferation and IL6 production

“…The bacterium Helicobacter pylori secretes cag A, a virulence factor whose uptake by epithelial cells can result in gastric cancer (Hatakeyama, 2004). Given that Ov ES products induce severe inflammation by up-regulation of cholangiocyte Toll-like receptor (TLR)4 mRNA expression, thereby promoting IL6 and IL-8 production (Ninlawan et al, 2010), it is reasonable to assume that internalized Ov ES products also promote a tumorigenic phenotype.…”

“…Opisthorchis viverrini metacercariae were obtained from naturally infected cyprinoid fish captured from an endemic area in Khon Kaen province, north-eastern Thailand, as described previously (Ninlawan et al, 2010). Briefly, fish were digested with pepsin-HCl, and after several washes with normal saline, metacercariae were collected, identified under a dissecting microscope and used to infect hamsters.…”

“…Normal human cholangiocytes (H69) were maintained in DMEM/HamF-12 (Gibco, USA) supplemented with insulin, adenine, epinephrine, T3-T, epidermal growth factors (EGF) and hydrocortisone (Ninlawan et al, 2010). Human cholangiocarcinoma (KKU-100 and KKU-M156) were maintained in Ham-F12 (Gibco) and human colon cancer (Caco-2) cell lines were maintained in DMEM containing L-gutamine and non-essential amino acids (Gibco) at 37°C, 5% CO 2 with 10% FCS (Gibco) containing 100 U/ml of penicillin and 100 μg/ml of streptomycin solution (Gibco).…”

An Interleukin-6 Receptor Polymorphism is Associated with Opisthorchiasis-Linked Cholangiocarcinoma Risk in Thailand

“…Among others, elevated O. viverrini specific interleukin-6 (IL-6) production has been reported from peripheral blood mononuclear cells from infected individuals with advanced periductal fibrosis and CCA (Sripa et al, 2009;Sripa et al, 2012b) as well as cholangiocytes cocultured with O. viverrini excretory-secretory products (Ninlawan et al, 2010). This pro-inflammatory cytokine may potentiate the proliferation and survival of cholangiocytes within local microenvironment (Sripa et al, 2012a).…”

Cytokine/Chemokine Secretion and Proteomic Identification of Upregulated Annexin A1 from Peripheral Blood Mononuclear Cells Cocultured with the Liver Fluke Opisthorchis viverrini

“…In human studies, elevated levels of IL-6 were found in PBMCs stimulated by O. viverrini excretory/secretory (ES) products (12) and in plasma (13) of opisthorchiasis patients with advanced periductal fibrosis. Moreover, treatment of a normal immortalized human cholangiocyte cell line with O. viverrini ES products also increased the expression of Toll-like receptor 4 (TLR-4) and subsequently stimulated the production of IL-6 and IL-8 (14). In addition, not only cytokines/chemokines but also other molecules, such as the protein peroxiredoxin 6 (Prdx6), a key regulator of the cellular redox balance, might play a role in host defense against O. viverrini infection (15).…”