Knockout of liver fluke granulin, Ov-grn-1, impedes malignant transformation during chronic infection with Opisthorchis viverrini

Journal Publication ResearchOnline@JCU
Chaiyadet, Sujittra;Tangkawattana, Sirikachorn;Smout, Michael J.;Ittiprasert, Wannaporn;Mann, Victoria H.;Deenonpoe, Raksawan;Arunsan, Patpicha;Loukas, Alex;Brindley, Paul J.;Laha, Thewarach
Michael SmoutAlex Loukas
Abstract

Infection with the food-borne liver fluke Opisthorchis viverrini is the principal risk factor for cholangiocarcinoma (CCA) in the Mekong Basin countries of Thailand, Lao PDR, Vietnam, Myanmar and Cambodia. Using a novel model of CCA, involving infection with gene-edited liver flukes in the hamster during concurrent exposure to dietary nitrosamine, we explored the role of the fluke granulin-like growth factor Ov-GRN-1 in malignancy. We derived RNA-guided gene knockout flukes (ΔOv-grn-1) using CRISPR/Cas9/gRNA materials delivered by electroporation. Genome sequencing confirmed programmed Cas9-catalyzed mutations of the targeted genes, which was accompanied by rapid depletion of transcripts and the proteins they encode. Gene-edited parasites colonized the biliary tract of hamsters and developed into adult flukes. However, less hepatobiliary tract disease manifested during chronic infection with ΔOv-grn-1 worms in comparison to hamsters infected with control gene-edited and mock-edited parasites. Specifically, immuno- and colorimetric-histochemical analysis of livers revealed markedly less periductal fibrosis surrounding the flukes and less fibrosis globally within the hepatobiliary tract during infection with ΔOv-grn-1 genotype worms, minimal biliary epithelial cell proliferation, and significantly fewer mutations of TP53 in biliary epithelial cells. Moreover, fewer hamsters developed high-grade CCA compared to controls. The clinically relevant, pathophysiological phenotype of the hepatobiliary tract confirmed a role for this secreted growth factor in malignancy and morbidity during opisthorchiasis.

Journal

PLoS Pathogens

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18

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1553-7374

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9

Pages Count

30

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Publisher

Public Library of Science

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DOI

10.1371/journal.ppat.1010839