Granulin Secreted by the Food-Borne Liver Fluke Opisthorchis viverrini Promotes Angiogenesis in Human Endothelial Cells.

Brandon Haugen,Michael J Smout,Anastas Popratiloff,Victoria H Mann,Shannon E Karinshak,Paul J Brindley,Alex Loukas

doi:10.3389/fmed.2018.00030

Abstract

The liver fluke Opisthorchis viverrini is a food-borne, zoonotic pathogen endemic to Thailand and adjacent countries in Southeast Asia. The adult developmental stage of the O. viverrini parasite excretes and secretes numerous proteins within the biliary tract including the gall bladder. Lesions caused by the feeding activities of the liver fluke represent wounds that undergo protracted cycles of healing and re-injury during chronic infection, which can last for decades. Components of the excretory/secretory (ES) complement released by the worms capably drive proliferation of bile duct epithelial cells and are implicated in establishing the oncogenic milieu that leads to bile duct cancer, cholangiocarcinoma. An ES protein, the secreted granulin-like growth factor termed Ov-GRN-1, accelerates wound resolution in mice and in vitro. To investigate angiogenesis (blood vessel development) that may contribute to wound healing promoted by liver fluke granulin and, by implication, to carcinogenesis during chronic opisthorchiasis, we employed an in vitro tubule formation assay (TFA) where human umbilical vein endothelial cells were grown on gelled basement matrix. Ten and 40 nM Ov-GRN-1 significantly stimulated angiogenesis as monitored by cellular proliferation and by TFA in real time. This demonstration of potent angiogenic property of Ov-GRN-1 bolsters earlier reports on the therapeutic potential for chronic non-healing wounds of diabetics, tobacco users, and the elderly and, in addition, showcases another of the hallmark of cancer characteristic of this carcinogenic liver fluke.

Highlights

Infection with the fish-borne liver flukes Opisthorchis viverrini, Opisthorchis felineus, and Clonorchis sinensis remains a major public health problem in East Asia and Eurasia with >40 million cases
We have shown that the granulin-like growth factor, Ov-GRN-1, secreted by the fluke is the dominant proliferative factor and is sufficient to drive wound healing [2, 7]
As the angiogenic assay with human umbilical vein endothelial cells (HUVECs) would be assessed at 12 h, for proliferation, we focused on the first 24 h and noted minor (5–11%) non-significant increase above values

Summary

Introduction

Infection with the fish-borne liver flukes Opisthorchis viverrini, Opisthorchis felineus, and Clonorchis sinensis remains a major public health problem in East Asia and Eurasia with >40 million cases. Eating undercooked fish infected with the metacercariae stage of the fluke leads to infection in humans and other mammals, such as cats and dogs [1]. The metacercarial stage of the parasite excysts in the duodenum and the juvenile fluke migrates into the bile ducts of the liver and matures over a month into an adult fluke, which grazes on biliary epithelia. The parasite develops within the snail, in turn releasing cercariae that seek out and penetrate the skin of a cyprinid fish, which encyst in the fish as a metacercaria, the infective stage for humans and other definitive host species [1]

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