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Publication
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| Research Title |
Diminished hepatobiliary disease during infection with CRISPR/Cas9-gene-edited liver flukes, Opisthorchis viverrini |
| Date of Distribution |
16 October 2019 |
| Conference |
| Title of the Conference |
Universal Academic Cluster International Autumn Conference |
| Organiser |
Universal Academic Cluster International Autumn Conference |
| Conference Place |
8th floor, inforRooms Tokyo, Hulic Shiba 4-chome Building, 4Chome-9-1 Shiba, Minato-ku |
| Province/State |
Tokyo , Japan |
| Conference Date |
16 October 2019 |
| To |
18 October 2019 |
| Proceeding Paper |
| Volume |
- |
| Issue |
- |
| Page |
- |
| Editors/edition/publisher |
- |
| Abstract |
Infection with the food-borne liver fluke Opisthorchis viverrini (Ov) causes cholangiocarcinoma (CCA). Whereas the causative agent for most cancers, including CCA in the West, remains obscure, the principal risk factor for CCA in Thailand is opisthorchiasis. We utilized this established link between infection and cancer to explore carcinogenesis of Ov-induced CCA. We targeted the liver fluke gene encoding granulin (Ov-grn-1) gene for knockout (KO) by deletion mutation of the coding region of the gene (Ov-grn-1). Both adult and infective larval flukes (newly excysted juveniles, or NEJ) were transfected with a plasmid encoding a guide RNA sequence specific for 20 nucleotides 5'- to a prototypic adjacent motif in exon-1 of Ov-grn-1 and also encoding the Cas9 nuclease of Streptococcus pyogenes. Illumina based deep sequencing of amplicon libraries from genomic DNAs from the worms demonstrated the presence of Cas9-catalyzed insertions/deletions (INDELs) within the Ov-GRN-1 locus, and tandem analyses by RT-PCR and Western blots revealed rapid depletion of Ov-GRN-1 transcripts and Ov-GRN-1 translates. Infection of hamsters with CRISPR/Cas9 gene edited NEJ enabled studies of liver fluke infection and biliary tract disease in hamsters. Following introduction of CRISPR/Cas9 nuclease plasmid into NEJ, marked reduction of Ov-GRN-1 transcripts was evident within days. The findings demonstrated transfection of NEJ with the gene editing plasmid, that KO abolished expression of Ov-GRN-1 by >95%, and infectivity of wild type (WT) and Ov-GRN-1 gene-edited NEJ for hamsters. Ov-GRN-1 KO flukes induced significantly less biliary tract disease. Ov-GRN-1 KO parasites were infectious, colonized the biliary tract, grew and developed, were active and motile, and induced a clinically relevant pathophysiological tissue phenotype that significantly differed from infection with WT liver flukes. |
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| Peer Review Status |
ไม่มีผู้ประเมินอิสระ |
| Level of Conference |
นานาชาติ |
| Type of Proceeding |
Abstract |
| Type of Presentation |
Poster |
| Part of thesis |
true |
| Presentation awarding |
false |
| Attach file |
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| Citation |
0
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Journal Publication
ไทย