Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.)
DNA vaccines are administered in the form of plasmid DNA (pDNA) carrying a strong promoter and the gene of interest. In this study we investigated the tissue distribution, cellular uptake and the fate of intravenously (i.v.) and intramuscularly (i.m.) injected pDNA in Atlantic cod ( Gadus morhua L.)...
Published in: | Journal of Experimental Biology |
---|---|
Main Authors: | , , , , |
Format: | Text |
Language: | English |
Published: |
Company of Biologists
2007
|
Subjects: | |
Online Access: | http://jeb.biologists.org/cgi/content/short/210/12/2091 https://doi.org/10.1242/jeb.003186 |
id |
fthighwire:oai:open-archive.highwire.org:jexbio:210/12/2091 |
---|---|
record_format |
openpolar |
spelling |
fthighwire:oai:open-archive.highwire.org:jexbio:210/12/2091 2023-05-15T15:27:49+02:00 Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.) Seternes, Tore Tonheim, Tom C. Løvoll, Marie Bøgwald, Jarl Dalmo, Roy A. 2007-06-15 00:00:00.0 text/html http://jeb.biologists.org/cgi/content/short/210/12/2091 https://doi.org/10.1242/jeb.003186 en eng Company of Biologists http://jeb.biologists.org/cgi/content/short/210/12/2091 http://dx.doi.org/10.1242/jeb.003186 Copyright (C) 2007, Company of Biologists Research Article TEXT 2007 fthighwire https://doi.org/10.1242/jeb.003186 2015-02-28T18:21:26Z DNA vaccines are administered in the form of plasmid DNA (pDNA) carrying a strong promoter and the gene of interest. In this study we investigated the tissue distribution, cellular uptake and the fate of intravenously (i.v.) and intramuscularly (i.m.) injected pDNA in Atlantic cod ( Gadus morhua L.). The anatomical distribution of pDNA was determined using both morphological and radiotracing methods. Cellular uptake and receptor specificity were studied in cultures of cod atrial endocardial endothelial cells (aEEC) and head kidney leukocytes. The short-term fate of the endocytosed pDNA in vivo and in vitro was investigated by Southern blot. Expression of the pDNA (R70pRomiLuc)-derived gene was investigated in cod tissues and cultures of cod aEEC by means of real-time RT-PCR and luciferase activity assay. 125I-labelled pDNA was rapidly eliminated from the blood by the aEEC of the cod heart atrium and ventricle. Co-injection of trace amounts of 125I-labelled pDNA with excess amounts of non-labelled pDNA or formaldehyde-treated albumin (FSA), a ligand for the cod EEC scavenger receptor, significantly inhibited the accumulation of the radiotracer in the heart. The organ to blood ratio of radioactivity after inhibition of the cod EEC scavenger receptor demonstrated that the radioactivity not taken up by the EEC remained in the blood. Fluorescence microscopy of tissue sections from cod injected with fluorescein-labelled pDNA confirmed intracellular uptake of pDNA by the endocardial cells of the atrium and ventricle. In purified cultures of cod aEEC the fluorescein-labelled pDNA was taken up in structures reminiscent of endosomal/lysosomal vesicles. Uptake of 125I-labelled pDNA in cultures of cod aEEC was specific. Incubation of cultures with 125I-labelled pDNA together with excess amounts of FSA and fucoidan, which are molecules also known to bind to the scavenger receptors, reduced the uptake of the pDNA by at least 70%. Mannan, a ligand for the mannose receptor, did not inhibit the uptake of 125I-labelled pDNA. ... Text atlantic cod Gadus morhua HighWire Press (Stanford University) Journal of Experimental Biology 210 12 2091 2103 |
institution |
Open Polar |
collection |
HighWire Press (Stanford University) |
op_collection_id |
fthighwire |
language |
English |
topic |
Research Article |
spellingShingle |
Research Article Seternes, Tore Tonheim, Tom C. Løvoll, Marie Bøgwald, Jarl Dalmo, Roy A. Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.) |
topic_facet |
Research Article |
description |
DNA vaccines are administered in the form of plasmid DNA (pDNA) carrying a strong promoter and the gene of interest. In this study we investigated the tissue distribution, cellular uptake and the fate of intravenously (i.v.) and intramuscularly (i.m.) injected pDNA in Atlantic cod ( Gadus morhua L.). The anatomical distribution of pDNA was determined using both morphological and radiotracing methods. Cellular uptake and receptor specificity were studied in cultures of cod atrial endocardial endothelial cells (aEEC) and head kidney leukocytes. The short-term fate of the endocytosed pDNA in vivo and in vitro was investigated by Southern blot. Expression of the pDNA (R70pRomiLuc)-derived gene was investigated in cod tissues and cultures of cod aEEC by means of real-time RT-PCR and luciferase activity assay. 125I-labelled pDNA was rapidly eliminated from the blood by the aEEC of the cod heart atrium and ventricle. Co-injection of trace amounts of 125I-labelled pDNA with excess amounts of non-labelled pDNA or formaldehyde-treated albumin (FSA), a ligand for the cod EEC scavenger receptor, significantly inhibited the accumulation of the radiotracer in the heart. The organ to blood ratio of radioactivity after inhibition of the cod EEC scavenger receptor demonstrated that the radioactivity not taken up by the EEC remained in the blood. Fluorescence microscopy of tissue sections from cod injected with fluorescein-labelled pDNA confirmed intracellular uptake of pDNA by the endocardial cells of the atrium and ventricle. In purified cultures of cod aEEC the fluorescein-labelled pDNA was taken up in structures reminiscent of endosomal/lysosomal vesicles. Uptake of 125I-labelled pDNA in cultures of cod aEEC was specific. Incubation of cultures with 125I-labelled pDNA together with excess amounts of FSA and fucoidan, which are molecules also known to bind to the scavenger receptors, reduced the uptake of the pDNA by at least 70%. Mannan, a ligand for the mannose receptor, did not inhibit the uptake of 125I-labelled pDNA. ... |
format |
Text |
author |
Seternes, Tore Tonheim, Tom C. Løvoll, Marie Bøgwald, Jarl Dalmo, Roy A. |
author_facet |
Seternes, Tore Tonheim, Tom C. Løvoll, Marie Bøgwald, Jarl Dalmo, Roy A. |
author_sort |
Seternes, Tore |
title |
Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.) |
title_short |
Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.) |
title_full |
Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.) |
title_fullStr |
Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.) |
title_full_unstemmed |
Specific endocytosis and degradation of naked DNA in the endocardial cells of cod (Gadus morhua L.) |
title_sort |
specific endocytosis and degradation of naked dna in the endocardial cells of cod (gadus morhua l.) |
publisher |
Company of Biologists |
publishDate |
2007 |
url |
http://jeb.biologists.org/cgi/content/short/210/12/2091 https://doi.org/10.1242/jeb.003186 |
genre |
atlantic cod Gadus morhua |
genre_facet |
atlantic cod Gadus morhua |
op_relation |
http://jeb.biologists.org/cgi/content/short/210/12/2091 http://dx.doi.org/10.1242/jeb.003186 |
op_rights |
Copyright (C) 2007, Company of Biologists |
op_doi |
https://doi.org/10.1242/jeb.003186 |
container_title |
Journal of Experimental Biology |
container_volume |
210 |
container_issue |
12 |
container_start_page |
2091 |
op_container_end_page |
2103 |
_version_ |
1766358222513373184 |