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CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation

The kidney is especially sensitive to diseases associated with overactivation of the complement system. While most of these diseases affect kidney glomeruli and the microvasculature, there is also evidence for tubulointerstitial deposition of complement factors. Complement inactivating factors on ce...

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Autores principales: Wieser, Matthias, Francisci, Teresa, Lackner, Daniel, Buerckstuemmer, Tilmann, Wasner, Kamilla, Eilenberg, Wolf, Stift, Anton, Wahrmann, Markus, Böhmig, Georg A., Grillari, Johannes, Grillari-Voglauer, Regina
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6453361/
https://www.ncbi.nlm.nih.gov/pubmed/30958843
http://dx.doi.org/10.1371/journal.pone.0214514
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author Wieser, Matthias
Francisci, Teresa
Lackner, Daniel
Buerckstuemmer, Tilmann
Wasner, Kamilla
Eilenberg, Wolf
Stift, Anton
Wahrmann, Markus
Böhmig, Georg A.
Grillari, Johannes
Grillari-Voglauer, Regina
author_facet Wieser, Matthias
Francisci, Teresa
Lackner, Daniel
Buerckstuemmer, Tilmann
Wasner, Kamilla
Eilenberg, Wolf
Stift, Anton
Wahrmann, Markus
Böhmig, Georg A.
Grillari, Johannes
Grillari-Voglauer, Regina
author_sort Wieser, Matthias
collection PubMed
description The kidney is especially sensitive to diseases associated with overactivation of the complement system. While most of these diseases affect kidney glomeruli and the microvasculature, there is also evidence for tubulointerstitial deposition of complement factors. Complement inactivating factors on cell membranes comprise CD55, CD59 and CD46, which is also termed membrane cofactor protein (MCP). CD46 has been described as localized to glomeruli, but especially also to proximal tubular epithelial cells (RPTECs). However, human cell culture models to assess CD46 function on RPTECs are still missing. Therefore, we here performed gene editing of RPTEC/TERT1 cells generating a monoclonal CD46(-/-) cell line that did not show changes of the primary cell like characteristics. In addition, factor I and CD46-mediated cleavage of C4b into soluble C4c and membrane deposited C4d was clearly reduced in the knock-out cell line as compared to the maternal cells. Thus, human CD46(-/-) proximal tubular epithelial cells will be of interest to dissect the roles of the epithelium and the kidney in various complement activation mediated tubulointerstitial pathologies or in studying CD46 mediated uropathogenic internalization of bacteria. In addition, this gives proof-of-principle, that telomerized cells can be used in the generation of knock-out, knock-in or any kind of reporter cell lines without losing the primary cell characteristics of the maternal cells.
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spelling pubmed-64533612019-04-19 CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation Wieser, Matthias Francisci, Teresa Lackner, Daniel Buerckstuemmer, Tilmann Wasner, Kamilla Eilenberg, Wolf Stift, Anton Wahrmann, Markus Böhmig, Georg A. Grillari, Johannes Grillari-Voglauer, Regina PLoS One Research Article The kidney is especially sensitive to diseases associated with overactivation of the complement system. While most of these diseases affect kidney glomeruli and the microvasculature, there is also evidence for tubulointerstitial deposition of complement factors. Complement inactivating factors on cell membranes comprise CD55, CD59 and CD46, which is also termed membrane cofactor protein (MCP). CD46 has been described as localized to glomeruli, but especially also to proximal tubular epithelial cells (RPTECs). However, human cell culture models to assess CD46 function on RPTECs are still missing. Therefore, we here performed gene editing of RPTEC/TERT1 cells generating a monoclonal CD46(-/-) cell line that did not show changes of the primary cell like characteristics. In addition, factor I and CD46-mediated cleavage of C4b into soluble C4c and membrane deposited C4d was clearly reduced in the knock-out cell line as compared to the maternal cells. Thus, human CD46(-/-) proximal tubular epithelial cells will be of interest to dissect the roles of the epithelium and the kidney in various complement activation mediated tubulointerstitial pathologies or in studying CD46 mediated uropathogenic internalization of bacteria. In addition, this gives proof-of-principle, that telomerized cells can be used in the generation of knock-out, knock-in or any kind of reporter cell lines without losing the primary cell characteristics of the maternal cells. Public Library of Science 2019-04-08 /pmc/articles/PMC6453361/ /pubmed/30958843 http://dx.doi.org/10.1371/journal.pone.0214514 Text en © 2019 Wieser et al http://creativecommons.org/licenses/by/4.0/ This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Wieser, Matthias
Francisci, Teresa
Lackner, Daniel
Buerckstuemmer, Tilmann
Wasner, Kamilla
Eilenberg, Wolf
Stift, Anton
Wahrmann, Markus
Böhmig, Georg A.
Grillari, Johannes
Grillari-Voglauer, Regina
CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation
title CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation
title_full CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation
title_fullStr CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation
title_full_unstemmed CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation
title_short CD46 knock-out using CRISPR/Cas9 editing of hTERT immortalized human cells modulates complement activation
title_sort cd46 knock-out using crispr/cas9 editing of htert immortalized human cells modulates complement activation
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6453361/
https://www.ncbi.nlm.nih.gov/pubmed/30958843
http://dx.doi.org/10.1371/journal.pone.0214514
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