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Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias
Abetalipoproteinemia (FHBL-SD1) and chylomicron retention disease (FHBL-SD3) are rare recessive disorders of lipoprotein metabolism due to mutations in MTTP and SAR1B genes, respectively, which lead to defective chylomicron formation and secretion. This results in lipid and fat-soluble vitamin malab...
Autores principales: | , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9921550/ https://www.ncbi.nlm.nih.gov/pubmed/36771214 http://dx.doi.org/10.3390/nu15030505 |
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author | Bordat, Claire Vairo, Donato Cuerq, Charlotte Halimi, Charlotte Peiretti, Franck Penhoat, Armelle Vieille-Marchiset, Aurélie Gonzalez, Teresa Michalski, Marie-Caroline Nowicki, Marion Peretti, Noël Reboul, Emmanuelle |
author_facet | Bordat, Claire Vairo, Donato Cuerq, Charlotte Halimi, Charlotte Peiretti, Franck Penhoat, Armelle Vieille-Marchiset, Aurélie Gonzalez, Teresa Michalski, Marie-Caroline Nowicki, Marion Peretti, Noël Reboul, Emmanuelle |
author_sort | Bordat, Claire |
collection | PubMed |
description | Abetalipoproteinemia (FHBL-SD1) and chylomicron retention disease (FHBL-SD3) are rare recessive disorders of lipoprotein metabolism due to mutations in MTTP and SAR1B genes, respectively, which lead to defective chylomicron formation and secretion. This results in lipid and fat-soluble vitamin malabsorption, which induces severe neuro-ophthalmic complications. Currently, treatment combines a low-fat diet with high-dose vitamin A and E supplementation but still fails in normalizing serum vitamin E levels and providing complete ophthalmic protection. To explore these persistent complications, we developed two knock-out cell models of FHBL-SD1 and FHBL-SD3 using the CRISPR/Cas9 technique in Caco-2/TC7 cells. DNA sequencing, RNA quantification and Western blotting confirmed the introduction of mutations with protein knock-out in four clones associated with i) impaired lipid droplet formation and ii) defective triglyceride (−57.0 ± 2.6% to −83.9 ± 1.6%) and cholesterol (−35.3 ± 4.4% to −60.6 ± 3.5%) secretion. A significant decrease in α-tocopherol secretion was also observed in these clones (−41.5 ± 3.7% to −97.2 ± 2.8%), even with the pharmaceutical forms of vitamin E: tocopherol-acetate and tocofersolan (α-tocopheryl polyethylene glycol succinate 1000). MTTP silencing led to a more severe phenotype than SAR1B silencing, which is consistent with clinical observations. Our cellular models thus provide an efficient tool to experiment with therapeutic strategies and will allow progress in understanding the mechanisms involved in lipid metabolism. |
format | Online Article Text |
id | pubmed-9921550 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-99215502023-02-12 Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias Bordat, Claire Vairo, Donato Cuerq, Charlotte Halimi, Charlotte Peiretti, Franck Penhoat, Armelle Vieille-Marchiset, Aurélie Gonzalez, Teresa Michalski, Marie-Caroline Nowicki, Marion Peretti, Noël Reboul, Emmanuelle Nutrients Article Abetalipoproteinemia (FHBL-SD1) and chylomicron retention disease (FHBL-SD3) are rare recessive disorders of lipoprotein metabolism due to mutations in MTTP and SAR1B genes, respectively, which lead to defective chylomicron formation and secretion. This results in lipid and fat-soluble vitamin malabsorption, which induces severe neuro-ophthalmic complications. Currently, treatment combines a low-fat diet with high-dose vitamin A and E supplementation but still fails in normalizing serum vitamin E levels and providing complete ophthalmic protection. To explore these persistent complications, we developed two knock-out cell models of FHBL-SD1 and FHBL-SD3 using the CRISPR/Cas9 technique in Caco-2/TC7 cells. DNA sequencing, RNA quantification and Western blotting confirmed the introduction of mutations with protein knock-out in four clones associated with i) impaired lipid droplet formation and ii) defective triglyceride (−57.0 ± 2.6% to −83.9 ± 1.6%) and cholesterol (−35.3 ± 4.4% to −60.6 ± 3.5%) secretion. A significant decrease in α-tocopherol secretion was also observed in these clones (−41.5 ± 3.7% to −97.2 ± 2.8%), even with the pharmaceutical forms of vitamin E: tocopherol-acetate and tocofersolan (α-tocopheryl polyethylene glycol succinate 1000). MTTP silencing led to a more severe phenotype than SAR1B silencing, which is consistent with clinical observations. Our cellular models thus provide an efficient tool to experiment with therapeutic strategies and will allow progress in understanding the mechanisms involved in lipid metabolism. MDPI 2023-01-18 /pmc/articles/PMC9921550/ /pubmed/36771214 http://dx.doi.org/10.3390/nu15030505 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Bordat, Claire Vairo, Donato Cuerq, Charlotte Halimi, Charlotte Peiretti, Franck Penhoat, Armelle Vieille-Marchiset, Aurélie Gonzalez, Teresa Michalski, Marie-Caroline Nowicki, Marion Peretti, Noël Reboul, Emmanuelle Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias |
title | Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias |
title_full | Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias |
title_fullStr | Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias |
title_full_unstemmed | Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias |
title_short | Validation of Knock-Out Caco-2 TC7 Cells as Models of Enterocytes of Patients with Familial Genetic Hypobetalipoproteinemias |
title_sort | validation of knock-out caco-2 tc7 cells as models of enterocytes of patients with familial genetic hypobetalipoproteinemias |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9921550/ https://www.ncbi.nlm.nih.gov/pubmed/36771214 http://dx.doi.org/10.3390/nu15030505 |
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