TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus Cells
Mechanical loading and inflammation interact to cause degenerative disc disease and low back pain (LBP). However, the underlying mechanosensing and mechanotransductive pathways are poorly understood. This results in untargeted pharmacological treatments that do not take the mechanical aspect of LBP...
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MDPI AG
2020-07-01
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| Series: | Cells |
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| Online Access: | https://www.mdpi.com/2073-4409/9/7/1736 |
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| author | Elena Cambria Matthias J. E. Arlt Sandra Wandel Olga Krupkova Wolfgang Hitzl Fabian S. Passini Oliver N. Hausmann Jess G. Snedeker Stephen J. Ferguson Karin Wuertz-Kozak |
| author_facet | Elena Cambria Matthias J. E. Arlt Sandra Wandel Olga Krupkova Wolfgang Hitzl Fabian S. Passini Oliver N. Hausmann Jess G. Snedeker Stephen J. Ferguson Karin Wuertz-Kozak |
| author_sort | Elena Cambria |
| collection | DOAJ |
| description | Mechanical loading and inflammation interact to cause degenerative disc disease and low back pain (LBP). However, the underlying mechanosensing and mechanotransductive pathways are poorly understood. This results in untargeted pharmacological treatments that do not take the mechanical aspect of LBP into account. We investigated the role of the mechanosensitive ion channel TRPV4 in stretch-induced inflammation in human annulus fibrosus (AF) cells. The cells were cyclically stretched to 20% hyperphysiological strain. TRPV4 was either inhibited with the selective TRPV4 antagonist GSK2193874 or knocked out (KO) via CRISPR-Cas9 gene editing. The gene expression, inflammatory mediator release and MAPK pathway activation were analyzed. Hyperphysiological cyclic stretching significantly increased the IL6, IL8, and COX2 mRNA, PGE2 release, and activated p38 MAPK. The TRPV4 pharmacological inhibition significantly attenuated these effects. TRPV4 KO further prevented the stretch-induced upregulation of IL8 mRNA and reduced IL6 and IL8 release, thus supporting the inhibition data. We provide novel evidence that TRPV4 transduces hyperphysiological mechanical signals into inflammatory responses in human AF cells, possibly via p38. Additionally, we show for the first time the successful gene editing of human AF cells via CRISPR-Cas9. The pharmacological inhibition or CRISPR-based targeting of TRPV4 may constitute a potential therapeutic strategy to tackle discogenic LBP in patients with AF injury. |
| first_indexed | 2024-03-10T18:19:46Z |
| format | Article |
| id | doaj.art-55ba19fd9cee4acc986df1334826d34c |
| institution | Directory Open Access Journal |
| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-03-10T18:19:46Z |
| publishDate | 2020-07-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj.art-55ba19fd9cee4acc986df1334826d34c2023-11-20T07:23:25ZengMDPI AGCells2073-44092020-07-0197173610.3390/cells9071736TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus CellsElena Cambria0Matthias J. E. Arlt1Sandra Wandel2Olga Krupkova3Wolfgang Hitzl4Fabian S. Passini5Oliver N. Hausmann6Jess G. Snedeker7Stephen J. Ferguson8Karin Wuertz-Kozak9Institute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandInstitute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandInstitute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandInstitute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandResearch Office (Biostatistics), Paracelsus Medical University, 5020 Salzburg, AustriaInstitute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandNeuro- and Spine Center, Hirslanden Klinik St. Anna, 6006 Lucerne, SwitzerlandInstitute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandInstitute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandInstitute for Biomechanics, ETH Zurich, 8093 Zurich, SwitzerlandMechanical loading and inflammation interact to cause degenerative disc disease and low back pain (LBP). However, the underlying mechanosensing and mechanotransductive pathways are poorly understood. This results in untargeted pharmacological treatments that do not take the mechanical aspect of LBP into account. We investigated the role of the mechanosensitive ion channel TRPV4 in stretch-induced inflammation in human annulus fibrosus (AF) cells. The cells were cyclically stretched to 20% hyperphysiological strain. TRPV4 was either inhibited with the selective TRPV4 antagonist GSK2193874 or knocked out (KO) via CRISPR-Cas9 gene editing. The gene expression, inflammatory mediator release and MAPK pathway activation were analyzed. Hyperphysiological cyclic stretching significantly increased the IL6, IL8, and COX2 mRNA, PGE2 release, and activated p38 MAPK. The TRPV4 pharmacological inhibition significantly attenuated these effects. TRPV4 KO further prevented the stretch-induced upregulation of IL8 mRNA and reduced IL6 and IL8 release, thus supporting the inhibition data. We provide novel evidence that TRPV4 transduces hyperphysiological mechanical signals into inflammatory responses in human AF cells, possibly via p38. Additionally, we show for the first time the successful gene editing of human AF cells via CRISPR-Cas9. The pharmacological inhibition or CRISPR-based targeting of TRPV4 may constitute a potential therapeutic strategy to tackle discogenic LBP in patients with AF injury.https://www.mdpi.com/2073-4409/9/7/1736mechanotransductioncyclic stretchingtransient receptor potential channelgene editinginterleukinslow back pain |
| spellingShingle | Elena Cambria Matthias J. E. Arlt Sandra Wandel Olga Krupkova Wolfgang Hitzl Fabian S. Passini Oliver N. Hausmann Jess G. Snedeker Stephen J. Ferguson Karin Wuertz-Kozak TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus Cells Cells mechanotransduction cyclic stretching transient receptor potential channel gene editing interleukins low back pain |
| title | TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus Cells |
| title_full | TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus Cells |
| title_fullStr | TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus Cells |
| title_full_unstemmed | TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus Cells |
| title_short | TRPV4 Inhibition and CRISPR-Cas9 Knockout Reduce Inflammation Induced by Hyperphysiological Stretching in Human Annulus Fibrosus Cells |
| title_sort | trpv4 inhibition and crispr cas9 knockout reduce inflammation induced by hyperphysiological stretching in human annulus fibrosus cells |
| topic | mechanotransduction cyclic stretching transient receptor potential channel gene editing interleukins low back pain |
| url | https://www.mdpi.com/2073-4409/9/7/1736 |
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