Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats.
BACKGROUND:There is a correlation between the endocannabinoid system and hepatic fibrosis based on the activation of CB1 and CB2 receptors; where CB1 has profibrogenic effects. Gene therapy with a plasmid carrying a shRNA for CB1 delivered by hydrodynamic injection has the advantage of hepatic tropi...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2020-01-01
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| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0228729 |
| _version_ | 1818925274341834752 |
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| author | Adriana Díaz-Rivera Alejandra Meza-Ríos Victoria Chagoya de Sánchez Gabriela Velasco-Loyden Leonel García-Benavides Luis F Jave-Suarez Hugo Christian Monroy-Ramirez Arturo Santos-García Juan Armendáriz-Borunda Ana Sandoval-Rodríguez |
| author_facet | Adriana Díaz-Rivera Alejandra Meza-Ríos Victoria Chagoya de Sánchez Gabriela Velasco-Loyden Leonel García-Benavides Luis F Jave-Suarez Hugo Christian Monroy-Ramirez Arturo Santos-García Juan Armendáriz-Borunda Ana Sandoval-Rodríguez |
| author_sort | Adriana Díaz-Rivera |
| collection | DOAJ |
| description | BACKGROUND:There is a correlation between the endocannabinoid system and hepatic fibrosis based on the activation of CB1 and CB2 receptors; where CB1 has profibrogenic effects. Gene therapy with a plasmid carrying a shRNA for CB1 delivered by hydrodynamic injection has the advantage of hepatic tropism, avoiding possible undesirable effects of CB1 pharmacological inhibition. OBJECTIVE:To evaluate hydrodynamics-based liver transfection in an experimental model of liver cirrhosis of a plasmid with the sequence of a shRNA for CB1 and its antifibrogenic effects. METHODS:Three shRNA (21pb) were designed for blocking CB1 mRNA at positions 877, 1232 and 1501 (pshCB1-A, B, C). Sequences were cloned in the pENTR™/U6. Safety was evaluated monitoring CB1 expression in brain tissue. The silencing effect was determined in rat HSC primary culture and CCl4 cirrhosis model. Hydrodynamic injection in cirrhotic liver was through iliac vein and with a dose of 3mg/kg plasmid. Serum levels of liver enzymes, mRNA levels of TGF-β1, Col IA1 and α-SMA and the percentage of fibrotic tissue were analyzed. RESULTS:Hydrodynamic injection allows efficient CB1 silencing in cirrhotic livers and pshCB1-B (position 1232) demonstrated the main CB1-silencing. Using this plasmid, mRNA level of fibrogenic molecules and fibrotic tissue considerably decrease in cirrhotic animals. Brain expression of CB1 remained unaltered. CONCLUSION:Hydrodynamics allows a hepatotropic and secure transfection in cirrhotic animals. The sequence of the shCB1-B carried in a plasmid or any other vector has the potential to be used as therapeutic strategy for liver fibrosis. |
| first_indexed | 2024-12-20T02:38:37Z |
| format | Article |
| id | doaj.art-dd83ede8704f46dbb743a58fb091fe0f |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-20T02:38:37Z |
| publishDate | 2020-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-dd83ede8704f46dbb743a58fb091fe0f2022-12-21T19:56:22ZengPublic Library of Science (PLoS)PLoS ONE1932-62032020-01-01152e022872910.1371/journal.pone.0228729Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats.Adriana Díaz-RiveraAlejandra Meza-RíosVictoria Chagoya de SánchezGabriela Velasco-LoydenLeonel García-BenavidesLuis F Jave-SuarezHugo Christian Monroy-RamirezArturo Santos-GarcíaJuan Armendáriz-BorundaAna Sandoval-RodríguezBACKGROUND:There is a correlation between the endocannabinoid system and hepatic fibrosis based on the activation of CB1 and CB2 receptors; where CB1 has profibrogenic effects. Gene therapy with a plasmid carrying a shRNA for CB1 delivered by hydrodynamic injection has the advantage of hepatic tropism, avoiding possible undesirable effects of CB1 pharmacological inhibition. OBJECTIVE:To evaluate hydrodynamics-based liver transfection in an experimental model of liver cirrhosis of a plasmid with the sequence of a shRNA for CB1 and its antifibrogenic effects. METHODS:Three shRNA (21pb) were designed for blocking CB1 mRNA at positions 877, 1232 and 1501 (pshCB1-A, B, C). Sequences were cloned in the pENTR™/U6. Safety was evaluated monitoring CB1 expression in brain tissue. The silencing effect was determined in rat HSC primary culture and CCl4 cirrhosis model. Hydrodynamic injection in cirrhotic liver was through iliac vein and with a dose of 3mg/kg plasmid. Serum levels of liver enzymes, mRNA levels of TGF-β1, Col IA1 and α-SMA and the percentage of fibrotic tissue were analyzed. RESULTS:Hydrodynamic injection allows efficient CB1 silencing in cirrhotic livers and pshCB1-B (position 1232) demonstrated the main CB1-silencing. Using this plasmid, mRNA level of fibrogenic molecules and fibrotic tissue considerably decrease in cirrhotic animals. Brain expression of CB1 remained unaltered. CONCLUSION:Hydrodynamics allows a hepatotropic and secure transfection in cirrhotic animals. The sequence of the shCB1-B carried in a plasmid or any other vector has the potential to be used as therapeutic strategy for liver fibrosis.https://doi.org/10.1371/journal.pone.0228729 |
| spellingShingle | Adriana Díaz-Rivera Alejandra Meza-Ríos Victoria Chagoya de Sánchez Gabriela Velasco-Loyden Leonel García-Benavides Luis F Jave-Suarez Hugo Christian Monroy-Ramirez Arturo Santos-García Juan Armendáriz-Borunda Ana Sandoval-Rodríguez Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats. PLoS ONE |
| title | Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats. |
| title_full | Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats. |
| title_fullStr | Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats. |
| title_full_unstemmed | Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats. |
| title_short | Hydrodynamics-based liver transfection achieves gene silencing of CB1 using short hairpin RNA plasmid in cirrhotic rats. |
| title_sort | hydrodynamics based liver transfection achieves gene silencing of cb1 using short hairpin rna plasmid in cirrhotic rats |
| url | https://doi.org/10.1371/journal.pone.0228729 |
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