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Delivery of Anti-IFNAR1 shRNA to Hepatic Cells Decreases IFNAR1 Gene Expression and Improves Adenoviral Transduction and Transgene Expression

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Abstract

Chronic liver injury leads to advanced fibrosis, cirrhosis, and hepatocellular carcinoma. Genetical cell treatment related to the use of adenovirus (Ads) has proven to be beneficial and efficient in the recovery of hepatic diseases. Nevertheless, they are highly immunogenic and trigger an immune response where interferons type 1 (IFN-I) play a very important role. Three shRNAs against the Interferon-1 receptor (IFNAR1) were designed and cloned in pENTR/U6 plasmid and amplified in DH5α cells. Huh7 cells were transfected with these plasmids in the presence or absence of 1 × 109 viral particles/ml of adenovirus containing the green fluorescent protein gene used as a reporter. Transfection with the shRNA plasmids partially inhibited the IFNAR1 expression. This inhibition substantially decreased antiviral response, demonstrated by the decrease of IFNAR1, IFN-α, and TNF-α gene expression, and the decrease at protein levels of IFNAR1, Protein kinase RNA-activated (PKR), and phosphorylated STAT1, allowing higher adenoviral transduction and transgene expression. Interestingly it was seen shRNA inhibited macrophage activation. These results suggest that the inhibition of the IFN-I pathway could be a strategy to minimize the immune response against Adenoviral vectors allowing higher Adenovirus transduction extending the transgene expression.

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Funding

Funding was provided by Universidad de Guadalajara (Grant No. 248576 PROGRAMA DE APOYO A LA MEJORA DE LAS CONDICIONES DE PRODUCCION DE LOS MIEMBROS DEL SNI (PROSNI) Y SNCA 2019)

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Authors and Affiliations

  1. Instituto de Investigación en Enfermedades Crónico-Degenerativas, Centro Universitario de Ciencias de la Salud, Universidad de Guadalajara, Sierra Mojada 950, Col. Independencia, C.P. 44340, Guadalajara, Jalisco, Mexico

    J. Guerrero-Rodríguez, G. Gutierrez-Silerio, B. Bastidas-Ramírez, C. Gurrola-Díaz, L. Gasca-Lozano & A. Salazar-Montes

  2. Universidad Autónoma de Zacatecas, Jardín Juárez #147, Centro Histórico, C.P. 98000, Zacatecas, Zacatecas, Mexico

    A. Cárdenas-Vargas

  3. Centro Universitario de Tonalá, Universidad de Guadalajara, Av. Nuevo Periférico No. 555 Ejido San José Tateposco, C.P. 45425, Tonalá, Jalisco, Mexico

    A. Sobrevilla-Navarro & L. Hernández-Ortega

  4. Instituto de Biología Molecular en Medicina y Terapia Génica, Centro Universitario de Ciencias de la Salud, Universidad de Guadalajara, Sierra Mojada 950, Col. Independencia, C.P. 44340, Guadalajara, Jalisco, Mexico

    J. Armendáriz-Borunda

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  1. J. Guerrero-Rodríguez
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  2. A. Cárdenas-Vargas
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  3. G. Gutierrez-Silerio
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Contributions

JG-R: He worked in Experimental phase performing. Cell culture, cloning and PCR. AC-V: Performed cell culture and flow cytometry. GG-S: Performed the flow cytometry analysis. AS-N: Performed the western-blot assays. BB-R: Contributed to the discussion of the results. LH-O: He designed the shRNA molecules and the cloning. CG-D: Analysis of the results. LG-L: Contributed to the discussion of the results. JA-B: Revision of the manuscript. AS-M: Administration of the project and manuscript writing.

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Correspondence to A. Salazar-Montes.

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Guerrero-Rodríguez, J., Cárdenas-Vargas, A., Gutierrez-Silerio, G. et al. Delivery of Anti-IFNAR1 shRNA to Hepatic Cells Decreases IFNAR1 Gene Expression and Improves Adenoviral Transduction and Transgene Expression. Mol Biotechnol 64, 413–423 (2022). https://doi.org/10.1007/s12033-021-00408-6

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  • DOI: https://doi.org/10.1007/s12033-021-00408-6

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