Several biomedical applications, such as xenotransplantation, require multiple genes simultaneously expressed in eukaryotic cells. Advances in genetic engineering technologies have led to the development of efficient polycistronic vectors based on the use of the 2A self-processing oligopeptide. The aim of this work was to evaluate the protective effects of the simultaneous expression of a novel combination of anti-inflammatory human genes, ENTPD1, E5NT and HO-1, in eukaryotic cells. We produced an F2A system-based multicistronic construct to express three human proteins in NIH3T3 cells exposed to an inflammatory stimulus represented by tumor necrosis factor alpha (TNF-α), a pro-inflammatory cytokine which plays an important role during inflammation, cell proliferation, differentiation and apoptosis and in the inflammatory response during ischemia/reperfusion injury in several organ transplantation settings. The protective effects against TNF-α-induced cytotoxicity and cell death, mediated by HO-1, ENTPD1 and E5NT genes were better observed in cells expressing the combination of genes as compared to cells expressing each single gene and the effect was further improved by administrating enzymatic substrates of the human genes to the cells. Moreover, a gene expression analyses demonstrated that the expression of the three genes has a role in modulating key regulators of TNF-α signalling pathway, namely Nemo and Tnfaip3, that promoted pro-survival phenotype in TNF-α injured cells. These results could provide new insights in the research of protective mechanisms in transplantation settings.

Cinti, A., De Giorgi, M., Chisci, E., Arena, C., Galimberti, G., Farina, L., et al. (2015). Simultaneous overexpression of functional human HO-1, E5NT and ENTPD1 protects murine fibroblasts against TNF-α-induced injury in vitro. PLOS ONE, 10(10) [10.1371/journal.pone.0141933].

Simultaneous overexpression of functional human HO-1, E5NT and ENTPD1 protects murine fibroblasts against TNF-α-induced injury in vitro

CINTI, ALESSANDRO
Primo
;
CHISCI, ELISA;GALIMBERTI, GLORIA;FARINA, LAURA;RIVOLTA, ILARIA;GAIPA, GIUSEPPE;CERRITO, MARIA GRAZIA;LAVITRANO, MARIALUISA
Penultimo
;
GIOVANNONI, ROBERTO
Ultimo
2015

Abstract

Several biomedical applications, such as xenotransplantation, require multiple genes simultaneously expressed in eukaryotic cells. Advances in genetic engineering technologies have led to the development of efficient polycistronic vectors based on the use of the 2A self-processing oligopeptide. The aim of this work was to evaluate the protective effects of the simultaneous expression of a novel combination of anti-inflammatory human genes, ENTPD1, E5NT and HO-1, in eukaryotic cells. We produced an F2A system-based multicistronic construct to express three human proteins in NIH3T3 cells exposed to an inflammatory stimulus represented by tumor necrosis factor alpha (TNF-α), a pro-inflammatory cytokine which plays an important role during inflammation, cell proliferation, differentiation and apoptosis and in the inflammatory response during ischemia/reperfusion injury in several organ transplantation settings. The protective effects against TNF-α-induced cytotoxicity and cell death, mediated by HO-1, ENTPD1 and E5NT genes were better observed in cells expressing the combination of genes as compared to cells expressing each single gene and the effect was further improved by administrating enzymatic substrates of the human genes to the cells. Moreover, a gene expression analyses demonstrated that the expression of the three genes has a role in modulating key regulators of TNF-α signalling pathway, namely Nemo and Tnfaip3, that promoted pro-survival phenotype in TNF-α injured cells. These results could provide new insights in the research of protective mechanisms in transplantation settings.
Articolo in rivista - Articolo scientifico
Multicistronic vector; 2A peptide; inflammation; heme oxygenase-1; Ectonucleotidases; Cytoprotection
English
Cinti, A., De Giorgi, M., Chisci, E., Arena, C., Galimberti, G., Farina, L., et al. (2015). Simultaneous overexpression of functional human HO-1, E5NT and ENTPD1 protects murine fibroblasts against TNF-α-induced injury in vitro. PLOS ONE, 10(10) [10.1371/journal.pone.0141933].
File in questo prodotto:
File Dimensione Formato  
journal.pone.0141933.pdf

accesso aperto

Descrizione: Cinti et al PLOS ONE 2015
Dimensione 788.29 kB
Formato Adobe PDF
788.29 kB Adobe PDF Visualizza/Apri

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/10281/92833
Citazioni
  • Scopus 6
  • ???jsp.display-item.citation.isi??? 6
Social impact