STIMULATORY AND INHIBITORY ACTION OF CYTOKINES ON THE REGULATION OF hCMV-IE PROMOTER ACTIVITY IN HUMAN ENDOTHELIAL CELLS

Abstract

Viral promoters are commonly used as regulatory elements in gene therapy vectors due to their strong activity in various cell lines in vitro. However, transgene expression under the control of viral promoters in vivo has been shown to be limited to a short period of time. Several mechanisms for the transient expression of the delivered transgene may be important including deletion of transduced cells or promoter downregulation. Recently we reported that cytokines may either decrease or increase the activity of the human cytomegalovirus (hCMV) promoter in monocytes depending on the differentiation status of the transduced cells. For many applications, the gene of interest has to be delivered into an inflammatory milieu (tumour, ischaemia/reperfusion, vector-induced inflammation etc.). In this report we investigated the influence of various inflammatory cytokines on the hCMV-IE promoter activity in transduced human primary endothelial cells (Huvec) in vitro, which may be the first target cells after gene transfer into different organs. Cultured cells were infected with an E1-deleted adenoviral vector encoding for E. coliβ-galactosidase (Adβ-gal) driven by the hCMV-IE promoter and incubated either with or without various cytokines. Our results indicate that interferon-γ (IFN-γ) and interleukin-10 (IL-10) downregulate promoter activity in endothelial cells whereas, in contrast, tumour necrosis factor (TNF-α), interleukin 1β (IL-1β) and interleukin 4 (IL-4) increased the promoter activity. These results suggest that inflammatory processes influence the in vivo expression of transferred viral promoter controlled genes of interest.