Immune modulation through RNA interference-mediated silencing of CD40 in dendritic cells.
RNA interference (RNAi) is an exciting mechanism for knocking down any target gene in transcriptional level. It is now clear that small interfering RNA (siRNA), a 19-21nt long dsRNA, can trigger a degradation process (RNAi) that specifically silences the expression of a cognate mRNA. Our findings in this study showed that down regulation of CD40 gene expression in dendritic cells (DCs) by RNAi culminated to immune modulation. Effective delivery of siRNA into DCs would be a reasonable method for the blocking of CD40 gene expression at the cell surface without any effect on other genes and cell cytotoxicity. The effects of siRNA against CD40 mRNA on the function and phenotype of DCs were investigated. The DCs were separated from the mice spleen and then cultured in vitro. By the means of Lipofectamine2000, siRNA was delivered to the cells and the efficacy of transfection was estimated by flow cytometry. By Annexine V and Propidium Iodide staining, we could evaluate the transfected cells viability. Also, the mRNA expression and protein synthesis were assessed by real-time PCR and flow cytometry, respectively. Knocking down the CD40 gene in the DCs caused an increase in IL-4 production, decrease in IL-12 production and allostimulation activity. All together, these effects would stimulate Th2 cytokines production from allogenic T-cells in vitro.
Karimi
Mohammad Hossein
MH
Immunology Department, Tarbiat Modares University, Tehran, Iran.
Ebadi
Padideh
P
Pourfathollah
Ali Akbar
AA
Soheili
Zahra Soheila
ZS
Samiee
Shahram
S
Ataee
Zahra
Z
Tabei
Seyyed Ziyaoddin
SZ
Moazzeni
Seyed Mohammad
SM
D000818:Animals; D019013:CD40 Antigens / Q000235:genetics* / Q000276:immunology; D002454:Cell Differentiation / Q000187:drug effects / Q000276:immunology; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology*; D020868:Gene Silencing / Q000276:immunology; D007371:Interferon-gamma / Q000276:immunology / Q000378:metabolism; D018664:Interleukin-12 / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000276:immunology; D015847:Interleukin-4 / Q000819:agonists / Q000096:biosynthesis / Q000276:immunology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D018418:Th2 Cells / Q000276:immunology / Q000378:metabolism
D019013:CD40 Antigens; D034741:RNA, Small Interfering; D018664:Interleukin-12; D015847:Interleukin-4; D007371:Interferon-gamma