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Updated: Jul 19, 2026
Development and Functional Characterization of Murine Tolerogenic Dendritic Cells
Published on: May 18, 2018
Effects of RNA interference on CD80 and CD86 expression in bone marrow-derived murine dendritic cells
1Department of General Surgery, Huashan Hospital, Fudan University, Shanghai, China.
Abstract:
To investigate whether RNA interference (RNAi) induced by small interfering RNA (siRNA) could suppress CD80 and CD86 expression in bone marrow-derived murine dendritic cells (DC). The bone marrow-derived DC of mice were separated and cultured in vitro, chemically synthesized siRNA were then transferred into the cells by LipofectAMINE 2000, and the siRNA transfection efficacy was assessed by both fluorescence microscope and flow cytometry. The mRNA expression and protein synthesis were analysed by real-time RT-PCR and flow cytometry. The cell viability of transfected DC was determined by annexin V and propidium iodine staining. Transfection of bone marrow-derived murine DC with a non-silencing FITC-labelled control siRNA demonstrated a high (71.86%) transfection efficiency without affecting cellular viability. CD80-1 siRNA was the most effective siRNA to block CD80 expression in three candidates. Similarly, CD86-3 siRNA was extraordinarily effective in repressing the expression of CD86. Cotransfection of siRNA specific to CD80 and CD86 can enhance gene silencing that is not affected by DC activation-inducing signals. CD80 and CD86 siRNA suppressed the expression of CD80 and CD86 to 31.05 +/- 2.41% and 25.43 +/- 0.85%, respectively, of the level in untreated cells (P < 0.05). siRNA is capable of triggering RNAi in bone marrow-derived DC; it can specifically and effectively knock down CD80 and CD86 gene expression. This approach is a useful tool by which costimulatory molecules of DC can be studied as well as a potential therapeutic option for allograft rejection.
Insights
Small interfering RNA (siRNA) effectively suppresses CD80 and CD86 expression in dendritic cells (DCs) via RNA interference (RNAi). This targeted gene silencing in DCs offers a promising therapeutic strategy for allograft rejection.
Area of Science:
- Immunology
- Molecular Biology
- Cell Biology
Background:
- Dendritic cells (DCs) play a crucial role in immune responses.
- CD80 and CD86 are key costimulatory molecules on DCs that regulate T-cell activation.
- Modulating DC costimulatory molecule expression is a potential therapeutic strategy for immune-related disorders.
Purpose of the Study:
- To investigate the efficacy of RNA interference (RNAi) using small interfering RNA (siRNA) to suppress CD80 and CD86 expression in murine bone marrow-derived dendritic cells (DCs).
Main Methods:
- Bone marrow-derived murine DCs were cultured in vitro.
- Chemically synthesized siRNA targeting CD80 and CD86 were transfected into DCs using LipofectAMINE 2000.
- Transfection efficiency was assessed by fluorescence microscopy and flow cytometry.
- mRNA and protein expression levels of CD80 and CD86 were analyzed using real-time RT-PCR and flow cytometry.
- Cell viability was determined by annexin V and propidium iodide staining.
Main Results:
- High transfection efficiency (71.86%) was achieved with FITC-labelled control siRNA without compromising cell viability.
- Specific siRNA sequences (CD80-1 and CD86-3) demonstrated high efficacy in suppressing CD80 and CD86 expression, respectively.
- Co-transfection of CD80 and CD86 siRNA resulted in significant gene silencing (to 31.05% and 25.43% of untreated levels, respectively, P < 0.05), independent of DC activation signals.
Conclusions:
- siRNA effectively induces RNA interference (RNAi) in bone marrow-derived DCs.
- This approach specifically and efficiently knocks down CD80 and CD86 gene expression.
- This method serves as a valuable tool for studying DC costimulatory molecules and holds potential for therapeutic applications in allograft rejection.
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