Induction of alloimmune tolerance in heart transplantation through gene silencing of TLR adaptors.

Toll-like receptors (TLRs) activate biochemical pathways that evoke activation of innate immunity, which leads to dendritic cell (DC) maturation and initiation of adaptive immune responses that provoke allograft rejection. We aimed to prolong allograft survival by selectively inhibiting expression of the common adaptors of TLR signaling, namely MyD88 and TRIF, using siRNA. In vitro we demonstrated that blocking expression of MyD88 and TRIF led to reduced DC maturation. In vivo treatment of recipients with MyD88 and TRIF siRNA significantly prolonged allograft survival in the BALB/c > C57BL6 cardiac transplant model. Moreover, the combination of MyD88 and TRIF siRNA along with a low dose of rapamycin further extended the allograft survival (88.8 ± 7.1 days). Tissue histopathology demonstrated an overall reduction in lymphocyte interstitium infiltration, vascular obstruction and hemorrhage in mice treated with MyD88 and TRIF siRNA vector plus rapamycin. Furthermore, treatment was associated with an increase in the numbers of CD4(+) CD25(+) FoxP3(+) regulatory T cells and Th2 deviation. To our knowledge, this study is the first demonstration of prolonging the survival of allogeneic heart grafts through gene silencing of TLR signaling adaptors, highlighting the therapeutic potential of siRNA in clinical transplantation.

Inhibition of allergic responses by CD40 gene silencing.

BACKGROUND: Gene silencing using small interfering RNA (siRNA) is a potent method of specifically knocking down molecular targets. Small interfering RNA is therapeutically promising, however, treatment of allergic diseases with siRNA has not been explored in vivo. The aim of this study was to evaluate therapeutic effects of CD40 siRNA on inhibition of allergic responses. METHODS: Mice sensitized with ovalbumin (OVA) and alum were treated with CD40 siRNA, scrambled siRNA, or phosphate buffer saline (PBS) alone, and then challenged intranasally with OVA. RESULTS: A significant reduction in nasal allergic symptoms was observed in the CD40 siRNA treated OVA-allergic mice compared to the controls of scrambled siRNA and PBS alone, which is correlated with the decrease of local eosinophil accumulation. CD40 siRNA treatment knocked down CD40 expression on dendritic cells (DCs) in vivo and impaired their antigen presenting function. Treatment with CD40 siRNA resulted in inhibition of OVA-specific T cell response and decrease of interleukin-4 (IL-4), IL-5, and interferon-gamma production from T cells stimulated with OVA. Administration of CD40 siRNA also suppressed CD40 expression on B cells, resulting in down-regulation of OVA-specific immunoglobulin E (IgE), IgG1, and IgG2a levels. Additionally, increased regulatory T cells were observed in the CD40 siRNA treated mice. CONCLUSIONS: The present study demonstrates a novel therapeutic use for siRNA in allergy. CD40 siRNA attenuated allergy through inhibition of DC and B cell functions and generation of regulatory T (Treg) cells.