Current therapies to block the sexual transmission of HIV-1 have utilized microbicidal or antiviral compounds to interfere with viral binding, internalization, or replication. These approaches, however, have met with only limited success. Thus, there is a critical demand to develop novel therapeutics to inhibit mucosal HIV-1 infection and transmission. We examined the efficacy of short interfering RNA (siRNA) molecules to silence expression of CD4 and CCR5 on cells in the reproductive tract of female humanized mice. We postulated that silencing expression of these receptors could inhibit HIV-1 binding and infection. Nanoparticles encapsulating siRNAs to silence CD4 and CCR5 expression, or an irrelevant siRNA, were instilled intravaginally in mice. Three days later the mice were challenged intravaginally with HIV-1. Results demonstrated significant reduction in HIV-1 DNA levels in the peripheral blood regardless of siRNA specificity. Elevated levels of both murine and human TNF-α transcripts were detected in the genital tracts of mice following siRNA delivery. These findings suggest that nucleic acid-based therapies may have the unintended, although highly advantageous, effect of triggering innate immune anti-viral mechanisms. Thus, the development of nucleic acid based-therapies such as the one using siRNA that exploits this potent additive anti-HIV-1 response while concurrently targeting specific HIV-1 recognition receptors, is highly desirable and could be used to supplement microbicidal or antiviral therapies.