This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. A major drawback of the NHP models is the inability to perform functional studies. While vast amounts of data can now be collected in NHPs using system-wide approaches, it is not possible to functionally characterize the role of interesting gene/pathways, via loss/gain of function studies. Approaches that allow in-vivo silencing in macaques would therefore be of immense value. Recently, the feasibility of this approach for NHPs has been demonstrated by the delivery of siRNAs encapsulated in cationic lipidated nanoparticles in-vivo. We propose to generate lipidoid like siRNAs specific to the rhesus SOCS3 (Suppressor of cytokine signaling 3). SOCS3 is crucial for modulating the expression of pro-inflammatory cytokine genes in response to Mtb infection. SOCS3 is expressed in rhesus macrophages infected with M. tuberculosis (Mtb) ex-vivo as well as in mature (week 12-13) NHP granulomas. We propose to infect four NHPs with Mtb and administer lipidoid siRNA five weeks post-infection. Through this approach, we expect to silence the expression of SOCS3 at a time when it is likely required by the host to reprogram pro-inflammatory response. The animals will be exposed to the siRNA via bronchoscopy or intravenous injection. The expression of SOCS3 will be temporally assessed in peripheral blood mononuclear cell population, temporal broncho-alveolar lavage cells, and in biopsied lung and LN tissue at critical times (pre-infection, 2 weeks, 4 weeks and 8 weeks post-infection). The animals will be necropsied at 12 weeks post-infection and the expression of SOCS3 will be assessed in terminal samples as well. If the in-vivo silencing of SOCS3 can be demonstrated, the microbiological, clinical, immunological and pathological correlates of TB infection in NHPs will be analyzed in this context.
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