MISSION^® In Vivo shRNA and Lentiviral Solutions

MISSION in vivo lentivirus (with or without shRNA) is designed to address the challenges of extending an RNAi study into an in vivo system. Lentivirus-delivered shRNAs in vivo has been accomplished by either single high-titer injections, or serial lower-dose administration. MISSION in vivo lentivirus is suitable for RNAi research in animals using multiple routes of delivery.

MISSION lentivirus can transduce a variety of organs and tissues in vivo through multiple routes of delivery. Figure 1 shows the results of a proof-of-principle experiment in mice using a lentivirus expressing far-red fluorescent protein.

MISSION lentivirus has been successful in direct intratumoral injections. A p55 knockout mouse was injected with 5x10⁵ LLC-1 (Mouse Lewis Lung Carcinoma) cells via a subcutaneous flank injection. The subsequent tumor was injected twice daily, for five days, with 20µL of 1.9x10⁷ TU/ml TurboGFP™-expressing lentivirus (Catalog Number SHC003V). Five days after the final injection, the tumors were biopsied, stained with TO-PRO®-3 (a nuclear stain) and imaged, see Figure 2.

MISSION shRNA has been shown to knockdown expression of Smad3 in direct intramuscular injections (Carlson et al., 2008). Older muscle satellite cells have a reduced muscle regeneration capacity compared to younger muscle satellite cells due to an imbalance in endogenous Smad3 and Notch expression. The authors of this paper hypothesized that this imbalance was due to an increase in Smad3 expression in older muscle satellite cells. When they injected Smad3 shRNA intramuscularly into damaged muscle tissue, the older muscle tissue was able to recover its regenerative capacity when assayed five days post-injection. The authors found that a balance between endogenous Smad3 and active Notch controls the regenerative competence of muscle satellite cells.

Figure 3 shows hematoxylin and eosin staining of young and old muscle tissue and the corresponding Hoechst staining (blue) and myosin heavy chain antibody staining (green). BrdU staining and the eMyHC antibody staining is shown in the offset images.

MISSION lentivirus has been successful in a xenograft mouse model. HeLa cells were transduced with far-red fluorescent protein (TagFP635) lentivirus and injected subcutaneously into a nude mouse. Figure 4 shows strong local fluorescence of the resulting HeLa cell tumor expressing far-red fluorescent protein in mice.

• Wiederschain, D., et al., Single-vector inducible lentiviral RNAi system for oncology target validation. Cell Cycle 8, 498-504 (2009). Pubmed ID 19177017.
• Krishnamachary, B., et al., Noninvasive Detection of Lentiviral-Mediated Choline Kinase Targeting in a Human Breast Cancer Xenograft. Cancer Res. 69, 3464-71 (2009). Pubmed ID 19336572
• Carlson, M.E., et al., Imbalance between pSmad3 and Notch induces CDK inhibitors in old muscle stem cells. Nature 454, 528-534 (2008). Pubmed ID 18552838
• Croyle, M.A., et al., PEGylation of a vesicular stomatitis virus G pseudotyped lentivirus vector prevents inactivation in serum. J Virol 78, 912-921 (2004). Pubmed ID 14694122
• Pan, D., et al., Biodistribution and toxicity studies of VSVG-pseudotyped lentiviral vector after intravenous administration in mice with the observation of in vivo transduction of bone marrow. Mol Ther 6, 19-29 (2002). Pubmed ID 12095299

For questions about the library, pricing and quotes or other concerns, please email us at: RNAi@sial.com. MISSION is a registered trademark of Sigma-Aldrich Biotechnology LP and Sigma-Aldrich Co. Label License. TO-PRO is a registered trademark of Molecular Probes, Inc. TurboGFP and TagFP635 are trademarks of Evrogen Co.