I would like to knockdown the expression of various ECM matrix proteins present in the sub-epithelial, ASM and sub-endothelial layers. Would inhalation of lentiviral particles achieve a stable knockdown that penetrates to all of these domains? Would systemic administration be more optimal?
ok... then i.p injection should be fine. Are you doing knockdown in adult mice or new-born pups?
In adult mice - in which tissue would you expect the knockdown to occur following i.p? I am primary interested in the airways so epithlium, ASM, basement membranes, pulmonary vascular endothelium etc.
well.. I havenot experimented in adult mice... but in p3-p7 pups, the effect of shRNA knockdown can be seen in retina...as my experience speaks!!!
Hope it helps.
For example Tyler Jacks laboratory has data that intranasal infection works at least with high-enough titer viruses - since you have shRNA the titer might not be a problem but the question is which proportion of the particular cell types you need to infect? The transduction efficiency is likely not very high but could be sufficient depending on what you want to do/analyze. It appears that viruses also reach lung endothelium. Check eg. these references:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2757265/
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2573216/?report=reader
http://www.fasebj.org/content/early/2013/06/14/fj.13-231225.full.pdf
I did via intranasal on 4 day old rat pups in different time intervals ( p4, p7 and p10), it was very successful. But you can also try intratracheal injection.
Intranasal works fairly well, you can also perform a tracheal instillation (but use small volumes of instillant to decrease the likelihood of killing your animal) if you are interested in getting into the airways. If you instill into the lung, the posterior segment of the right upper lobe is the usual spot for the aspirate to flood; you can remove this section and perform a western to ascertain some degree of knock-down (realize this section will contain multiple cell types so you will not have a high degree of efficiency.
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