I plan to deplete alveolar macrophages in BALB/c mice using clodronate liposomes in order to evaluate their implication in pneumovirus infections. To my knowledge, the best solution is intranasal injection of 100 µl of clodronate liposomes per mouse 48 hours before viral infections. However, it seems to be difficult for animals to receive a single administration of 100 µl without risk of suffocation. Therefore, I'd like to know whether we can devide for example this volume in a half (50 µl/times, 2 times). Furthermore, do clodronate liposomes induce any side effects such as respiratory troubles, weight loss, mortality, etc. in mice?
Thanks a lot for your answers.
Hi Vuong,
I will suggest you to use 50 ul only, to avoid suffocaion of mice, but u can double the amount of clodronate to get the complete depletion of Alv Macrophages.
Best,
Hello Vuong,
Normally, 30 ul of inoculant is preferred as higher volumes of fluid in the lungs can slightly alter the tissue environment (through increases in the number of recruited monocytes for instance). BALB/c mice are more susceptible to lung neutrophilia than C57BL/6 mice so you have to be careful to avoid too high doses - as increased lung neutrophils is a side effect which will confound your infection studies (so to me, 100 ul sounds too high and potentially quite damaging).
We are doing clodronate depletion through the intranasal route and report 80-100% depletion of BAL alveolar macrophages by day 3 post administration. The cells will take a few days to all undergo apoptosis. I would say to try a range of doses from 30 ul onwards, and check at day 2 and day 3 (day 3 is better because presumably, lungs have had one more day to recover from any effects of fluid administration to the lungs).
We have done it with 50 ul, and with 30x2 ul, the results were similar: we managed to eliminate only about 75-80% of macs.
The first publication on alveolar macrophage depletion in mice was from Theo Thepen in our lab:
Thepen, T., Van Rooijen, N., Kraal, G. 1989. Alveolar macrophage elimination in vivo is associated with an increase in pulmonary immune responses in mice. J. Exp. Med. 170, 499-509.
Since, a lot of publications did follow this protocol. You could e.g. perform a search in the publications/references list of www.Clodronateliposomes.org by searching for alveolar macrophage or lung.
Another administration route was intranasal administration. However in that case material may be spoiled via the oesophagos. (see de Haan et al Immunology).
Please see also FAQ (frequently asked questions) in the above website.
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