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Won’t somebody, please, think of the mice?

07, Sep, 2016

By Tom Pruen

A recent study purports to have demonstrated that “Chronic electronic cigarette exposure in mice induces features of COPD in a nicotine-dependent manner”.

The test on mice that was conducted was:

“Mice were exposed to aerosolised phosphate-buffered saline, nicotine-free or nicotine-containing e-cigarette solution, 1-hour daily for 4 months.”

And the conclusion reached:

“Exposure to inhaled nicotine-containing e-cigarette fluids triggered effects normally associated with the development of COPD including cytokine expression, airway hyper-reactivity and lung tissue destruction.”

However, while 4 months seems like a reasonable duration for a chronic exposure study, the modelling of chronic exposure was actually extremely poor.

The mice were exposed for 1 hour per day, to 0.4ml of saline solution, nicotine free e-liquid or 18mg/ml e-liquid.

One hour per day is not a good replication of a chronic exposure. This is perhaps fortunate, since 0.4ml of 18mg/ml solution is also a terrible replication of a chronic exposure.

No reliable data exists on the concentration of nicotine that is dangerous to mice(such as the concentration that kills 50% of those exposed, referred to as an LC50), but the oral dose that kills 50% of those exposed (LD50)is 3.3mg/kg[i].  The type of mice used in the study have an average weight of about 22g, so the fatal oral dose for a single mouse is about 0.07mg.

0.4ml of 18mg/ml nicotine solution contains 7.2mg of nicotine.

Sadly, no information is provided on the volumes of the mouse exposure cages, or flow rates, or anything else that might allow a concentration to be calculated, but assuming (largely because the mice were not wholly euthanized by the exposure) that the 0.4ml was delivered consistently over the course of the hour, rather than all at once, this is still 0.12mg/min.

It is hard to imagine these being happy mice, although since there is no reporting of behavioural observations or (oddly) death rates, the extent of this can only be speculative.

It would seem, however that this study is a better model for repeated acute toxicity than chronic effects, since rather than a low level exposure over a large period of time; it used a short period of very high exposure and repeated it for a long period.

(Chronic exposure is considered to be “Continued exposure or exposures occurring over an extended period of time, or a significant fraction of the test species' or of the group of individuals', or of the population's life-time.”[ii] While acute exposure is “Exposure of short duration.”[iii]).

As if this were not enough of an issue, the nicotine aerosol was administered to the mice as a whole body exposure. In contrast to the claims made in the study about inhalation effects, nicotine (and flavoured carrier) would have been deposited in the mouse’s environment, and on its fur. No effort appears to have been made to estimate the exposure through this route, but this would also seem to have been a significant route of exposure, particularly through grooming.


Additionally, as well as the likely inappropriateness of the nicotine doses, an exposure of 0.4ml of liquid to a 22g mouse is equivalent to an exposure of 18ml/kg of body weight. This is equivalent to exposing a 70kg human to 1.2 litres of e-liquid. Given that an average vaper consumes less than 10ml of liquid per day, this is wildly excessive.  (it should also be noted that vapers do not fill rooms with vapour and sit in them). A dose representative of human exposure would have been  0.14ml/kg, or about 0.003ml/day for a 22g mouse.

Interestingly, despite exceeding the human exposure by two orders of magnitude, the non-nicotine containing liquid did not induce any effects that were significantly different from the saline control. This would seem to largely assuage concerns that the propylene glycol or glycerol that forms the majority of e-liquids has significant effects when inhaled.

It would also seem to rule out any local toxicity from the flavourings used, but since these were not reported, this still doesn’t provide us with any useful information.

If the researchers actually wanted to demonstrate a dose-effect response, then it would have been vital to have used more than one nicotine dose for example using exposures of 0.1, 0.2, 0.3 and 0.4ml/day, although these would still be levels that are poorly representative of human exposures. If nicotine was the cause of the changes in lung function and structure, this would have been demonstrated by an increase in effect with an increase in exposure. It’s also entirely possible that there would be a threshold below which no effect would have occurred (or been too small to measure). 

As it is, this study demonstrates that whole body exposure to nicotine doses which are likely to be near toxic in mice cause changes in lung tissue (they may also cause changes in other organs, but these do not appear to have been examined). Its design precludes determining if this is a dose or duration dependant effect, and the exposures used bear no resemblance or relevance to human exposures. It doesn’t report any effects on the mice during the test (did they show odd behaviours? Did they lose weight? How many died?)

If a significantly redesigned and improved replicate of this study were made, it might be able to provide us with all kinds of interesting data, but as performed, it tells us that poisoning mice is bad for the mice – and that’s about all.

In any scientific experiment, it is important to design the experiment, and the methods used, to maximise the information that can be gained. This is even more imperative when live subjects are involved, be they mice or men.

In this case I feel sorry for the mice whose tiny little lives were not only lost, but wasted.


[i] From Lazutka et al. 1969, and referenced widely, e.g. http://www.cdc.gov/niosh/idlh/54115.html

[ii] https://sis.nlm.nih.gov/enviro/iupacglossary/glossaryc.html#chronicexposure

[iii] https://sis.nlm.nih.gov/enviro/iupacglossary/glossarya.html#acuteexposure


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