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Scratch test -- wound healing disrupted by smoke but not by Vype e-cigarette vapor

June 25, 2017

A new laboratory study reveals that cigarette smoke completely prevented wound healing at concentrations over 20% in a wound healing assay, whereas e-cigarette vapour had no effect, even at 100% concentration and double the amount of nicotine relative to smoke (see Figure 1).

The scratch test involves growing a layer of endothelial cells in the lab -- these are cells that line the inside of blood vessels -- creating a wound/scratch and observing how long it takes to heal. The wound heals normally when exposed cells are untreated or when they are exposed to e-cigarette vapour, but not when exposed to cigarette smoke. The results are published today in Toxicology Letters (DOI is 10.1016/j.toxlet.2017.06.001).

'Our results suggest that chemicals in cigarette smoke that inhibit wound healing are either absent from e-cigarette vapour or present in concentrations too low for us to detect an effect,' says Dr James Murphy, Head of Reduced Risk Substantiation at British American Tobacco.

It is thought that the presence of damaged endothelial cells, which have an impaired ability to repair, may be a factor in the development of heart disease. Smoking is known to be a risk factor for the development of heart disease.

The basic steps of the test involve creating a wound in a single layer of cells grown in the lab, capturing images of the beginning and at regular intervals during the 'healing' process, as the cells move together, and then comparing the images (see Figure 2).

In this way, it is possible to measure the ability of a tissue to repair an artificial injury in the presence of various substances. To repair the wound created by a scratch, cells must move into the wound and close the gap, and it's the rate at which they do it that the test measures.

Scientists at British American Tobacco used the scratch test to compare the effects of smoke extract from a reference cigarette (3R4F) and vapour extract from two commercial e-cigarettes, Vype ePen (a closed modular device) and Vype eStick (a cig-a-like device) on the wound healing process.

When a person smokes or vapes, water-soluble chemicals pass into circulation and interact with endothelial cells lining blood vessels. So toextracts -- the water-soluble fraction -- of smoke or vapour. Aqueous extracts were obtained by bubbling puff-matched amounts of smoke or vapour through cell-growth medium to produce a stock that could be diluted into various concentrations. Smoke extract was then assessed at concentrations from 0% to 30%. To ensure that e-cigarette extracts were tested at equivalent and higher nicotine concentrations than smoke (as possibly experienced by a heavy vaper), vapour was tested at concentrations between 40% and 100% (over twice the nicotine).

Immediately after the wound was made, the cells were immersed in smoke or vapour extract for 20 hours. Smoke decreased cell migration rate in a concentration-dependent manner, completely inhibiting movement of cells towards the wound at concentrations over 20%. In stark contrast, vapour from both types of e-cigarette had no effect -- cells could migrate into the wounded area, as normal, even at 100% concentration and double the amount of nicotine.

Previous research has shown that Vype e-cigarette vapour has 92-99% fewer toxicants than cigarette smoke. Many in the public health community believe e-cigarettes offer great potential for reducing the projected public health impact of smoking. Public Health England, an executive body of the UK Department of Health, published a report saying that the current expert estimate is that using e-cigarettes is around 95% safer than smoking cigarettes.

R&D at British American Tobacco

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