The cellular model is subjected to cellular vitality tests carried out with the aid of “AlamarBlue®”, a reagent based on a very simple and reliable process. A small blue-colored molecule (Resazurina) penetrates the cell membrane and comes into contact with the internal environment. The process that normally take place inside a healthy cell is able to convert Resazurina into a different pink-colored molecule (Resoufina).
The more healthy is the cell, the more effective is the process; on the contrary, the process will be less effective if the cell is living a state of suffering.
Once the time for a complete reaction of the cell expires, the amount of Resorufin (the fluorescent form) is measured by reading the fluorescence using a spectrophotometer, thus it will be possible to measure the cellular vitality level of the model and the consequent degree of toxicity of the initial liquid.
The organs (for example the lungs) are composed of tissues, that are composed of cells.
Cells form a tissue through the creation of stable connections between them, just as bricks are welded together in a supporting wall. These connections are therefore essential to guarantee the structure of the tissue, that is necessary to guarantee the functionality of the whole organ.
The reduction of the adhesion between cells leads to a flaking of the tissue, and a consequent reduction of the functionality of the organ (in this specific case the organism manifests a respiratory pathology).
The degree of adhesion between cells, related to the functionality of the tissue, can be measured as the ability of cells to prevent the passage of current. To measure it, two electrodes are applied to the cellular structure: the greater the current passage detected, the greater the disintegration of the tissue and therefore the greater the toxic effect of the substances in contact with the cells.
In the presence of a stress factor (cigarette smoke, alcohol, viruses, pathogens, parasites, etc.) the cells of the human body react by releasing molecules that mediate the inflammatory process. Among the most important are those belonging to the “IL” interleukin family, in particular the IL-6 and IL-8.
Inflammation is a very complex process, controlled by numerous factors, which can involve entire organs. The purpose of the inflammatory process is to draw the attention of the immune system and to activate the most competent cells to solve the problem instead of the stress factor. This process normally takes some time and results in the complete healing (as an example, think about the regeneration of a wound).
If the stress factor persists for a long time, in addition to the direct effects due to a chronic exposure to this factor, also additional side effects can arise due to the continuous inflammatory response that the body puts in place to defend itself. Actually, an overstimulated immune system can start to hit the cells of the same organism, with negative consequences on health.
Numerous studies have already amply demonstrated that the traditional cigarette has a high pro-inflammatory power due to various substances present in the smoke and to the frequent exposure due to the smoker’s activity.
To assess if the e-liquid vapors can cause the same effects of smoking on lung tissue, we measure the quantities of IL-8 by tests based on the enzyme immunoassay (ELISA) both in the apical compartment (where pulmonary epithelial cells develop ) and in the basal compartment (where vascular endothelium cells develop).
The current Tobacco Product Directive (TPD), which includes alternative products such as electronic cigarettes and their liquids, requires manufacturer to conduct their own assessment on the presence of some toxic substances in the emissions. Each Member State is responsible for establishing the parameters to be checked and the criteria to determine the danger. Currently, France and the United Kingdom provided national guidelines or technical standards, that envisage the determination of the presence of x potentially toxic chemicals. In particular, France (through the AFNOR standards) requires the analysis of nicotine, diacetyl, aldehydes and heavy metals; while the United Kingdom (through the MHRA agency) requires the analysis of: nicotine, acetaldehyde, acrolein, formaldehyde and based on the liquid / device combination, diethylene glycol, ethylene glycol, diacetyl, 2,3 pentanedione, and the two nitrosamines (NNN and NNK), for a total of 10 parameters.
To protect the health of its consumers at most, FlavourArt tested 23 parameters in each product put on the European market. In the ClearStream Onward program, 22 of the 23 parameters tested are displayed: nicotine is not shown, as it’s present by definition thus it does not represent an added risk factor.
