E-Fluid Induces Inflammation Responses in Lung Epithelial Cells

By Hayley E. Tartell
Discussions
2015, Vol. 12 No. 1 | pg. 2/2 |

Discussion

Given the data, activation of JNK and NF-κB suggests that signaling associated with inflammation occurs when the cells are exposed to E-cigarette vapor. ERK had a relatively short period of activation, implying less intense signaling. It can be deduced from the data that JNK's prolonged activation implies more intense signaling. The data indicates that p38 has a prolonged level of activation and, accordingly, more intense signaling. Therefore, as indicated by the results, nicotine enhances the activation of the aforementioned MAP kinases.

Figure 7. Western blots of targets.

Figure 7. Western blots of targets.

Cigarette smoke activates MAP kinases (Mercer & D'Armiento , 2006, p. 137-150). This study was proposed to detennine whether nicotine alone could activate these MAP kinases as well. The data show that E-liquid containing nicotine activates ERK, JNK, and p38 MAP kinases. The next step would be to repeat the experiment to substantiate the results as well as to determine whether the protein's level of activation is greater when treated with E-liquid containing nicotine, E-liquid without nicotine, or nicotine alone. Greater activation may imply greater inflammation and protease activity. Additionally, studies in animals would further enhance the results established in this study and suggest the potential human implications of using E-cigarettes.

Figure 8. Semi-quantitative analysis of Western blots using the ratio of total protein to actin. PBS is used as a control. Silencing PP2A produces (a) less of the protein MMP-9, (b) a 25% increase in protein levels from PBS to E-liquid with nicotine (statistically significant; p < 0.05), (c) an increase in TLR3 protein levels. and (d) an increase in p-Src 416.

Figure 8. Semi-quantitative analysis of Western blots using the ratio of total protein to actin. PBS is used as a control. Silencing PP2A produces (a) less of the protein MMP-9, (b) a 25% increase in protein levels from PBS to E-liquid with nicotine (statistically significant; p < 0.05), (c) an increase in TLR3 protein levels. and (d) an increase in p-Src 416.

Figure 9. The amount of DNA binding detected during the transcription factor assay was proportional to the activity of NF-κB and AP-1. In graph (a), a significant difference (p < 0.05) suggests that NF-κB was activated.

Figure 9. The amount of DNA binding detected during the transcription factor assay was proportional to the activity of NF-κB and AP-1. In graph (a), a significant difference (p < 0.05) suggests that NF-κB was activated.

MMP-9 is a protease (Foronjy et al., 2008, p. 1149-57). As mentioned before, proteases play a key role in the development of COPD ("Protease", 2013). MMP-9 levels were unaltered by E-liquid in this study. However, the extracellular levels and activity of this important protease were not determined. Despite not identifying a change in intracellular MMP-9, this mechanism cannot be fully eliminated because MMP-9 gene expression and protease activity must be fully investigated to draw a conclusion.

The data also indicate that when cells are treated with concentrated E-liquid, NF-κB is activated. When cells are treated with cigarette smoke, MMP-9 gene expression and levels increase and ultimately contribute to the development of emphysema (Thorley & Tetley, 2007, p. 409428).

Dr. Foronjy (unpublished data) recently found that when primary lung epithelial cells are treated with E-liquid, cytokine and protease gene expression (e.g. MMP-7, MMP-9, Cathepsin Lt, TLR3, and c-Src) rises. This study hypothesizes that when cells are treated with 1% E-liquid, MMP-9 levels increase. The cells utilized in this study are not a primary cell line and expression of MMP-9 may not be the same as primary cells. Nevertheless, the results of either study must be verified through repeated experimentation and clinical samples.

When cells are exposed to cigarette smoke extract, ERK, JNK, or p38 is silenced and MMP9 levels drop (Mehra et al., 2012). In future experiments ERK, p38, and JNK could be knocked down using siRNAs, and MMP-9, EGRt, TLR3, and p-Src 416 levels could be analyzed. These experiments could determine whether the aforementioned MAP kinases are integral to protein synthesis. Cigarette smoke induces cells to produce more proteases and inflammatory signaling (Mercer & D'Armiento, 2006, p. 137-150). Future studies ought to attempt to show whether E-liquid induces cells to produce more proteases and inflammatory signaling such as Cathepsin Lt, MMP-7, TLR3, and c-Src. This study determined that the use of nicotine delivery via E-cigarettes may impact lung inflammation and consequently lead to disease initiation and progression.


Publisher's Note

A substantially similar version of this article previously appeared in Student Pulse (2015) Vol. 07 No. 05. That article is available here: Are E-Cigarettes a Safe Alternative to Smoking?


Acknowledgments

First and foremost, I would like to thank Dr. Foronjy for his time and enthusiasm in supervising this project. Moreover, I thank Dr. Geraghty and Dr. Turino for their guidance and encouragement. Finally, I want to thank my family for their unconditional love and support.


References

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Foronjy R., Nkyimbeng T., Wallace A., Thankachen J., Okada, Y., Lemaitre V., D'Armiento J. (2008). Transgenic expression of matrix metalloproteinase-9 causes adult-onset emphysema in mice associated with the loss of alveolar elastin. Am J Physiol Lung Cell Mol Physio. 294(6),L1149-57. doi: 10.1152/ajplung.00481.2007. Epub 2008 Apr 11.

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