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. 2017 Mar 6:11:669-676.
doi: 10.2147/DDDT.S125843. eCollection 2017.

Protective influence of hyaluronic acid on focal adhesion kinase activity in human skin fibroblasts exposed to ethanol

Magdalena Donejko  1 , Edyta Rysiak  2 , Elżbieta Galicka  1 , Robert Terlikowski  3 , Edyta Katarzyna Głażewska  1 , Andrzej Przylipiak  1
Affiliations

Protective influence of hyaluronic acid on focal adhesion kinase activity in human skin fibroblasts exposed to ethanol

Magdalena Donejko et al. Drug Des Devel Ther. .

Abstract

Aim: The aim of this study was to evaluate the effect of ethanol and hyaluronic acid (HA) on cell survival and apoptosis in cultured human skin fibroblasts. Regarding the mechanism of ethanol action on human skin fibroblasts, we investigated cell viability and apoptosis, expression of focal adhesion kinase (FAK), and the influence of HA on those processes.

Materials and methods: Studies were conducted in confluent human skin fibroblast cultures that were treated with 25 mM, 50 mM, and 100 mM ethanol or with ethanol and 500 µg/mL HA. Cell viability was examined using methyl thiazolyl tetrazolium (MTT) assay and NC-300 Nucleo-Counter. Imaging of the cells using a fluorescence microscope Pathway 855 was performed to measure FAK expression.

Results: Depending on the dosage, ethanol decreased cell viability and activated the process of apoptosis in human skin fibroblasts. HA prevented the negative influence of ethanol on cell viability and prevented apoptosis. The analysis of fluorescence imaging using BD Pathway 855 High-Content Bioimager showed the inhibition of FAK migration to the cell nucleus, depending on the increasing concentration of ethanol.

Conclusion: This study proves that downregulation of signaling pathway of FAK is involved in ethanol-induced apoptosis in human skin fibroblasts. The work also indicates a protective influence of HA on FAK activity in human skin fibroblasts exposed to ethanol.

Keywords: apoptosis; ethanol; focal adhesion kinase; hyaluronic acid; skin fibroblast.

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Conflict of interest statement

Disclosure The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
Viability of the cells in MTT assay; cultured human skin fibroblasts were exposed for 24 hours to 25 mM, 50 mM, or 100 mM ethanol or to 25 mM, 50 mM, or 100 mM ethanol +500 µg/mL HA. *P<0.05. Abbreviations: HA, hyaluronic acid; MTT, methyl thiazolyl tetrazolium.
Figure 2
Figure 2
Fluorescence labeling of FAK in human skin fibroblasts exposed for 24 hours to 25 mM, 50 mM, or 100 mM ethanol. Notes: Pictures of fluorescing cells from microscope Nicon Eclipse Ti/C Plus. (A) Cellular nuclei stained with Hoechst dye (green). (B) Immunodetection of FAK using antibodies to FAK labeled with FITC (red). (C) Overlapped pictures with labeled cellular nuclei and with labeled FAK. Scale bar =20 µm. Abbreviations: FAK, focal adhesion kinase; FITC, fluorescein isocyanate.
Figure 3
Figure 3
Fluorescence labeling of FAK in human skin fibroblasts exposed to 25 mM, 50 mM, or 100 mM ethanol +500 µg/mL HA. Notes: Pictures of fluorescing cells from microscope Nicon Eclipse Ti/C Plus. (A) Cellular nuclei stained with Hoechst dye (green). (B) Immunodetection of FAK using antibodies to FAK labeled with FITC (red). (C) Overlapped pictures with labeled cellular nuclei and with labeled FAK. Scale bar =20 µm. Abbreviations: FAK, focal adhesion kinase; FITC, fluorescein isocyanate; HA, hyaluronic acid.
Figure 4
Figure 4
Percent of apoptotic cells in contact-inhibited human skin fibroblast cultures exposed for 24 hours to 25 mM, 50 mM, or 100 mM ethanol or to 25 mM, 50 mM, or 100 mM ethanol +500 µg/mL HA. Note: Measurement was performed in Nucleo-Counter® NC-300™ apparatus. Abbreviation: HA, hyaluronic acid.
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References

    1. Ranzer MJ, Chen L, DiPietro LA. Fibroblast function and wound breaking strength is impaired by acute ethanol intoxication. Alcohol Clin Exp Res. 2011;35(1):83–90. - PMC - PubMed
    1. Seth D, D’Souza El-Guindy NB, Apte M, et al. Alcohol, signaling, and ECM turnover. Alcohol Clin Exp Res. 2010;34(1):4–18. - PubMed
    1. Neuman MG, Haber JA, Malkiewicz IM, Cameron RG, Katz GG, Shear NH. Ethanol signals for apoptosis in cultured skin cells. Alcohol. 2002;26(3):179–190. - PubMed
    1. Neuman MG, Nanau RM, Oruña L, Coto G. In vitro anti-inflammatory effects of hyaluronic acid in ethanol-induced damage in skin cells. J Pharm Pharm Sci. 2011;14(3):425–437. - PubMed
    1. Mello T, Ceni E, Surrenti C, Galli A. Alcohol induced hepatic fibrosis: role of acetaldehyde. Mol Aspects Med. 2008;29:17–21. - PubMed

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