Antimicrobial properties of olive oil phenolic compounds and their regenerative capacity towards fibroblast cells

https://doi.org/10.1016/j.jtv.2021.03.003Get rights and content

Highlights

  • Stimulated the proliferative capacity of fibroblasts, increasing migration and expression of the aforementioned genes.

  • Treatment with luteolin, apigenin, ferulic, coumaric acid or caffeic acid compounds inhibited the growth of different microorgananisms.

  • The phenolic compounds have a biostimulatory effect on regeneration, differentiation, and migration of fibroblasts and exert antibacterial activity.

  • Phenolic compounds may have a strong therapeutic effect on wound recovery.

Abstract

Some micronutrients of vegetable origin are considered potentially useful as wound-healing agents because they can increase fibroblast proliferation and differentiation.

The aim of this study

was to evaluate the regenerative effects of selected olive oil phenolic compounds on cultured human fibroblasts and explore their antimicrobial properties.

Material and methods

The CCD-1064Sk fibroblast line was treated for 24 h with 10−6M luteolin, apigenin, ferulic, coumaric acid or caffeic acid, evaluating the effects on cell proliferation by using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) spectrophotometric assay; the migratory capacity by the scratch assay and determining the expression of Fibroblast Growth Factor (FGF), Vascular Endothelial Growth Factor (VEGF), Transforming Growth Factor- β1 (TGFβ1), Platelet Derived Growth Factor (PDGF), and Collagen Type I (COL-I) genes by real-time polymerase chain reaction. The antimicrobial capacity of the polyphenols was evaluated by the disc diffusion method.

Results

All compounds except for ferulic acid significantly stimulated the proliferative capacity of fibroblasts, increasing their migration and their expression of the aforementioned genes. With respect to their antimicrobial properties, treatment with the studied compounds inhibited the growth of Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, Proteus spp., and Candida Albicans.

Conclusions

The phenolic compounds in olive oil have a biostimulatory effect on the regeneration capacity, differentiation, and migration of fibroblasts and exert major antibacterial activity. According to the present findings, these compounds may have a strong therapeutic effect on wound recovery.

Introduction

Skin is the largest organ of the body (surface area of 1.5–2 m) and comprises three main layers: the epidermis, dermis, and hypodermis [1]. Damage to the skin activates mechanisms to close the wound and recover its functional status. Wound healing is a complex process characterized by inflammatory, proliferative, and remodeling phases [2]. Hemostasia at the start of the inflammatory phase involves fibrin clot formation and coagulation, for which platelets are largely responsible [3]. Platelets also release growth factors, including transforming growth factor-β (TGF-β), platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), and epidermal growth factor (EGF), which are essential for the regeneration of damaged skin [2,4]. Leukocytes (neutrophils, monocytes, and macrophages) then migrate to the tissue, attracted by histamines, growth factors, and proinflammatory cytokines, among other molecules [5,6]. These leukocytes release chemotactic factors that promote the arrival of various cell populations to the damaged area [7,8]. These include dermal fibroblasts, which are derived from dermal progenitors [9] and are largely responsible for producing the elements that compose the extracellular matrix (e.g., collagen and fibronectin), contributing to the formation of granulation tissue [10]. These fibroblasts can sometimes differentiate towards myofibroblasts, whose main functions are related to wound contraction [11]. The proliferative phase is completed by angiogenesis and re-epithelialization processes [12,13]. During the remodeling phase, the matrix stabilizes and its constitutive elements are reorganized, with the degradation of fibroblast-secreted type III collagen and the increased production of type I collagen [14].

Wound healing can be affected by local factors such as oxygenation [15,16] or infections [17,18] and by systemic factors such as age [19,20], stress [21,22], diabetes [23,24], obesity [25,26], drug consumption [[27], [28], [29]], or nutrition [[30], [31], [32]]. Thus, various micronutrients have been found to influence wound healing, including vitamins A [33], C [34], and E [35], whose antioxidant capacity has been associated with greater fibroblast proliferation and differentiation and increased collagen and hyaluronic acid production, and whose deficiency has been associated with reduced angiogenic activity and greater capillary fragility [36,37]. The phenolic compounds of olive oil have also demonstrated antioxidant capacity through their action as chain breakers, donating hydrogen radicals to alkylperoxyl radicals [38,39] produced by lipid oxygenation and giving rise to the formation of stable derivatives during the reaction. These properties have attracted the attention of the food and drug industry to the phenolic compounds in olive oil as possible nutraceuticals, with the potential to protect against chronic, degenerative, and oxidative stress-related diseases [[40], [41], [42]].

However, despite the myriad of potential health benefits of olive oil, there is a gap of data published on the possible effects of olive oil phenolic fraction on fibroblasts, especially over the last two year. Despite of in vitro studies limitations our results provide new knowledge about molecular mechanism of action of olive oil phenolic compounds on fibroblasts, which suggest that these compounds could be useful to enhance wound healing. Thus, the objective of this study was to evaluate the regenerative effects of selected olive oil phenolic compounds on cultured human fibroblasts and explore their antimicrobial properties.

Section snippets

Chemical products and reagents

Standards of apigenin, luteolin, caffeic acid, p-coumaric acid and ferulic acid were purchased from Sigma-Aldrich (St. Louis, MO). Stock solutions of phenolic compounds were prepared in ethanol and stored at −20 °C. All solvents were of analytical or HPLC grade (Sigma-Aldrich) and water was of Milli-Q quality (Millipore Corp, Bedford, MA, USA).

Cell culture

The CCD-1064Sk typified human fibroblast cell line was purchased from American Type Cultures Collection (ATCC, Manassas, VA). It was kept in Dulbecco's

Effect of olive oil phenolic compounds on fibroblast proliferation

Fig. 1 depicts the proliferative effect on CCD-1064SK human epithelial fibroblast cell line at 24 h after treatment with the studied phenolic compounds, which significantly increased the proliferative capacity of fibroblasts (p < 0.0001) versus controls by 38–81.2%, depending on the compound in question. Coumaric acid achieved the highest increase (81.2% vs. controls), followed by caffeic acid (77.1%), ferulic acid (65.0%), apigenin (55.8%), and luteolin (38.0%).

Effect of phenolic compounds on the gene expression of fibroblasts

Fig. 2 depicts the results of

Discussion

Wound regeneration can be enhanced by cell biostimulation or bacterial load control, among other approaches [47]. The results of this study demonstrate the regenerative potential of extra virgin olive oil phenolic compounds and their promotion of wound healing through the biostimulation of fibroblasts, augmenting their proliferative capacity and migration, increasing their expression of VEGF, PDGF, IGF, TGFβ1, and Col-I, and exerting antimicrobial activity to reduce the bacterial load.

Funding sources

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

Declaration of competing interest

We wish to draw the attention of the Editor to the following facts which may be considered as potential conflicts of interest and to significant financial contributions to this work.

Authors confirm that there are no known conflicts of interest associated with this publication and there has been no significant financial support for this work that could have influenced its outcome.

Acknowledgments

This study was supported by the research group BIO277 (Junta de Andalucía) and Department of Nursing (University of Granada).

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