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Fabrication and Operation of an Oxygen Insert for Adherent Cellular Cultures

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Ozone mediators effect on "in vitro" scratch wound closure.

Giuseppe Valacchi¹, Claudia Sticozzi¹, Iacopo Zanardi²

¹a Department of Life Sciences and Biotechnology , University of Ferrara , Ferrara , Italy ;

Free Radical Research

|August 4, 2016

Summary

Ozonated saline enhances wound healing by stimulating keratinocyte proliferation. The combined action of 4-hydroxynonenal (HNE) and hydrogen peroxide (H2O2) mimics this effect, activating protective cellular pathways.

Keywords:

Cutaneous tissues H2O2 HNE oxidative stress wound healing

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Area of Science:

Biomedical Science
Wound Healing Research
Oxidative Stress Mechanisms

Background:

Low-dose ozone therapy shows benefits in wound healing, primarily due to bactericidal and pro-oxidant effects.
Ozone's effects are mediated by oxidative molecules like 4-hydroxynonenal (HNE) and hydrogen peroxide (H2O2), formed from reactions with biological systems.
The precise cellular mechanisms behind ozone's positive impact on wound closure remain largely uninvestigated.

Purpose of the Study:

To investigate the effects of varying non-toxic doses of ozonated saline on an in vitro wound scratch model using human keratinocytes.
To identify the key molecular mediators responsible for the wound healing benefits of ozonated saline.

Main Methods:

Utilized an in vitro wound scratch model with human keratinocytes.
Administered ozonated saline at concentrations from 2 to 300 μM.
Assessed cell proliferation using BrdU assay and PCNA protein levels.
Investigated the roles of HNE and H2O2, individually and in combination, to replicate ozonated saline effects.
Analyzed Nrf2 pathway activation via nuclear translocation and HO1 gene expression.

Main Results:

Ozonated saline significantly improved in vitro wound healing by enhancing keratinocyte proliferation.
Neither HNE nor H2O2 alone significantly improved wound closure, but their combination demonstrated a positive effect.
Nrf2 pathway activation was observed, evidenced by nuclear translocation of Nrf2 and increased HO1 gene expression.

Conclusions:

Ozonated saline effectively promotes wound closure in vitro by stimulating keratinocyte proliferation.
The beneficial effects of ozonated saline on wound healing are attributed to a combination of molecules, with HNE and H2O2 playing crucial roles.
Activation of the Nrf2 pathway is involved in the cellular response to ozonated saline treatment.