Effects of Hypericin on Cultured Primary Normal Human Dermal Fibroblasts Under Increased Oxidative Stress

Ioanna A Anastasiou; Panagiotis Sarantis; Ioanna Eleftheriadou; Konstantinos N Tentolouris; Iordanis Mourouzis; Michalis V Karamouzis; Konstantinos Pantos; Nikolaos Tentolouris

doi:10.1177/15347346231212332

Effects of Hypericin on Cultured Primary Normal Human Dermal Fibroblasts Under Increased Oxidative Stress

Int J Low Extrem Wounds. 2023 Nov 13:15347346231212332. doi: 10.1177/15347346231212332. Online ahead of print.

Authors

Ioanna A Anastasiou^{1

2}, Panagiotis Sarantis³, Ioanna Eleftheriadou¹, Konstantinos N Tentolouris^{1

2}, Iordanis Mourouzis², Michalis V Karamouzis³, Konstantinos Pantos², Nikolaos Tentolouris¹

Affiliations

¹ Diabetes Center, First Department of Propaedeutic Internal Medicine, Medical School, National and Kapodistrian University of Athens, Laiko General Hospital, Athens, Greece.
² Department of Pharmacology, Medical School, National and Kapodistrian University of Athens, Athens, Greece.
³ Molecular Oncology Unit, Department of Biological Chemistry, Medical School, National and Kapodistrian University of Athens, Athens, Greece.

PMID: 37956650
DOI: 10.1177/15347346231212332

Abstract

Introduction: Wound healing is a dynamic process that begins with inflammation, proliferation, and cell migration of a variety of fibroblast cells. As a result, identifying possible compounds that may improve fibroblast cell wound healing capacity is crucial. Hypericin is a natural quinine that has been reported to possess a wide range of pharmacological profiles, including antioxidant and anti-inflammatory, activities. Herein we examined for the first time the effect of hypericin on normal human dermal fibroblasts (NHDFs) under oxidative stress.

Methods: NHDF were exposed to different concentrations of hypericin (0-20 μg/mL) for 24 h. For the oxidative stress evaluation, H₂O₂ was used as a stressor factor. Cell viability and proliferation levels were evaluated. Immunohistochemistry and flow cytometry were performed to assess cell apoptosis levels and with confocal microscopy we identified the mitochondrial superoxide production under oxidative stress and after the treatment with hypericin. Scratch assay was performed under oxidative stress to evaluate the efficacy of hypericin in wound closure. To gain an insight into the molecular mechanisms of hypericin bioactivity, we analyzed the relative expression levels of genes involved in oxidative response and in wound healing process.

Results: We found that the exposure of NHDF to hypericin under oxidative stress resulted in an increase in cell viability and ATP levels. We found a decrease in apoptosis and mitochondrial superoxide levels after treatment with hypericin. Moreover, treatment with hypericin reduced wound area and promoted wound closure. The levels of selected genes showed that hypericin upregulated the levels of antioxidants genes. Moreover, treatment with hypericin in wound under oxidative stress downregulated the levels of proinflammatory cytokines, and metalloproteinases; and upregulated transcription factors and extracellular matrix genes.

Conclusion: These findings indicated that hypericin possesses significant in vitro antioxidant activity on NHDF and provide new insights into its potential beneficial role in the management of diabetic ulcers.

Keywords: antioxidant; in vitro; oxidative stress; primary human dermal fibroblasts; wound healing.