Skin aging is a complex biological process significantly driven by Reactive Oxygen Species (ROS) and mitochondrial dysfunction. The skin, as the body’s largest organ, constantly renews itself and requires substantial adenosine triphosphate (ATP) produced by mitochondria. During aging, ROS are produced as natural by-products of oxidative phosphorylation. These ROS activate signaling pathways, leading to the upregulation of matrix metalloproteinases (MMPs), which degrade essential extracellular matrix (ECM) proteins like collagen and elastin. This degradation, combined with ROS inhibiting collagen synthesis, leads to the appearance of dry, thin, and finely wrinkled skin. Mitochondrial DNA damage by ROS further impairs mitochondrial efficiency, creating a vicious cycle of dysfunction and damage.
To counteract this critical pathway of skin aging, the study explored the potential of cytoglobin (CYGB) and extracts from Tropaeolum majus (nasturtium). CYGB is a globin protein reported to be involved in oxygen homeostasis and redox monitoring. It reversibly binds O2 and has peroxidatic activity, consuming hydrogen peroxide and lipid peroxides, thereby protecting cells from oxidative stress. Furthermore, T. majus has historically been used in traditional medicine for its anti-inflammatory properties, and a compound identified in the plant exhibits bioactivity against ROS. Given the suggested actions of CYGB against ROS, the objective was to evaluate whether a T. majus extract that upregulates CYGB could protect against various markers of skin aging, including mitochondrial function, ROS production, and ECM composition. Researchers also hypothesized that the extract could restore natural killer (NK) cell cytotoxicity, potentially offering a senolytic effect.
Methods
Primary human dermal fibroblasts and keratinocytes were cultured and treated with 0.5% v/v T. majus extract (prepared by Silab). CYGB synthesis was detected using immunohistochemical analysis. Cellular and mitochondrial ROS levels were quantified using flow cytometry and confocal microscopy, and mitochondrial activity was assessed via a Seahorse extracellular flux analyzer. Extracellular matrix components (collagen and elastin) were evaluated using immunolabeling, and transcriptomic analysis was performed via bulk RNA-seq on young and elderly keratinocytes. Finally, the impact on immune function was determined using a CD107a degranulation assay to measure NK cell activity in peripheral blood mononuclear cells (PBMCs).
Key Findings
The study yielded several crucial findings regarding the anti-aging mechanisms of the T. majus extract:
• CYGB Expression Stimulation: Treatment of aged human fibroblasts with T. majus extract increased cytoglobin synthesis in a dose-dependent manner, with the 0.25% v/v concentration resulting in a 56% increase.
• ROS Reduction: Overexpression of CYGB in dermal fibroblasts reduced both cytosolic and mitochondrial ROS concentrations. Similarly, treatment with the extract reduced ROS in keratinocytes and decreased ROS levels by 57% in keratinocytes exposed to blue light.
• Mitochondrial Efficiency Improvement: The T. majus extract treatment significantly improved mitochondrial efficiency in normal human keratinocytes (NHKS), resulting in a 57% increase in basal respiration rate and a 40% increase in ATP production.
• ECM Restoration and Cell Proliferation: The extract stimulated the synthesis of key ECM components in fibroblasts, increasing collagen levels by 215% and elastin concentrations by 158%. Furthermore, it stimulated proliferation rates in keratinocytes (22% increase) and human dermal fibroblasts (42% increase).
• Transcriptional Regulation: RNA-seq analysis revealed that the extract downregulated genes involved in the terminal differentiation of keratinocytes, specifically those related to keratinization and the cornified envelope (e.g., LCE family members).
• NK Cell Activity Restoration: The extract stimulated the activity of Natural Killer (NK) cells, increasing activity by 84% when the cells were cultured under hypoxic conditions (which otherwise reduced activity by 80%).
This research demonstrated that treating skin cells with the T. majus extract exerts comprehensive anti-aging effects by modulating cellular physiology. The novelty of this research lies in strongly suggesting that the observed beneficial actions—reduction of ROS, improved mitochondrial efficiency, and replenished collagen and elastin levels—are primarily mediated by the extract’s ability to increase Cytoglobin (CYGB) synthesis. This positions CYGB as a protein with potentially potent anti-aging action in the skin by regulating energy production and ROS defense.
A significant future implication of this work is the potential for the extract to stimulate the clearance of senescent cells. The finding that the extract restores NK cell cytotoxicity, which is necessary for eliminating senescent cells, strongly suggests a potential senolytic effect. Future studies should investigate this senolytic action by measuring NK cell activity when incubated specifically with senescent fibroblasts following treatment. Additionally, confirmation of the regulatory pathway could involve measuring MMP levels to confirm that the observed increase in collagen and elastin synthesis is mediated by a reduction in MMP activity.
Link to the study: https://www.mdpi.com/2079-9284/12/5/230
