Radical idea: Resveratrol shows intracellular antioxidant properties in normal human fibroblasts

Keypoints:

"In general, we're looking at molecules that have a great safety record and which appear to be the kinds of molecules we can easily incorporate into clinical use.

Fruit of the vine

Dr. Brody says resveratrol shows promise for several reasons. A phytoalexin, resveratrol is a natural defense of higher plants against infections and nutrient deprivation. It occurs most commonly in the skin of grapes and in red wines that use grape skins in the fermentation process, Dr. Brody tells Dermatology Times.

Resveratrol first captured researchers' attention as a possible explanation for what's known as "the French Paradox."

"Many French citizens are engaged in activities that most of us know to be harmful, such as cigarette smoking and alcohol intake. However, the French are known to have longer lives than we would expect," says Jared Jagdeo, M.D., assistant clinical instructor of dermatology, SUNY Downstate.

Researchers, therefore, hypothesize that resveratrol found in red wine might provide a protective effect - as has been shown in many cardiovascular studies - that can explain their longer lifespans, he says.

Topical absorption

Furthermore, Dr. Jagdeo says resveratrol shows promise as both a topical preventive agent and a chemopreventive compound in studies involving cutaneous neoplasms.

In addition to these properties, resveratrol has demonstrated the most efficient topical absorption among red wine polyphenols in a murine skin cancer model (Soleas GJ et al. Clin Biochem. 2002 Mar; 35(2):119-124).

It also has shown chemopreventive and chemotherapeutic properties in a murine skin cancer model, Dr. Brody says.

Research

To help characterize resveratrol's intracellular antioxidant properties in human skin cells, researchers cultured normal human skin fibroblast AG13145 primary cells in DMEM with 10 percent fetal bovine serum (both made by Gibco) in a humidified incubator at 37 degrees Celsius with an atmosphere containing 5 percent CO₂.

Next, researchers seeded the cells in tissue culture dishes before pretreating them for four hours with resveratrol concentrations of 0.01 percent, 0.001 percent and 0.0001 percent (Topix Pharmaceuticals). The investigators also left one group of cultured cells untreated for control purposes.

Investigators then washed the cultured cells with a phosphate buffer solution (PBS) and exposed them to 1.2 mM (millimolar) hydrogen peroxide (H₂O₂, Sigma) in DMEM for 30 minutes to generate intracellular ROS. Next, researchers again washed the cells with PBS and processed them for flow cytometry analysis.

In this analysis, Dr. Jagdeo says, "As we increased the resveratrol concentrations, we were able to decrease the amount of free radical damage caused by hydrogen peroxide to normal human skin cells."

Intravital staining results supported these findings, with nearly half of the H₂O₂-upregulated fibroblasts staining positive for dihydrorhodamine-identified free radicals, versus a negligible proportion of those treated with 0.01 percent resveratrol and H₂O₂.

"In this study," Dr. Brody says, "resveratrol did exactly what we had hoped it would do. It seems to protect cells against ROS challenges. It also prevents them from accumulating other kinds of cell damage that we associate with either earlier cell senescence or cancer."

Clinical testing to come

"A model is exactly that - it's something that we assume looks like the real-world entity we're trying to mimic, but we don't know that it does" until after clinical trials, Dr. Brody says.

At press time, Drs. Jagdeo, Brody and their colleagues were conducting additional in vitro studies with resveratrol and other antioxidants.

"We're adding to the creativity of our model system, trying to separate out what we consider to be markers of youth and markers of age to see if, in fact, these molecules act differently in 'young' cells versus 'old' cells," Dr. Brody says.

The research team is "very close" to beginning human clinical trials, he adds.

In addition to the necessary laboratory facilities, Dr. Brody says SUNY Downstate's association with Topix Pharmaceuticals provides the ability to translate lessons from the laboratory into products.

Disclosure: Topix Pharmaceuticals provided the resveratrol used in this study.