Inhibition of fibroblast proliferation in vitro using low-level infrared light-emitting diodes.

BACKGROUND: Scars, including hypertrophic and keloidal-type scars, may occur after burns, trauma, or surgery. Despite several treatment options available for scars, few effective, noninvasive modalities exist. Recently, a few small clinical studies revealed the possible benefit of red and infrared (IR) low-level light therapy (LLLT) in scar treatment. One of the important features of scars is proliferation of dermal fibroblasts, but in vitro data regarding the effects of light-emitting diode (LED)-generated IR light on human skin fibroblasts is lacking. OBJECTIVE: To evaluate the effect of IR LLLT generated using LEDs on fibroblast proliferation and viability in vitro. METHODS AND MATERIALS: Irradiation of normal human skin fibroblasts using IR LED panels was performed in vitro, and modulation of proliferation and viability was quantified using Trypan blue dye exclusion assay. RESULTS: Fluences of 80, 160 and 320 J/cm(2) resulted in statistically significantly less fibroblast proliferation than in controls, without statistically significantly less cellular viability. CONCLUSION: IR LLLT can effectively inhibit fibroblast proliferation in vitro without altering viability and holds promise for the treatment of scars.

Inhibition of fibroblast proliferation in vitro using red light-emitting diodes.

BACKGROUND: Red light is part of the visible light spectrum. The effects of light-emitting diode (LED)-generated red light on human skin are not well-characterized. OBJECTIVE: To study the effect of red LED-generated low-level light therapy (LLLT) on fibroblast proliferation and viability in vitro. METHODS AND MATERIALS: Irradiation of normal human skin fibroblasts using red LED panels was performed in vitro, and modulation of proliferation and viability was quantified using trypan blue dye exclusion assay. RESULTS: Statistically significant decreases in cell proliferation were noted at the following fluences (time): 160 J/cm2 (30 minutes, 34 seconds), 320 J/cm2 (61 minutes, 07 seconds) and 640 J/cm2 (122 minutes, 14 seconds) (Figure 1). Irradiation at the 160- (98.5 ± 1.2%) and 320-J/cm2 (98.0 ± 3.1%) doses did not significantly alter viability. CONCLUSION: At certain fluences, red LLLT can effectively inhibit fibroblast proliferation in vitro without altering viability and holds promise for the treatment of scars and other proliferative skin diseases.

Reduction of facial redness with resveratrol added to topical product containing green tea polyphenols and caffeine.

BACKGROUND/OBJECTIVE: Many topical formulations include antioxidants to improve the antioxidant capability of the skin. This study evaluated the ability of a unique combination of antioxidants including resveratrol, green tea polyphenols, and caffeine to reduce facial redness. METHODS: Subjects (n=16) presenting with facial redness applied the resveratrol-enriched product twice daily to the entire face. Reduction in redness was evaluated by trained staff members and dermatology house staff officers. Evaluators compared clinical photographs and spectrally enhanced images taken before treatment and at 2-week intervals for up to 12 weeks. RESULTS: 16 of 16 clinical images showed improvement and 13 of 16 spectrally enhanced images were improved. Reduction in facial redness continued to evolve over the duration of the study period but was generally detectable by 6 weeks of treatment. Adverse effects were not observed in any subject. CONCLUSION: The skin product combination of resveratrol, green tea polyphenols, and caffeine safely reduces facial redness in most patients by 6 weeks of continuous treatment and may provide further improvement with additional treatment.

The active natural anti-oxidant properties of chamomile, milk thistle, and halophilic bacterial components in human skin in vitro.

The number of skin cancers continues to rise, accounting for approximately 40% of all cancers reported in the United States and approximately 9,500 deaths per year. Studies have shown reactive oxygen species (ROS) type free radicals are linked to skin cancer and aging. Therefore, it is important for us to identify agents that have anti-oxidant properties to protect skin against free radical damage. The purpose of this research is to investigate the anti-oxidant properties of bisabolol, silymarin, and ectoin that are components from chamomile, milk thistle, and halophilic bacteria, respectively. We measured the ability of bisabolol, silymarin, and ectoin to modulate the hydrogen peroxide (H2O2)-induced upregulation of ROS free radicals in normal human skin fibroblasts in vitro. Using a flow cytometry-based assay, we demonstrated that varying concentrations of these natural components were able to inhibit upregulation of H2O2-generated free radicals in human skin fibroblasts in vitro. Our results indicate components of chamomile, milk thistle, and halophilic bacteria exhibit anti-oxidant capabilities and warrant further study in clinical trials to characterize their anti-cancer and anti-aging capabilities.

Caffeine protects human skin fibroblasts from acute reactive oxygen species-induced necrosis.

Oxidative damage by reactive oxygen species (ROS) plays a major role in aging and carcinogenesis. Little is known about either the effects of acute ROS in necrosis and inflammation of skin or the therapeutic agents for prevention and treatment. Previously, our laboratory identified caffeine as an inhibitor of hydrogen peroxide (H2O2)-generated lipid peroxidation products in human skin fibroblasts. Here, we study effects of caffeine on acute ROS-mediated necrosis. Human skin fibroblasts were incubated with caffeine, followed by H2O2 challenge. Flow cytometry was used to analyze cell morphology, counts, apoptosis and necrosis, and ROS. We found that caffeine protects from H2O2 cell damage at lower (0.01 mM) and intermediate (0.1 mM) doses. The beneficial effects of caffeine appear to be mediated by a mechanism other than antioxidant function.

Complementary antioxidant function of caffeine and green tea polyphenols in normal human skin fibroblasts.

The study of free radicals is particularly relevant in the context of human skin carcinogenesis and photoaging because of these oxidants' ability to induce DNA mutations and produce lipid peroxidation byproducts, including 4-hydroxy-2-nonenal (HNE). Therefore, it is important to identify and evaluate agents with the ability to modulate intracellular free radicals and HNE. The purpose of this research is to investigate the ability of antioxidants green tea polyphenols (GTPs) and caffeine, alone and in combination, to modulate the hydrogen peroxide (H2O2)-induced upregulation of reactive oxygen species (ROS) free radicals and HNE in normal human skin fibroblast WS-1 cells in vitro. GTPs and caffeine were selected for evaluation because these compounds have demonstrated antioxidative properties in various skin models. Furthermore, GTPs and caffeine share a close natural botanical association as caffeine is present in green tea, as well. Hydrogen peroxide is a well-known generator of free radicals that is produced during endogenous and UV-induced oxidation processes in human skin and was used to upregulate ROS and HNE in normal human fibroblast WS-1 cells. Using a flow cytometry-based assay, the results demonstrate that at 0.001% concentration, green tea polyphenols alone, and in combination with 0.1 mM caffeine, inhibited the upregulation of H2O2-generated free radicals and HNE in human skin fibroblasts in vitro. Caffeine alone demonstrated limited anti-oxidant properties.

Green tea extract protects human skin fibroblasts from reactive oxygen species induced necrosis.

Oxidative damage by reactive oxygen species (ROS) plays a major role in skin aging, carcinogenesis and inflammation. Little is known about the protective effects of green tea extract (GTE) on toxic ROS-induced skin death. We use an in vitro model of normal human skin fibroblasts (AG13145) to study the effects of green tea extract (GTE) on hydrogen peroxide (H(2)O(2)) induced necrosis. Cell morphology, numbers, apoptosis, necrosis, and ROS were assessed by epifluorescence microscopy and flow cytometry. This study demonstrates that GTE protected from H(2)O(2)-induced necrosis in a dose-dependent manner, with highest dose GTE (100 ng/mL) resulting in the most protection from necrosis, as assessed by improved cell morphology, increased cell numbers, and decreased necrosis. The protective effects of GTE on H(2)O(2)-induced necrosis appear to be mediated directly by decreasing intracellular ROS. The present study suggests that pretreatment with high doses of GTE could protect from toxic ROS-induced injury of skin in the clinical setting. However, additional studies are necessary to determine the clinical utility of GTE for decreasing skin cell ROS, necrosis and inflammation.

Dose-dependent antioxidant function of resveratrol demonstrated via modulation of reactive oxygen species in normal human skin fibroblasts in vitro.

The study of free radicals is particularly relevant in the context of human skin carcinogenesis and photoaging because of their ability to induce DNA mutations and damaging lipid peroxidation byproducts. Therefore, it is important to identify and evaluate agents with the ability to modulate intracellular free radicals. Significant interest exists in evaluating the chemotherapeutic and anti-oxidant properties of resveratrol (trans-3,4',5-trihydroxystilbene). Resveratrol is a phytoalexin, a naturally occurring compound derived from the skin of grapes and other plants. Resveratrol was selected for evaluation because of demonstrated chemopreventive and chemotherapeutic properties in a murine skin cancer model and other human cancer models through a variety of mechanisms. However, the intracellular anti-oxidant properties of resveratrol on free radicals in human skin cells in vitro is not well characterized. The purpose of this research is to investigate the ability of resveratrol to modulate the hydrogen peroxide-induced upregulation of reactive oxygen species (ROS) free radicals in normal human skin fibroblast cells in vitro. Hydrogen peroxide is a well known generator of free radicals that occurs during endogenous and UV-induced oxidation processes in the human skin and was used to upregulate ROS in normal human skin fibroblast cells. Using a flow cytometry-based assay, the results demonstrate highly significant (P<0.001) dose-dependent reduction of intracellular hydrogen peroxide-upregulated ROS by resveratrol at 0.01%, 0.001% and 0.0001% concentrations in human skin fibroblasts in vitro.

Sirtuins in dermatology: applications for future research and therapeutics.

Sirtuins are a family of seven proteins in humans (SIRT1-SIRT7) that are involved in multiple cellular processes relevant to dermatology. The role of sirtuins in other organ systems is established. However, the importance of these proteins in dermatology is less defined. Recently, sirtuins gained international attention because of their role as "longevity proteins" that may extend and enhance human life. Sirtuins function in the cell via histone deacetylase and/or adenosine diphosphate ribosyltransferase enzymatic activity that target histone and non-histone substrates, including transcription regulators, tumor suppressors, structural proteins, DNA repair proteins, cell signaling proteins, transport proteins, and enzymes. Sirtuins are involved in cellular pathways related to skin structure and function, including aging, ultraviolet-induced photoaging, inflammation, epigenetics, cancer, and a variety of cellular functions including cell cycle, DNA repair and proliferation. This review highlights sirtuin-related cellular pathways, therapeutics and pharmacological targets in atopic dermatitis, bullous dermatoses, collagen vascular disorders, psoriasis, systemic lupus erythematosus, hypertrophic and keloid scars, cutaneous infections, and non-melanoma and melanoma skin cancer. Also discussed is the role of sirtuins in the following genodermatoses: ataxia telangiectasia, Cowden's syndrome, dyskeratosis congenita, Rubenstein-Taybi, Werner syndrome, and xeroderma pigmentosum. The pathophysiology of these inherited diseases is not well understood, and sirtuin-related processes represent potential therapeutic targets for diseases lacking suitable alternative treatments. The goal of this review is to bring attention to the dermatology community, physicians, and scientists, the importance of sirtuins in dermatology and provide a foundation and impetus for future discussion, research and pharmacologic discovery.

Transcranial red and near infrared light transmission in a cadaveric model.

BACKGROUND AND OBJECTIVE: Low level light therapy has garnered significant interest within the past decade. The exact molecular mechanisms of how red and near infrared light result in physiologic modulation are not fully understood. Heme moieties and copper within cells are red and near infrared light photoreceptors that induce the mitochondrial respiratory chain component cytochrome C oxidase, resulting in a cascade linked to cytoprotection and cellular metabolism. The copper centers in cytochrome C oxidase have a broad absorption range that peaks around 830 nm. Several in vitro and in vivo animal and human models exist that have demonstrated the benefits of red light and near infrared light for various conditions. Clinical applications for low level light therapy are varied. One study in particular demonstrated improved durable functional outcomes status post-stroke in patients treated with near infrared low level light therapy compared to sham treatment [1]. Despite previous data suggesting the beneficial effect in treating multiple conditions, including stroke, with low level light therapy, limited data exists that measures transmission in a human model. STUDY DESIGN/MATERIALS AND METHODS: To investigate this idea, we measured the transmission of near infrared light energy, using red light for purposes of comparison, through intact cadaver soft tissue, skull bones, and brain using a commercially available LED device at 830 nm and 633 nm. RESULTS: Our results demonstrate that near infrared measurably penetrates soft tissue, bone and brain parenchyma in the formalin preserved cadaveric model, in comparison to negligible red light transmission in the same conditions. CONCLUSION: These findings indicate that near infrared light can penetrate formalin fixed soft tissue, bone and brain and implicate that benefits observed in clinical studies are potentially related to direct action of near infrared light on neural tissue.

Green tea (Camelia sinensis) suppresses B cell production of IgE without inducing apoptosis.

Green tea (Camelia sinensis) is known to possess biological properties that are antioxidative and antimutagenic. Recent studies demonstrated beneficial effects of green tea in inflammatory allergy. However, the effect of green tea on anti-allergic activity/IgE responses in vitro has not been studied. U266 myeloma cells (2 x 10(6)/ml), which secrete IgE, were cultured for 0-72 hr with or without green tea extract (1-300 ng/ml), and IgE levels in the supernatants were determined (24-72 hr) by ELISA. The effects of green tea extract on U266 cell numbers, viability, and apoptosis were studied by flow cytometry. High levels of IgE produced by U266 cells were observed at 24, 48, and 72 hr (1.3 +/- 0.3 x 10(3), 1.7 +/- 0.3 x 10(3), 2.8 +/- 0.4 x 10(3) IU/ml, respectively). Addition of green tea extract either as (a) a single dose, or (b) repeated daily doses, suppressed IgE production with increasing suppression over time (up to 90%; p <0.05); the suppression was dose-dependent with the highest concentrations resulting in the greatest suppression. The suppression of IgE production by green tea extract was not mediated by apoptosis or cell death. This study demonstrates that green tea extract has immunoregulatory effects on human IgE responses in vitro.

Intralesional interferon alpha-2b in the treatment of basal cell carcinoma and squamous cell carcinoma: revisited.

BACKGROUND: Intralesional interferon (IFN) alpha-2b has been shown to be a safe and effective mode of treatment for basal cell carcinoma and squamous cell carcinoma. Multiple studies published in the 1980s through the early 1990s have demonstrated the efficacy of intralesional interferon in the treatment of these malignancies. Unfortunately, this modality appears to be underused. OBJECTIVE: This article serves to remind dermatologists that in addition to cryotherapy, electrodesiccation, and surgical excision, intralesional IFN-alpha is an important part of the armamentarium in the treatment of nonmelanoma skin cancers. METHODS: In addition to a review of the literature, we present eight cases in seven patients successfully treated with intralesional IFN for basal cell carcinoma and squamous cell carcinoma. CONCLUSIONS: Its nonsurgical approach and excellent cosmetic results make IFNalpha-2b an attractive option for patients and an important alternative when other treatment modalities are impractical or contraindicated.