Andrew Mamalis

The role of microRNAs in skin fibrosis

Fibrotic skin disorders may be debilitating and impair quality of life. There are few effective treatment options for cutaneous fibrotic diseases. In this review, we discuss our current understanding of the role of microRNAs (miRNAs) in skin fibrosis. miRNAs are a class of small, non-coding RNAs involved in skin fibrosis. These small RNAs range from 18 to 25 nucleotides in length and modify gene expression by binding to target messenger RNA (mRNA), causing degradation of the target mRNA or inhibiting the translation into proteins. We present an overview of the biogenesis, maturation and function of miRNAs. We highlight miRNA’s role in key skin fibrotic processes including: transforming growth factor-beta signaling, extracellular matrix deposition, and fibroblast proliferation and differentiation. Some miRNAs are profibrotic and their upregulation favors these processes contributing to fibrosis, while anti-fibrotic miRNAs inhibit these processes and may be reduced in fibrosis. Finally, we describe the diagnostic and therapeutic significance of miRNAs in the management of skin fibrosis. The discovery that miRNAs are detectable in serum, plasma, and other bodily fluids, and are relatively stable, suggests that miRNAs may serve as valuable biomarkers to monitor disease progression and response to treatment. In the treatment of skin fibrosis, anti-fibrotic miRNAs may be upregulated using mimics and viral vectors. Conversely, profibrotic miRNAs may be downregulated by employing anti-miRNAs, sponges, erasers and masks. We believe that miRNA-based therapies hold promise as important treatments and may transform the management of fibrotic skin diseases by physicians.

Laser and Light-based Treatment of Keloids – A Review

Keloids are an overgrowth of fibrotic tissue outside the original boundaries of an injury and occur secondary to defective wound healing. Keloids often have a functional, aesthetic, or psychosocial impact on patients as highlighted by quality‐of‐life studies.

Targeting the PD-1 pathway: a promising future for the treatment of melanoma

Advanced melanoma presents a significant therapeutic challenge to clinicians. Many therapies for metastatic melanoma are limited by low response rates, severe toxicities, and/or relatively short response duration. Cancer immunotherapies that act as immune-checkpoint inhibitors to block the localized immune suppression mechanisms utilized by tumors are undergoing development and clinical trials. A clinically relevant immune escape mechanism in melanoma is the activation of the programmed cell death-1 (PD-1) receptor on infiltrating T cells. Activating PD-1 triggers an immune checkpoint resulting in inhibition of T cells directed against melanoma antigens and prevents the immune system from combating the melanoma. In Phase I clinical trials, two anti-PD1 therapies, Nivolumab and MK-3475, that block the PD-1 receptor to enable T cell killing have demonstrated objective tumor responses in patients with advanced melanoma. The purpose of this review is to present the available clinical evidence on anti-PD-1 and anti-PD-L1 immunotherapy for the treatment of advanced melanoma. We also discuss limitations associated with anti-PD-1 therapy. The blockade of the PD-1–PD-L1 pathway has shown promising results in clinical trials and has revolutionized melanoma immunotherapy.

Light emitting diode-generated blue light modulates fibrosis characteristics: fibroblast proliferation, migration speed, and reactive oxygen species generation.

Blue light is part of the visible light spectrum that does not generate harmful DNA adducts associated with skin cancer and photoaging, and may represent a safer therapeutic modality for treatment of keloid scars and other fibrotic skin diseases. Our laboratory previously demonstrated that light‐emitting diode (LED) red and infrared light inhibits proliferation of skin fibroblasts. Moreover, different wavelengths of light can produce different biological effects. Furthermore, the effects of LED blue light (LED‐BL) on human skin fibroblasts are not well characterized. This study investigated the effects of LED‐BL on human skin fibroblast proliferation, viability, migration speed, and reactive oxygen‐species (ROS) generation.

NADPH oxidase enzymes in skin fibrosis: molecular targets and therapeutic agents.

Fibrosis is characterized by the excessive deposition of extracellular matrix components eventually resulting in organ dysfunction and failure. In dermatology, fibrosis is the hallmark component of many skin diseases, including systemic sclerosis, graft-versus-host disease, hypertrophic scars, keloids, nephrogenic systemic fibrosis, porphyria cutanea tarda, restrictive dermopathy and other conditions. Fibrotic skin disorders may be debilitating and impair quality of life. There are few FDA-approved anti-fibrotic drugs; thus, research in this area is crucial in addressing this deficiency. Recent investigations elucidating the pathogenesis of skin fibrosis have implicated endogenous reactive oxygen species produced by the multicomponent nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) enzyme complex. In this review, we discuss Nox enzymes and their role in skin fibrosis. An overview of the Nox enzyme family is presented and their role in the pathogenesis of skin fibrosis is discussed. The mechanisms by which Nox enzymes influence specific fibrotic skin disorders are also reviewed. Finally, we describe the therapeutic approaches to ameliorate skin fibrosis by directly targeting Nox enzymes with the use of statins, p47phox subunit modulators, or GKT137831, a competitive inhibitor of Nox enzymes. Nox enzymes can also be targeted indirectly via scavenging ROS with antioxidants. We believe that Nox modulators are worthy of further investigation and have the potential to transform the management of skin fibrosis by dermatologists.

Oxidative Stress and Skin Fibrosis

Fibrosis is defined as increased fibroblast proliferation and deposition of extracellular matrix components with potential clinical ramifications including organ dysfunction and failure. Fibrosis is a characteristic finding of various skin diseases which can have life-threatening consequences. These implications call for research into this topic as only a few treatments targeting fibrosis are available. In this review, we discuss oxidative stress and its role in skin fibrosis. Recent studies have implicated the importance of oxidative stress in a variety of cellular pathways directly and indirectly involved in the pathogenesis of skin fibrosis. The cellular pathways by which oxidative stress affects specific fibrotic skin disorders are also reviewed. Finally, we also describe various therapeutic approaches specifically targeting oxidative stress to prevent skin fibrosis. We believe oxidative stress is a relevant target, and understanding the role of oxidative stress in skin fibrosis will enhance knowledge of fibrotic skin diseases and potentially produce targeted therapeutic options.

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.

Temperature-dependent impact of thermal aminolaevulinic acid photodynamic therapy on apoptosis and reactive oxygen species generation in human dermal fibroblasts.

Actinic keratoses (AKs) are generally accepted as common precursor lesions to invasive squamous cell carcinoma. Photodynamic therapy (PDT) is a common, in‐office, field therapy modality used in the treatment of AKs. Clinical and laboratory observations have demonstrated that temperature modulation can affect PDT efficacy.

Light-emitting diode-generated red light inhibits keloid fibroblast proliferation

BACKGROUND Red light is part of the visible light spectrum that does not generate DNA adducts associated with skin cancer and photoaging and may represent a safer therapeutic modality for treatment of keloid scars and other fibrotic skin diseases. Our laboratory previously demonstrated that light-emitting diode–generated red light (LED-RL) inhibits proliferation of skin fibroblasts. The effects of LED-RL on keloidal skin are not well characterized. OBJECTIVE To determine the effect of LED-RL on keloid-derived fibroblast proliferation and viability in vitro. METHODS Irradiation of primary keloid–derived human skin fibroblasts using LED-RL panels was performed in vitro, and modulation of proliferation and viability was quantified using trypan blue dye exclusion assay. Statistical analysis was performed using analysis of variance to compare treatment arms and the Student t-test to compare each treatment arm with the paired bench control arm. RESULTS Keloid fibroblasts treated with LED-RL 240, 320, and 480 J/cm2 demonstrated statistically significant dose-dependent decreases in relative proliferation rate of 12.4%, 16.5%, and 28.9%, respectively, compared with matched nonirradiated controls (p < .05) and did not significantly alter viability relative to the matched nonirradiated controls. CONCLUSION Light-emitting diode–generated red light can inhibit keloid fibroblast proliferation in a dose-dependent manner without altering viability. Light-emitting diode–generated red light has the potential to contribute to the treatment of keloids and other fibrotic skin diseases and is worthy of further translational and clinical investigation.

Resveratrol Prevents High Fluence Red Light-Emitting Diode Reactive Oxygen Species-Mediated Photoinhibition of Human Skin Fibroblast Migration.

Background Skin fibrosis is a significant medical problem that leads to a functional, aesthetic, and psychosocial impact on quality-of-life. Light-emitting diode-generated 633-nm red light (LED-RL) is part of the visible light spectrum that is not known to cause DNA damage and is considered a safe, non-invasive, inexpensive, and portable potential alternative to ultraviolet phototherapy that may change the treatment paradigm of fibrotic skin disease. Objective The goal of our study was to investigate the how reactive oxygen species (ROS) free radicals generated by high fluence LED-RL inhibit the migration of skin fibroblasts, the main cell type involved in skin fibrosis. Fibroblast migration speed is increased in skin fibrosis, and we studied cellular migration speed of cultured human skin fibroblasts as a surrogate measure of high fluence LED-RL effect on fibroblast function. To ascertain the inhibitory role of LED-RL generated ROS on migration speed, we hypothesized that resveratrol, a potent antioxidant, could prevent the photoinhibitory effects of high fluence LED-RL on fibroblast migration speed. Methods High fluence LED-RL generated ROS were measured by flow cytometry analysis using dihydrorhodamine (DHR). For purposes of comparison, we assessed the effects of ROS generated by hydrogen peroxide (H2O2) on fibroblast migration speed and the ability of resveratrol, a well known antioxidant, to prevent LED-RL and H2O2 generated ROS-associated changes in fibroblast migration speed. To determine whether resveratrol could prevent the high fluence LED-RL ROS-mediated photoinhibition of human skin fibroblast migration, treated cells were incubated with resveratrol at concentrations of 0.0001% and 0.001% for 24 hours, irradiated with high fluences LED-RL of 480, 640, and 800 J/cm2. Results High fluence LED-RL increases intracellular fibroblast ROS and decreases fibroblast migration speed. LED-RL at 480, 640 and 800 J/cm2 increased ROS levels to 132.8%, 151.0%, and 158.4% relative to matched controls, respectively. These LED-RL associated increases in ROS were prevented by pretreating cells with 0.0001% or 0.001% resveratrol. Next, we quantified the effect of hydrogen peroxide (H2O2)-associated ROS on fibroblast migration speed, and found that while H2O2–associated ROS significantly decreased relative fibroblast migration speed, pretreatment with 0.0001% or 0.001% resveratrol significantly prevented the decreases in migration speed. Furthermore, we found that LED-RL at 480, 640 and 800 J/cm2 decreased fibroblast migration speed to 83.0%, 74.4%, and 68.6% relative to matched controls, respectively. We hypothesized that these decreases in fibroblast migration speed were due to associated increases in ROS generation. Pretreatment with 0.0001% and 0.001% resveratrol prevented the LED-RL associated decreases in migration speed. Conclusion High fluence LED-RL increases ROS and is associated with decreased fibroblast migration speed. We provide mechanistic support that the decreased migration speed associated with high fluence LED-RL is mediated by ROS, by demonstrating that resveratrol prevents high fluence LED-RL associated migration speed change. These data lend support to an increasing scientific body of evidence that high fluence LED-RL has anti-fibrotic properties. We hypothesize that our findings may result in a greater understanding of the fundamental mechanisms underlying visible light interaction with skin and we anticipate clinicians and other researchers may utilize these pathways for patient benefit.