Multiphoton Microscopy of Collagen Structure in Ex Vivo Human Skin Following Electrochemical Therapy.

OBJECTIVES: Injury to healthy dermis and the dermoepidermal junction initiates a robust healing process consisting of fibrous tissue overgrowth, collagen deposition, and scar formation. The conventional management of scars and other skin injuries has largely relied upon surgical soft tissue transfer to resurface and/or replace damaged and dysmorphic tissue with new skin. However, these strategies are invasive, expensive, and may further exacerbate integumentary injury. In this study, we examine the creation of in situ redox generated pH changes in fresh human skin. We believe this process of "electrochemical therapy" (ECT) leads to changes in collagen matrix structure. Our objective is to map local tissue pH landscapes and image changes in collagen structure of non-injured skin following ECT. STUDY DESIGN: Ex vivo human study involving ECT of human skin. METHODS: Remnant fresh ex vivo human facial skin from facelift operations was enveloped in saline-soaked gauze for a maximum of 2 hours prior to ECT and imaging. ECT was performed by inserting platinum-plated needle electrodes connected to a DC power supply. Voltage (4, 5, or 6 V) and time (3, 4, or 5 minutes) were varied systematically. High frequency ultrasound (25 MHz) was performed immediately after ECT on each sample. Treated samples were also imaged using multiphoton microscopy (MPM) with second harmonic generation (SHG) to specifically visualize collagen fibers in the dermis. The pH landscapes were mapped using indicator dyes in bisected specimens and the MPM images were compared with histologic findings. RESULTS: Above 4 V and 3 minutes, a profound reduction in dermal collagen SHG signal was observed at the anode. Although there was less blunting of SHG signal seen at the cathode, a decrease in the fluorescence of the dermoepidermal junction was observed. The pH application suggests ECT spatial selectivity and a direct relationship between voltage and application time. Ultrasound demonstrated gas formation between the anode and cathode, which is consistent with ECT's mechanism of action. Importantly, these electrochemical changes occurred without disrupting dermal and epidermal histologic architecture. CONCLUSION: ECT alters tissue pH leading to dermal collagen structural change. These results offer additional insight into the translational potential of ECT to locally remodel the soft-tissue matrix. Future directions aim to expand into a skin injury model to determine if similar collagen effects are observed in vivo. ECT is incredibly inexpensive (~$5) and may be a means to treat soft tissue injuries using simple needle-based devices and DC battery power supplies. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc.

Electrochemical treatment of ex vivo human abdominal skin and potential use in scar management: A pilot study.

INTRODUCTION: Scar treatments aim to address pathologic collagen deposition; however, they can be expensive or difficult to control. Electrochemical therapy (ECT) offers a simple alternative treatment. The purpose of this study is to examine the acid-base and histological changes in ex vivo human abdominal skin following ECT. METHODS: Forty-two ex vivo human panniculus tissue sections collected from six individuals were tumesced with normal saline. ECT was performed by inserting two platinum needle electrodes connected to a DC power supply into each specimen. Voltage was varied (3-6 V) and applied for 5 minutes. Each specimen was sectioned across both electrode insertion sites and immediately stained with pH sensitive dye. The width of dye color change for each dosimetry pair was calculated. Hematoxylin and eosin staining was used to evaluate samples. RESULTS AND DISCUSSION: ECT caused a spatially localised and dose-dependent increased area of acidic and basic pH around the anode and cathode, respectively. A significantly greater mean width of pH change was generated at the cathode compared to the anode in all treatment groups. Histological evaluation displayed broad condensation and hyalinisation of dermal collagen. CONCLUSION: ECT triggered dermal pH alterations and changed the underlying structural framework of the specimen. This technology may serve as a low-cost, minimally invasive local soft-tissue remodeling technique with potential application in scar management. LEVEL OF EVIDENCE: 5. LAY SUMMARY: Electrochemical therapy is a novel treatment that causes spatially selective dermal injury in areas of interest. This study measures the effects of electrochemical therapy when applied to abdominal skin. Electrochemical therapy appears to have beneficial effects by causing a highly localised reduction in collagen content or local softening of tissue, which is consistent with other studies on scar therapies, including chemexfoliation, radiofrequency technologies, and lasers. However, electrochemical therapy can be performed at a fraction of the costs of these aforementioned modalities.

Electrochemolipolysis of Human Adipose Tissue.

Importance: Body fat contouring procedures have increasingly grown in popularity over the years. As such, there is a need for inexpensive, minimally invasive, and simple fat reduction/contouring technique. Objective: To examine the acid-base and histological changes in ex vivo human adipose tissue after electrochemolipolysis (ECL). Design, Setting, and Participants: Panniculus tissue specimens obtained after abdominoplasty procedures were tumesced with normal saline. Two platinum needle electrodes were inserted into each sample and connected to a DC power supply. Voltage (3-6 V) was varied and applied for 5 min. Specimens were sectioned through a sagittal midline across both electrode insertion sites and immediately stained with pH-sensitive dye. A numerical algorithm was used to calculate the area of the dye color change for each dosimetry pair. Samples were also evaluated utilizing light microscopy (hematoxylin and eosin). An ex vivo human adipose tissue model was used for evaluating the effects of ECL. Results: Acidic and basic pH was appreciated surrounding the anode and cathode insertion sites, respectively. The effect was spatially localized and dose dependent. Statistical analysis of these data showed no significant difference between the mean area of the pH disturbance generated at the anode compared with the cathode at 3 V for 5 min (6.04 mm2 vs. 2.95 mm2, p = 0.40, 95% CI -4.8 to 11). A significantly greater area of pH disruption was generated at the cathode versus the anode in groups 4 V for 5 min (14.7 mm2 vs. 5.00 mm2, p = 0.032, 95% CI 0.93-19), 5 V for 5 min (15.5 mm2 vs. 6.72 mm2, p = 0.019, 95% CI 1.6-16), and 6 V for 5 min (22.5 mm2 vs. 10.0 mm2, p = 0.047, 95% CI 0.22-25). Acute structural changes in adipocytes were observed in all specimens. Vascular damage with adjacent adipocyte necrosis was prominent at the cathode site in group 6 V for 5 min. Conclusions and Relevance: ECL at the studied dosimetry parameters induced acid and base changes in human adipose tissue, suggesting its potential use in nonsurgical fat reduction as an ultralow cost alternative to current lipolytic devices and pharmaceuticals. Level of Evidence: NA.

Electrochemical degradation and saponification of porcine adipose tissue.

Body contouring achieved via subcutaneous adipose tissue reduction has notably advanced over the past century, from suction assisted lipectomy to techniques with reduced degrees of invasiveness including laser, radiofrequency, high frequency focused ultrasound, cryolipolysis, and drug-based injection approaches. These costly techniques have focused on damaging adipocyte cell membranes, hydrolyzing triglycerides (TGs), or inducing apoptosis. Here, we present a simple, low-cost technique, termed electrochemical lipolysis (ECLL). During ECLL, saline is injected into the subcutaneous adipose tissue, followed by insertion of needle electrodes and application of an electrical potential. Electrolysis of saline creates localized pH gradients that drive adipocyte death and saponification of TGs. Using pH mapping, various optical imaging techniques, and biochemical assays, we demonstrate the ability of ECLL to induce acid and base injury, cell death, and the saponification of triglycerides in ex vivo porcine adipose tissue. We define ECLL's potential role as a minimally-invasive, ultra-low-cost technology for reducing and contouring adipose tissue, and present ECLL as a potential new application of an emerging electrochemical redox based treatment modality.

The biophysical effects of localized electrochemical therapy on porcine skin.

BACKGROUND: Minimally-invasive methods to treat scars address a common pathway of altering collagen structure, leading to collagen remodeling. OBJECTIVE: In this study, we employed in situ redox chemistry to create focal pH gradients in skin, altering dermal collagen, in a process we refer to as electrochemical therapy (ECT). The effects of ECT to induce biochemical and structural changes in ex vivo porcine skin were examined. METHODS: During ECT, two platinum electrodes were inserted into fresh porcine skin, and following saline injection, an electrical potential was applied. pH mapping, high frequency ultrasonography, and two photon excitation microscopy and second harmonic generation (SHG) microscopy were used to evaluate treatment effects. Findings were correlated with histology. RESULTS: Following ECT, pH mapping depicted acid and base production at anode and cathode sites respectively, with increasing voltage and application time. Gas formation during ECT was observed with ultrasonography. Anode sites showed significant loss of SHG signal, while cathode sites showed disorganized collagen structure with fewer fibrils emitting an attainable signal. Histologically, collagen denaturation at both sites was confirmed. CONCLUSION: We demonstrated the production of in situ acid and base in skin occurring via ECT. The effects chemically and precisely alter collagen structure through denaturation, giving insight on the potential of ECT as a simple, low-cost, and minimally-invasive means to remodel skin and treat scars.