Structural and molecular characteristics of weight-bearing volar skin can be reconstituted by micro skin tissue column grafting.

For patients with lower limb amputations, prostheses are immensely helpful for mobility and the ability to perform job-related or recreational activities. However, the skin covering the amputation stump is typically transposed from adjacent areas of the leg and lacks the weight-bearing capacity that is only found in the specialized skin covering the palms and soles (a.k.a. volar skin). As a result, the skin tissue in direct contact with the prosthesis frequently breaks down, leading to the development of painful sores and other complications that limit, and often preclude, the use of prostheses. Transplanting volar skin onto amputation stumps could be a solution to these problems, but traditional skin transplantation techniques cause substantial morbidity at the donor site, such as pain and scarring, which are especially problematic for volar skin given the critical functional importance of the volar skin areas. We previously developed the technology to collect and engraft full-thickness skin tissue while avoiding long-term donor site morbidity, by harvesting the skin in the form of small (~0.5 mm diameter) cores that we termed "micro skin tissue columns" (MSTCs), so that each donor wound is small enough to heal quickly and without clinically appreciable scarring or other long-term abnormalities. The goal of this study was to establish whether a similar approach could be used to confer the structural and molecular characteristics of volar skin ectopically to other skin areas. In a human-to-mouse xenograft model, we show the long-term persistence of various human plantar MSTC-derived cell types in the murine recipient. Then in an autologous porcine model, we harvested MSTCs from the bottom of the foot and transplanted them onto excision wounds on the animals' trunks. The healing processes at both the donor and graft sites were monitored over 8 weeks, and tissue samples were taken to verify volar-specific characteristics by histology and immunohistochemistry. The volar donor sites were well-tolerated, healed rapidly, and showed no signs of scarring or any other long-term defects. The graft sites were able to maintain volar-specific histologic features and expression of characteristics protein markers, up to the 8-week duration of this study. These results suggest that MSTC grafting could be a practical approach to obtain autologous donor volar skin tissue, confer volar skin characteristics ectopically to nonvolar skin areas, improve the load-bearing capacity of amputation stump skin, and ultimately enhance mobility and quality-of-life for lower limb amputees.

Noninvasive treatment of cutaneous neurofibromas (cNFs): Results of a randomized prospective, direct comparison of four methods.

BACKGROUND: People with Neurofibromatosis Type 1 (NF1) suffer disfigurement and pain when hundreds to thousands of cutaneous neurofibromas (cNFs) appear and grow throughout life. Surgical removal of cNFs under anesthesia is the only standard therapy, leaving surgical scars. OBJECTIVE: Effective, minimally-invasive, safe, rapid, tolerable treatment(s) of small cNFs that may prevent tumor progression. METHODS: Safety, tolerability, and efficacy of 4 different treatments were compared in 309, 2-4 mm cNFs across 19 adults with Fitzpatrick skin types (FST) I-IV: radiofrequency (RF) needle coagulation, 755 nm alexandrite laser with suction, 980 nm diode laser, and intratumoral injection of 10 mg/mL deoxycholate. Regional pain, clinical responses, tumor height and volume (by 3D photography) were assessed before, 3 and 6 months post-treatment. Biopsies were obtained electively at 3 months. RESULTS: There was no scarring or adverse events > grade 2. Each modality significantly (P < .05) reduced or cleared cNFs, with large variation between tumors and participants. Alexandrite laser and deoxycholate were fast and least painful; 980 nm laser was most painful. Growth of cNFs was not stimulated by treatment(s) based on height and volume values at 3 and 6 months compared to baseline. LIMITATIONS: Intervention was a single treatment session; dosimetry has not been optimized. CONCLUSIONS: Small cNFs can be rapidly and safely treated without surgery.

Optical, flow, and thermal analysis of a phototherapy extracorporeal membrane oxygenator for treating carbon monoxide poisoning.

BACKGROUND: Extracorporeal membrane oxygenators (ECMO) are currently utilized to mechanically ventilate blood when lung or lung and heart function are impaired, like in cases of acute respiratory distress syndrome (ARDS). ARDS can be caused by severe cases of carbon monoxide (CO) inhalation, which is the leading cause of poison-related deaths in the United States. ECMOs can be further optimized for severe CO inhalation using visible light to photo-dissociate CO from hemoglobin (Hb). In previous studies, we combined phototherapy with an ECMO to design a photo-ECMO device, which significantly increased CO elimination and improved survival in CO-poisoned animal models using light at 460, 523, and 620 nm wavelengths. Light at 620 nm was the most effective in removing CO. OBJECTIVE: The aim of this study is to analyze the light propagation at 460, 523, and 620 nm wavelengths and the 3D blood flow and heating distribution within the photo-ECMO device that increased CO elimination in CO-poisoned animal models. METHODS: Light propagation, blood flow dynamics, and heat diffusion were modeled using the Monte Carlo method and the laminar Navier-Stokes and heat diffusion equations, respectively. RESULTS: Light at 620 nm propagated through the device blood compartment (4 mm), while light at 460 and 523 nm only penetrated 48% to 50% (~2 mm). The blood flow velocity in the blood compartment varied with regions of high (5 mm/s) and low (1 mm/s) velocity, including stagnant flow. The blood temperatures at the device outlet for 460, 523, and 620 nm wavelengths were approximately 26.7°C, 27.4°C, and 20°C, respectively. However, the maximum temperatures within the blood treatment compartment rose to approximately 71°C, 77°C, and 21°C, respectively. CONCLUSIONS: As the extent of light propagation correlates with efficiency in photodissociation, the light at 620 nm is the optimal wavelength for removing CO from Hb while maintaining blood temperatures below thermal damage. Measuring the inlet and outlet blood temperatures is not enough to avoid unintentional thermal damage by light irradiation. Computational models can help eliminate risks of excessive heating and improve device development by analyzing design modifications that improve blood flow, like suppressing stagnant flow, further increasing the rate of CO elimination.

Total ice content and lipid saturation determine adipose tissue cryolipolysis by injection of ice-slurry.

OBJECTIVES: Cryolipolysis uses tissue cooling to solidify lipids, preferentially damaging lipid-rich cells. Topical cooling is popular for the reduction of local subcutaneous fat. Injection of biocompatible ice-slurry is a recently introduced alternative. We developed and verified a quantitative model that simulates the heat exchange and phase changes involved, offering insights into ice-slurry injection for treating subcutaneous fat. METHODS: Finite element method was used to model the spatial and temporal progression of heat transfer between adipose tissue and injected ice-slurry, estimating dose-response relationships between properties of the slurry and size of tissue affected by cryolipolysis. Phase changes of both slurry and adipose tissue lipids were considered. An in vivo swine model was used to validate the numerical solutions. Oils with different lipid compositions were exposed to ice-slurry in vitro to evaluate the effects of lipid freezing temperature. Microscopy and nuclear magnetic resonance (NMR) were performed to detect lipid phase changes. RESULTS: A ball of granular ice was deposited at the injection site in subcutaneous fat. Total injected ice content determines both the effective cooling region of tissue, and the duration of tissue cooling. Water's high latent heat of fusion enables tissue cooling long after slurry injection. Slurry temperature affects the rate of tissue cooling. In swine, when 30 ml slurry injection at -3.5°C was compared to 15 ml slurry injection at -4.8°C (both with the same total ice content), the latter led to almost twice faster tissue cooling. NMR showed a large decrease in diffusion upon lipid crystallization; saturated lipids with higher freezing temperatures were more susceptible to solidification after ice-slurry injection. CONCLUSIONS: Total injected ice content determines both the volume of tissue treated by cryolipolysis and the cooling duration after slurry injection, while slurry temperature affects the cooling rate. Lipid saturation, which varies with diet and anatomic location, also has an important influence.

Injectable ice slurry for reducing pericardial adipose tissue.

OBJECTIVES: Excess pericardial adipose tissue (PAT) is associated with a higher risk of cardiovascular diseases. Currently, available methods for reducing PAT volume include weight loss through diet and exercise, weight loss with medications, and bariatric surgery. However, these methods are all limited by low patient compliance to maintain the results. We have developed an injectable ice slurry that could selectively target and reduce subcutaneous adipose tissue volume. The aim of this study was to investigate the feasibility and safety of using injectable slurry to selectively reduce PAT volume in a preclinical large animal model. METHODS: PAT in Yucatan swine was injected with slurry or room temperature control solution. All animals were imaged with baseline chest computed tomography (CT) before slurry injection and at 2 months after injection to quantify PAT volume. Specimens from injected and noninjected PAT were harvested for histology. RESULTS: Slurry treatment of PAT was well tolerated in all animals. Slurry-induced selective cryolipolysis in treated PAT. CT imaging showed decrease in PAT volume in treated area at 8 weeks posttreatment compared to baseline, that was significantly different from control solution treated group (median [range]: -29.66 [-35.07 to -27.92]% vs. -1.50 [-11.69 to 8.69]% in control animals respectively, p < 0.05). CONCLUSIONS: This study demonstrated that slurry injection into PAT is feasible in a large animal model. Slurry injection was safe and effective in inducing selective cryolipolysis in PAT and reducing PAT volume. Slurry reduction of PAT could potentially serve as a novel treatment for cardiovascular diseases.

Local vasoregulative interventions impact drug concentrations in the skin after topical laser-assisted delivery.

INTRODUCTION: The ability of ablative fractional lasers (AFL) to enhance topical drug uptake is well established. After AFL delivery, however, drug clearance by local vasculature is poorly understood. Modifications in vascular clearance may enhance AFL-assisted drug concentrations and prolong drug dwell time in the skin. Aiming to assess the role and modifiability of vascular clearance after AFL-assisted delivery, this study examined the impact of vasoregulative interventions on AFL-assisted 5-fluorouracil (5-FU) concentrations in in vivo skin. METHODS: 5-FU uptake was assessed in intact and AFL-exposed skin in a live pig model. After fractional CO2 laser exposure (15 mJ/microbeam, 5% density), vasoregulative intervention using topical brimonidine cream, epinephrine solution, or pulsed dye laser (PDL) was performed in designated treatment areas, followed by a single 5% 5-FU cream application. At 0, 1, 4, 48, and 72 h, 5-FU concentrations were measured in 500 and 1500 µm skin layers by mass spectrometry (n = 6). A supplemental assessment of blood flow following AFL ± vasoregulation was performed using optical coherence tomography (OCT) in a human volunteer. RESULTS: Compared to intact skin, AFL facilitated a prompt peak in 5-FU delivery that remained elevated up to 4 hours (1500 µm: 1.5 vs. 31.8 ng/ml [1 hour, p = 0.002]; 5.3 vs. 14.5 ng/ml [4 hours, p = 0.039]). However, AFL's impact was transient, with 5-FU concentrations comparable to intact skin at later time points. Overall, vasoregulative intervention with brimonidine or PDL led to significantly higher peak 5-FU concentrations, prolonging the drug's dwell time in the skin versus AFL delivery alone. As such, brimonidine and PDL led to twofold higher 5-FU concentrations than AFL alone in both skin layers by 1 hour (e.g., 500 µm: 107 ng/ml [brimonidine]; 96.9 ng/ml [PDL], 46.6 ng/ml [AFL alone], p ≤ 0.024), and remained significantly elevated at 4 hours (p ≤ 0.024). A similar pattern was observed for epinephrine, although trends remained nonsignificant (p ≥ 0.09). Prolonged 5-FU delivery was provided by PDL, resulting in sustained drug deposition compared to AFL alone at both 48 and 72 hours in the superficial skin layer (p ≤ 0.024). Supporting drug delivery findings, OCT revealed that increases in local blood flow after AFL were mitigated in test areas also exposed to PDL, brimonidine, or epinephrine, with PDL providing the greatest, sustained reduction in flow over 48 hours. CONCLUSION: Vasoregulative intervention in conjunction with AFL-assisted delivery enhances and prolongs 5-FU deposition in in vivo skin.

Hyperbaric phototherapy augments blood carbon monoxide removal.

BACKGROUND AND OBJECTIVES: Carbon monoxide (CO) poisoning is responsible for nearly 50,000 emergency department visits and 1200 deaths per year. Compared to oxygen, CO has a 250-fold higher affinity for hemoglobin (Hb), resulting in the displacement of oxygen from Hb and impaired oxygen delivery to tissues. Optimal treatment of CO-poisoned patients involves the administration of hyperbaric 100% oxygen to remove CO from Hb and to restore oxygen delivery. However, hyperbaric chambers are not widely available and this treatment requires transporting a CO-poisoned patient to a specialized center, which can result in delayed treatment. Visible light is known to dissociate CO from carboxyhemoglobin (COHb). In a previous study, we showed that a system composed of six photo-extracorporeal membrane oxygenation (ECMO) devices efficiently removes CO from a large animal with CO poisoning. In this study, we tested the hypothesis that the application of hyperbaric oxygen to the photo-ECMO device would further increase the rate of CO elimination. STUDY DESIGN/MATERIAL AND METHODS: We developed a hyperbaric photo-ECMO device and assessed the ability of the device to remove CO from CO-poisoned human blood. We combined four devices into a "hyperbaric photo-ECMO system" and compared its ability to remove CO to our previously described photo-ECMO system, which was composed of six devices ventilated with normobaric oxygen. RESULTS: Under normobaric conditions, an increase in oxygen concentration from 21% to 100% significantly increased CO elimination from CO-poisoned blood after a single pass through the device. Increased oxygen pressure within the photo-ECMO device was associated with higher exiting blood PO2 levels and increased CO elimination. The system of four hyperbaric photo-ECMO devices removed CO from 1 L of CO-poisoned blood as quickly as the original, normobaric photo-ECMO system composed of six devices. CONCLUSION: This study demonstrates the feasibility and efficacy of using a hyperbaric photo-ECMO system to increase the rate of CO elimination from CO-poisoned blood. This technology could provide a simple portable emergency device and facilitate immediate treatment of CO-poisoned patients at or near the site of injury.

Veno-venous extracorporeal blood phototherapy increases the rate of carbon monoxide (CO) elimination in CO-poisoned pigs.

BACKGROUND AND OBJECTIVES: Carbon monoxide (CO) inhalation is the leading cause of poison-related deaths in the United States. CO binds to hemoglobin (Hb), displaces oxygen, and reduces oxygen delivery to tissues. The optimal treatment for CO poisoning in patients with normal lung function is the administration of hyperbaric oxygen (HBO). However, hyperbaric chambers are only available in medical centers with specialized equipment, resulting in delayed therapy. Visible light dissociates CO from Hb with minimal effect on oxygen binding. In a previous study, we combined a membrane oxygenator with phototherapy at 623 nm to produce a "mini" photo-ECMO (extracorporeal membrane oxygenation) device, which improved CO elimination and survival in CO-poisoned rats. The objective of this study was to develop a larger photo-ECMO device ("maxi" photo-ECMO) and to test its ability to remove CO from a porcine model of CO poisoning. STUDY DESIGN/MATERIALS AND METHODS: The "maxi" photo-ECMO device and the photo-ECMO system (six maxi photo-ECMO devices assembled in parallel), were tested in an in vitro circuit of CO poisoning. To assess the ability of the photo-ECMO device and the photo-ECMO system to remove CO from CO-poisoned blood in vitro, the half-life of COHb (COHb-t1/2 ), as well as the percent COHb reduction in a single blood pass through the device, were assessed. In the in vivo studies, we assessed the COHb-t1/2 in a CO-poisoned pig under three conditions: (1) While the pig breathed 100% oxygen through the endotracheal tube; (2) while the pig was connected to the photo-ECMO system with no light exposure; and (3) while the pig was connected to the photo-ECMO system, which was exposed to red light. RESULTS: The photo-ECMO device was able to fully oxygenate the blood after a single pass through the device. Compared to ventilation with 100% oxygen alone, illumination with red light together with 100% oxygen was twice as efficient in removing CO from blood. Changes in gas flow rates did not alter CO elimination in one pass through the device. Increases in irradiance up to 214 mW/cm2 were associated with an increased rate of CO elimination. The photo-ECMO device was effective over a range of blood flow rates and with higher blood flow rates, more CO was eliminated. A photo-ECMO system composed of six photo-ECMO devices removed CO faster from CO-poisoned blood than a single photo-ECMO device. In a CO-poisoned pig, the photo-ECMO system increased the rate of CO elimination without significantly increasing the animal's body temperature or causing hemodynamic instability. CONCLUSION: In this study, we developed a photo-ECMO system and demonstrated its ability to remove CO from CO-poisoned 45-kg pigs. Technical modifications of the photo-ECMO system, including the development of a compact, portable device, will permit treatment of patients with CO poisoning at the scene of their poisoning, during transit to a local emergency room, and in hospitals that lack HBO facilities.

Selective Cryolysis of Melanocytes: Critical Temperature and Exposure Time to Induce Selective Pigmentary Loss in Yucatan Pig Skin.

BACKGROUND AND OBJECTIVES: Cryotherapy for melanocytic lesions is often accompanied by collateral damage to the surrounding skin, resulting in skin necrosis and scarring. Adipocytes, like melanocytes, are neural crest-derived cells. Adipocytes have been shown to be more sensitive to cold exposure than their neighboring cells of ectodermal origin, such as epidermal keratinocytes. Such differential sensitivity to cold exposure has led to the development of novel treatment modalities, like cryolipolysis, to selectively target a cell type while sparing neighboring cells. STUDY DESIGN/MATERIALS AND METHODS: In this study, we investigated the roles of controlled skin freezing, tissue temperature, and exposure time in inducing selective loss of melanocytes and skin depigmentation in swines. RESULTS: The results of our study demonstrated that contact cooling of the skin surface causes selective loss of epidermal melanocytes when the tissue temperature reaches -7.5°C or cooler with an exposure time of 10 minutes or longer, leading to partial skin depigmentation in swine skin. Longer exposures combined with colder temperature exposure led to more complete depigmentation in the treated skin surface. CONCLUSION: Cold-sensitivity of melanocytes can be harnessed to selectively remove melanocytes while sparing surrounding keratinocytes. The results from this study demonstrated that improved clinical treatments specifically targeting melanocytic lesions is possible using skin cooling to achieve tissue temperatures capable of inducing selective loss of melanocytes without skin necrosis or scarring. Additional studies are needed to optimize the treatment conditions to prolong the selective removal of melanocytes. Lasers Surg. Med. © 2020 Wiley Periodicals LLC.

Construction and validation of UV-C decontamination cabinets for filtering facepiece respirators.

We present evidence-based design principles for three different UV-C based decontamination systems for N95 filtering facepiece respirators (FFRs) within the context of the SARS-CoV-2 outbreak of 2019-2020. The approaches used here were created with consideration for the needs of low- and middle-income countries (LMICs) and other under-resourced facilities. As such, a particular emphasis is placed on providing cost-effective solutions that can be implemented in short order using generally available components and subsystems. We discuss three optical designs for decontamination chambers, describe experiments verifying design parameters, validate the efficacy of the decontamination for two commonly used N95 FFRs (3M, #1860 and Gerson #1730), and run mechanical and filtration tests that support FFR reuse for at least five decontamination cycles.

Significant skin-tightening by closure of fractional ablative laser holes.

BACKGROUND AND OBJECTIVE: Ablative fractional laser treatment uses thousands of very small laser beam wounds to damage a fraction of the skin, which stimulates tissue remodeling. Each open micro-wound heals without scarring, but the amount of skin tightening achieved is limited. This animal study was performed to test the hypothesis that immediate temporary closure of fractional laser wounds could increase skin tightening after fractional ablative laser treatment. MATERIALS AND METHODS: Four adult swine were used for the study; 98 square test sites (3 × 3 cm) were tattooed on the abdomen and flanks of each pig. An ablative fractional Erbium:YAG laser (Sciton Profile, Sciton Inc, Palo Alto, CA) was used to treat the test areas. A laser micro-spot fluence of 375 J/cm2 was delivered in 150-250 microseconds pulses, resulting in an array of ablation channels extending 1.5 mm deep into the skin, with a spot size of 250 µm, with 10% treatment density. Immediately following laser exposure the resulting holes were closed using a stretched elastic adhesive dressing, which, when applied, recoiled and compressed the diameter of the ablation holes. The compressive dressings were removed after 7 days. This procedure was compared to removing the same amount of skin (10%) mechanically by specially designed 19 gauge coring needles, as well as to the same laser and coring methods without compression closure. Area and shape of test sites were measured by digital photography before and 28 days after treatment. Data analysis included compensation for animal growth, as measured by increase in the area of the untreated control sites. RESULTS: All treated and control sites healed within a week, without scarring evident at 28 days. Laser treatment combined with compressive wound closure caused significant shrinkage at 28 days compared with untreated control sites. The treated skin area was reduced by 11.5% (P = 0.0001). Needle coring with wound closure produced similar, significant shrinkage (8%, P < 0.0021), whereas laser and needle coring treatment without closure did not result in significant area reduction (P = 0.1289) compared with untreated control sites. CONCLUSION: Significant skin tightening can be achieved by immediate temporary non-invasive wound closure after short pulse Er:YAG fractional ablative laser treatment, as well as after mechanically removing skin with a coring needle. This approach may improve skin tightening after ablative laser treatments. Further clinical studies are necessary to confirm successful application in humans. Lasers Surg. Med. 50:64-69, 2018. © 2017 Wiley Periodicals, Inc.

Pulmonary Phototherapy for Treating Carbon Monoxide Poisoning.

RATIONALE: Carbon monoxide (CO) exposure is a leading cause of poison-related mortality. CO binds to Hb, forming carboxyhemoglobin (COHb), and produces tissue damage. Treatment of CO poisoning requires rapid removal of CO and restoration of oxygen delivery. Visible light is known to effectively dissociate CO from Hb, with a single photon dissociating one CO molecule. OBJECTIVES: To determine whether illumination of the lungs of CO-poisoned mice causes dissociation of COHb from blood transiting the lungs, releasing CO into alveoli and thereby enhancing the rate of CO elimination. METHODS: We developed a model of CO poisoning in anesthetized and mechanically ventilated mice to assess the effects of direct lung illumination (phototherapy) on the CO elimination rate. Light at wavelengths between 532 and 690 nm was tested. The effect of lung phototherapy administered during CO poisoning was also studied. To avoid a thoracotomy, we assessed the effect of lung phototherapy delivered to murine lungs via an optical fiber placed in the esophagus. MEASUREMENTS AND MAIN RESULTS: In CO-poisoned mice, phototherapy of exposed lungs at 532, 570, 592, and 628 nm dissociated CO from Hb and doubled the CO elimination rate. Phototherapy administered during severe CO poisoning limited the blood COHb increase and improved the survival rate. Noninvasive transesophageal phototherapy delivered to murine lungs via an optical fiber increased the rate of CO elimination while avoiding a thoracotomy. CONCLUSIONS: Future development and scaling up of lung phototherapy for patients with CO exposure may provide a significant advance for treating and preventing CO poisoning.

Fractional laser-assisted drug delivery: Active filling of laser channels with pressure and vacuum alteration.

BACKGROUND AND OBJECTIVE: Ablative fractional laser (AFXL) is rapidly evolving as one of the foremost techniques for cutaneous drug delivery. While AFXL has effectively improved topical drug-induced clearance rates of actinic keratosis, treatment of basal cell carcinomas (BCCs) has been challenging, potentially due to insufficient drug uptake in deeper skin layers. This study sought to investigate a standardized method to actively fill laser-generated channels by altering pressure, vacuum, and pressure (PVP), enquiring its effect on (i) relative filling of individual laser channels; (ii) cutaneous deposition and delivery kinetics; (iii) biodistribution and diffusion pattern, estimated by mathematical simulation. METHODS: Franz diffusion chambers (FCs) were used to evaluate the PVP-technique, comparing passive (AFXL) and active (AFXL + PVP) channel filling. A fractional CO2-laser generated superficial (225 µm;17.5 mJ/channel) and deep (1200 µm; 130.5 mJ/channel) channels, and PVP was delivered as a 3-minutes cycle of 1 minute pressure (+1.0 atm), 1 minute vacuum (-1.0 atm), and 1 minute pressure (+1.0 atm). Filling of laser channels was visualized with a colored biomarker liquid (n = 12 FCs, n = 588 channels). Nuclear magnetic resonance quantified intracutaneous deposition of topically applied polyethylene glycol (PEG400) over time (10 minutes, 1 hour, and 4 hours), investigated with (n = 36 FCs) and without (n = 30 FCs) PVP-filling. Two-dimensional mathematical simulation was used to simulate intradermal biodistribution and diffusion at a depth of 1,000 µm. RESULTS: Active filling with application of PVP increased the number of filled laser channels. At a depth of 1,000 µm, filling increased from 44% (AFXL) to 94% with one PVP cycle (AFXL + PVP; P < 0.01). Active filling greatly enhanced intracutaneous deposition of PEG400, resulting in a rapid delivery six-folding uptake at 10 minutes (AFXL 54 µg/ml vs. AFXL + PVP 303 µg/ml, P < 0.01). AFXL alone generated an inhomogeneous uptake of PEG400, which greatly improved with active filling, resulting in a uniform uptake within the entire tissue. CONCLUSION: Active filling with PVP secures filling of laser channels and induces a deeper, greater, more rapid delivery than conventional AFXL. This delivery technique has promise to improve treatment efficacy for medical treatments of dermally invasive lesions, such as BCCs.

Micro-fractional, directional skin tightening: A porcine model.

BACKGROUND AND OBJECTIVE: Skin changes are among the most visible signs of aging. Fractional ablative lasers improve skin quality by making small skin wounds that heal rapidly without scarring. While they improve skin texture and discoloration, there is minimal effect on skin laxity. This study was performed to assess skin shrinkage performed by removing multiple small full-thickness skin columns with coring needles combined with wound closure. MATERIALS AND METHODS: In 5 swine 116 squares (3 cm(2) ) were demarcated for treatment and control sites. In treatment sites 10% of the skin was removed by full-thickness skin coring needles (19 gauge) and afterwards closed and compressed with an elastic adhesive dressing. This procedure was compared to puncturing the skin with standard hypodermic needles (without tissue removal) and subsequent closure with compressive dressing. Area and shape of sites were measured before and 28 days after treatment. RESULTS: Test and control sites healed within a week without scarring. Coring with wound closure caused significant shrinkage after 28 days. The treated skin area was reduced by 9% (P < 0.0001) and the direction of shrinkage was influenced by the direction of wound closure. Coring without wound closure and puncturing the skin without tissue removal produced an insignificant 3% decrease in area. CONCLUSION: Significant minimally invasive skin tightening in a preferred direction can be achieved by removing skin with coring needles followed by wound closure. The direction of shrinkage is influenced by the direction of micro-hole closure, irrespective of the skin tension lines. This approach may allow reshaping the skin in a desired direction without scarring.

Light-activated sealing of skin wounds.

BACKGROUND AND OBJECTIVES: We have developed a light-activated technology for rapidly sealing skin surgical wounds called photochemical tissue bonding (PTB). The goals of this study were to evaluate parameters influencing PTB in order to optimize its clinical efficacy and to determine whether PTB can be used to seal wounds in moderately to highly pigmented skin. STUDY DESIGN/MATERIALS AND METHODS: Application of Rose Bengal (RB) followed by exposure to 532 nm was used to seal linear incisions (1.5 mm deep, 2 cm long) in lightly pigmented (Yorkshire) and darkly pigmented (Yucatan) swine skin. The force required to open the seal (the bonding strength) was measured by in situ tensiometry. Reflectance spectra, epidermal transmission spectra, and histology were used to characterize the skin. The relationships of RB concentration and fluence to bonding strength were established in Yorkshire skin. Surface temperature was measured during irradiations and cooling was used while sealing incisions in Yucatan skin. Monte Carlo simulations were carried out to estimate the effect of epidermal melanin on the power absorbed in the dermis at the incision interface. RESULTS: The lowest fluence, 25 J/cm(2), delivered at an irradiance of 0.5 W/cm(2) substantially increased the bonding strength (∼ 10-fold) compared to controls in Yorkshire swine skin. Increasing the fluence to 100 J/cm(2) enhanced bonding strength by a further 1.5-fold. Application of 0.1% RB for 2 minutes produced the greatest bonding strength using 100 J/cm(2) and limited the penetration of RB to an ∼ 50 µm band on the dermal incision wall. Reflectance spectra indicated that Yorkshire skin had minimal melanin and that Yucatan skin was a good model for highly pigmented human skin. In Yucatan skin, the bonding strength increased 1.7-fold using 0.1% RB and 200 J/cm(2) at 1.5 W/cm(2) with cooling and epinephrine. Monte Carlo simulation indicated that absorption of 532 nm light by epidermal melanin in dark skin decreased the power absorbed along the incision in the dermis by a factor of 2.7. CONCLUSIONS: These results suggest that in lightly pigmented skin the PTB treatment time can be shortened without compromising the bonding strength. Sealing incisions using PTB in moderately and highly pigmented skin will require a careful balance of irradiance and cooling.

Use of Photodynamic Therapy and Sterile Water to Target Adipose Tissue.

BACKGROUND: Neither photodynamic therapy (PDT) nor sterile water has not been well studied for the treatment of adipose tissue. OBJECTIVE: This investigation studied 2 different modalities, verteporfin PDT and sterile water, on adipose tissue compared with control. MATERIALS AND METHODS: Four light-skinned pigs were used. Test sites received verteporfin PDT or sterile water injection. Control sites received injection of verteporfin without PDT, normal saline injection, no intervention, exposure to laser only, or insertion of a needle or cannula only. Sites were evaluated clinically, by ultrasound, and with histology 4 to 6 weeks after treatment. RESULTS: There was a decrease in adipose tissue by ultrasound after verteporfin PDT (15%, p < .001) and sterile water (2%, p = .23). Verteporfin without PDT showed a decrease in adipose tissue (17%, p = .21). All other control sites showed an increase in adipose tissue. Histologically, verteporfin PDT and sterile water showed moderate damage (median Grade 2, p < .001) 4 to 6 weeks after intervention. CONCLUSION: Verteporfin decreased adipose tissue after treatment. Sterile water injection had a statistically significant effect on adipose tissue histologically but did not substantially decrease the adipose tissue by ultrasound 4 to 6 weeks after intervention. Longer follow-up may be needed.

Pretreatment with ablative fractional laser changes kinetics and biodistribution of topical 5-aminolevulinic acid (ALA) and methyl aminolevulinate (MAL).

BACKGROUND AND OBJECTIVES: 5-Aminolevulinic acid (ALA) and methyl aminolevulinate (MAL) are porphyrin precursors used topically for photodynamic therapy (PDT). Previous studies have established that ablative fractional laser (AFXL) increases topical drug uptake. We evaluated kinetics and biodistribution of ALA- and MAL-induced porphyrins on intact and disrupted skin due to AFXL. MATERIALS AND METHODS: Two Yorkshire swine were exposed to CO2 AFXL (10.6 µm, 1,850 µm ablation depth) and subsequent topical application of ALA and MAL cream formulations (20%, weight/weight). Porphyrin fluorescence was quantified by digital fluorescence photography (30, 90, and 180 minutes) and fluorescence microscopy at specific skin depths (180 minutes). RESULTS: Porphyrins gradually formed over time, differently on intact and AFXL-disrupted skin. On intact skin (no AFXL), fluorescence photography showed that MAL initially induced higher fluorescence than ALA (t = 30 minutes MAL 21.1 vs. ALA 7.7 au, t = 90 minutes MAL 39.0 vs. ALA 26.6 (P < 0.009)) but reached similar intensities for long-term applications (t = 180 minutes MAL 56.6 vs. ALA 52 au, P = ns). AFXL considerably enhanced porphyrin fluorescence from both photosensitizers (P < 0.05). On AFXL-exposed skin, MAL expressed higher fluorescence than ALA for short-term application (t = 30 minutes, AFXL-MAL 26.4 vs. AFXL-ALA 14.1 au, P < 0.001), whereas ALA over time overcame MAL and induced the highest fluorescence intensities obtained (t = 180 minutes, AFXL-MAL 98.6 vs. AFXL-ALA 112.0 au, P < 0.001). In deep skin layers, fluorescence microscopy showed higher fluorescence in hair follicle epithelium for ALA than MAL (t = 180 minutes, 1.8 mm, AFXL-MAL 35.3 vs. AFXL-ALA 46.7 au, P < 0.05). CONCLUSIONS: AFXL changes kinetics and biodistribution of ALA and MAL. It appears that AFXL-ALA favors targeting deep structures.

Laser Ablation Facilitates Implantation of Dynamic Self-Regenerating Cartilage for Articular Cartilage Regeneration.

OBJECTIVES: This study investigated a novel strategy for improving regenerative cartilage outcomes. It combines fractional laser treatment with the implantation of neocartilage generated from autologous dynamic Self-Regenerating Cartilage (dSRC). METHODS: dSRC was generated in vitro from harvested autologous swine chondrocytes. Culture was performed for 2, 4, 8, 10, and 12 weeks to study matrix maturation. Matrix formation and implant integration were also studied in vitro in swine cartilage discs using dSRC or cultured chondrocytes injected into CO2 laser-ablated or mechanically punched holes. Cartilage discs were cultured for up to 8 weeks, harvested, and evaluated histologically and immunohistochemically. RESULTS: The dSRC matrix was injectable by week 2, and matrices grew larger and more solid with time, generating a contiguous neocartilage matrix by week 8. Hypercellular density in dSRC at week 2 decreased over time and approached that of native cartilage by week 8. All dSRC groups exhibited high glycosaminoglycan (GAG) production, and immunohistochemical staining confirmed that the matrix was typical of normal hyaline cartilage, being rich in collagen type II. After 8 weeks in cartilage lesions in vitro, dSRC constructs generated a contiguous cartilage matrix, while isolated cultured chondrocytes exhibited only a sparse pericellular matrix. dSRC-treated lesions exhibited high GAG production compared to those treated with isolated chondrocytes. CONCLUSIONS: Isolated dSRC exhibits hyaline cartilage formation, matures over time, and generates contiguous articular cartilage matrix in fractional laser-created microenvironments in vitro, being well integrated with native cartilage.

Vitamin K3 (Menadione) is a multifunctional microbicide acting as a photosensitizer and synergizing with blue light to kill drug-resistant bacteria in biofilms.

Cutaneous bacterial wound infections typically involve gram-positive cocci such as Staphylococcus aureus (SA) and usually become biofilm infections. Bacteria in biofilms may be 100-1000-fold more resistant to an antibiotic than the clinical laboratory minimal inhibitory concentration (MIC) for that antibiotic, contributing to antimicrobial resistance (AMR). AMR is a growing global threat to humanity. One pathogen-antibiotic resistant combination, methicillin-resistant SA (MRSA) caused more deaths globally than any other such combination in a recent worldwide statistical review. Many wound infections are accessible to light. Antimicrobial phototherapy, and particularly antimicrobial blue light therapy (aBL) is an innovative non-antibiotic approach often overlooked as a possible alternative or adjunctive therapy to reduce antibiotic use. We therefore focused on aBL treatment of biofilm infections, especially MRSA, focusing on in vitro and ex vivo porcine skin models of bacterial biofilm infections. Since aBL is microbicidal through the generation of reactive oxygen species (ROS), we hypothesized that menadione (Vitamin K3), a multifunctional ROS generator, might enhance aBL. Our studies suggest that menadione can synergize with aBL to increase both ROS and microbicidal effects, acting as a photosensitizer as well as an ROS recycler in the treatment of biofilm infections. Vitamin K3/menadione has been given orally and intravenously worldwide to thousands of patients. We conclude that menadione/Vitamin K3 can be used as an adjunct to antimicrobial blue light therapy, increasing the effectiveness of this modality in the treatment of biofilm infections, thereby presenting a potential alternative to antibiotic therapy, to which biofilm infections are so resistant.

An antimicrobial blue light device to manage infection at the skin-implant interface of percutaneous osseointegrated implants.

Antimicrobial blue light (aBL) is an attractive option for managing biofilm burden at the skin-implant interface of percutaneous osseointegrated (OI) implants. However, marketed aBL devices have both structural and optical limitations that prevent them from being used in an OI implant environment. They must be handheld, preventing even irradiation of the entire skin-implant interface, and the devices do not offer sufficient optical power outputs required to kill biofilms. We present the developmental process of a unique aBL device that overcomes these limitations. Four prototypes are detailed, each being a progressive improvement from the previous iteration as we move from proof-of-concept to in vivo application. Design features focused on a cooling system, LED orientation, modularity, and "sheep-proofing". The final prototype was tested in an in vivo OI implant sheep model, demonstrating that it was structurally and optically adequate to address biofilm burdens at the skin-implant of percutaneous OI implants. The device made it possible to test aBL in the unique OI implant environment and compare its efficacy to clinical antibiotics-data which had not before been achievable. It has provided insight into whether or not continued pursual of light therapy research for OI implants, and other percutaneous devices, is worthwhile. However, the device has drawbacks concerning the cooling system, complexity, and size if it is to be translated to human clinical trials. Overall, we successfully developed a device to test aBL therapy for patients with OI implants and helped progress understanding in the field of infection management strategies.