DOI: 10.1002/lsm.23536
A B S T R A C T
Abstracts
B a s i c S c i e n c e –Animal‐Based P r e c l i n i c a l S t u d i e s
ABLATIVE FRACTIONAL LASER EXPOSURE AS AN ADJUVANT FOR IMMUNE CHECKPOINT THERAPY OF COLON CANCER IN A MICE MODEL
Masayoshi Kawakubo, Dieter MansteinCutaneous Biology Research Center, Massachusetts General Hospital, Boston, MA, USA
Background: Currently ablative fractional photother- molysis (aFP) with CO2laser is used for a wide variety of dermatological indications. This study demonstrates the utility of aFP for oncological indications.
Study Design/Materials and Method: We used a fractional CO2 laser and checkpoint molecules such as anti‐PD‐1 inhibitor and OX40 agonist for a treatment of a tumor established bilaterally by the CT26 wild type (CT26WT) murine colon carcinoma cell line, treating tumors on one‐side and observing both tumors. We made eight groups such as control, PD‐1, OX40, PD‐1 + OX40, aFP only, aFP + PD‐1, aFP + OX40, and aFP + PD‐1 + OX40 groups. Each group contains 12 mice.
Results: Interestingly, all tumors in triple treat- ments such as aFP + anti‐PD‐1 + OX40 agonist group grew significantly slower and were cured completely on both aFP‐treated and untreated sides. Flow cytometric analysis showed the triple treatments elicited an increase of granzyme B+ CD8+ T cells in untreated contralateral tumor due to synergistic effect of aFP treatment and the checkpoint molecules, including induction of dendritic cells (DCs) in aFP‐ treated tumor and in drainage lymph node, and OX40+ CD8+ T cells in the lymph node. When anti‐ CD8 antibody was used for ablation of the CD8+ T cells, triple therapy‐mediated tumor regression and survival was abrogated, meaning CD8+ T cells play critical role for therapeutic benefit.
Conclusion: We have demonstrated that aFP is able to augment the systemic antitumor adaptive immunity induced by checkpoint molecules such as anti‐PD‐1 inhibitor and OX40 agonist. This study demonstrated a potential of adjuvant aFP treatments in combina- tion with the immunotherapy for optimal cancer therapy.
A B L A T I V E F R A C T I O N A L L A S E R T R E A T M E N T O F B A S A L C E L L C A R C I N O M A I N C R E A S E S E A R L Y I M M U N E I N F I L T R A T I O N A N D P O L A R I Z E S M A C R O P H A G E S T O W A R D S A W O U N D ‐HEALING P H E N O T Y P E
Martin Wiinberg, Ditte E. Jæhger, Thomas L. Andresen, Merete Haedersdal, Uffe H. Olesen
Technical University of Denmark, Kongens Lyngby, Denmark; Department of Dermatology and Wound Healing Centre, Copenhagen University Hospital, Bispebjerg, Copenhagen, Denmark
Background: Ablative fractional laser (AFL) treatment shows therapeutic potential in various murine cancer models. Our study explores the early innate immune response following AFL treatment in a clinically‐relevant basal cell carcinoma (BCC) model.
Study Design/Materials and Method: Ptch1+/– K14‐ CreER2 p53fl/fl‐mice were injected with tamoxifen to delete p53 (tumor suppressor gene) in basal cells of the hair follicles to initialize the BCC tumor formation. Mice were subsequently whole‐body irradiated to further enhance the tumor formation process. BCC tumors became visible 2–4 months after induction. Mice (n = 12) were randomized into two groups—an untreated group (total of 15 tumors) and an AFL group (total of 13 tumors). AFL‐group received a single CO2 laser dose (100 mJ/microbeam, 5% density covering the tumor area) on every tumor. Tumors were excised 5 days after treatment for ex vivoflow cytometry analysis. Data were analyzed blinded.
Results: Flow cytometry analysis of BCC tumors showed twofold higher immune infiltration in the AFL‐
treated group than in the untreated group (p = 0.0063).
AFL‐treated tumors displayed a sixfold increase in neutrophils (defined as CD11b + F4/80‐Ly6G+) per gram tissue compared to untreated tumors (p = 0.0065).
No difference in the number of macrophages (defined as CD11b + F4/80 + Ly6G−) was observed between groups (p = 0.1491). However, AFL treatment induced a change in macrophage phenotype markers with an increase in the percentage of Arginase‐1+ macrophages (p = 0.0006) and lowered MHC class II signal on macrophages (p = 0.0106), both characteristics of a wound‐healing phenotype.
Lasers Surg Med. 2022;54:S5–S112. wileyonlinelibrary.com/journal/lsm © 2022 Wiley Periodicals LLC
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S5Conclusion: Overall, AFL treatment results in a significant early immune infiltration of tumors with abundant neutrophil infiltration. Specifically, we observed a change in macrophages towards a wound‐healing phenotype after AFL treatment. The results suggest that the early immune response in BCC tumors to AFL may be a combination of an innate inflammatory response and an anti‐inflammatory wound‐healing response.
C O M B I N A T I O N T H E R A P Y F O R T R E A T M E N T O F S P O N T A N E O U S C H O R O I D A L
N E O V A S C U L A R I Z A T I O N I N M I C E
Rupesh Singh, Bela Anand‐Apte, Zoe MillerCleveland Clinic Foundation, Cleveland, OH, USA Background: Choroidal neovascularization (CNV), a characteristic of neovascular age‐related macular degeneration (nAMD), is a vision‐threatening disease of the elderly. In this study, we examined a nanomedicine
—NIR laser combination therapy for treating CNV in a mouse model of nAMD.
Study Design/Materials and Method: Eight 1‐month‐
old spontaneous choroidal neovascularization (sCNV) transgenic mice (JR5558) were used in this study. The sCNV mice develop CNV lesions in the retina/choroid spontane- ously between P10‐15. The animals were divided into two groups, treatment and control. The treatment group received a single tail vein injection of 50 µl Indocyanine green (ICG)‐ attached gold nanorods solution followed by 810 nm laser exposure for three mins (power: 40 mW, beam diameter:
2 mm). Fluorescein angiography (FA) using confocal scanning laser ophthalmoscopy (cSLO) (Heidelberg SPEC- TRALIS; Heidelberg Engineering) was performed on Days 1, 7, and 10 with intraperitoneal injection of 40 µl sodium fluorescein following treatment. The FA images were taken at ~3 and ~6 minutes postinjection for early and late phase circulation with a 55o retina lens. The FA images were quantified by measuring the lesion area with built‐in software. Volumetric optical coherence tomography (OCT) (Envisu R2210 UHR; Leica Microsystems Inc.) was performed in enhanced depth imaging mode at three‐time points (Days 1 and 10). The OCT images were captured using 55o mouse retina lens with 1000 a‐scan × 100 b‐scans × 5‐frame averaging. OCT images were analyzed for cross‐sectional lesion volume.
Results: The CNV lesions in the treatment group showed a significant decrease in dye leakage, as seen in fluorescein angiography images. The OCT imaging revealed faster healing of CNV lesions in the treatment group when compared with untreated control group.
Conclusion: The ICG attached gold nanorods assisted near‐infrared combination therapy has significant therapeu- tic beneficial effects for retinal‐choroidal neovascularization.
The proposed photodynamic‐photothermal nanomedicine based approach is a promising therapeutic for nAMD.
E N H A N C I N G M E T A B O L I S M T H R O U G H S T I M U L A T I O N O F B R O W N I N G O F W H I T E A D I P O S E T I S S U E B Y F R A C T I O N A L
P H O T O T H E R M O L Y S I S
Nunciada Salma, Daniel Karasik, Michael Wang‐Evers, Tuanlian Luo, Dieter Manstein
Cutaneous Biology Research Center, Massachusetts General Hospital, Charlestown, MA, USA
Background: Obesity is a chronic condition thought to be caused by excess energy intake relative to energy expenditure, leading to weight gain and impacting minority populations disproportionately. Brown adipose tissue (BAT) can convert chemical energy to heat through a process of uncoupled respiration (nonshivering thermogenesis), which is mediated by the uncoupling protein 1 (UCP1) expressed in the mitochondria of brown adipocytes. It has been found that defective or insufficient BAT is associated with obesity. Herein, we analyzed levels of brown adipose tissue in mice following ablative fractional photothermolysis treatment.
Study Design/Materials and Method: For our study, 22‐week‐old male C57BL/6J mice underwent exposure to ablative fractional photothermolysis treatment of 30% of their body surface area, being sure to avoid the extremities, head, tail, and ventral surface. After laser exposure, mice were individually housed in cages for 5 days. Levels of norepinephrine in serum and UCP1 by immunohistochemistry of adipose tissue were measured in mice at the end of the experiment.
Results: Serum levels of noradrenaline were signifi- cantly increased in mice following fractional photother- molysis exposure. This sympathetic stimulation was linked with stimulation of browning of white adipocytes, demonstrated by a subsequently elevated expression of UCP1 in laser‐treated compared to control mice.
Conclusion: Given the current knowledge of WAT, BAT, beige fat, and their relation to metabolism, laser treatment is a possible new method of modulating metabo- lism through increasing BAT and/or BAT activity and consequently of treating obesity and metabolic disorders.
I N V I V O D E R M A L D E L I V E R Y O F B L E O M Y C I N W I T H E L E C T R O N I C P N E U M A T I C I N J E C T I O N : D R U G V I S U A L I Z A T I O N A N D
Q U A N T I F I C A T I O N W I T H M A S S S P E C T R O M E T R Y
Liora Bik, Martijn van Doorn, Anders Hansen, Christian Janfelt, Uffe H. Olesen, Merete Haedersdal, Catharina M.
Lerche, Kristoffer Hendel
Erasmus MC, University Medical Center Rotterdam, Rotterdam, Netherlands; University of Copenhagen, Copenhagen, Denmark; Bispebjerg Hospital, University
of Copenhagen, Copenhagen, Denmark; Department of Dermatology and Wound Healing Centre, Copenhagen University Hospital, Bispebjerg, Copenhagen, Denmark;
Bispebjerg University Hospital, Copenhagen, Denmark;
Copenhagen University Hospital, Bispebjerg and Frederiksberg, Copenhagen, Denmark
Background: Intralesional bleomycin (BLM) admin- istration by needle injection is effective for keloids and common warts but has significant drawbacks, including treatment‐related pain and highly operator‐depended success rates. Electronic pneumatic injection (EPI) is a promising, less painful, needle‐free delivery method that potentially enables more precise and controlled dermal drug delivery. Here, we aimed to explore the cutaneous pharmacokinetics, biodistribution patterns and tolerabil- ity of BLM administered by EPI in vivo.
Study Design/Materials and Method: In a pig model, EPI with BLM or saline (SAL) were evaluated after 1, 48, and 216 hours. Mass spectrometry quantification and imaging were used to assess BLM concentrations and biodistribution patterns in skin biopsies. Tolerability was assessed by scoring local skin reactions (LSR) and measuring transepidermal water loss (TEWL).
Results: Directly after BLM injection a peak concen- tration of 109.2 µg/cm3 (43.9–175.2) was measured in skin biopsies. After 9 days BLM was undetectable. EPI resulted in a focal BLM biodistribution in the mid‐ dermal delivery zone resembling a triangular shape. Mild LSRs were resolved spontaneously and TEWL was unaffected.
Conclusion: BLM administered by EPI resulted in quantifiable and focal mid‐dermal distribution of BLM.
The high skin bioavailability holds a great potential for clinical effects and warrants further evaluation in future human studies.
L A S E R T R E A T M E N T A N D A N T I ‐ P D ‐1 IMMUNOTHERAPY
P R O L O N G S U R V I V A L I N A M O U S E M O D E L O F S Q U A M O U S C E L L C A R C I N O M A S
Rikke L. Christensen, Silje H. Omland, Catharina M.
Lerche, Peter A. Philipsen, Uffe H. Olesen, Merete Haedersdal
Bispebjerg University Hospital, Copenhagen, Denmark; Department of Dermatology and Wound Healing Centre, Copenhagen University Hospital, Bispebjerg, Copenhagen, Denmark
Background: PD‐1 checkpoint inhibitors are approved as systemic immunotherapy for advanced and metastatic cutaneous squamous cell carcinoma (SCC). In this explorative study, we aimed to examine the tumor response to treatment with ablative fractional laser (AFL) alone, anti‐PD‐1 antibody alone and the two in combination in an SCC mouse model.
Study Design/Materials and Method: Mice from a hairless SCC mouse model (C3.Cg‐Hrhr/TifBomTac) with ultraviolet radiation‐induced tumors at an advanced stage of 4 mm (Day 0) were divided into four treatment groups: untreated controls (n = 17), AFL alone (n = 15), mouse‐specific anti‐PD‐1‐antibody alone (n = 18) and AFL + anti‐PD‐1‐antibody (n = 15). AFL was given as a single treatment (100 mJ/mb, 5% density) at Day 0, while mice treated with anti‐PD‐1‐antibody were injected 200 µg intraperitoneally at Day 0, 2, 4, 6, and 8 after treatment initiation. The tumors were measured twice a week until they reached the endpoint of 8 mm. Tumor growth rate, tumor clearance and survival curves represented outcome measures. Data for a study of intervention at an earlier tumor stage (2 mm tumors) will be presented.
Results: Treatment of advanced SCC tumors with AFL and anti‐PD‐1‐antibody as monotherapies resulted in tumor clearance of one tumor in each treatment group, whereas three tumors were cleared by the combination treatment. A significantly slower tumor growth rate was observed for all three treatment groups, compared with untreated controls (p = 0.01).
Conclusion: Monotherapy and combination treat- ment with AFL and anti‐PD‐1‐antibody of advanced stage tumors in a murine SCC model resulted in similar tumor responses including increased survival time and slower tumor growth rate.
R E P E A T E D E X P O S U R E S T O
F R A C T I O N A L C O 2 L A S E R R E D U C E S K I N P H O T O D A M A G E A N D
S Q U A M O U S C E L L C A R C O N O M A S I N A P R E C L I N I C A L M O D E L
Uffe H. Olesen, Kevin Jacobsen, Catharina M. Lerche, Merete Haedersdal
Copenhagen University Hospital, Bispebjerg and Frederiksberg, Copenhagen, Denmark
Background: Ablative fractional laser (AFL) exerts an impact on squamous cell carcinoma (SCC) growth. We hypothesized that repeated AFL treatment of ultraviolet radiation (UVR)‐exposed skin could reduce photodam- age and prevent formation of new SCC. Thus, we aimed to investigate the prophylactic potential of AFL in a preclinical UVR‐induced SCC model.
Study Design/Materials and Method: Hairless C3.Cg‐ Hrhr/TifBomTac mice (n = 50) received continued UVR, that is, three standard erythema doses three times per week (from Day 0), and subsequently randomized into two groups (n = 25). Within a dorsal area of 2 × 3 cm, one group served as untreated control and one group received biweekly CO2AFL (10 mJ/mb, 5% density) for a total of 16 weeks (n = 9 treatments) starting at Day 30.
Time to occurrence of first tumor within the treatment area was recorded. In a subset of mice (n = 10),
photodamage was clinically assessed as skin tightness and pigmentation homogeneity at Day 150 (arbitrary scales, 1–5) by three blinded evaluators. Epidermal thickness was evaluated by line‐field optical coherence tomography.
Results: The AFL treated group showed delayed SCC tumor development compared to control mice with a median time to first tumor of 160 days versus 148 days for controls (p = 0.001). Further, AFL increased skin tightness (p = 0.008) and induced a more homogenous pigmentation (p = 0.008) compared to photodamaged control skin. In addition, AFL treated mice displayed thinner epidermis (45 vs. 95 µm, p = 0.016).
Conclusion: In conclusion, repeated AFL treatment of UVR‐exposed skin reduced photodamage substan- tially and significantly delayed SCC tumor formation, which indicates a potential for AFL in prevention of SCC.
T H E E F F E C T S O F I N T E N S E P U L S E D L I G H T A N D A B L A T I V E E R B I U M L A S E R T R E A T M E N T O N T H E E X P R E S S I O N O F
T R A N S F O R M I N G G R O W T H F A C T O R A N D M A T R I X
M E T A L L O P R O T E I N A S E S I N R A B B I T S
Jiayi Feng
Plastic Surgery Hospital, Peking Union Medical College and Chinese Academy of Medical Science, Beijing, China
Background: To investigate the effects of intense pulsed light (IPL) and ablative erbium laser on the expression of transforming growth factor (TGF), matrix metalloproteinases (MMP), and collagen regeneration on dermis of rabbits.
Study Design/Materials and Method: Backs of 10 New Zealand rabbits were divided into four treatment areas:
IPL normal energy group, IPL high energy group, erbium laser normal energy group, and erbium laser high energy group. Hematoxylin and eosin (HE) staining was performed immediately after first treatment and 2 weeks after thefirst treatment. The TGF‐β1 and MMP‐1 immunohistochemical staining was performed on 2 weeks after each treatment and 3 months after the whole treatment monthly.
Results: In HE staining, collagen fibers in the superficial dermis of IPL high energy group and erbium laser group were thickened and arranged tightly. The expression of TGF‐β1 was significantly higher than that of baseline in each group in a dose‐dependent manner.
The peak of MMP‐1 appeared at 2 weeks after the second treatment, and the increase in the erbium laser group was significantly higher than that in the IPL group.
Conclusion: IPL and ablative erbium laser can promote collagen remodeling in a dose‐dependent manner. High energy of ablative erbium laser has the most significant effect on upregulation of TGF‐β1 and downregulation of MMP‐1, suggesting that it has stronger ability to stimulate collagen regeneration and can achieve better results.
B a s i c S c i e n c e –Disinfection
INTELLIGENT UVC DISINFECTION SYSTEM
Sandeep Korupolu, William A. Farinelli, Chhavi Goenka, R. Rox Anderson, Yakir Levin, Martin Purschke
Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Quincy, MA, USA; Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Danvers, MA, USA; Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Cambridge, MA, USA;
Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA; Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Los Angeles, CA, USA
Background: The coronavirus pandemic has height- ened awareness of air and surface disinfection to prevent transmission of infectious disease. While SARS‐CoV‐2 is transmitted primarily through the air, surface transmis- sion does occur. Many other microbial infections are known to be transmitted in this way. Therefore, surface disinfection has become part of cleaning protocols in medical environments and public places. This frequently involves manual cleaning using chemicals and disposable applicators, resulting in labor costs, medical waste, and possible inhalational injury. Ultraviolet‐C light (UVC, 200–280 nm) is known for its antibacterial and antiviral properties. Nucleic acids have an absorption peak around 260 nm; absorption of UVC results in damage to DNA and RNA. Microbial killing is fluence dependent; increasing irradiance decreases time required to achieve sufficient killing.
Study Design/Materials and Method: We built a UVC robot that is capable of efficiently disinfecting vertical and horizontal surfaces in an automated fashion.
By incorporating sensors to minimize the distance between the UVC sources and target surfaces, irradiance is maximized, and decontamination time is minimized.
The robot disinfects horizontal and vertical surfaces in three dimensions. The device is composed of afixed high output vertical lamp, and a horizontal lamp that is capable of motion in two dimensions (up–down and forward–backward) relative to the base. Sensors attached to the base enable the robot to follow a predesignated
path around a room such that the apparatus is brought as close as possible to contaminated surfaces.
Results: For testing, we used bacillus stearothermo- philus spore strips and required a 6‐log kill. Exposure time was varied by adjusting the speed at which the robot traveled. A 6‐log kill was achieved at a speed of 0.125 ft/s.
Conclusion: We built an intelligent UVC device that can disinfect all surfaces in a typical outpatient clinical room very efficiently in 4.5 minutes. The device has potential application in many environments.
B a s i c S c i e n c e –Imaging and D i a g n o s t i c s
CELL ‐ACQUIRING FALLOPIAN ENDOSCOPE FOR DETECTION OF OVARIAN CANCER VIA REFLECTANCE IMAGING, FLUORESCENCE IMAGING, AND CELL COLLECTION
Dominique Galvez, Ricky Cordova, Kelli Kiekens, Andrew Rocha, William Drake, Photini Rice, John Heusinkveld, Jennifer K. Barton
James C. Wyant College of Optical Sciences, University of Arizona, Tucson, AZ, USA; University of Arizona Biomedical Engineering, Tucson, AZ, USA;
University of Arizona, Tucson, AZ, USA
Background: Ovarian cancer is the deadliest gyneco- logical cancer, with a 5‐year survival rate of less than 50%. It is believed that most cases of high‐grade serous ovarian carcinoma originate as serous tubal intraepithe- lial carcinoma (STIC) lesions in the fallopian tube epithelium.
Study Design/Materials and Method: The Cell‐ Acquiring Fallopian Endoscope (CAFE) was developed to optically detect STIC lesions and collect cells for analysis. To help physicians locate potentially pathologi- cal tissue, the CAFE visualizes alterations of the autofluorescence or presence of a targeted fluorescent contrast agent. Upon identification of a potential STIC lesion, a scrape biopsy is used to collect cells from the region of interest. Thefirst generation of CAFE featured a 0.79 mm outer diameter with a pull‐wire for articula- tion of the endoscope tip that could achieve a 25 mm bend radius. The CAFE used multiple laser wavelengths (405, 488, 520, 642 nm) for multispectral fluorescence and white light imaging. Optical fiber with a 100 µm diameter illuminated the tissue, and a 3000‐element fiber bundle collected the light. A quarter‐circle shaped nitinol wire with a 178 µm diameter was spun against the tissue to collect cells.
Results: The CAFE achieved an imaging resolution of 88 µm at a 5 mm distance, and 45° fullfield of view in air.
When tested on ex vivo porcine tissue, hemocytometry
counts demonstrated that on the order of 105cells could be collected per biopsy.
Conclusion: A second‐generation endoscope is in progress with a 10k‐element fiber bundle to increase resolution, a more cost‐effective stainless steel U‐shaped wire to collect cells, and a slightly larger outer diameter of 1.02 mm to accommodate the new components. The second‐generation CAFE has now completed tests in ex vivo porcine and human tissue that demonstrate collected cell counts comparable to those captured with the first‐generation device, as well as measurable improvements in image resolution.
C L I N I C A L I M P L I C A T I O N O F T H E R E G I O N A L T H I C K N E S S O F T H E L O W E R ‐FACIAL SKIN,
S U P E R F I C I A L F A T , A N D S U P E R F I C I A L M U S C U L O ‐ A P O N E U R O T I C S Y S T E M O N H I G H ‐INTENSITY FOCUSED U L T R A S O U N D T R E A T M E N T
Soon‐Hyo Kwon, Chang‐Hun Huh, Jung Won Shin, Jung‐Im Na, Gun Young Ahn
Kyung Hee University, Seongnam, South Korea;
Seoul National University Bundang Hospital, Seong- nam, South Korea; Chungdam Gowoonsesang Derma- tology Clinic, Seoul, South Korea
Background: Knowledge of the topographic thickness of the skin and soft tissues is necessary when performing a high‐intensity focused ultrasound (HIFU) procedure.
Thermal tissue injury to the superficial musculo‐
aponeurotic system (SMAS) or deeper can injure the facial nerve and its branches. The objective of this study is to demonstrate the topographic thickness of the lower facial skin, superficial fat, and SMAS.
Study Design/Materials and Method: The ultrasound data of 200 healthy patients who underwent lower facial rejuvenation were retrospectively reviewed.
Results: The mean age was 41.1 ± 13.7 years (range, 19–76 years). The jowl had thinner skin, thicker superficial fat, and deeper superficial and deep margins of the SMAS than the preauricle or lower cheek. The thickness of the superficial fat decreased with age, especially on the preauricle, lower cheek, and jowl.
Women had thicker superficial fat than men on the preauricle and lower cheek. The superficial and deep margins of the SMAS were located more superficially in old and male patients with a slim facialfigure than in young and female patients with a chubby facial figure.
Conclusion: The presentfindings provide anatomical information regarding the superficial fat and SMAS, which is useful in determining the focal penetration depth of HIFU treatment for lower‐face rejuvenation.
F E A S I B I L I T Y O F N O N ‐IMAGING S E C O N D H A R M O N I C
G E N E R A T I O N M E A S U R E M E N T S T O I D E N T I F Y C O L O N T U M O R T I S S U E
Hasina Shir, Jenna Montague, Dominique Galvez, Travis W. Sawyer, Valentine Nfonsam, Jennifer K.
Barton
University of Arizona Biomedical Engineering, Tucson, AZ, USA; James C. Wyant College of Optical Sciences, University of Arizona, Tucson, AZ, USA;
University of Arizona, Tucson, AZ, USA
Background: Globally, colorectal cancer ranks second in cancer‐related mortalities. Effective screening which detects colon cancer at stage one can lead to a 92% 5‐year survival rate. The most commonly‐used colon cancer screening method is colonoscopy. A current challenge in colonoscopy is differentiating cancerous versus normal or benign tissue.
Study Design/Materials and Method: Collagen is a key structural protein which is altered during cancer devel- opment. It produces a strong second harmonic genera- tion (SHG) signal, and analysis of SHG images shows promise to identify a variety of cancers. However, SHG imaging typically utilizes complex and expensive scan- ning elements that are particularly difficult to implement in flexible endoscopes. This study sought to determine the feasibility of using nonimaging, randomly sampled point intensity SHG measurements to distinguish normal tissue from tumor and tumor‐adjacent tissue. Unstained, formalin‐fixed paraffin‐embedded sections of tumor, normal, and tumor‐adjacent tissue from 12 patients with colorectal cancer were imaged using SHG, a technique in which two photons are absorbed and remitted as one through scattering by nonaxisymmetric molecules. Imag- ing parameters were: 850 nm excitation with constant power, 400–430 nm emission, image size 1024 × 1024 pixels over 425 × 425 µm. Images were intensity thre- sholded to exclude noncollagen signal. The total mean SHG signal from normal samples was significantly greater than tumor‐adjacent samples (p = 0.014) and tumor samples (p = 0.007). Then, image pixels were randomly selected at a logarithmically increasing sample size using MATLAB.
Results: We found that the average intensity of a random sampling of 1000 pixels was not significantly different than the average intensity of the entire 1M+
pixel image.
Conclusion: This finding suggests that a simple continuously sampling, point‐measuring SHG system may provide information equivalent to a scanning system that obtains full images. Therefore, nonimaging, random‐sampling SHG intensity measurements may be sufficient to distinguish colorectal tumor and tumor‐
adjacent from normal tissue.
N O V E L M E T H O D T O E V A L U A T E T H E E F F E C T O F I N T R I N S I C A G I N G A N D S U N E X P O S U R E O N S K I N M O R P H O L O G Y B Y O P T I C A L C O H E R E N C E T O M O G R A P H Y
Vidyalakshmi Anilkumar, Felix Hilge, Michael Wang‐ Evers, Malte Casper, Hinnerk Schulz‐Hildebrandt, Dieter Manstein
Cutaneous Biology Research Center, Massachusetts General Hospital, Boston, MA, USA; Laboratory for Functional Optical Imaging, Columbia University, New York City, NY, USA; University of Lübeck, Lübeck, Germany
Background: Skin aging is caused by intrinsic aging as well as extrinsic factors like exposure to ultraviolet light and is widely known to induce morphological changes to the skin's surface. Although a variety of skin rejuvena- tion products and procedures exist, the evaluation of their endpoints solely relies on the patients' qualitative perception which is highly subjective. For research applications, DIN/ISO parameters Ra (arithmetical mean deviation) and Rmax (maximum peak‐to‐peak height of profile) have been previously used for quantification of the skin's surface morphology, however they neglect the skin's micro‐structure. Hence, they are not suitable for a conclusive assessment. A novel method for evaluating the morphology utilizing the area of the geometric patterns in the micro‐structure was introduced as an alternative to DIN/ISO parameters and yielded promising results. In this study, we evaluate skin aging using additional characteristics of the geometric patterns in the micro‐structure.
Study Design/Materials and Method: The surface data was obtained from the forearms of 14 young (age: 22–27) and 12 elderly (age: 66–78) healthy females (Fitzpatrick type: 1–3) using a commercially available optical coherence tomography (OCT) device. The volumetric OCT data were converted into 2D surface projections after tilt compensation.
Subsequently, the elements of the skin's micro‐ structure were segmented, and the circularity of the polygons was compared with respect to the age of the subject and anatomical sites.
Results: According to our study, the morphology of old cohort subjects significantly differs from that of young cohort subjects, whose micro‐structure is pre- dominated by triangles, whereas quadrilaterals prevail among the old group. Further, the micro‐structure elements of the young group showed a higher circularity compared to the elderly subjects.
Conclusion: We demonstrated that our surface morphology evaluation method is a reliable alternative for traditional roughness parameters. Furthermore, our data suggest it has the capacity to assess the efficacy of cosmetic and dermatological treatments.
U S E O F O P T I C A L C O H E R E N C E T O M O G R A P H Y ( O C T ) I N
A S S E S S M E N T O F D I A B E T I C S K I N W O U N D C H A R A C T E R I S T I C S A N D B L O O D F L O W
Shyon Parsa, Christine E. Wamsley, Paul Kim, Jeffrey M. Kenkel, Yucel Akgul
University of Texas Southwestern, Dallas, TX, USA Background: Optical coherence tomography (OCT) allows for measurements of skin including arithmetic mean roughness (Ra), depth of roughness (Rz), epidermal thickness, dermal‐epidermal junction (DEJ) visualization, attenuation coefficient, and blood flow, but the utility of the device in determining properties of the normal and wounded skin in diabetic patients is unknown.
Study Design/Materials and Method: This single‐site, nonrandomized, noncontrolled, observational study used an OCT device to acquire skin images at 1305 nm, creating real‐time image of patient skin and wounds 1–2 mm under the skin surface. Vertical B‐scan, en‐face, and 3D images were produced to calculate surface and DEJ roughness, the optical attenuation coefficient, a measure of light absorption and scattering, and blood flow metrics. Ra is defined as he mean of the absolute amounts of all variances in the roughness profile. Rz is calculated as the highest absolute change in skin roughness from the mean skin surface.
Results: In subjects with diabetes there was an increase in both the Ra (p.0001) of the wounded versus the control skin. In subjects without diabetes, there was an increase in both the Ra (p = .03) values of the wounded versus the control skin. When comparing blood flow down to a depth of 0.6 mm across all subjects, there was an average decrease in blood flow of 63.47% from control to wounded skin. In subjects with diabetes, this decrease was 75.66%. In the subjects without diabetes the decrease was 54.96%. The change in vasculature between control and wounded skin was associated with a p < 0.005.
Conclusion: The OCT displayed an increase in the Ra and Rz values and an average decrease in blood flow between the wounded skin and the control. The device additionally determined a difference in the Ra and Rz values of both diabetic and healthy subjects' skin and a more pronounced decrease in blood flow in the wounds of patients with diabetes as opposed to those without.
Basic Science –Laser‐Assisted Drug Delivery ADVANCED IMAGING TECHNIQUES FOR VISUALIZATION OF LASER ASSISTED DRUG DELIVERY
Haley Marks, Martin Buta, Joshua Z. Glahn, Neera Nathan, Maryanne Senna, Dieter Manstein
Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
Background: Laser‐assisted drug delivery (LADD) can enhance topical drug uptake by disrupting the stratum corneum, providing microchannels into the dermis surrounded by a coagulation zone and a potential reservoir for sustained drug release. However, few analytical techniques exist to provide accurate and reliable 2D mapping of small molecule drugs in biologi- cal tissue. Herein, we explore advanced lifetime micros- copy and vibrational spectroscopy imaging approaches for visualizing drug distribution in skin.
Study Design/Materials and Method: Using an intrin- sically fluorescent drug, we first evaluated macroscale luminescence photography, traditional intensity‐based fluorescence microscopy, and fluorescence lifetime imag- ing microscopy (FLIM) for their efficacy in visualizing LADD. Second, for topical drug formulations which had negligible intrinsic fluorescence, Raman spectroscopy was used for “chemical fingerprinting” of both the drug and vehicle structures. Formulations were placed over a 1 × 1 cm area of ex vivo human skin after ablative fractional laser treatment, and biopsies taken 24 hours later were cryosectioned and left unstained for imaging.
Results: As FLIM depends on thefluorescence lifetime of the molecule and not the concentration, it was most accurate for isolating the correct region of interest around each laser hole and determining depth of penetration. For quantification, traditional intensity‐based microscopic images are more accurate than unfiltered macroscopic photographs, though photographs are still advantageous as they can be obtained easily at the point‐of‐care. Raman spectroscopy is accurate for isolating regions of drug vs vehicle only, however, long acquisition times resulting in sample burning limit its applicability for whole‐slide imaging.
Conclusion: This study suggests promise for emerging imaging methods for visualizing LADD of topical formulations, however, accurate quantification is still currently limited to intrinsically fluorescent drugs. For nonfluorescent drugs, Raman spectroscopy was success- ful in differentiating between normal tissue and areas containing drug versus vehicle only, but future work is still needed to increase the field of view and reduce aquation times.
C O 2 F R A C T I O N A L A B L A T I V E L A S E R ‐ASSISTED DRUG
D E L I V E R Y O F S Y N T H E T I C A L P H A M E L A N O C Y T E ‐STIMULATING H O R M O N E L E A D S T O I N C R E A S E D P I G M E N T A T I O N A R E A I N A
H Y P E R T R O P H I C S C A R P O R C I N E M O D E L
Bonnie C. Carney, Mary A. Oliver, Sanjana Kurup, John W. Keyloun, Lauren T. Moffatt, Jeffrey W. Shupp, Taryn E. Travis
Firefighters' Burn and Surgical Research Laboratory, MedStar Health Research Institute, Washington, DC, USA; The Burn Center, MedStar Washington Hospital Center, Washington, DC, USA
Background: One symptom of hypertrophic scar (HTS) that can develop after burn injury is dyschromia with hyperpigmentation and hypopigmentation. There are limited treatments for these conditions. Previously, we showed there is no expression of alpha melanocyte‐ stimulating hormone (a‐MSH) in hypopigmented scars, and if these melanocytes are treated with synthetic a‐MSH in vitro, they respond by repigmenting. The current study tested the same hypothesis in the in vivo environment using laser‐assisted drug delivery.
Study Design/Materials and Method: HTSs were created in red Duroc pigs. At Day 70 (Pre), they were treated with CO2 fractional ablative laser (FLSR).
Synthetic a‐MSH was delivered as a topical solution (n = 6). Control scars received no treatment (n = 4). Scars were treated and examined weekly through Week 5.
Digital images and punch biopsies of hyper, hypo, and normally pigmented scar and skin were collected. Digital pictures were analyzed with ImageJ by tracing the area of hyperpigmentation. Epidermal sheets were obtained from punch biopsies through dispase separation and RNA was isolated. Quantitative reverse‐transcription polymerase chain reaction was run for melanogenesis‐ related genes. Two‐way analysis of variance with multiple comparisons and Sidak's correction compared the groups.
Results: The areas of hyperpigmentation were varia- ble before treatment. Therefore, data are represented as fold‐change where each scar was normalized to its own Pre. Hyperpigmented areas gradually increased each week, reaching 1.51‐fold over Pre by Week 5. At each timepoint, change in area of hyperpigmentation was greater in the treated versus the control (1.19 ± 0.03 vs. 1.02 ± 0.08, 1.23 ± 0.06 vs. 1.03 ± 0.03, 1.44 ± 0.08 vs.
1.11 ± 0.03, 1.44 ± 0.09 vs. 1.15 ± 0.05, 1.51 ± 0.11 vs.
1.06 ± 0.07). This difference was significant at Week 3 (p = 0.0001). Expression of melanogenesis genes were increased after treatment.
Conclusion: A clinically‐relevant FLSR treatment method can be combined with topical delivery of synthetic a‐MSH to increase the area of pigmentation in hypo‐pigmented HTS. Future studies will aim to optimize drug delivery timing and dosing.
E N H A N C E D L A S E R ‐ASSISTED D R U G D E L I V E R Y U S I N G A T O P I C A L F O R M U L A T I O N ‐ S A T U R A T E D P R E S S U R E D R E S S I N G
Martin Buta, Haley Marks, Neera Nathan, Branko Bojovic, Dieter Manstein
Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA; Cutaneous Biology Research Center, Massachusetts General Hospital, Charlestown, MA, USA
Background: Laser‐assisted drug delivery (LADD) utilizes an ablative fractional laser (AFL) to produce microchannels in the skin to overcome the epidermal barrier and augment topical drug uptake. New and innovative delivery and analytical techniques are needed to control and evaluate the spatial distribution and concentration of topical drugs within biological tissues. Drug‐saturated pressure dressings have recently shown promise for improving drug uptake after LADD, however, the effect has not yet been qualified. This study investigates the application of a green‐emissive fluores- cent analog‐soaked pressure dressing for visualizing topical formulations applied to laser‐treated skin.
Study Design/Materials and Method: A nonadherent polyester film‐coated bandage was submerged in a solution containing a water‐soluble green‐emissive fluor- ophore suspended in phosphate‐buffered saline buffer.
The soaked bandage was then applied to 1 × 1 cm areas of ex vivo human skin both treated and untreated with a CO2AFL and covered with a transparent, waterproof, sterile film dressing. A 1 × 1 cm area treated with the laser but no topical fluorophore served as a control.
Macroscale luminescence photography was utilized to measure ultraviolet light‐induced fluorescence intensity from the surface of the skin and biopsies were cryosec- tioned for depth of penetration analysis.
Results: Macroscale luminescence photography dem- onstrated significant localization of topical fluorophore within microchannels for samples with and without the pressure dressing. Greater uniformity was observed for those using the pressure dressing, andfluorescence intensity also correlated strongly with depth of drug penetration.
Conclusion: This study demonstrates a feasible method to augment the depth of penetration and establish a correlate for concentration using saturated pressure dressings on AFL‐treated skin. While the use of a saturated pressure dressing allows for more control of drug distribution and relative concentration, additional microscopic techniques are still needed to more accu- rately quantify topical drug delivery and concentration within skin after AFL.
L A S E R ‐ASSISTED TOPICAL D E L I V E R Y O F A N A N T I B O D Y V I Z U A L I Z E D B Y E X V I V O C O N F O C A L M I C R O S C O P Y
Rikke L. Christensen, Vinzent K. Ortner, Merete Haedersdal, Uffe H. Olesen
Department of Dermatology and Wound Healing Centre, Copenhagen University Hospital, Bispebjerg, Copenhagen, Denmark
Background: The skin barrier generally limits the topical delivery of hydrophilic molecules. Ablative fractional laser (AFL) has facilitated cutaneous drug uptake of smaller hydrophilic compounds in several studies. In this descriptive study using ex vivo confocal microscopy, we aimed to evaluate whether ablative fractional CO2 laser at low settings could facilitate the uptake of large hydrophilic compounds such as antibodies.
Study Design/Materials and Method: The hydrophilic molecules ATTO 647N (746 g/mol) and fluorescently labeled antibody nivolumab (150,000 g/mol) were topi- cally applied on intact and CO2AFL (2.5 mJ/mb and 5%
density) treated ex vivo pig skin (n = 3–4 samples per intervention). The molecules were incubated on the skin for 20 hours in a Franz cell setup followed by visualization of the biodistribution in vertical tissue sections by ex vivo confocal microscopy. Optical coher- ence tomography was used to assess laser‐induced skin barrier disruption.
Results: AFL‐induced minimal skin barrier disrup- tion, with laser channels penetrating into epidermis.
Biodistribution of ATTO 647N resulted in a homoge- nous, horizontal band‐like pattern reaching a depth of approximately 1000 µm. Moreover, our study revealed similar delivery of the larger antibody nivolumab, compared with ATTO 647N. Thus, nivolumab also distributed in a band‐like biodistribution pattern reach- ing a comparable tissue depth as ATTO 647N. Neither ATTO 647N nor nivolumab penetrated intact skin.
Conclusion: AFL at low pulse energy and low density facilitates cutaneous delivery of hydrophilic compounds with similar biodistribution for small and large size molecules.
L A S E R ‐ASSISTED TOPICAL D E L I V E R Y O F V I S M O D E G I B R E D U C E S H E D G E H O G
E X P R E S S I O N I N H U M A N B A S A L C E L L C A R C I N O M A S
Kristian Kaaber Pedersen, Katrine Togsverd‐Bo, Edyta Biskup, Gael Clergeaud Veiga, Anni Lisbeth Linnet Nielsen, Thomas L. Andresen, Merete Haedersdal, Uffe H. Olesen
Department of Dermatology and Wound Healing Centre, Copenhagen University Hospital, Bispebjerg, Copenhagen, Denmark; Copenhagen University Hospital—Herlev and Gentofte, Copenhagen, Denmark;
Technical University of Denmark, Kongens Lyngby, Denmark
Background: Oral administration of the hedgehog inhibitor vismodegib is used to treat basal cell carcino- mas (BCC) but limited by frequent systemic adverse effects. Topical uptake of vismodegib is enhanced in pig skin following a combination of ablative fractional laser
(AFL) and vismodegib microemulsion. This open‐label clinical study aimed to investigate the biological response to laser‐assisted topical vismodegib treatment in human BCCs.
Study Design/Materials and Method: Fifteen patients with at least one histologically verified nodular BCC (>7mm) were included. At baseline, we delineated BCCs with optical coherence tomography before taking biopsies. We then exposed BCCs from 12 patients to CO2‐AFL (10600 nm, 40 mJ/beam, 5% density) and vismodegib emulsion (3.8 mg/ml) in custom hydrocolloid wells. After 4 hours, excess emulsion was removed, while wells were left in place until follow‐up on Days 3–4.
At follow‐up, we assessed local skin reactions in all topically treated patients, followed by sampling of tumor biopsies for quantitative polymerase chain reaction (six patients), and serum and tumor biopsies for mass spectrometry detection of vismodegib (12 patients).
Separately, serum samples and tumor biopsies from three patients receiving oral treatment were obtained, likewise for evaluation of vismodegib concentrations.
Results: Messenger RNA expression levels of hedge- hog genes GLI1, GLI2, PTCH1, and PTCH2 were investigated in seven tumors from six patients. Laser‐ assisted topical vismodegib decreased all four genes compared to baseline. GLI1, GLI2, and PTCH2 were significantly reduced with median reductions of 50.7%–72.9% (p = 0.047–0.031), while the median reduc- tion of PTCH1 was nonsignificant; 30.5% (p = 0.265).
The treatment resulted in mild local skin reactions (moderate erythema and mild edema). Intratumoral vismodegib concentrations for topical and oral vismo- degib will also be presented.
Conclusion: AFL‐assisted topical delivery of vismo- degib reduces expression of hedgehog pathway genes in human BCCs following a single application with an acceptable safety profile.
L O C A L V A S O R E G U L A T I V E
I N T E R V E N T I O N S I M P A C T D R U G C O N C E N T R A T I O N S I N S K I N
A F T E R T O P I C A L L A S E R ‐ASSISTED D E L I V E R Y
Emily Wenande, Sarat Chandra Gundavarapu, Joshua Tam, Carina N. Thomas, William A. Farinelli, Benjamin Vakoc, R. Rox Anderson, Merete Haedersdal
Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA; Department of Dermatology and Wound Healing Centre, Copenhagen University Hospital, Bispebjerg, Copenhagen, Denmark
Background: The ability of ablative fractional lasers (AFLs) 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 dwell time in 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.
Study Design/Materials and Method: 5‐FU uptake was assessed in intact‐ and AFL‐exposed skin in a live pig model. After fractional CO2 laser exposure (15 mJ/
mb, 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 hours, 5‐FU concentrations were measured in 500 and 1500 µm skin layers by mass spectrometry (n = 6). Supplemental assessment of blood flow following AFL ± vasoregula- tion 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 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 (i.e., 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 greatest, sustained reduction in flow over 48 hours.
Conclusion: Vasoregulative intervention in conjunc- tion with AFL‐assisted delivery enhances and prolongs 5‐FU deposition in in vivo skin.
B a s i c S c i e n c e –Laser‐Tissue I n t e r a c t i o n
PHOTOTHERMAL LASER
PULSEWIDTHS ARE NOT RELATED TO THEIR TARGET'S THERMAL
RELAXATION TIMES. WHY THIS APPROACH IS INCORRECT
Michael J. Murphy, Per‐Arne TorstenssonDermalase Training Services, Glasgow, United King- dom; Dermalase Training Services, Gothenburg, Sweden Background: Since the inception of selective photo- thermolysis the idea of using a target's thermal relaxation time (TRT—its cooling time) has been used to determine laser energy pulsewidths. This relaxation time is defined as the time taken for the peak elevated temperature to fall to 50% of the peak value and is dependent on the target's dimensions, geometry and the local thermal diffusivity.
Study Design/Materials and Method: The reason any photothermal approach is adopted, when using lasers, is to irreversibly denature the target cell proteins using heat energy. For this process to succeed requires a suitable temperature applied for an appropriate time—the “dena- turation” time. This time does not equal the relaxation time, or the pulsewidth. A computer analysis of the pulsewidth versus TRT versus denaturation time in the laser treatment of blood vessels and hair follicles was carried out to study the processes in detail.
Results: Changing the applied fluence and/or the pulsewidth directly changes the denaturation time, and hence the clinical outcomes, but not change the relaxa- tion times, for any given target.
Conclusion: It is clear that the cooling time, or rate, of the target proteins cannot determine their denaturation time. This report discusses how using TRTs to determine pulsewidths is incorrect and how this should be properly calculated based on the intrinsic thermal response of the proteins.
T H E R M A L E V A L U A T I O N O F A R E D E S I G N E D 1 0 M M
R A D I O F R E Q U E N C Y H A N D P I E C E
Sean T. DohertyBoston Center for Facial Rejuvenation, Boston, MA, USA
Background: Radiofrequency devices are often used noninvasively to heat up the skin to stimulate collagen and rejuvenate the overall appearance. The importance of evaluating a handpiece's capabilities to safely heat the skin while maintaining a therapeutic temperature. Two separate studies were designed to test the new 10 mm radiofrequency handpiece first for its overall safety and then it's overall clinical capabilities.
Study Design/Materials and Method: The first study involved enrolling three subjects treated on both sides of their forehead in separate treatments. Temperature was recorded from both a handpiece reading as well as FLIR thermal imaging camera during the entire testing protocol. Thefirst safety test involved maximizing device settings and leaving the handpiece in one spot on the skin. The second scenario involved subjects being treated as normal until they reached a therapeutic temperature where the treating physician then discontinued moving
the handpiece to confirm the device would not continue heating the skin past a safe temperature. The second study involved similar thermal measurements as six patients were enrolled and treated on both sides of their forehead for a total treatment time of 10 minutes at a therapeutic temperature of at least 40°C.
Results: For the first safety testing scenario, no subject exceeded a maximum temperature of 37.8°C.
For the second safety testing scenario there was a singular test case where the temperature reached 45.6°C but the RF output of the device was already disabled due to lack of handpiece movement. No adverse events were reported across all testing outside of the expected mild erythema immediately after treatment. The second study was able to keep all 12 testing scenarios were able to keep subjects at an average treatment area temperature of at least 41.2°C for a full 10 minutes.
Conclusion: The new 10 mm radiofrequency hand- piece has successfully implemented additional accelerom- eter technology to increase patient safety while still maintaining its ability to reach and maintain clinically efficacious skin temperatures.
B a s i c S c i e n c e –Light Therapy
DESIGN OPTIMIZATION OF A DEVICE FOR EXTRACORPOREAL BLOOD
PHOTOTHERAPY: LIGHT
PROPAGATION AND THERMAL LOAD ANALYSIS
Anastasia Goulopoulos, Dr. Anna Fischbach, Walfre Franco, Edidiong Etim
University of Massachusetts Lowell, Lowell, MA;
Massachusetts General Hospital, Boston, MA, USA Background: The leading cause of poison‐related deaths in the United States is carbon monoxide (CO) inhalation. CO binds to hemoglobin (Hb) and displaces oxygen, reducing oxygen delivery to tissues. The optimal treatment for CO poisoning in patients with normal lung function is administering hyperbaric oxygen. However, hyperbaric chambers are only available in medical centers with specialized equipment, resulting in delayed therapy. Visible light dissociates CO from Hb.
Study Design/Materials and Method: In previous studies, we combined a membrane oxygenator with phototherapy using different wavelengths to design a photo‐ECMO (extracorporeal membrane oxygen- ation) device, which improved CO elimination and survival in CO‐poisoned animal models. This study aims to use computational modeling to optimize the design of the photo‐ECMO device. In particular, the study focuses on 460 (blue), 532 (green), and 620 (red) nm wavelengths.
Results: The deposition of light and the thermal load on blood circulating through the device were calculated
using Monte Carlo and heat convection and diffusion models. All wavelengths effectively dissociate CO from Hb and improve CO elimination. However, light at a wavelength of 620 nm penetrates deeper and generates the lowest thermal load.
Conclusion: Hence, further design optimization of photo‐ECMO devices should consider light at red wavelengths as the optical source for photodissociation.
B a s i c S c i e n c e –Optical Imaging DELINEATING BASAL CELL CARCINOMA USING OPTICAL POLARIZATION IMAGING
Tyler Iorizzo, Peter Jermain, Anna N. Yaroslavsky Advanced Biophotonics Laboratory, University of Massachusetts Lowell, Lowell, MA, USA
Background: Basal cell carcinoma (BCC) is one of the most common forms of human cancer. This type of cancer is curable by surgery if detected and treated early. However, visual contrast of the lesions is often poor, which complicates preoperative tumor delinea- tion and precise excision. We developed Optical Polarization Imager (OPI) that aids in preoperative tumor demarcation by registering tumor‐induced disruption of dermal collagen. The goal of this study was to design a handheld OPI prototype and test it in the pilot trial with Mohs surgery as the gold standard.
Study Design/Materials and Method: The miniatur- ized OPI prototype was designed, constructed, and tested using resolution targets and by imaging normal human skin of healthy volunteers. Then patients scheduled for Mohs Micrographic Surgery were enrolled in the IRB approved pilot clinical study.
The OPI illuminated surgical sites with linearly polarized light. Copolarized light was filtered from light reflected from the skin's surface, allowing cross‐
polarized light to be imaged. Spectrally encoded images were recorded, utilizing 440 nm to image dermal collagen, and 640 or 680 nm for background correction. To determine the accuracy of OPI in determining tumor boundaries, images were com- pared to histopathology processed during Mohs surgeries.
Results: The newly designed handheld device weighed 0.3 kg. It incorporated a light and flexible 3D‐printed spacer design, and a small CMOS camera. The imager provided 32 × 23 mm2field of view, a lateral resolution of 12 µm, and image acquisition time of less than 1 s. Nine subjects with BCCs were enrolled in this study. Tumor boundaries predicted by OPI correlated with the histo- pathology in eight cases. In one case, attenuation of the imaged signal by blood prevented adequate image analysis.
Conclusion: Overall, OPI imaging yielded promising results and demonstrated high potential for preoperative tumor delineation.
D E S I G N A N D D E V E L O P M E N T O F A N E W D E V I C E F O R E S T I M A T I O N O F S K I N P I G M E N T A T I O N
Chhavi Goenka, Karen Kagha, Sandeep Korupolu, William A. Farinelli, Jungyeon You, Sara Moradi Tuchayi, Sidharth Chand, Brian Hibler, Alena Blaise, Savannah Richard, Katherine Vogel, Margaret Funk, R.
Rox Anderson
Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
Background: Accurate estimation of skin melanin content is critical to light‐based healthcare, especially in dermatological care. While the Fitzpatrick scale has been used widely for skin classification, skin color estimation was not its purpose. There is a pressing need for a new scale and associated methodologies that can be used for better skin melanin estimation.
We have developed a new skin classification scale and imaging method with a simple implementation that can be used with any smartphone. We have also developed image analysis algorithms that aid the classification process and reduce human error. In this talk we will discuss the design of the device, the algorithms and future work.
Study Design/Materials and Method: The smartphone serves as our imaging and computational device. A three‐ dimensional (3D) printed structure serves as a mount for the smartphone and the skin melanin scale. The structure is printed using ASA plastic, a commonly used material for 3D printing. We use an off the shelf polarizer to remove specular reflection and the phone's flash as the light source. The device and algorithms have been characterized using standard color swatches and tested through a clinical trial of 40 participants. The measure- ments were verified using the current gold standards for skin color estimation, a reflectance spectrometer and a commercial colorimeter.
Results: The smartphone‐based device, along with the scale and algorithms, was successful in estimating the relative melanin content of test color swatches and various skin colors in characterization tests and in a clinical trial.
Conclusion: In this talk we will present a smart- phone attachment for use in investigations that require accurate skin melanin content estimation. The design greatly simplifies the production and implementation of a skin scale device without compromising perform- ance. The easy reproducibility and simple operation will encourage a broad use of this device in clinical settings.
EFFECTS ON DYNAMIC ‐MICROSCOPIC OCT SIGNALS AFTER MANIPULATION OF KERATINOCYTE METABOLISM IN AN IN ‐VITRO SKIN MODEL BY
TEMPERATURE AND METABOLIC REAGENTS
Felix Hilge, Michael Evers, Reginald Birngruber, Nunciada Salma, Gereon Huettmann, Hinnerk Schulz‐Hildebrandt, Dieter Manstein
Cutaneous Biology Research Center, Massachusetts General Hospital, Boston, MA, USA; University of Lübeck, Luebeck, Germany; Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA
Background: Microscopic optical coherence tomogra- phy (OCT) is a three‐dimensional, noninvasive imaging modality. Utilizing a broadband supercontinuum light source in combination with high numerical aperture optics leads to isotropic resolution of few micrometers.
However, microscopic OCT does not yield (sub‐) cellular contrast since most biological samples express homoge- nous scattering properties within a small imaging area.
Dynamic‐microscopic OCT (dmOCT) is a technique which exploits dynamic changes of the scattering behavior in metabolically active samples. Recording a time series of B‐scans followed by analysis of the intensity fluctuations in the frequency domain, high‐
resolution images with subcellular contrast can be acquired. The underlying cellular processes responsible for those intensity fluctuations and hence the dynamic signals are not fully identified yet.
Study Design/Materials and Method: In this study, we show the effects of different temperatures on dmOCT images of an in vitro human skin model. In addition, we present the response of the samples to two different metabolic reagents, namely carbonyl cyanide‐p‐trifluoro methoxyphenylhydrazone (FCCP) and 2‐deoxy‐D‐glucose (2‐DG), affecting separate pathways of adenosine tri- phosphate (ATP) production, after different time periods of exposure to the substance.
Results: We demonstrate that the medium and high‐ frequency fluctuations decrease in dmOCT signals after the sample was stored in a low‐temperature environment (+4°C) as well as the recovery of these dynamic signals after removal from those conditions. Also, we show differences in the dmOCT signal depending on which ATP synthesis pathway was targeted by the metabolic reagent.
Conclusion: The dependence of the dynamic signals on temperature could indicate metabolic dependency on the dmOCT signal but is also expected by the temperature dependence of diffusive processes. However, changes in the dmOCT signals after affecting the ATP synthesis pathway by the metabolic reagent clearly suggest that the dmOCT signals are influenced by ATP‐dependent pro- cesses. This could allow the analysis of the current metabolic state of the keratinocytes by dmOCT.
VISUALIZATION OF ENERGY ‐BASED DEVICE ‐INDUCED THERMAL TISSUE ALTERATIONS USING BIMODAL EX ‐ VIVO CONFOCAL MICROSCOPY WITH DIGITAL STAINING
Gabriella Fredman, Rikke L. Christensen, Vinzent K.
Ortner, Merete Haedersdal
Department of Dermatology and Wound Healing Centre, Copenhagen University Hospital, Bispebjerg, Copenhagen, Denmark; Bispebjerg University Hospital, Copenhagen, Denmark
Background: Ex‐vivo confocal microscopy (EVCM) enables examination of tissue alterations immediately after treatment with energy‐based devices (EBD). This proof‐of‐concept study aimed to describe EBD‐induced tissue effects in ex vivo porcine skin after treatment with microneedle radiofrequency (MNRF) and ablative frac- tional CO2‐laser (AFL) using EVCM.
Study Design/Materials and Method: Ex‐vivo porcine skin was treated with MNRF and AFL. Three cryosections from each intervention were stained with acridine orange (AO) and scanned with EVCM. Reflectance confocal microscopy (RCM, 638 nm) and fluorescence confocal microscopy (FCM, 488 nm) images were captured and evaluated individually, after image fusion, and after digital hematoxylin and eosin (H&E) staining.
Results: Bimodal EVCM was able to visualize EBD‐ induced thermal alterations in porcine skin. In RCM mode, the full width and depth of the vertically aligned microscopic treatment zones (MTZ) were displayed with clear demarca- tion to surrounding intact skin. In FCM mode, the ablation of the epidermis after AFL was prominent in contrast to the almost intact epidermis observed in MNRF treated skin.
In fusion mode, fluorescence signal from AO marked the surrounding coagulation zone (CZ) from both interventions, with enhanced discrimination between ablation and coagu- lation. Digitally H&E stained images closely resembled conventional histopathology, but proved superior in terms of visualization of the CZ.
Conclusion: Bimodal EVCM with digital H&E stain- ing facilitates the identification and qualitative evalua- tion of thermal alterations induced by treatment with EBD. By providing high‐resolution images comparable to standard histology, EVCM is a useful tool in the research and development of EBD to visualize and evaluate device‐tissue interactions.
B a s i c S c i e n c e –Photobiomodulation MODELING OF EXTRAORAL
PHOTOBIOMODULATION THERAPY FOR ORAL MUCOSITIS
Anna N. Yaroslavsky, Tyler Iorizzo, Amy Juliano, Ather Adnan, Stephen Sonis, Christine Duncan, Wendy London, Nathaniel Treister
Advanced Biophotonics Laboratory, University of Massachusetts Lowell, Lowell, MA, USA; Massachu- setts Eye and Ear, Harvard Medical School, Boston, MA, USA; Texas A&M Health Science Center, College of Medicine, Houston, TX, USA; Division of Oral Medicine and Dentistry, Brigham and Women's Hospital, Boston, MA, USA; Dana‐Farber/Boston Children's Cancer and Blood Disorders Center, Harvard Medical School, Boston, MA, USA
Background: Photobiomodulation therapy (PBMT) is a preventative/treatment method for oral mucositis (OM) that may develop in patients undergoing high dose chemotherapy, hematopoietic cell transplantation, as well as head and neck chemoradiation. The goal of this study was to model light propagation to determine safe and efficacious extraoral treatment parameters.
Study Design/Materials and Method: Monte Carlo method was used to calculate light distribution andfinite difference method to solve the heat transfer equation.
The model accounted for the geometry of emitter, optical, thermal, and morphological tissue properties, as well as possible changes of the optical properties during the therapy. Tissue type and thickness were determined from MRI archival images. Propagation of 850 nm light was simulated at 399 mW/cm2. The optical and thermal properties of skin, fat, muscle, cartilage, and blood, were taken from the literature and assigned to each pixel individually. Spatially resolved fluence rate, absorbed power, temperature increase and thermal damage were evaluated.
Results: The results of the study demonstrate that at 850 nm and 399 mW/cm2incident power, there was neither temperature increase nor damage to the tissues. Dose delivered to mucosa varied over several orders of magni- tude, with the median dose ranging between 0.18 and 2.4 J/
cm2. Modeling revealed that total tissue thickness was the greatest factor that affected light attenuation. No correla- tion was found between light attenuation and patients' age or gender. Light transmission through the neck was negligible due to the presence of large blood vessels. As presence of blood hinders light penetration, treatment sites should be chosen to avoid major blood vessels.
Conclusion: Our results indicate that extraoral PBMT is safe and shows promise for the prevention and treatment of OM.
T H E H A I R G R O W T H P R O M O T I O N I N M I C E B Y
P H O T O B I O M O D U L A T I O N T H E R A P Y A T D I F F E R E N T P A R A M E T E R S
Chang‐Hun Huh, Tae‐Rin Kwon, Dong Wook Moon, Jungkwan Lee
Seoul National University Bundang Hospital, Seongnam, South Korea; LG Electronics, Seoul, South Korea
Background: Photobiomodulation therapy (PBMT) devices for hair loss have been popular for many years.
But most treatment settings are decided by empirically, no objectively accepted parameter has been proposed.
We would like to find optimal parameter of PBMT for hair growth.
Study Design/Materials and Method: A dome‐shaped LD/LED module was applied to C57BL/6 mice and each device has different LD modules in numbers, power and treatment times. A negative control and 3% minoxidil positive control were coinvestigated. 3 mW LDs in 5, 10, and 15 mm spacing, 4.5 mW LD in 10 mm spacing were applied three times orfive times in a week.
Results: The dome‐shaped LD/LED modules effec- tively induce telogen‐to‐anagen transition earlier and show increased hair follicle density and diameter. The effect was greater with decreasing spacing and increasing dose and treatment number. Immunohistochemistry analysis shows an increased β‐catenin/sonic hedgehog (Shh) expression like the minoxidil and increases the proliferation rate offibroblast growth factor‐7 (KGF).
Conclusion: Increasing module numbers, powers, and treatment number may increase the effect of PBMT in hair regrowth.
V I A B I L I T Y A N D M M P ‐2 S Y N T H E S I S B Y G I N G I V A L
F I B R O B L A S T S E X P O S E D T O L O W ‐ L E V E L L A S E R T H E R A P Y
Larissa Miotto, Lais Medeiros Cardoso, Taisa Nogueira Pansani, Carlos Alberto de‐Souza‐Costa, Fernanda Basso
Araraquara Dental School—UNESP, Araraquara, SP, Brazil
Background: Oral mucositis is the main acute complication of head and neck cancer treatment. Its pathogenesis is not fully comprehensive but is related to increased expression of cytokines and metalloproteinases (MMPs). Modulation of these inflammatory mediators can result in accelerated repair. Photobiomodulation can be used as a treatment, however, despite the standard- ization of protocols, there is still no consensus on the parameters.
Study Design/Materials and Method: The object of this study was to assess the viability and synthesis of MMP‐2 by cells submitted to photobiomodulation (PBM) with low‐level laser (LLL) at distinct energy doses. A primary culture of gingivalfibroblasts (GF) was obtained from voluntary donors according to the Human Research Ethics Committee (CAAE: 74823317.0.0000.5416). Cells seeded in wells of 24‐well plates received PBM by using a LaserTABLE device (780 nm, 0.025 W, 2 cm2), at 0.5, 2, and 4 J/cm2and then exposed or not to TNF‐a (100 ng/
ml). Cells treated with TNF‐a or no treated were used as positive and negative controls, respectively. The synthesis
of MMP‐2 was evaluated by ELISA immunoassay and the cell viability by MTT assay (n = 8; ANOVA and Tukey; a = 5%).
Results: PBM with LLL at 4 J/cm2 resulted in the highest GF viability and the lowest rates of MMP‐2 synthesis (p < 0.05).
Conclusion: It was concluded that PBM with LLLT at the dose of 4 J/cm2 is capable of inhibiting MMP‐2 synthesis and increasing the viability of GF, which may favor the gingival healing process.
B a s i c S c i e n c e –Plasma Analysis of L i v e r a n d L i p i d P r o files
SAFETY OF SIMULTANEOUS APPLICATION OF HIFEM AND SYNCHRONIZED RF FOR FAT
REDUCTION IN MULTIPLE ZONES ON THE SAME DAY VERIFIED BY PLASMA ANALYSIS OF LIVER AND LIPID
PROFILES
Robert A. WeissMaryland Laser, Skin & Vein Institute, Hunt Valley, MD, USA
Background: High‐intensity focused electromagnetic field (HIFEM) and synchronized radiofrequency (RF) technologies have been widely accepted for body contouring. This study verifies the safety of simultaneous therapy when multiple modalities on several body parts are delivered on the same day.
Study Design/Materials and Method: Seven human subjects (24–50 years, BMI 22.4–27.11 kg/m2) underwent four consecutive HIFEM + RF procedures on the abdo- men, lateral thighs, and inner thighs on the same day.
Each body part was treated for 30 minutes with HIFEM intensity at a maximal tolerable level with simultaneous RF at 100%. The primary outcome of the study was blood plasma analysis of liver function (AST, ALT, GGT, ALP) and lipid profile (cholesterol, HDL, LDL, TG), with fasting blood drawn before the treatment, 1 hour, 24–48 hours, and 1‐month after the last treatment. Furthermore, digital photographs of the treated body parts were taken, together with satisfaction and comfort questionnaires.
Results: Obtained values were compared to laboratory standards. No abnormal fluctuation in parameters was observed. Average measured levels of liver markers ranged as follows: AST 16.3–17.1 IU/l, ALT 9.9–10.6 IU/l, GGT 9.1–11.6 IU/l, and ALP 64.0–70.3 IU/l, which corre- sponded to normal reference ranges. Average levels of lipid markers were TG 0.6–0.8 mmol/L, HDL 1.8–2.0 mmol/L, LDL 2.5–2.6 mmol/L, and total choles- terol 4.6–4.8 mmol/L, all within normal limits. After the 1‐
month follow‐up, all seven subjects were satisfied with their results, includingfive patients strongly satisfied with
