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Objectives: Fractional ablative CO 2 laser (FLSR) is used to treat hypertrophic scars (HTSs) resulting from burn injuries, which are characterized by factors, such as erythema, contracture, thickness, and symptoms of pain and itch. Traditionally, waiting a year after injury for scar maturation before starting laser treatment has been recommended; however, the potential benefits of earlier intervention have gained popularity. Still, the optimal timing for beginning laser intervention in patients with HTSs remains uncertain. This study aims to evaluate the ideal timing for the initiation of FLSR for HTSs using several qualitative and quantitative assessment measures. It was hypothesized that early intervention would lead to similar improvement trends as later intervention, however, would be more ideal due to the shortened time without symptom relief for patients., Methods: Patients who received three or more laser treatment sessions and completed both pre- and posttreatment evaluations were included in this analysis (n = 69). FLSR treatment was administered at 4-8-week intervals. Patients starting treatment before 6 months after injury were classified as the early-stage intervention group and those beginning treatment at 6-12 months after injury were classified as the late-stage intervention group. Demographic data, including the age of patients at the time of first treatment, age of scars at the time of first treatment, biological sex, ethnicity, Fitzpatrick skin type, and use of laser-assisted drug delivery, were collected by retrospective chart review. Patients were evaluated on six subjective scales and objectively for scar stiffness with durometry. For all scales, higher scores indicate worse scars. A two-way ANOVA, Student's t-test, and Mann-Whitney U-test were used to compare scores from the pre- to posttreatment evaluations., Results: There were no significant differences between the groups for any of the demographic or scar-specific variables; thus, differences in outcome can be attributed to the timing of intervention. Both groups demonstrated an improvement in scars with treatment over time (p < 0.05). Both early- and middle-stage initiation showed scar symptom improvement in five out of six scales. In the late-stage intervention, the Patient and Observer Scar Assessment Scale-Patient average score did not show improvement. In the early-stage intervention, the Vancouver Scar Scale total did not show improvement. Quantitative evaluation of scar stiffness by durometry did not show symptom improvement in either group. The Scar Comparison Scale demonstrated the greatest improvement across groups., Conclusion: Laser treatment led to scar improvement in at least one scale at each stage of initiation. Both intervention timelines resulted in equivalent outcomes, and early intervention should be considered when initiating FLSR treatment in burn scars to alleviate symptoms earlier., (© 2024 The Author(s). Lasers in Surgery and Medicine published by Wiley Periodicals LLC.)

Background: Existing animal models for testing therapeutics in the skin are limited. Mouse and rat models lack similarity to human skin in structure and wound healing mechanism. Pigs are regarded as the best model with regards to similarity to human skin; however, these studies are expensive, time-consuming, and only small numbers of biologic replicates can be obtained. In addition, local-regional effects of treating wounds that are closely adjacent to one-another with different treatments make assessment of treatment effectiveness difficult in pig models. Therefore, here, a novel nude mouse model of xenografted porcine hypertrophic scar (HTS) cells was developed. This model system was developed to test if supplying hypo-pigmented cells with exogenous alpha melanocyte stimulating hormone (α-MSH) will reverse pigment loss in vivo ., Methods: Dyschromic HTSs were created in red Duroc pigs. Epidermal scar cells (keratinocytes and melanocytes) were derived from regions of hyper-, hypo-, or normally pigmented scar or skin and were cryopreserved. Dermal fibroblasts (DFs) were isolated separately. Excisional wounds were created on nude mice and a grafting dome was placed. DFs were seeded on day 0 and formed a dermis. On day 3, epidermal cells were seeded onto the dermis. The grafting dome was removed on day 7 and hypo-pigmented xenografts were treated with synthetic α-MSH delivered with microneedling. On day 10, the xenografts were excised and saved. Sections were stained using hematoxylin and eosin hematoxylin and eosin (H&E) to assess xenograft structure. RNA was isolated and quantitative real-time polymerase chain reaction (qRT-PCR) was performed for melanogenesis-related genes TYR , TYRP1 , and DCT ., Results: The seeding of HTSDFs formed a dermis that is similar in structure and cellularity to HTS dermis from the porcine model. When hyper-, hypo-, and normally-pigmented epidermal cells were seeded, a fully stratified epithelium was formed by day 14. H&E staining and measurement of the epidermis showed the average thickness to be 0.11 ± 0.07 µm vs. 0.06 ± 0.03 µm in normal pig skin. Hypo-pigmented xenografts that were treated with synthetic α-MSH showed increases in pigmentation and had increased gene expression of TYR , TYRP1 , and DCT compared to untreated controls (TYR: 2.7 ± 1.1 vs. 0.3 ± 1.1; TYRP1: 2.6 ± 0.6 vs. 0.3 ± 0.7; DCT 0.7 ± 0.9 vs. 0.3 ± 1-fold change from control; n = 3)., Conclusions: The developed nude mouse skin xenograft model can be used to study treatments for the skin. The cells that can be xenografted can be derived from patient samples or from pig samples and form a robust dual-skin layer containing epidermis and dermis that is responsive to treatment. Specifically, we found that hypo-pigmented regions of scar can be stimulated to make melanin by synthetic α-MSH in vivo ., Competing Interests: The authors declare no conflict of interest., (© 2024 The Author(s). Published by IMR Press.)

Dyschromic hypertrophic scar (HTS) is a common sequelae of burn injury, however, its mechanism has not been elucidated. This work is a histological study of these scars with a focus on rete ridges. Rete ridges are important for normal skin physiology, and their absence or presence may hold mechanistic significance in post-burn HTS dyschromia. It was posited that hyper-, and hypo-pigmented areas of scars have different numbers of rete ridges. Subjects with dyschromic burn hypertrophic scar were prospectively enrolled (n = 44). Punch biopsies of hyper-, hypo-, and normally pigmented scar and skin were collected. Biopsies were paraffin embedded, sectioned, stained with H&E, and imaged. The number of rete ridges were investigated. Burn hypertrophic scars that healed without autografts were first investigated. The number of rete ridges was higher in normal skin compared to HTS that was either hypo- (p < 0.01) or hyper-pigmented (p < 0.001). This difference was similar despite scar pigmentation phenotype (p = 0.8687). Autografted hyper-pigmented scars had higher rete ridge ratio compared to non-autografted hyper-pigmented HTS (p < 0.0001). Burn hypertrophihc scars have fewer rete ridges than normal skin. This finding may explain the decreased epidermal adherence to underlying dermis associated with hypertrophic scars. Though, contrary to our hypothesis, no direct link between the extent of dyschromia and rete ridge quantity was observed, the differences in normal skin and hypertrophic scar may lead to further understanding of dyschromic scars., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2023 Elsevier Ltd and International Society of Burns Injuries. All rights reserved.)

Objectives: Dyschromia is an understudied aspect of hypertrophic scar (HTS). The use of topical tacrolimus has successfully shown repigmentation in vitiligo patients through promotion of melanogenesis and melanocyte proliferation. It was hypothesized that HTSs treated with topical tacrolimus would have increased repigmentation compared to controls., Methodology: Full-thickness burns in red Duroc pigs were either treated with excision and meshed split-thickness skin grafting or excision and no grafting, and these wounds formed hypopigmented HTSs (n = 8). Half of the scars had 0.1% tacrolimus ointment applied to the scar twice a day for 21 days, while controls had no treatment. Further, each scar was bisected with half incurring fractional ablative CO 2 laser treatment before topical tacrolimus application to induce laser-assisted drug delivery (LADD). Pigmentation was evaluated using a noninvasive probe to measure melanin index (MI) at Days 0 (pretreatment), 7, 14, and 21. At each timepoint, punch biopsies were obtained and fixed in formalin or were incubated in dispase. The formalin-fixed biopsies were used to evaluate melanin levels by H&E staining. The biopsies incubated in dispase were used to obtain epidermal sheets. The ESs were then flash frozen and RNA was isolated from them and used in quantitative reverse transcription polymerase chain reaction for melanogenesis-related genes: Tyrosinase (TYR), TYR-related protein-1 (TYRP1), and dopachrome tautomerase (DCT). Analysis of variance test with Šídák's multiple comparisons test was used to compare groups., Results: Over time, within the grafted HTS and the NS group, there were no significant changes in MI, except for Week 3 in the -Tacro group. (+Tacro HTS= pre = 685.1 ± 42.0, w1 = 741.0 ± 54.16, w2 = 750.8 ± 59.0, w3 = 760.9 ± 49.8) (-Tacro HTS= pre = 700.4 ± 54.3, w1 = 722.3 ± 50.7, w2 = 739.6 ± 53.2, w3 = 722.7 ± 50.5). Over time, within the ungrafted HTS and the NS group, there were no significant changes in MI. (+Tacro HTS= pre = 644.9 ± 6.9, w1 = 661.6 ± 3.3, w2 = 650.3 ± 6.2, w3 = 636.3 ± 7.4) (-Tacro HTS= pre = 696.8 ± 8.0, w1 = 695.8 ± 12.3, w2 = 678.9 ± 14.0, w3 = 731.2 ± 50.3). LADD did not lead to any differential change in pigmentation compared to the non-LADD group. There was no evidence of increased melanogenesis within the tissue punch biopsies at any timepoint. There were no changes in TYR, TYRP1, or DCT gene expression after treatment., Conclusion: Hypopigmented HTSs treated with 0.1% tacrolimus ointment with or without LADD did not show significantly increased repigmentation. This study was limited by a shorter treatment interval than what is known to be required in vitiligo patients for repigmentation. The use of noninvasive, topical treatments to promote repigmentation are an appealing strategy to relieve morbidity associated with dyschromic burn scars and requires further investigation., (© 2024 Wiley Periodicals LLC.)

Mechanisms and timing of hypertrophic scar (HTS) improvement with laser therapy are incompletely understood. Epidermal keratinocytes influence HTS through paracrine signaling, yet they are understudied compared to fibroblasts. It was hypothesized that fractional ablative CO2 laser scar revision (FLSR) would change the fibrotic histoarchitecture of the epidermis in HTS. Duroc pigs (n = 4 FLSR and n = 4 controls) were injured and allowed to form HTS. HTS and normal skin (NS) were assessed weekly by noninvasive skin probes measuring trans-epidermal water loss (TEWL) and biopsy collection. There were 4 weekly FLSR treatments. Immediate laser treatment began on day 49 postinjury (just after re-epithelialization), and early treatment began on day 77 postinjury. Punch biopsies from NS and HTS were processed and stained with H&E. Epidermal thickness and rete ridge ratios (RRR) were measured. Gene and protein expression of involucrin (IVL) and filaggrin (FIL) were examined through qRT-PCR and immunofluorescent (IF) staining. After treatment, peeling sheets of stratum corneum were apparent which were not present in the controls. TEWL was increased in HTS vs NS at day 49, indicating decreased barrier function (P = .05). In the immediate group, TEWL was significantly decreased at week 4 (P < .05). The early group was not significantly different from NS at the prelaser timepoint. After four sessions, the epidermal thickness was significantly increased in treated scars in both FLSR groups (immediate: P < .01 and early: P < .001, n = 8 scars). Early intervention significantly increased RRR (P < .05), and immediate treatment trended toward an increase. There was no increase in either epidermal thickness or RRR in the controls. In the immediate intervention group, there was increased IVL gene expression in HTS vs NS that decreased after FLSR. Eight scars had upregulated gene expression of IVL vs NS levels pretreatment (fold change [FC] > 1.5) compared to four scars at week 4. This was confirmed by IF where IVL staining decreased after FLSR. FIL gene expression trended towards a decrease in both interventions after treatment. Changes in epidermal HTS histoarchitecture and expression levels of epidermal differentiation markers were induced by FLSR. The timing of laser intervention contributed to differences in TEWL, epidermal thickness, and RRR. These data shed light on the putative mechanisms of improvement seen after FLSR treatment. Resolution of timing must be further explored to enhance efficacy. An increased understanding of the difference between the natural history of HTS improvement over time and interventional-induced changes will be critical to justifying the continued approved usage of this treatment., (© The Author(s) 2023. Published by Oxford University Press on behalf of the American Burn Association. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objectives: One symptom of hypertrophic scar (HTS) that can develop after burn injury is dyschromia with hyper- and hypopigmentation. There are limited treatments for these conditions. Previously, we showed there is no expression of alpha melanocyte stimulating hormone (α-MSH) in hypopigmented scars, and if these melanocytes are treated with synthetic α-MSH in vitro, they respond by repigmenting. The current study tested the same hypothesis in the in vivo environment using laser-assisted drug delivery (LADD)., Methods: HTSs were created in red Duroc pigs. At Day 77 (pre), they were treated with CO 2 fractional ablative laser (FLSR). Synthetic α-MSH was delivered as a topical solution dissolved in  l-tyrosine (n = 6, treated). Control scars received LADD of  l-tyrosine only (n = 2, control). Scars were treated and examined weekly through Week 4. 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. qRT-PCR was run for melanogenesis-related genes: tyrosinase (TYR), tyrosinase-related protein-1 (TYRP1), and dopachrome tautomerase (DCT). Two-way ANOVA with multiple comparisons and Dunnett's correction compared the groups., Results: The areas of hyperpigmentation were variable before treatment. Therefore, data is represented as fold-change where each scar was normalized to its own pre value. Within the LADD of NDP α-MSH + l-tyrosine group, hyperpigmented areas gradually increased each week, reaching 1.3-fold over pre by Week 4. At each timepoint, area of hyperpigmentation was greater in the treated versus the control (1.04 ± 0.05 vs. 0.89 ± 0.08, 1.21 ± 0.07 vs. 0.98 ± 0.24, 1.21 ± 0.08 vs. 1.04 ± 0.11, 1.28 ± 0.11 vs. 0.94 ± 0.25; fold-change from pre-). Within the treatment group, pretreatment, levels of TYR were decreased -17.76 ± 4.52 below the level of normal skin in hypopigmented scars. After 1 treatment, potentially due to laser fractionation, the levels decreased to -43.49 ± 5.52. After 2, 3, and 4 treatments, there was ever increasing levels of TYR to almost the level of normally pigmented skin (-35.74 ± 15.72, -23.25 ± 6.80, -5.52 ± 2.22 [p < 0.01, Week 4]). This pattern was also observed for TYRP1 (pre = -12.94 ± 1.82, Week 1 = -48.85 ± 13.25 [p < 0.01], Weeks 2, 3, and 4 = -34.45 ± 14.64, -28.19 ± 4.98, -6.93 ± 3.05 [p < 0.01, Week 4]) and DCT (pre = -214.95 ± 89.42, Week 1 = -487.93 ± 126.32 [p < 0.05], Weeks 2, 3, and 4 = -219.06 ± 79.33, -72.91 ± 20.45 [p < 0.001], -76.00 ± 24.26 [p < 0.001]). Similar patterns were observed for scars treated with LADD of  l-tyrosine alone without NDP α-MSH. For each gene, in hyperpigmented scar, levels increased at Week 4 of treatment compared to Week 1 (p < 0.01)., Conclusions: A clinically-relevant FLSR treatment method can be combined with topical delivery of synthetic α-MSH and l-tyrosine to increase the area of pigmentation and expression of melanogenesis genes in hypopigmented HTS. LADD of  l-tyrosine alone leads to increased expression of melanogenesis genes. Future studies will aim to optimize drug delivery, timing, and dosing., (© 2023 Wiley Periodicals LLC.)

Hypertrophic scar (HTS) formation is a common challenge for patients after burn injury. Dermal microvascular endothelial cells (DMVECs) are an understudied cell type in HTS. An increase in angiogenesis and microvessel density can be observed in HTS. Endothelial dysfunction may play a role in scar development. This study aims to generate a functional and expression profile of HTS DMVECs. We hypothesize that transcript and protein-level responses in HTS DMVECs differ from those in normal skin (NS). HTSs were created in red Duroc pigs. DMVECs were isolated using magnetic-activated cell sorting with ulex europaeus agglutinin 1 (UEA-1) lectin. Separate transwell inserts were used to form monolayers of HTS DMVECs and NS DMVECs. Cell injury was induced and permeability was assessed. Gene expression in HTS DMVECS versus NS DMVECs was measured. Select differentially expressed genes were further investigated. HTS had an increased area density of dermal microvasculature compared to NS. HTS DMVECs were 17.59% less permeable than normal DMVECs (p < 0.05). After injury, NS DMVECs were 28.4% and HTS DMVECs were 18.8% more permeable than uninjured controls (28.4 ± 4.8 vs 18.8 ± 2.8; p = 0.11). PCR array identified 31 differentially expressed genes between HTS and NS DMVECs, of which 10 were upregulated and 21 were downregulated. qRT-PCR and ELISA studies were in accordance with the array. DMVECs expressed a mixed profile of factors that can contribute to and inhibit scar formation. HTS DMVECs have both a discordant response to cellular insults and baseline differences in function, supporting their proposed role in scar pathology. Further investigation of DMVECs is warranted to elucidate their contribution to HTS pathogenesis., (© 2022. The Author(s).)

Laser treatment of burn scar has increased in recent years. Standard components of scar evaluation during laser scar revision have yet to be established. Patients who began laser scar revision from January 2018 to 2020, underwent at least three treatments, and completed evaluations for each treatment were included. Patients underwent fractional ablative carbon dioxide laser scar revision and pre- and postprocedure scar evaluations by a burn rehabilitation therapist, including Patient and Observer Scar Assessment Scale, Vancouver Scar Scale, our institutional scar comparison scale, durometry, and active range of motion measurements. Twenty-nine patients began laser scar revision and underwent at least three treatments with evaluations before and after each intervention. All patients improved in at least one scar assessment metric after a single laser treatment. After the second and third treatments, all patients improved in at least three scar assessment metrics. Range of motion was the most frequently improved. Durometry significantly improved after the third treatment. Patients and observers showed some agreement in their assessment of scar, but observers rated overall scar scores better than patients. Patients acknowledged substantial scar improvement on our institutional scar comparison scale. Burn scar improves with fractional ablative laser therapy in a range of scar ages and skin types, as early as the first session. Improvements continue as additional sessions are performed. This work suggests baseline evaluation components for patients undergoing laser and a timeline for expected clinical improvements which may inform conversations between patients and providers when considering laser for the symptomatic hypertrophic scar., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Burn Association. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Wound healing can result in various outcomes, including hypertrophic scar (HTS). Pigs serve as models to study wound healing as their skin shares physiologic similarity with humans. Yorkshire (Yk) and Duroc (Dc) pigs have been used to mimic normal and abnormal wound healing, respectively. The reason behind this differential healing phenotype was explored here., Methods: Excisional wounds were made on Dc and Yk pigs and were sampled and imaged for 98 days. PCR arrays were used to determine differential gene expression. Vancouver Scar Scale (VSS) scores were given. Re-epithelialization was analyzed. H&E, Mason's trichrome, and immunostains were used to determine cellularity, collagen content, and blood vessel density, respectively., Results: Yk wounds heal to a "port wine" HTS, resembling scarring in Fitzpatrick skin types (FST) I-III. Dc wounds heal to a dyspigmented, non-pliable HTS, resembling scarring in FST IV-VI. Gene expression during wound healing was differentially regulated versus uninjured skin in 40/80 genes, 15 of which differed between breeds. Yk scars had a higher VSS score at all time points. Yk and Dc wounds had equivalent re-epithelialization, collagen disorganization, and blood vessel density., Conclusions: Our findings demonstrate that Dc and Yk pigs can produce HTS. Wound creation and healing were consistent among breeds, and differences in gene expression were not sufficient to explain differences in resulting scar phenotype. Both pig breeds should be used in animal models to investigate novel therapeutics to provide insight into a treatment's effectiveness on various skin types., Competing Interests: The authors declare no conflicts of interest., (© 2021 The Authors. Animal Models and Experimental Medicine published by John Wiley & Sons Australia, Ltd on behalf of The Chinese Association for Laboratory Animal Sciences.)

There are limited treatments for dyschromia in burn hypertrophic scars (HTSs). Initial work in Duroc pig models showed that regions of scar that are light or dark have equal numbers of melanocytes. This study aims to confirm melanocyte presence in regions of hypo- and hyper-pigmentation in an animal model and patient samples. In a Duroc pig model, melanocyte presence was confirmed using en face staining. Patients with dyschromic HTSs had demographic, injury details, and melanin indices collected. Punch biopsies were taken of regions of hyper-, hypo-, or normally pigmented scar and skin. Biopsies were processed to obtain epidermal sheets (ESs). A subset of ESs were en face stained with melanocyte marker, S100β. Melanocytes were isolated from a different subset. Melanocytes were treated with NDP α-MSH, a pigmentation stimulator. mRNA was isolated from cells, and was used to evaluate gene expression of melanin-synthetic genes. In patient and pig scars, regions of hyper-, hypo-, and normal pigmentation had significantly different melanin indices. S100β en face staining showed that regions of hyper- and hypo-pigmentation contained the same number of melanocytes, but these cells had different dendricity/activity. Treatment of hypo-pigmented melanocytes with NDP α-MSH produced melanin by microscopy. Melanin-synthetic genes were upregulated in treated cells over controls. While traditionally it may be thought that hypopigmented regions of burn HTS display this phenotype because of the absence of pigment-producing cells, these data show that inactive melanocytes are present in these scar regions. By treating with a pigment stimulator, cells can be induced to re-pigment., Competing Interests: The authors declare no conflicts of interest.

Upon healing, burn wounds often leave hypertrophic scars (HTSs) marked by excess collagen deposition, dermal and epidermal thickening, hypervascularity, and an increased density of fibroblasts. The Galectins, a family of lectins with a conserved carbohydrate recognition domain, function intracellularly and extracellularly to mediate a multitude of biological processes including inflammatory responses, angiogenesis, cell migration and differentiation, and cell-ECM adhesion. Galectin-1 (Gal-1) has been associated with several fibrotic diseases and can induce keratinocyte and fibroblast proliferation, migration, and differentiation into fibroproliferative myofibroblasts. In this study, Gal-1 expression was assessed in human and porcine HTS. In a microarray, galectins 1, 4, and 12 were upregulated in pig HTS compared to normal skin (fold change = +3.58, +6.11, and +3.03, FDR <0.01). Confirmatory qRT-PCR demonstrated significant upregulation of Galectin-1 (LGALS1) transcription in HTS in both human and porcine tissues (fold change = +7.78 and +7.90, P <.05). In pig HTS, this upregulation was maintained throughout scar development and remodeling. Immunofluorescent staining of Gal-1 in human and porcine HTS showed significantly increased fluorescence (202.5 ± 58.2 vs 35.2 ± 21.0, P <.05 and 276.1 ± 12.7 vs 69.7 ± 25.9, P <.01) compared to normal skin and co-localization with smooth muscle actin-expressing myofibroblasts. A strong positive correlation (R = .948) was observed between LGALS1 and Collagen type 1 alpha 1 mRNA expression. Gal-1 is overexpressed in HTS at the mRNA and protein levels and may have a role in the development of scar phenotypes due to fibroblast over-proliferation, collagen secretion, and dermal thickening. The role of galectins shows promise for future study and may lead to the development of a pharmacotherapy for treatment of HTS., (© 2020 The Wound Healing Society.)

Burn injuries frequently result in hypertrophic scars (HTSs), specifically when excision and grafting are delayed due to limited resources or patient complications. In patient populations with dark baseline pigmentation, one symptom of HTS that often occurs is dyspigmentation. The mechanism behind dyspigmentation has not been explored, and, as such, prevention and treatment strategies for this morbidity are lacking. The mechanism by which cells make pigment is controlled at the apex of the pathway by pro-opiomelanocortin (POMC), which is cleaved to its products alpha-melanocyte-stimulating hormone (α-MSH) and adrenocorticotropin hormone (ACTH). α-MSH and ACTH secreted by keratinocytes bind to melanocortin 1 receptor (MC1R), expressed on melanocytes, to initiate melanogenesis. POMC protein expression is upregulated in hyperpigmented scar compared to hypopigmented scar by an unknown mechanism in a Duroc pig model of HTS. POMC RNA levels, as well as the POMC gene promoter methylation status were investigated as a possible mechanism. DNA was isolated from biopsies obtained from distinct areas of hyper- or hypopigmented scar and normal skin. DNA was bisulfite-converted, and amplified using two sets of primers to observe methylation patterns in two different CpG islands near the POMC promoter. Amplicons were then sequenced and methylation patterns were evaluated. POMC gene expression was significantly downregulated in hypopigmented scar compared to normal skin, consistent with previously reported protein expression levels. There were significant changes in methylation of the POMC promoter; however, none that would account for the development of hyper- or hypopigmentation. Future work will focus on other areas of POMC transcriptional regulation., (© American Burn Association 2019. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Reactive oxygen species (ROS) can damage macromolecules if not appropriately neutralized by ROS scavengers. The balance between ROS and ROS scavengers is essential to prevent the accumulation of damage in healthy tissues. This balance is perturbed in hypertrophic scar (HTS)., Materials and Methods: Full-thickness wounds were created on the flanks of Duroc pigs at day 0 that developed into HTS (n = 4). Wounds and HTSs were biopsied weekly for 135 d. Total transcriptome microarrays were conducted with focused ROS scavenger analysis. Confirmatory quantitative reverse transcription polymerase chain reaction and immunofluorescence of ROS scavengers: superoxide dismutase 1, microsomal glutathione S-transferase 1, and peroxiredoxin 6 were performed throughout wound healing and HTS development., Results: Total transcriptome microarray analysis identified over 25 ROS scavenger genes that were significantly downregulated in HTS at all time points compared with basal level controls (BL) (FDR<0.01; fold change > or <2). Ingenuity pathway analysis identified multiple ROS scavenging pathways involved in HTS (P < 0.01). Quantitative reverse transcription polymerase chain reaction of representative scavengers confirmed and expanded this finding to the initial phases of wound healing (P < 0.05, n = 4). The protein products of the genes were lower in wound and HTS tissues compared with BL., Conclusions: A balance between ROS production and scavenging must be maintained for normal wound healing, which is perturbed in wounds that heal to form HTSs. We postulate that endogenous scavengers can be administered as a prophylactic or post-treatment to rebalance ROS and attenuate symptoms of scar., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Although pigment synthesis is well understood, relevant mechanisms of psychologically debilitating dyspigmentation in nascent tissue after cutaneous injuries are still unknown. Here, differences in genomic transcription of hyper- and hypopigmented tissue relative to uninjured skin were investigated using a red Duroc swine scar model. Transcription profiles differed based on pigmentation phenotypes with a trend of more upregulation or downregulation in hyper- or hypopigmented scars, respectively. Ingenuity Pathway Analysis of significantly modulated genes in both pigmentation phenotypes showed pathways related to redox, metabolic, and inflammatory responses were more present in hypopigmented samples, while those related to stem cell development differentiation were found mainly in hyperpigmented samples. Cell-cell and cell-extracellular matrix interactions and inflammation responses were predicted (z-score) active in hyperpigmented and inactive in hypopigmented. The proinflammatory high-mobility group box 1 pathway showed the opposite trend. Analysis of differentially regulated mutually exclusive genes showed an extensive presence of metabolic, proinflammatory, and oxidative stress pathways in hypopigmented scars, while melanin synthesis, glycosaminoglycans biosynthesis, and cell differentiation pathways were predominant in hyperpigmented scar. Several potential therapeutic gene targets have been identified., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Hypertrophic scar (HTS) occurs frequently after burn injury. Treatments for some aspects of scar morbidity exist, however, dyspigmentation treatments are lacking due to limited knowledge about why scars display dyschromic phenotypes. Full thickness wounds were created on duroc pigs that healed to form dyschromic HTS. HTS biopsies and primary cell cultures were then used to study pigmentation signaling. Biopsies of areas of both pigment types were taken for analysis. At the end of the experiment, melanocyte-keratinocyte cocultures were established from areas of differential pigmentation. Heterogeneously dyspigmented scars formed with regions of hyperpigmentation and hypopigmentation. Melanocytes were present in both pigment types measured by S100β quantitative real time-polymerase chain reaction (qRT-PCR) and immunostaining, and visualized by dendritic cell presence in primary cultures. P53 expression was not different between the two pigment types. Hyperpigmented scars had upregulated levels of proopiomelanocortin (POMC), adrenocorticotropic hormone (ACTH), α-melanocyte stimulating hormone (α-MSH), stem cell factor (SCF), and c-KIT and melanocortin 1 receptors (MC1R) compared to hypopigmented regions. Many genes involved in dyspigmentation were differentially regulated by microarray analysis including MITF, TYR, TYRP1, and DCT. MiTF expression was not different upon further exploration, but TYR, TYRP1, and DCT were upregulated in intact biopsies measured by qRT-PCR and confirmed by immunostaining. This is the first work to confirm the presence of melanocytes in hypopigmented scar using qRT-PCR and primary cell culture. An understanding of the initial steps in dyspigmentation signaling, as well as the downstream effects of these signals, will inform treatment options for patients with scars and provide insight to where pharmacotherapy may be directed.

Introduction: The value of compression studies and applications in hypertrophic scar (HTS) treatment is often undermined due to the lack of ideal controls, patient compliance, and clear action mechanisms., Objective: This study assesses the genome-wide compression effects on scars under well-controlled conditions., Materials and Methods: An automated pressure delivery system (APDS) applied controlled doses of pressure to scars in a red Duroc swine HTS model. Full-thickness wounds were created by a skin grafting instrument on each animal's bilateral flanks and were observed through reepithelialization and scar development. On day 70, the APDSs were mounted on the developed scars; right flank scars received a pressure of 30 mm Hg, while left flank scars received APDSs with no pressure (sham) for 2 weeks. A genome-wide assessment of compression effect on transcription in scar specimens before (early), shortly after (mid), and long after (late) compression initiation were performed., Results: Analysis of early-phase biopsies showed similar transcriptome profiles, which diverged thereafter in gene numbers and functions between compression- and sham-treated scars in the mid phase. The majority of these changes persisted in the late-phase scar samples. Canonical pathway analysis of differentially regulated genes resulted in an almost identical list of pathways during the early phase prior to compression. In the mid and late phases after compression, many of the identified pathways shifted in significance, and new pathways such as calcium signaling and cholesterol synthesis emerged., Conclusions: Compression modulates transcription and affects multiple biological functions associated with an improved scar appearance.

Approximately three quarters of major thermal burn injury (MThBI) survivors suffer from hypertrophic scarring (HTS) and over half experience chronic pain or itch. In survivors of MThBI, HTS and chronic pain or itch are considered one of the greatest unmet challenges of postburn injury care and psychosocial reintegration. Although scarring, itch, and pain have been clinically associated, there are no prospective, multisite studies examining tissue autograft site pain or itch and scar outcomes. The authors collected a representative cohort (n = 56) of MThBI survivors who received autografting within 14 days of injury and evaluated graft-site pain or itch severity (0-10 Numeric Rating Scale) and HTS using a validated scar photograph assessment scale 6 months following MThBI. Given that stress is known to influence wound healing, the authors also assessed the relationship between previous trauma exposure, peritraumatic stress, preburn overall health (SF-12), scarring, and chronic pain or itch severity using Spearman's correlation. Association between HTS and chronic pain or itch was significant in a linear regression model adjusted for age, sex, and ethnicity (β = 0.2, P = .033 for pain, β = 0.2, P = .019 for itch). Results indicate that prior trauma exposure is inversely correlated (r = -.363, P = .030) with scar severity, but not pain or itch severity 6 months after MThBI. Study results suggest that preburn chronic pain or itch is associated with pathological scarring 6 months following MThBI. Results also indicate that stress may improve scarring after MThBI. Further work to understand the mechanisms that underlie both HTS and chronic pain or itch and their relationship to chronic stress is critical to the development of novel therapies to assist burn survivors recover.

At the 2016 State of the Science meeting, clinicians and burn survivors met to discuss the advances in scar prevention, evaluation and treatment. While emerging evidence exists to support pressure garment treatment of scars and the use of silicone gel, further research is necessary to better delineate indications duration and efficacy of established therapies and to develop and test badly needed new treatments. More accurate and objective assessment of burn depth would assist in the prevention and identification of wounds requiring customized surgery. Laser treatment of scar while rapidly gaining popularity, still lacks high quality evidence as to its efficacy. The psychological impact of burn scars on the recovering patient is poorly appreciated and increased interaction with our patients is needed to more fully understand and address the impact on health related quality of life of their burn scars.

Beneficial effects of pressure therapy for hypertrophic scars have been reported, but the mechanisms of action are not fully understood. This study evaluated elastin and its contribution to scar pliability. The relationship between changes in Vancouver Scar Scale (VSS) scores of pressure-treated scars and differential regulation of elastin was assessed. Hypertrophic scars were created and assessed weekly using VSS and biopsy procurement. Pressure treatment began on day 70 postinjury. Treated scars were compared with untreated shams. Treatment lasted 2 weeks, through day 84, and scars were assessed weekly through day 126. Transcript and protein levels of elastin were quantified. Pressure treatment resulted in lower VSS scores compared with sham-treated scars. Pliability (VSSP) was a key contributor to this difference. At day 70 pretreatment, VSSP = 2. Without treatment, sham-treated scars became less pliable, while pressure-treated scars became more pliable. The percentage of elastin in scars at day 70 was higher than in uninjured skin. Following treatment, the percentage of elastin increased and continued to increase through day 126. Untreated sham scars did not show a similar increase. Quantification of Verhoeff-Van Gieson staining corroborated the findings and immunofluorescence revealed the alignment of elastin fibers. Pressure treatment results in increased protein level expression of elastin compared with sham-untreated scars. These findings further characterize the extracellular matrix's response to the application of pressure as a scar treatment, which will contribute to the refinement of rehabilitation practices and ultimately improvements in functional and psychosocial outcomes for patients.

Background: The effects of pressure on hypertrophic scar are poorly understood. Decreased extracellular matrix deposition is hypothesized to contribute to changes observed after pressure therapy. To examine this further, collagen composition was analyzed in a model of pressure therapy in hypertrophic scar., Materials and Methods: Hypertrophic scars created on red Duroc swine (n = 8) received pressure treatment (pressure device mounting and delivery at 30 mm Hg), sham treatment (device mounting and no delivery), or no treatment for 2 wk. Scars were assessed weekly and biopsied for histology, hydroxyproline quantification, and gene expression analysis. Transcription levels of collagen precursors COL1A2 and COL3A1 were quantified using reverse transcription-polymerase chain reaction. Masson trichrome was used for general collagen quantification, whereas immunofluorescence was used for collagen types I and III specific quantification., Results: Total collagen quantification using hydroxyproline assay showed a 51.9% decrease after pressure initiation. Masson trichrome staining showed less collagen after 1 (P < 0.03) and 2 wk (P < 0.002) of pressure application compared with sham and untreated scars. Collagen 1A2 and 3A1 transcript decreased by 41.9- and 42.3-fold, respectively, compared with uninjured skin after pressure treatment, whereas a 2.3- and 1.3-fold increase was seen in untreated scars. This decrease was seen in immunofluorescence staining for collagen types I (P < 0.001) and III (P < 0.04) compared with pretreated levels. Pressure-treated scars also had lower levels of collagen I and III after pressure treatment (P < 0.05) compared with sham and untreated scars., Conclusions: These results demonstrate the modulation of collagen after pressure therapy and further characterize its role in scar formation and therapy., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The duroc pig has been described as a promising animal model for use in the study of human wound healing and scar formation. However, little is known about the presence and chronology of the fibrocyte cell population in the healing process of these animals. Wounds known to form scar were created on red duroc swine (3" x 3") with a dermatome to a total depth of either 0.06 inches or 0.09 inches. These wounds were allowed to heal completely and biopsies were done at scheduled time points during the healing process. Biopsies were formalin fixed and paraffin embedded for immunohistochemical analysis. Porcine reactive antibodies to CD-45 and procollagen-1 and a human reactive antibody to LSP-1 were used to detect the presence of fibrocytes in immunohistochemistry, an immunocytochemistry. Initial immunohistochemical studies showed evidence of a biphasic presence of fibrocytes. Pigs with 0.06 inches deep wounds showed positive staining for CD-45 and LSP-1 within highly cellular areas at days 2 and 4 after wounding. Additional animals with 0.09 inches deep wounds showed positive staining within similar areas at days 56, 70, and 113 after wounding. There was no immunohistochemical evidence of fibrocytes in skin biopsies taken at days 14, 28, or 42. Procollagen-1 staining was diffused in all samples. Cultured cells were stained for CD-45, LSP-1, and procollagen-1 by immunocytochemistry. These data confirm that fibrocytes are indeed present in this porcine model. We conclude that these cells are present after initial wounding and later during scar formation and remodeling. We believe that this is an evidence of a biphasic presence of fibrocytes, first as an acute response to skin wounding followed by later involvement in the remodeling process, prompted by continued inflammation in a deep partial thickness wound.

Using a validated swine model of human scar formation, hyperpigmented and hypopigmented scar samples were examined for their histological and optical properties to help elucidate the mechanisms and characteristics of dyspigmentation. Full-thickness wounds were created on the flanks of red Duroc pigs and allowed to heal. Biopsies from areas of hyperpigmentation, hypopigmentation, and uninjured tissue were fixed and embedded for histological examination using Azure B and primary antibodies to S100B, HMB45, and α-melanocyte-stimulating hormone (α-MSH). Spatial frequency domain imaging (SFDI) was then used to examine the optical properties of scars. Hyperpigmentation was first noticeable in healing wounds around weeks 2 to 3, gradually becoming darker. There was no significant difference in S100B staining for the presence of melanocytes between hyperpigmented and hypopigmented scar samples. Azure B staining of melanin was significantly greater in histological sections from hyperpigmented areas than in sections from both uninjured skin and hypopigmented scar (P < .0001). There was significantly greater staining for α-MSH in hyperpigmented samples compared with hypopigmented samples (P = .0121), and HMB45 staining was positive for melanocytes in hyperpigmented scar. SFDI at a wavelength of 632 nm resulted in an absorption coefficient map correlating with visibly hyperpigmented areas of scars. In a red Duroc model of hypertrophic scar formation, melanocyte number is similar in hyperpigmented and hypopigmented tissues. Hyperpigmented tissues, however, show a greater amount of melanin and α-MSH, along with immunohistochemical evidence of stimulated melanocytes. These observations encourage further investigation of melanocyte stimulation and the inflammatory environment within a wound that may influence melanocyte activity. Additionally, SFDI can be used to identify areas of melanin content in mature, pigmented scars, which may lead to its usefulness in wounds at earlier time points before markedly apparent pigmentation abnormalities.

Compression therapy has long been a standard treatment for hypertrophic scar prevention. However, due to the lack of objective, quantitative assessments, and measurements of scar severity, as well as the lack of a self-operated, controllable, and precise pressure delivery technique, limited concrete evidence exists, demonstrating compression therapy's efficacy. We have designed and built an automatic pressure delivery system to apply and maintain constant pressure on scar tissue in an animal model. A force sensor positioned on a compression plate reads the imposed force in real-time and sends the information to a feedback system controlling two position actuators. The actuators move accordingly to maintain a preset value of pressure onto the skin. The system was used in an in vivo model of compression therapy on hypertrophic scars. It was shown that the system was capable of delivering a constant pressure of 30 mmHg on scar wounds for a period of two weeks, and that phenotypic changes were seen in the wounds.