Burn Wound Progression Research Articles

Locally Activated Mitophagy Contributes to a 'Built-In' Protection Against Early Burn-Wound Progression in Rats Following a Regulation of HIF-1α

Backgrounds: Deep burn-wounds undergo a dynamic progression in the initial or periburn area after insults. The zone of stasis has a higher risk of deterioration and is considered a salvageable target. Few studies have explored the role of mitochondrial damage in this process and potential “built-in” self-defense within the human body. Methods: A classic “comb” scald rat model was established. Histological and blood-flow observation were processed based on hematoxylin-eosin staining and laser analysis. Oxidative and apoptotic status were analyzed by commercial kits. Transmission electron microscope, immunofluorescence staining, and western blot were applied to detect the mitophagy happened in the zone of stasis and potential regulators. Adenovirus-based gene-silence contributed to determine the role of HIF-1α as a regulatory mediator. Findings: We found that burn insults caused typical ischemia and histological deterioration in the zone of stasis, in parallel with increases in oxidative stress and apoptosis. Mitochondrial damage was also involved in the aforementioned changes. Furthermore, we detected typical mitophagy in burn-wounds, which was contradictory to the burn-wound conversion. HIF-1α expression was closely related to the level of mitophagy, while BNIP3 and PARKIN are involved downstream. Interpretation: Accordingly, burn-induced mitochondrial impairment contributes to the mobilization of injurious mechanisms in the zone of stasis and that mitophagy provides a more beneficial way to protect against burn-wound progression via the elimination of damaged mitochondria. Our findings offer insights into mitochondrial quality control in burn-wound progression and suggest the novel concept that HIF-1α may be a potential therapeutic target due to its possible regulatory effects on BNIP3- or PARKIN-mediated mitophagy. Funding Statement: This work was supported by the following grants: National Natural Science Foundation of China (NSFC) Grants 81671909, 81901958 and Zhejiang Provincial Natural Science Foundation of China Grants: LY18H150004, LY19H150004, LY20H150010. Declaration of Interests: The authors declare no competing financial interests. Ethics Approval Statement: The present study was performed according to protocols approved by the Committee on Animal Care and Use of the Second Affiliated Hospital Zhejiang University School of Medicine (2019-331), and strictly followed the National Institutes of Health Guidelines for the Care and Use of Laboratory Animals.

Modeling early thermal injury using an ex vivo human skin model of contact burns

BackgroundEarly mechanisms underlying the progressive tissue death and the regenerative capability of burn wounds are understudied in human skin. A clinically relevant, reproducible model for human burn wound healing is needed to elucidate the early changes in the human burn wound environment. This study reports a reproducible contact burn model on human skin that explores the extent of tissue injury and healing over time, and defines the inter-individual variability in human skin to enable use in mechanistic studies on burn wound progression and healing. MethodsUsing a customized burn device, contact burns of various depths were created on human skin by two operators and were evaluated for histologic depth by three raters to determine reproducibility. Early burn wound progression and wound healing were also evaluated histologically after the thermally injured human skin was cultured ex vivo for up to 14 days. ResultsBurn depths were reproducibly generated on human skin in a temperature- or time-dependent manner. No significant difference in operator-created or rater-determined depth was observed within each patient sample. However, significant inter-individual variation was identified in burn depth in ten patient samples. Burn-injured ex vivo human skin placed into culture demonstrated differential progression of cell death and collagen denaturation for high and low temperature contact burns, while re-epithelialization was observed in superficial burn wounds over a period of 14 days. ConclusionThis model represents an invaluable tool to evaluate the inter-individual variability in early burn wound progression and wound healing to complement current animal models and enhance the translation of preclinical research to improvements in patient care.

Safety profile of atorvastatin in the role of burn wound injury conversion

BackgroundAtorvastatin could be beneficial in the treatment of burn patients to prevent burn wound progression from partial to full thickness. Our primary aim is to evaluate the safety of atorvastatin in burn patients. MethodsSingle center retrospective chart review of burn patients receiving atorvastatin during admission May 2016–May 2019 with historic controls was performed. Demographics, burn total body surface area, atorvastatin doses, creatinine phosphokinase, aspartate aminotransferase levels and adverse events were analyzed. Results48 burn patients received atorvastatin during admission. Nine patients experienced elevated CK or AST levels during admission, but did not correlate with timing of atorvastatin administration and were comparable to levels in control patients. No adverse events associated with atorvastatin were identified. ConclusionsAtorvastatin administered to patients with burn injuries was not associated with any adverse events or attributable lab abnormalities. We believe that atorvastatin is safe to use in patients with burns and can be safely studied to determine the drug’s effect on the prevention of burn wound conversion.

A novel human ex-vivo burn model and the local cooling effect of a bacterial nanocellulose-based wound dressing

BackgroundBurn wound progression is a significant problem as burns initially thought to be superficial can actually become full thickness over time. Cooling is an efficient method to reduce burn wound conversion. However, if the cooling agent is below room temperature, depending on the wound size the patient is at risk of hypothermia. Additionally, tissue perfusion is reduced leading to an aggravation of burn wound progression. We investigated if wound dressings based on non-pre-cooled bacterial nanocellulose (BNC) with a high water content cool a burn just by evaporation and reduce the intradermal damages in the skin. Material and methodsIn a human ex-vivo model, skin explants underwent contact burns using a 100 °C hot steel block. The burned areas were divided into two groups of which one was cooled with a BNC-based wound dressing. Intradermal temperature probes measured temperature in cooled and uncooled burn sites over 24 h. For histological assessments of the burned areas biopsies were taken at different time points. High mobility group box-1 (HMBG1) staining served as marker for cell vitality and necrosis in the different skin layers. ResultsIntradermal temperature measurement showed that application of the BNC-based wound dressing reduced temperature significantly in burned skin. This cooling effect resulted in a maximum temperature difference of 6.4 ± 1.9 °C and a significant mean reduction of the area under the curve in the first hour after burn of 62% (p < 0.0001). The histological results showed less necrosis and less dermal-epidermal separation in the cooled areas. The HMGB1 staining revealed more vital cells in the cooled group than in the uncooled group. ConclusionBased on our results, BNC-based wound dressings cool a burn. Intradermal temperature as well as thermal damage of the tissue was reduced. The tested BNC-based wound dressing can be used without pre-cooling to cool a burn as well as to reduce the burn BNC-based wound progression through its evaporation cooling effect.

Comparison of a topical surfactant and a topical antibiotic in the rat comb burn model

BackgroundBurn injury progression in the secondary zone of ischemia is common leading to delayed healing and increased scarring. We hypothesized that a topical surfactant, would reduce burn injury progression in a validated rat comb burn model compared with topical antibiotic ointment. MethodsWe created 40 comb burns on 20 rats which were randomized to daily topical application of the surfactant or a triple antibiotic ointment. The comb burns consisted of 4 full thickness burns with 3 unburned interspaces between the 4 burns. These unburned interspaces represented the zone of ischemia, and when left untreated, generally progress to full thickness necrosis within several days. Comb burns were assessed daily for the presence of gross necrosis of the interspaces. At 7 days the comb burns were excised and blindly evaluated for the presence of histological evidence of necrosis. The study had 80% power to detect a 25% difference in the percentages of necrotic interspaces on day 7. ResultsThere were no differences in the percentages of histologically necrotic interspaces at 7 days in burns treated with the surfactant or antibiotic ointment (85% [95%CI, 74 to 92] vs. 75% [95%CI, 63 to 84]; mean difference 10% [95%CI −4 to 24]). There were also no between group differences in the percentages of grossly necrotic interspaces on any of the seven days of the experiment. The surfactant remained intact and adherent while the antibiotic had been absorbed at each daily dressing change. ConclusionsA topical surfactant did not reduce injury progression in the rat comb burn model when compared with antibiotic ointment. The surfactant was more durable than the antibiotic ointment.

Antioxidants Reduce Tissue Necrosis in The Zone of Stasis: Review of Burn Wound Conversion

Summary: Severe burns are devastating condition identified by loss of hemodynamic stability and intravascular volume. Adequate fluid replacement, nutritional support, and immediate wound grafting can reduce the risk of infection and mortality. Oxidative stress was shown to have significant role in the burn wound conversion, which happens when the zone of stasis can’t be salvaged and progresses to necrosis. Decreasing the level of oxidative stress early may be fundamental in reducing burn injury progression into deeper tissue. Several animal studies have demonstrated the advance of antioxidant supplementation for burns outcomes. Approach to this salvageable burn tissue is a breakthrough for new directions in burn management. Antioxidant supplementations was proven to prevent burn conversion on the ischemic zone. Administering antioxidant post-burn is linked with less progression of burn depth and inflammatory cytokine release, which alleviates burn-related morbidity and mortality and improves patient’s quality of life. To date, no clinical trials have been done to reproduce similar outcomes of this ROS-scavenging therapy as successfully observed in murine models. Antioxidant supplementation is a promising treatment avenue to halt burn wound conversion following severe burns.&#x0D; Keywords: Burn wound, wound conversion, burn management, antioxidant

Injectable Enzyme-Based Hydrogel Matrix with Precisely Oxidative Stress Defense for Promoting Dermal Repair of Burn Wound.

Burn wound healing remains a challenging health problem worldwide due to the lack of efficient and precise therapy. Inherent oxidative stress following burn injury is importantly responsible for prolonged inflammation, fibrotic scar, and multiple organ failure. Herein, a bioinspired antioxidative defense system coupling with in situ forming hydrogel, namely, multiresponsive injectable catechol-Fe3+ coordination hydrogel (MICH) matrix, is engineered to promote burn-wound dermal repair by inhibiting tissue oxidative stress. This MICH matrix serves as the special traits of "Fe-superoxide dismutases," small molecular antioxidant (vitamin E), and extracellular matrix (ECM) in alleviating cellular oxidative damage, which demonstrates precise scavenging on reactive oxygen species (ROS) of different cellular locations, blocking lipid peroxidation and cell apoptosis. In in vivo burn-wound treatment, this MICH promptly integrates with injured surrounding tissue to provide hydration microenvironment and physicochemical ECM for burn wounds. Importantly, the MICH matrix suppresses tissue ROS production, reducing the inflammatory response, prompting re-epithelization and neoangiogenesis during wound healing. Meanwhile, the remodeling skin treated with MICH matrix demonstrates low collagen deposition and normal dermal collagen architecture. Overall, the MICH prevents burn wound progression and enhances skin regeneration, which might be a promising biomaterial for burn-wound care and other disease therapy induced by oxidative stress.

642 The Use of Laser Speckle Contrast Imaging in Assessing Depth and Progression of Burn Wounds

Abstract Introduction The ability of laser speckle contrast imaging (LSCI) to provide real-time images of blood flow makes this modality appealing in the assessment of burn wounds, particularly for clinicians making treatment decisions based on burn wound depth and presumed progression. Here we present 2 preclinical studies that used LSCI to assess wound progress, both immediately and months after injury. Methods LSCI images were taken 10-40cm away from the wound and captured with a 1388x1038-pixel CCD camera. In the first study, LSCI images were captured prior to and immediately following creation of superficial partial-thickness (SPTB, 10s), deep partial-thickness (DPTB, 15s), and full-thickness burns (FTB, 20s), and on post-burn day (PBD) 1, 2, and 3. In the second study, LSCI images were obtained before and after DPTB creation and on PBD 7, 14, 21, 28, 60, 90, and 120. Results 92 wounds from 9 swine were included. Speckle data was normalized to control sites and converted to percentages ([speckle wound/speckle control] x 100), producing speckle percentage of control (SPOC) which quantifies the relative decrease or increase in speckle output (vascularity). SPOC was significantly decreased for all burn times on PBD 0, 1, and 2. By PBD 3, only DPTB and FTB remained diminished (p=0.028 and p=0.005, respectively), and FTB SPOC was significantly less than the SPTB (p=0.015). In the second study, SPOC showed an increase post-debridement on PBD 7, noted as post-debridement day (PDD) 0. SPOC continued to increase significantly to a peak at PDD 7 (p&amp;lt; 0.0001) and remained elevated until PDD 28. By PDD 60, SPOC was no longer significantly increased. Conclusions LSCI is a reliable method for analyzing burn depth and wound progression in the preclinical setting. LSCI data shows an immediate decrease in vascularity at all burn depths immediately following burn creation, followed by a peak in vascularity on PDD 7, with a trend back to normal by PDD 60. Applicability of Research to Practice The correlation of wound bed vascularity based on LSCI to known data on burn depth and progression suggests that LSCI could be a useful measurement tool in the clinical setting for the provider determining wound viability.

Leptin and fractalkine: novel subcutaneous cytokines in burn injury.

ABSTRACTBurn injury is a pathology underpinned by progressive and aberrant inflammation. It is a major clinical challenge to survival and quality of life. Although the complex local and disseminating pathological processes of a burn injury ultimately stem from local tissue damage, to date relatively few studies have attempted to characterise the local inflammatory mediator profile. Here, cytokine content and associated transcriptional changes were measured in rat skin for three hours immediately following induction of a scald-type (60°C, 2 min) burn injury model. Leptin (P=0.0002) and fractalkine (P=0.0478) concentrations were significantly elevated post-burn above pre-burn and control site values, coinciding with the development of burn site oedema and differential expression of leptin mRNA (P=0.0004). Further, gene sequencing enrichment analysis indicated cytokine-cytokine receptor interaction (P=1.45×10−6). Subsequent behavioural studies demonstrated that, following subcutaneous injection into the dorsum of the paw, both leptin and fractalkine induced mechanical allodynia, heat hyperalgesia and the recruitment of macrophages. This is the first report of leptin elevation specifically at the burn site, and the first report of fractalkine elevation in any tissue post-burn which, together with the functional findings, calls for exploration of the influence of these cytokines on pain, inflammation and burn wound progression. In addition, targeting these signalling molecules represents a therapeutic potential as early formative mediators of these pathological processes.

The study of the impact of herbal medicinal products in soft dosage forms on the burn wound progression in the experiment

The treatment of burns with topical preparations, namely ointments, which consist of herbal ingredients, becomes relevant among the population due to the fact that this dosage form is the most convenient to apply as self-medication; and biologically active substances that are part of these medicines are less toxic, and at the same time, they are not less effective than synthetic drugs against burn damages.Aim. To study the wound-healing effect of drugs containing biologically active herbal ingredients using the model of burn wound.Materials and methods. The study was conducted on 24 white female rats weighing 200-240 g. The experimental animals were divided into 4 groups: Group 1 – pathology (n = 6); Group 2 – animals treated with Wundahyl ointment (n = 6); Group 3 – animals treated with Spasatel Forte ointment (n = 6); Group 4 – animals treated with 2 % thiotriazoline ointment (n = 6).Results. When treating the burn wounds with Wundahyl ointment and 2 % thiotriazoline ointment the wound-healing progression was better. The burn wound area for the following two weeks of observation (Days 14-21) decreased faster compared to the application of Spasatel Forte ointment and much faster in compared to the group of pathology. On Day 28, the burn wound was epithelized completely.Conclusions. According to the results of the experiment, the effect of Wundahyl ointment was the most pronounced in relation to all the soft dosage forms under study.

Wound Healing Effect of Nonthermal Atmospheric Pressure Plasma Jet on a Rat Burn Wound Model: A Preliminary Study.

Worldwide, an estimated 6 million patients seek medical attention for burns annually. Various treatment methods and materials have been investigated and developed to enhance burn wound healing. Recently, a new technology, plasma medicine, has emerged to offer new solutions in wound care. As the development of plasma medicine has shown benefit in wound healing, we aimed to assess the effects of plasma medicine on burn wounds. To investigate the effectiveness of a nonthermal atmospheric pressure plasma jet (NAPPJ) for burn wound treatment on a brass comb burn wound rat model. Burn wounds were made by applying a preheated brass comb (100°C) for 2 minutes, which resulted in four full-thickness burn wounds separated by three interspaces. Interspaces were exposed to NAPPJ treatment for 2 minutes and morphological changes and neutrophil infiltration were monitored at 0, 4, and 7 days post-wounding. The percentage of necrotic interspace was higher in the control group than in the plasma-treated group (51.8 ± 20.5% vs 31.5 ± 19.0%, P < .001). Moreover, the exposure of interspace to NAPPJ greatly reduced the number of infiltrating neutrophils. In addition, the percentage of interspace that underwent full-thickness necrosis in the plasma-treated group was smaller than that in the control group (28% vs 67%). NAPPJ exposure on interspaces has a positive effect on burn wounds leading to wound healing by reducing burn injury progression.

Severe Burn Injury Progression and Phasic Changes of Gene Expression in Mouse Model.

Patients with severe burns are susceptible to infectious complications including burn-site infections and sepsis. The purpose of this study was to explore the pathologic development of burn injury in a mouse model and to screen genes dysregulated at different time points on the basis of gene expression microarrays. Differential expression analysis identified a total 223 genes that related to only time progression independent of burn injury and 214 genes with aberrant expression due to burn injury. Weighted gene co-expression network analysis (WGCNA) of the 214 genes obtained seven gene modules which named as red, blue, turquoise, green, brown, yellow, and gray module, and the blue module was found to be significantly associated with severe burn injury progression, and in which several genes were previously reported being associated with inflammation and immune response, such as interleukin IL-6, IL-8, and IL-1b. Functional enrichment analysis indicated significant enrichment of biological processes that related to metabolism and catabolism, and pathways of proteasome, notch signaling and cell cycle. This result supports a phase progression of severe burn with gene expression changes and interpretation of biological processes in mouse.

Notoginsenoside R1 protects human keratinocytes HaCaT from LPS-induced inflammatory injury by downregulation of Myd88.

Burn injury is a gigantic challenge in public health which brings multiple negative effects to patients both in physical and spiritual aspects. Inflammation plays vital roles in the progression of burn injury, and our study investigated whether notoginsenoside R1 (NGR1) alleviated lipopolysaccharide (LPS)-induced human keratinocyte HaCaT cell inflammatory injury. Inflammatory injury was induced by LPS in HaCaT cells. Stimulated cells were then treated by NGR1 in different concentrations. Cell viability and cell apoptosis were detected by Cell Counting Kit-8 and flow cytometry, respectively. The concentration of tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6) was measured by enzyme-linked immunosorbent assay (ELISA). The accumulated levels of apoptosis-related proteins (caspase-3 and caspase-9), nuclear factor κB (NF-κB), p38 mitogen-activated protein kinase (p38MAPK) signal pathways–related proteins (p65, IκBα, and p38MAPK), and myeloid differentiation primary response 88 (MyD88) were examined by western blot. Transfection was used to alter the expression of MyD88. We found that LPS stimulated HaCaT cells and induced cell inflammation, evidenced by decreasing cell viability, increasing cell apoptosis, and elevating TNF-α and IL-6 expressions. Then, we found that NGR1 reversed the results by enhancing cell viability, inhibiting cell apoptosis, and reducing TNF-α and IL-6 expressions. In addition, NGR1 decreased the phosphorylation of p65, IκBα, and p38MAPK, which increased by LPS. Moreover, NGR1 negatively regulated the expression of MyD88, and transfection with pMyD88 led to the opposite results with what showed by NGR1 in LPS-stimulated HaCaT cells. To sum up, NGR1 alleviates LPS-induced HaCaT cell inflammatory injury by downregulation of MyD88, as well as inactivation of NF-κB and p38MAPK signal pathways.

Role of infrared thermography in the assessment of burn wounds treated with and without hyperbaric oxygen therapy

Aim: The aim is to study the role of infrared thermography in the assessment of burns wound treated with and without hyperbaric oxygen (HBO) therapy. Burn wound progression is assessed with clinical evaluation and infrared thermography for the patients subjected to HBO therapy with controls. Materials and Methods: This study was carried out in the plastic surgery department, from November 2018 to February 2019. A total of 21 patients with Burns were subjected to clinical evaluation and thermographic study. All patients were treated according to standard burns treatment protocol, and photographic data of clinical and thermographic imagings were sequentially recorded. These clinical and radiographic images were preserved for analysis. In 10 patients, HBO therapy was used as an adjunct for burns wound management and results were evaluated and rest of the patients was used as control. Results: Of 21 patients, with all patients' burns patients assessment done clinically and infrared thermography, 10 patients were subjected to hyperbaric therapy in random fashion and 11 patients were not subjected to HBO therapy. The duration of wound healing and wound progression to deeper planes were reduced in the patients subjected to hyperbaric therapy, which was evident both clinically and, by thermographic assessment. No complications were noted in patients subjected to HBO therapy. Conclusions: Infrared thermography can be used in association with clinical examination to assess the various types of burns wound and could be utilized for understanding the dynamic changes that happen in burns wound due to changes in the treatment protocol with evolving adjunct managements.

Prevention of Burn Wound Progression by Mesenchymal Stem Cell Transplantation: Deeper Insights Into Underlying Mechanisms.

Burns are dynamic wounds that may present a progressive expansion of necrosis into the initially viable zone of stasis. Therefore, salvage of this zone is a major subject of focus in burn research. The beneficial effects of mesenchymal stem cells (MSCs) on the survival of the zone of stasis have been previously documented. However, many gaps still exist in our knowledge regarding the underlying protective mechanisms. Hence, this study was designed to evaluate the pathophysiological basis of MSCs in the prevention of burn wound progression. Wistar rats received thermal trauma on the back according to the "comb burn" model. Animals were randomly divided into sham, control, and stem cell groups with sacrifice and analysis at 72 hours after the burn. The stasis zones were evaluated using histochemistry, immunohistochemistry, biochemistry, real-time polymerase chain reaction assay, and scintigraphy to evaluate the underlying mechanisms. Gross evaluation of burn wounds revealed that vital tissue percentage of the zone of stasis was significantly higher in the stem cell group. Semiquantitative grading of the histopathologic findings showed that MSCs alleviated burn-induced histomorphological alterations in the zone of stasis. According to CC3a staining and expression analysis of Bax (B-cell leukemia 2-associated X) and Bcl-2 (B-cell leukemia 2) genes, MSCs attenuated increases in apoptosis postburn. In addition, these transplants showed an immunomodulatory effect that involves reduced neutrophilic infiltration, down-regulation of proinflammatory cytokines (tumor necrosis factor α, interleukin 1β [IL-1β], and IL-6), and up-regulation of the anti-inflammatory cytokine IL-10 in the zone of stasis. Burn-induced oxidative stress was significantly relieved with MSCs, as shown by increased levels of malondialdehyde, whereas the expression and activity of the antioxidant enzyme superoxide dismutase were increased. Finally, MSC-treated interspaces had enhanced vascular density with higher expression levels for vascular endothelial growth factor A, platelet-derived growth factor, fibroblast growth factor, and transforming growth factor β. Gamma camera images documented better tissue perfusion in animals treated with MSCs. The protective effects of MSCs are mediated by the inhibition of apoptosis through immunomodulatory, antioxidative, and angiogenic actions.

Glucocorticoids suppress fibroblast apoptosis in an in vitro thermal injury model

The wounds of full- and deep partial-thickness burns result in hypertrophic scars and lead to skin contracture more severely than those of superficial partial-thickness burns. Therefore, preventing burn progression may help improve the aesthetic and functional outcomes after healing. Although a number of studies have focused on elucidating the underlying mechanisms of and preventing burn wound progression, it is still difficult to rescue burned dermis unless early tangential excision is performed. To investigate the underlying mechanisms of and prevent cell death of heat-injured fibroblasts, we developed an in vitro experimental model of heat-injured fibroblasts. We confirmed that heating at 55°C for 30s caused fibroblast necrosis immediately after heating, whereas heating at 46°C for 30s induced apoptosis 24h after heating. We also found that the supplementation of 100ng/ml betamethasone to the culture medium after heating decreased the number of apoptotic cells and increased that of live cells. Our studies suggest that glucocorticoids suppress apoptosis of heat-injured fibroblasts and may be useful for preventing burn wound progression.

The Time Course Pathological Changes After Burn Injury.

In the present study, we aimed to explore the time course pathological changes after burn injury. The time course microarray data of burn injury from the Gene Expression Omnibus (GEO) was further analyzed through bioinformatics analysis. The differential expression genes (DEGs) were identified in the early-stage vs. control groups, middle-stage vs. control groups, and early-stage vs. middle-stage groups after burn injury, followed by pathway enrichment analysis. Gene modules associated with burn injury progression were identified through weighted gene co-expression network analysis (WGCNA), and hub genes were identified via network topology analysis. There were a total of 745 DEGs in the early vs. control group, 1104 DEGs in mid vs. control, and 61 DEGs in early vs. mid group. The significant pathways enriched by DEGs in the middle stage were also enriched by DEGs in the early stage. Immunodeficiency was a significant pathway specific for the DEGs in the early stage. There were 19 overlapped genes, such as myeloperoxidase, transcobalamin, and interferon-induced protein with tetratricopeptide repeats 1, among DEGs in early vs. control, middle vs. control, and early vs. middle groups. WGCNA identified three gene modules that were significantly associated with burn injury progression. Furthermore, we identified several gene modules and biological processes that might be associated with burn injury progression, and such results may be beneficial in understanding the underlying mechanisms and developing novel drugs.

Nicorandil reduces burn wound progression by enhancing skin blood flow

We assessed whether nicorandil, an adenosine triphosphate-sensitive K + channel opener, reduces burn wound progression in a rat comb burn model. A total of 24 rats were used. Following thermal injury, one dose of nicorandil (10 or 30 mg/kg) was administered intragastrically twice daily for 3 days. At days 1 and 3 after injury, skin was harvested for histopathological examination and protein isolation. Rats treated with the 10-mg/kg and 30-mg/kg doses of nicorandil exhibited significantly increased tissue survival in the zone of stasis at days 1 and 3 after injury. The 10-mg/kg and 30-mg/kg nicorandil doses also significantly increased skin perfusion in the zone of stasis at days 1 and 3 after injury. At 30 mg/kg, nicorandil significantly reduced hypoxia-inducible factor-1α expression in the zone of stasis at day 1 after injury and reduced inflammatory responses in the zone of stasis. The latter effect included decreased polymorphonuclear neutrophil leukocyte infiltration and interleukin-1β release at day 1 after injury. At 30 mg/kg, nicorandil also significantly reduced expression of nuclear factor-κB p65, akey transcriptional factor in the regulation of inflammatory mediators, in the zone of stasis at day 1 after injury. Our study demonstrates that a 30 mg/kg dosing schedule of nicorandil increases tissue survival in the zone of stasis by attenuating ischemia-reperfusion injury. This effect is mediated by the enhancement of skin blood flow and reduction in the inflammatory response. Therefore, our findings suggest that nicorandil has potential clinical applications for patients with burns.

Blood vessel occlusion with erythrocyte aggregates causes burn injury progression-microvasculature dilation as a possible therapy.

Burns are dynamic injuries characterized by progressive tissue death and continuous severe pain over the course of several days. The extent of burn injury progression determines the ultimate patient outcome. Initial burns result in a central zone of necrosis surrounded by a potentially viable zone of ischemia. Several mechanisms have been proposed to explain injury progression, including oxidant and cytokine stress resulting from either ischemia/reperfusion and/or inflammation, but no proven therapy has emerged. To address the unmet need to limit burn injury progression, the root cause of this process must be delineated. For this reason, we have recently focused on post-burn blood vessel occlusion, currently ascribed to microthrombi. We have found that blood vessel occlusion is initially, mainly and persistently caused by erythrocyte aggregation. Although thermal-induced cell necrosis is the immediate cause of cell death, apoptotic cells from persistent ischemia/anoxia, admixed with inflammatory cells, form a band between viable and nonviable tissue 24hours later. The delayed cell death by apoptosis appears to be the main attractant for inflammatory cells. Finally, we posit that fibrinogen elevation arising from inflammation provides stimulus for additional erythrocyte aggregation, further extending blood vessel occlusion. In our view this persistent occlusion with resultant prolonged tissue ischemia/anoxia, not ischemia/reperfusion, is the root cause of burn injury progression concomitant with associated severe and persistent pain. Epiviosamines, a new class of peptides, appear to selectively dilate microvasculature, and may provide therapy for burn injury progression.

Evidence for modulation of heat induced changes in erythrocytes by cytochalasin D: potential role for oxygen nanobubbles in aggregation

Thermal burn injury is associated with an increase in erythrocyte (RBC) aggregation (rouleaux) prior to fibrin clot formation; unresolved rouleaux occlude vessels and contribute to thermal burn injury progression (Clark, 2013). RBC aggregation is reversible, yet the mechanism is unknown. Current models do not include all biophysical phenomena, including the role of the cytoskeleton, and sudden oxygen release from hemoglobin. We previously proposed that thermally induced membrane deformation creates nucleation sites for excess oxygen to coalesce, i.e., form nanobubbles, which may contribute to surface tension induced aggregation. To begin to address the role of the cytoskeleton in membrane deformation through changes in the band 3‐ankyrin/adducin bridge and spectrin‐actin interactions, we have disrupted actin polymerization with cytochalasin D (cytoD). We hypothesized that disrupting the cytoskeleton would alter the apparent presence of oxygen nanobubbles on singlet RBCs. Our experimental model examines both size and greyscale appearance of RBCs with sudden changes in temperature. Temperatures of 41–49 C cause shifts in brighter greyscale and larger apparent RBC size, consistent with light diffraction through concentrated gas bubbles that dissipates with time or return to 37 C (Blasco, 2017). Washed murine RBCs (obtained through a tissue sharing program) were suspended in saline (PBS) containing 1% EtOH, 0.5 uM CytoD (in 0.1% EtOH) or control, and incubated at 37°C prior to use. RBCs settled to a monolayer within an enclosed microchannel at 37°C (heated stage). The stage temperature rapidly heated from 37°C to test temperature (37, 39, 41, 43, 45, 47, or 49°C) and we imaged at 2 Hz for 70 seconds (60X, SWI). Singlet RBCs were followed over time, measuring greyscale and apparent size of the cell (outer rim) and core (central biconcave) (ImageJ). The initial changes were seen during the temperature shift and included increased core greyscale and apparent size of 20–40% between 41–47°C, with corresponding changes in cell size. Over the next several seconds there was temperature dependent fluctuations in size and greyscale of up to 50%. Cells in EtOH were similar overall to controls. CytoD prevented changes in greyscale and in size, and suppressed further fluctuations until higher temperatures where those treated cells had formed echinocytes. The data are consistent with gas bubbles forming at the RBC membrane, causing diffraction of the light, in a temperature dependent fashion. As destabilization of the cytoskeleton prevented these changes, we conclude that the cytoskeleton contributes to membrane shape change necessary for oxygen nanobubble formation with heat.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.