Association of a 40-Gene Expression Profile With Risk of Metastatic Disease Progression of Cutaneous Squamous Cell Carcinoma (cSCC) and Specification of Benefit of Adjuvant Radiation Therapy.

PURPOSE: Adjuvant radiation therapy (ART) for cutaneous squamous cell carcinoma (cSCC) is recommended based on a number of wide-ranging clinicopathologic features, which encompass a broad array of patients. The 40-gene expression profile (40-GEP) test classifies cSCC tumors into low (Class 1) or higher (Class 2A) or highest (Class 2B) risk of nodal and/or distant metastasis. This study's hypotheses are 1) local recurrence is associated with metastatic disease progression, and 2) 40-GEP, by identifying high risk for metastasis, could predict a metastasis-specific benefit from ART. METHODS: Samples were obtained from 920 patients (ART-untreated: 496 Class 1, 335 Class 2A, 33 Class 2B; ART-treated: 11 Class 1, 35 Class 2A, 10 Class 2B) who were matched on clinical risk factors and stratified by ART status, to create 49 matched patient strata. To control for the variety of characteristics and treatment selection bias, randomly sampled pairs of matched ART and non-ART patients comprising 10,000 resampled cohorts were each analyzed for 5-year metastasis-free survival and predicted time to metastatic event. RESULTS: Of 96 patients experiencing local recurrence, 56.3% experienced metastasis; of those experiencing both, 88.9% had local recurrence before (75.9%) or concurrently (13.0%) with metastasis. After matching for clinicopathological risk, median 5-year disease progression rates for resampled cohorts demonstrated approximately 50% improvement for Class 2B ART-treated as compared to ART-untreated cohorts. Class 2B ART-treated cohorts had a 5-fold delay in predicted time to metastatic event and deceleration of disease progression as compared to ART-untreated cohorts (Kolmogorov-Smirnov test, p<0.01); this was not observed for Class 1 or 2A patients (p>0.05 for each). No risk factor or staging system combined with ART status identified groups that would benefit from ART as well as 40-GEP. CONCLUSION: 40-GEP identifies patients at highest risk of nodal/distant metastasis who may derive greatest benefit from ART, as well as patients who may have clinical indications for ART but are at low risk of metastasis. Compared to current guidelines, 40-GEP could provide greater specificity concerning the benefit of ART in individual patients.

Integrating the 40-Gene Expression Profile (40-GEP) Test Improves Metastatic Risk-Stratification Within Clinically Relevant Subgroups of High-Risk Cutaneous Squamous Cell Carcinoma (cSCC) Patients.

INTRODUCTION: The validated 40-gene expression profile (40-GEP) test independently stratifies risk of regional or distant metastasis for cutaneous squamous cell carcinoma (cSCC) tumors with high-risk clinicopathologic features. This study evaluated the stratification of risk by the 40-GEP test in a large cohort of tumors with one or more high-risk factors and in clinically relevant subgroups, including tumors within National Comprehensive Cancer Network (NCCN) high- and very-high-risk groups, lower-stage BWH T1 and T2a tumors, and patients > 65 years old. METHODS: This multicenter (n = 58) performance study of the 40-GEP included 897 patients. Kaplan-Meier analyses were performed to assess risk stratification profiles for 40-GEP Class 1 (low), Class 2A (higher) and Class 2B (highest) risk groups, while nested Cox regression models were used to compare risk prediction of clinicopathologic risk classification systems versus risk classification systems in combination with 40-GEP. RESULTS: Patients classified as 40-GEP Class 1, Class 2A, or Class 2B had significantly different metastatic risk profiles (p < 0.0001). Integrating 40-GEP results into models with individual clinicopathologic risk factors or risk classification systems (Brigham and Women's Hospital, American Joint Committee on Cancer Staging Manual, 8th Edition) and NCCN demonstrated significant improvement in accuracy for prediction of metastatic events (ANOVA for model deviance, p < 0.0001 for all models). CONCLUSION: The 40-GEP test demonstrates accurate, independent, clinically actionable stratification of metastatic risk and improves predictive accuracy when integrated into risk classification systems. The improved accuracy of risk assessment when including tumor biology via the 40-GEP test ensures more risk-aligned, personalized patient management decisions.

Post-traumatic stress disorder in burn patients - A large database analysis.

INTRODUCTION: Post-traumatic stress disorder (PTSD) afflicts a significant portion of burn patients. This study aims to analyze the morbidity, prevalence, and treatment of PTSD in the burn population. METHODS: Using the TriNetX database, we identified burned patients > 18 years of age without (A) or with (B) a PTSD diagnosis. Patients were then stratified by percent of total body surface area (TBSA) burned. Morbidity and mortality was analyzed in each cohort. Prevalence and pharmacologic treatments for PTSD were analyzed from 2002 to 2022. RESULTS: PTSD incidence increased from 2.4% (n = 2281) in patients with < 10% to 3.1% (n = 542) in 10-30%, 7.4% (n = 285) in 30-59%, and 5.3% (n = 90) in > 60% TBSA burned. In patients with < 60% TBSA burned, PTSD diagnosis increased the risk of depression (p = <0.0003) and anxiety (p = <0.0001). In those with < 30% TBSA burned, PTSD diagnosis also increased risk of insomnia (p = <0.0001) and pruritus (p = 0.0211 for TBSA <10% and 0.0059 for TBSA 10-29%). PTSD diagnosis was associated with a decreased risk of mortality in patients with > 30% TBSA burned (p = 0.0179 for TBSA 30-59% and p = 0.0089 for TBSA >60%). From 2002 to 2022, the prevalence of PTSD in all burn patients was relatively stable between 2.2% and 3.2%. We found an increase in the use of serotonergic agents and prazosin for the treatment of PTSD during this timeframe. CONCLUSION: PTSD is not uncommon in the burn population, and those with burns and concomitant PTSD have an increased risk of morbidity. Screening and preventative measures to reduce morbidity and early implementation of care in burned patients with PTSD are indicated.

Retrospective outcomes analysis of tracheostomy in a paediatric burn population.

INTRODUCTION: Previous analyses of tracheostomy in paediatric burns was hindered by a lack of multi-institution or nationwide analysis. This study aims to explore the effects of tracheostomy in paediatric burn patients in such an analysis. De-identified data was obtained from the TriNetX Research Network database. METHODS: Two cohorts were identified using ICD and CPT codes: paediatric burn patients with tracheostomy (cohort 1) and paediatric burn patients without tracheostomy (cohort 2). Cohorts were matched according to age at diagnosis and pulmonary condition, specifically influenza and pneumonia, respiratory failure, acute upper respiratory infection, and pulmonary collapse. Cohorts were also matched for age at burn diagnosis and surface area burned. Several parameters including infection following a procedure, sepsis, volume depletion, respiratory disorders, laryngeal disorders, pneumonia, and other metrics were also compared. RESULTS: A total of 152 patients were matched according to age and pulmonary condition. Cohort 1 and cohort 2 had a mean age of 4.45 ± 4.06 and 4.39 ± 3.99 years, respectively. Matched patients with tracheostomy had a higher risk for pneumonia, respiratory failure, other respiratory disorders, diseases of the vocal cord and larynx, sepsis, volume depletion, pulmonary edema, and respiratory arrest. The risk ratios for these outcomes were 2.96, 3.5, 3.13, 3.9, 2.5, 2.5, 3.3, and not applicable. Analysis of longitudinal outcomes of paediatric burn patients with tracheostomy vs. those without demonstrated the tracheostomy cohort suffered much worse morbidity and experienced higher health burden across several metrics. CONCLUSION: The potential benefits of tracheostomy in paediatric burn patients should be weighed against these outcomes.

Real-World Evidence Shows Clinicians Appropriately Use the Prognostic 40-Gene Expression Profile (40-GEP) Test for High-Risk Cutaneous Squamous Cell Carcinoma (cSCC) Patients.

Treatment decisions for patients with cutaneous squamous cell carcinoma (cSCC) are traditionally based upon clinicopathologic risk factors and staging systems. Due to the accuracy limitations of these resources in predicting poor outcomes, there is a clinically significant need for more accurate methods of risk assessment. The 40-gene expression profile (40-GEP) test was developed to augment metastatic risk prediction of high-risk cSCC patients and has been validated in two independent, multi-center studies involving over 1,000 patients. This study substantiates that the 40-GEP is appropriately utilized by clinicians and that the personalized risk-stratification results are impactful in guiding risk-aligned patient management.

Galunisertib Exerts Antifibrotic Effects on TGF-ß-Induced Fibroproliferative Dermal Fibroblasts.

Dermal fibroblasts in pathological scars secrete constitutively elevated levels of TGF-ß, signaling the transcription of fibrotic genes via activin-like kinase 5 (ALK5). In the present study, we examine the antifibrotic effects of galunisertib, a small-molecule inhibitor of ALK5, on fibroproliferative dermal fibroblasts in an in vitro model of wound healing. We induced fibrosis in human dermal fibroblasts with exogenous TGF-ß and performed cellular proliferation assays after treatment with varying concentrations of galunisertib. Dermal fibroblast proliferation was diminished to homeostatic levels without cytotoxicity at concentrations as high as 10 µM. An in vitro scratch assay revealed that galunisertib significantly enhanced cellular migration and in vitro wound closure beginning 24 h post-injury. A gene expression analysis demonstrated a significant attenuation of fibrotic gene expression, including collagen-1a, alpha-smooth muscle actin, fibronectin, and connective tissue growth factor, with increased expression of the antifibrotic genes MMP1 and decorin. Protein synthesis assays confirmed drug activity and corroborated the transcription findings. In summary, galunisertib simultaneously exerts antifibrotic effects on dermal fibroblasts while enhancing rates of in vitro wound closure. Galunisertib has already completed phase II clinical trials for cancer therapy with minimal adverse effects and is a promising candidate for the treatment and prevention of pathological cutaneous scars.

Role of Exosomes in Dermal Wound Healing: A Systematic Review.

Cell-based therapy imparts its therapeutic effects through soluble GFs and vesicular bodies such as exosomes. A systematic review with a meta-analysis of preclinical studies was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses and the modified Stroke Therapy Academic Industry Roundtable guidelines to identify exosomes as an archetype biological therapy for dermal wound healing and to provide guidelines for the concentrations to be used in preclinical studies. A total of 51 rodent studies were included in the systematic review and 9 were included in the meta-analysis section. Three independent reviewers cross-screened eligibility and selected studies for quality assessment from 3,064 published studies on exosomes and wound healing. The mean quality scores for all studies were 5.08 ± 0.752 and 5.11 ± 1.13 for systematic review and meta-analysis, respectively. Exosome effects were reported to have the highest efficacy at 7 days (OR = 1.82, 95% confidence interval = 0.69‒2.95) than at 14 days (OR = 2.29, 95% confidence interval = 0.01‒4.56) after administration. Exosomes were reported to regulate all phases of skin wound healing, mostly by the actions of circulating microRNA. The outcome of this review may be used to guide preclinical and clinical studies on the role of exosomes in wound healing.

Cafeteria Diet Impacts the Body Weight and Energy Expenditure of Brown Norway Rats in an Apparent Age Dependent Manner, but Has no Effect on Muscle Anabolic Sensitivity to Nutrition.

While obesity blunts the ability of muscle to mount a protein synthetic response to an amino acid infusion in older adults, it is unclear if this insensitivity to nutrition persists long term and in response to complete foods is unknown. To address this, young (2 months old) and old (17-20 months old) Brown Norway rats were randomized to receive either chow or a 12 wk diet of calorie-dense human foods. At wk 10 drinking water was supplemented with 2% heavy water, followed 2 weeks later by a flooding dose of [2H5]-phenylalanine and an oral leucine bolus, allowing the short and long-term effects of age and diet on muscle protein synthesis rates to be determined. The experimental diet increased energy intake in both young (7.4 ± 0.9%) and old (18.2 ± 1.8%) animals (P < 0.01), but only led to significant increases in body weight in the former (young: 10.2 ± 3.0% (P < 0.05) and old: 3.1 ± 5.1% (NS) vs. age-matched controls). Notably, energy expenditure in response to the cafeteria diet was increased in old animals only (chow: 5.1 ± 0.4; cafe: 8.2 ± 1.6 kcal.kg b.w-1.h-1; P < 0.05). Gastrocnemius protein fractional synthetic rates in response to either an acute leucine bolus or two weeks of feeding were equivalent across groups irrespective of age or diet. Rodents in old age appear capable of preventing weight gain in response to a calorie-dense diet by increasing energy expenditure while maintaining the anabolic sensitivity of muscle to nutrition; the mechanisms of which could have important implications for the aging obese human.

Metal chelation reduces skin epithelial inflammation and rescues epithelial cells from toxicity due to thermal injury in a rat model.

BACKGROUND: One of the most pervasive complications of burn injury is wound progression, characterized by continuous tissue destruction in untreated wounds, which leads to wound infection, inflammation, oxidative stress and excessive scar formation. We determined whether additional tissue destruction could be attenuated with Livionex formulation (LF) lotion, which contains a metal-chelating agent and reduces inflammation in burn wounds. METHODS: We subjected male Sprague Dawley rats to a 2% total body surface area (TBSA) burn using a brass comb model and topically applied LF lotion (containing ethylenediaminetetraacetic acid and methyl sulfonyl methane) to the affected area every 8 hours over 3 days. Inflammatory cytokine levels, cell apoptosis and wound healing were compared in LF lotion-treated and untreated rats. Statistical analysis was performed using a one-way analysis of variance in conjunction with Tukey's post-hoc test. RESULTS: Serum inflammatory cytokines were not detectable after 3 days, suggesting that small burn wounds induce only an immediate, localized inflammatory response. Microscopy revealed that LF lotion improved burn site pathology. Deoxynucleotidyl transferase biotin-d-UTP nick-end labeling staining showed reduced cell death in the LF-treated samples. LF lotion prevented the spread of tissue damage, as seen by increased amounts of Ki-67-positive nuclei in the adjacent epidermis and hair follicles. Tumor necrosis factor-alpha, interleukin-6 and inducible nitric oxide synthase levels in LF-treated skin sections from burned rats were comparable to the levels observed in unburned control sections, indicating that LF lotion reduces inflammation in and around the burn site. CONCLUSIONS: These results establish LF lotion as a therapeutic agent for reducing inflammatory stress, cell death and tissue destruction when applied immediately after a burn injury. Further studies of LF lotion on large TBSA burns will determine its efficacy as an emergency treatment for reducing long-term morbidity and scarring.

Thermal injury initiates pervasive fibrogenesis in skeletal muscle.

Severe burn injury induces a myriad of deleterious effects to skeletal muscle, resulting in impaired function and delayed recovery. Following burn, catabolic signaling and myofiber atrophy are key fiber-intrinsic determinants of weakness; less well understood are alterations in the interstitial environment surrounding myofibers. Muscle quality, specifically alterations in the extracellular matrix (ECM), modulates force transmission and strength. We sought to determine the impact of severe thermal injury on adaptation to the muscle ECM and quantify muscle fibrotic burden. After a 30% total body surface area dorsal burn, spinotrapezius muscle was harvested from mice at 7 (7d, n = 5), 14 (14d, n = 4), and 21 days (21d, n = 4), and a sham control group was also examined (Sham, n = 4). Expression of transforming growth factor-ß (TGFß), myostatin, and downstream effectors and proteases involved in fibrosis and collagen remodeling were measured by immunoblotting, and immunohistochemical and biochemical analyses assessed fibrogenic cell abundance and collagen deposition. Myostatin signaling increased progressively through 21 days postburn alongside fibrogenic/adipogenic progenitor cell expansion, with abundance peaking at 14 days postburn. Postburn, elevated expression of tissue inhibitor of matrix metalloproteinase 1 supported collagen remodeling resulting in a net accumulation of muscle collagen content. Collagen accumulation peaked at 14 days postburn but remained elevated through 21 days postburn, demonstrating minimal resolution of burn-induced fibrosis. These findings highlight a progressive upregulation of fibrogenic processes following burn injury, eliciting a fibrotic muscle phenotype that hinders regenerative capacity and is not resolved with 21 days of recovery.

Adipose-derived stem cells improve grafted burn wound healing by promoting wound bed blood flow.

BACKGROUND: Researchers have explored the use of adipose-derived stem cells (ASCs) as a cell-based therapy to cover wounds in burn patients; however, underlying mechanistic aspects are not completely understood. We hypothesized that ASCs would improve post-burn wound healing after eschar excision and grafting by increasing wound blood flow via induction of angiogenesis-related pathways. METHODS: To test the hypothesis, we used an ovine burn model. A 5 cm2 full thickness burn wound was induced on each side of the dorsum. After 24 hours, the burned skin was excised and a 2 cm2 patch of autologous donor skin was grafted. The wound sites were randomly allocated to either topical application of 7 million allogeneic ASCs or placebo treatment (phosphate-buffered saline [PBS]). Effects of ASCs culture media was also compared to those of PBS. Wound healing was assessed at one and two weeks following the application of ASCs. Allogeneic ASCs were isolated, cultured and characterized from non-injured healthy sheep. The identity of the ASCs was confirmed by flow cytometry analysis, differentiation into multiple lineages and gene expression via real-time polymerase chain reaction. Wound blood flow, epithelialization, graft size and take and the expression of vascular endothelial growth factor (VEGF) were determined via enzyme-linked immunosorbent assay and Western blot. RESULTS: Treatment with ASCs accelerated the patch graft growth compared to the control (p < 0.05). Topical application of ASCs significantly increased wound blood flow (p < 0.05). Expression of VEGF was significantly higher in the wounds treated with ASCs compared to control (p < 0.05). CONCLUSIONS: ASCs accelerated grafted skin growth possibly by increasing the blood flow via angiogenesis induced by a VEGF-dependent pathway.

Current Approaches Targeting the Wound Healing Phases to Attenuate Fibrosis and Scarring.

Cutaneous fibrosis results from suboptimal wound healing following significant tissue injury such as severe burns, trauma, and major surgeries. Pathologic skin fibrosis results in scars that are disfiguring, limit normal movement, and prevent patient recovery and reintegration into society. While various therapeutic strategies have been used to accelerate wound healing and decrease the incidence of scarring, recent studies have targeted the molecular regulators of each phase of wound healing, including the inflammatory, proliferative, and remodeling phases. Here, we reviewed the most recent literature elucidating molecular pathways that can be targeted to reduce fibrosis with a particular focus on post-burn scarring. Current research targeting inflammatory mediators, the epithelial to mesenchymal transition, and regulators of myofibroblast differentiation shows promising results. However, a multimodal approach addressing all three phases of wound healing may provide the best therapeutic outcome.

Buprenorphine-Sustained Release Alters Hemodynamic Parameters in a Rat Burn Model.

BACKGROUND: It has been previously shown that anesthesia and analgesia can affect outcomes in the rat burn model and that buprenorphine alleviated pain without drastically altering the outcomes of interest. Recently, the use of a sustained release (SR) formulation of buprenorphine has been promoted over conventional buprenorphine. In this study, we assessed whether buprenorphine-SR altered hemodynamic parameters in our rat model of severe burn injury. MATERIALS AND METHODS: Adult male Sprague-Dawley rats were randomized to receive either conventional buprenorphine (0.05 mg/kg) or buprenorphine-SR (1 mg/kg). Buprenorphine-SR was administered 24 h before the experiment. Buprenorphine was administered on the day of experiment. These groups were further randomized to control or scald burn (60% of total body surface area). Systolic and diastolic blood pressure (SBP, DBP) and heart rate (HR) were measured using a noninvasive blood pressure system before receiving analgesia and after 72 h. RESULTS: As expected, HR was significantly higher after burn injury regardless of analgesic (P <0.0001). Both SBP and DBP were significantly decreased in burned animals receiving conventional buprenorphine (P < 0.0001), but neither was altered in the buprenorphine-SR-treated burned animals. However, SBP, DBP, and HR were significantly increased after 72 h in control animals receiving buprenorphine-SR (P < 0.0001). CONCLUSIONS: These data indicate that buprenorphine-SR alters the hemodynamic response to injury and may not be an appropriate choice for a model of severe burn injury. If this analgesic is used, investigators must cautiously form conclusions, especially in experimental conditions that would be expected to alter cardiac hemodynamics.

Reduced Postburn Hypertrophic Scarring and Improved Physical Recovery With Yearlong Administration of Oxandrolone and Propranolol.

BACKGROUND: Massive burns induce a hypermetabolic response that leads to total body wasting and impaired physical and psychosocial recovery. The administration of propranolol or oxandrolone positively affects postburn metabolism and growth. The combined administration of oxandrolone and propranolol (OxProp) for 1 year restores growth in children with large burns. Here, we investigated whether the combined administration of OxProp for 1 year would reduce scarring and improve quality of life compared with control. STUDY DESIGN: Children with large burns (n = 480) were enrolled into this institutional review board-approved study; patients were randomized to control (n = 226) or administration of OxProp (n = 126) for 1 year postburn. Assessments were conducted at discharge and 6, 12, and 24 months postburn. Scar biopsies were obtained for histology. Physical scar assessments and patient reported outcome measures of physical and psychosocial function were obtained. RESULTS: Reductions in cellularity, vascular structures, inflammation, and abnormal collagen (P < 0.05) occurred in OxProp-treated scars. With OxProp, scar severity was attenuated and pliability increased (both P < 0.05). Analyses of patient-reported outcomes showed improved general and emotional health within the OxProp-treated group (P < 0.05). CONCLUSIONS: Here, we have shown improvements in objective and subjective measures of scarring and an increase in overall patient-reported physical function. The combined administration of OxProp for up to a year after burn injury should be considered for the reduction of postburn scarring and improvement of long-term psychosocial outcomes in children with massive burns.

ß-Adrenergic Receptor Trafficking, Degradation, and Cell Surface Expression Are Altered in Dermal Fibroblasts from Hypertrophic Scars.

Burn trauma elevates catecholamines for up to 2 years and causes hypertrophic scarring. Propranolol, a nonspecific ß1-, ß2-adrenergic receptor (AR) inverse agonist, counters the hypermetabolic response to elevated catecholamines and may decrease hypertrophic scarring by an unknown mechanism. We investigated the effect of burn injury on ß1-, ß2-, and ß3-AR expression, trafficking, and degradation in human dermal fibroblasts from hypertrophic scar [HSF], non-scar fibroblasts, and normal fibroblasts. We also investigated the modulation of these events by propranolol. Catecholamine-stimulated cAMP production was lower in HSFs and non-scar fibroblasts than in normal fibroblasts. ß1- and ß2-AR cell surface expression was lowest in HSFs, but propranolol increased cell surface expression of these receptors. Basal ß2-AR ubiquitination was higher in HSFs than non-scar or normal fibroblasts, suggesting accelerated receptor degradation. ß-AR degradation was mainly driven by lysosomal-specific polyubiquitination at Lys-63 in normal fibroblasts and HSFs, which was abrogated by propranolol. Propranolol also targeted ß-AR to the proteasome in HSFs. Confocal imaging showed a lack of ß2-AR-GFP trafficking to lysosomal compartments in catecholamine-stimulated HSFs. These data suggest that burn trauma alters the expression, trafficking, and degradation of ß-ARs in dermal fibroblasts, which may then affect fibroblast responses to propranolol.

Burn Trauma Acutely Increases the Respiratory Capacity and Function of Liver Mitochondria.

BACKGROUND: A complete understanding of the role of the liver in burn-induced hypermetabolism is lacking. We investigated the acute effect of severe burn trauma on liver mitochondrial respiratory capacity and coupling control as well as the signaling events underlying these alterations. METHODS: Male BALB/c mice (8-12 weeks) received full-thickness scald burns on ∼30% of the body surface. Liver tissue was harvested 24 h postinjury. Mitochondrial respiration was determined by high-resolution respirometry. Citrate synthase activity was determined as a proxy of mitochondrial density. Male Sprague-Dawley rats received full-thickness scald burns to ∼60% of the body surface. Serum was collected 24 h postinjury. HepG2 cells were cultured with serum-enriched media from either sham- or burn-treated rats. Protein levels were analyzed via western blot. RESULTS: Mass-specific (P = 0.01) and mitochondrial-specific (P = 0.01) respiration coupled to ATP production significantly increased in the liver after burn. The respiratory control ratio for ADP (P = 0.04) and the mitochondrial flux control ratio (P = 0.03) were elevated in the liver of burned animals. Complex III and Complex IV protein abundance in the liver increased after burn by 17% and 14%, respectively. Exposure of HepG2 cells to serum from burned rats increased the pAMPKα:AMPKα ratio (P < 0.001) and levels of SIRT1 (P = 0.01), Nrf2 (P < 0.001), and PGC1α (P = 0.02). CONCLUSIONS: Severe burn trauma augments respiratory capacity and function of liver mitochondria, adaptations that augment ATP production. This response may be mediated by systemic factors that activate signaling proteins responsible for regulating cellular energy metabolism and mitochondrial biogenesis.

Biventricular differences in ß-adrenergic receptor signaling following burn injury.

Burn injury detrimentally affects the myocardium, primarily due to over-activation of ß-adrenergic receptors (ß-AR). Autopsy reports from our institution reveal that patients often suffer from right ventricle (RV) failure. Since burn injury affects ß-AR signaling in the left ventricle (LV), we proposed that ß-AR signaling may also be altered in the RV. A rodent model with a scald burn of 60% of the total body surface area was used to test this hypothesis. Ventricles were isolated 7 days post-burn. We examined the expression of ß-ARs via Western blotting and the mRNA expression of downstream signaling proteins via qRT-PCR. Cyclic adenosine monophosphate (cAMP) production and protein kinase A (PKA) activity were measured in membrane and cytosolic fractions, respectively, using enzyme immunoassay kits. ß1-AR protein expression was significantly increased in the RV following burn injury compared to non-burned RV but not in the LV (p = 0.0022). In contrast, ß2-AR expression was unaltered among the groups while Gαi expression was significantly higher in the LV post-burn (p = 0.023). B-arrestin-1 and G-protein coupled receptor kinase-2 mRNA expression were significantly increased in the left ventricle post-burn (p = 0.001, p<0.0001, respectively). cAMP production and PKA activity were significantly lower in the LV post-burn (p = 0.0063, 0.0042, respectively). These data indicate that burn injury affects the ß-AR signaling pathway in the RV independently of the LV. Additionally, non-canonical ß-AR signaling may be activated in the RV as cAMP production and PKA activity were unchanged despite changes in ß1-AR protein expression.

Inducible satellite cell depletion attenuates skeletal muscle regrowth following a scald-burn injury.

KEY POINTS: Severe burns result in significant skeletal muscle cachexia that impedes recovery. Activity of satellite cells, skeletal muscle stem cells, is altered following a burn injury and likely hinders regrowth of muscle. Severe burn injury induces satellite cell proliferation and fusion into myofibres with greater activity in muscles proximal to the injury site. Conditional depletion of satellite cells attenuates recovery of myofibre area and volume following a scald burn injury in mice. Skeletal muscle regrowth following a burn injury requires satellite cell activity, underscoring the therapeutic potential of satellite cells in the prevention of prolonged frailty in burn survivors. ABSTRACT: Severe burns result in profound skeletal muscle atrophy; persistent muscle atrophy and weakness are major complications that hamper recovery from burn injury. Many factors contribute to the erosion of muscle mass following burn trauma, and we have previously shown concurrent activation and apoptosis of muscle satellite cells following a burn injury in paediatric patients. To determine the necessity of satellite cells during muscle recovery following a burn injury, we utilized a genetically modified mouse model (Pax7CreER -DTA) that allows for the conditional depletion of satellite cells in skeletal muscle. Additionally, mice were provided 5-ethynyl-2'-deoxyuridine to determine satellite cell proliferation, activation and fusion. Juvenile satellite cell-wild-type (SC-WT) and satellite cell-depleted (SC-Dep) mice (8 weeks of age) were randomized to sham or burn injury consisting of a dorsal scald burn injury covering 30% of total body surface area. Both hindlimb and dorsal muscles were studied at 7, 14 and 21 days post-burn. SC-Dep mice had >93% depletion of satellite cells compared to SC-WT (P < 0.05). Burn injury induced robust atrophy in muscles located both proximal and distal to the injury site (∼30% decrease in fibre cross-sectional area, P < 0.05). Additionally, burn injury induced skeletal muscle regeneration, satellite cell proliferation and fusion. Depletion of satellite cells impaired post-burn recovery of both muscle fibre cross-sectional area and volume (P < 0.05). These findings support an integral role for satellite cells in the aetiology of lean tissue recovery following a severe burn injury.

Characterization of Adipose-Derived Stem Cells Following Burn Injury.

Severe burns induce a prolonged inflammatory response in subcutaneous adipose tissue that modulates signaling in adipose-derived stem cells (ASCs), which hold potential for healing burn wounds or generating skin substitutes. Using a 60% rat scald burn model, we conducted a series of experiments to determine which cells isolated from the adipose tissue produced inflammatory mediators and how these changes affect ASC fate and function. The stromal vascular fraction (SVF), adipocytes, and ASCs were isolated from adipose tissue at varying times up to 4 weeks postburn and from non-injured controls. Endpoints included inflammatory marker expression, expression of ASC-specific cell-surface markers, DNA damage, differentiation potential, and proliferation. Inflammatory marker expression was induced in adipocytes and the SVF at 24 and 48 h postburn; expression of inflammatory marker mRNA transcripts and protein returned to normal in the SVF isolated 1 week postburn. In enriched ASCs, burns did not alter cell-surface expression of stem cell markers, markers of inflammation, differentiation potential, or proliferative ability. These results suggest that adipocytes and the SVF produce large quantities of inflammatory mediators, but that ASCs do not, after burns and that ASCs are unaffected by burn injury or culturing procedures.. They also suggest that cells isolated over 48 h after injury are best for cell culture or tissue engineering purposes.

Effects of the nephrilin peptide on post-burn glycemic control, renal function, fat and lean body mass, and wound healing.

The mechanisms underlying the effects of severe burn trauma are not well understood. We previously demonstrated the ability of nephrilin peptide (an iron-binding peptide believed to enter cells through iron-uptake pathways) to suppress aspects of the neuroinflammatory response in a rat scald model, as well as sepsis mortality in a mouse model. This study explores the effect of nephrilin on other clinically relevant outcomes in the rat scald model. In a rat scald model, animals were treated with nephrilin either in week 1 or week 2 post-burn. Measurements were made of serum glucose and creatinine as well as wound area by planimetry and body composition by DEXA. Given the potential role of iron, results were analyzed both for the entire cohort of animals and for the normoferremic (>100 ug/dL serum iron) subset of animals. Nephrilin improved body composition, wound healing, kidney function, and glycemic control. The first two effects were significant in normoferremic but not in hypoferremic animals suggesting an effect of iron status on burn injury outcomes. Nephrilin treatment modulates a number of relevant variables in the rat scald model.