Third-degree Scald Research Articles

Effects of different low-dose of insulin glargine on antioxidation of organs in burned rats with delayed resuscitation

To study the antioxidant protective effects of different low-dose of insulin glargine on organs of burned rats with delayed resuscitation. Forty male Sprague-Dawley (SD) rats were randomly divided into sham group, delayed resuscitation control group, and insulin glargine 0.5, 1.0, and 2.0 U groups, with 8 rats in each group. The rats were immersed in hot water (95.0±0.5) centigrade for 15 s to establish the third-degree scald model with 30% total body surface area. The rats in the sham group were immersed in a 37 centigrade water bath for 15 s. Insulin glargine (0.5, 1.0, 2.0 U×kg-1×d-1) was injected subcutaneously in corresponding insulin glargine group 2 hours after injury, and the same amount of normal saline was injected intraperitoneally in the delayed resuscitation control group. Intraperitoneal injection of normal saline 40 mL/kg simulated delayed resuscitation 6 hours after injury in all groups. Abdominal aortic blood samples, heart and kidney tissue were collected immediately after simulating burn in the sham group, and 24 hours after burn in other four groups. The blood glucose, myocardial enzymes [lactate dehydrogenase (LDH), creatine kinase (CK), α-hydroxybutyrate dehydrogenase (α-HBDH), and aspartate aminotransferase (AST)] and renal function indexes [blood urea nitrogen (BUN) and serum creatinine (SCr)] were measured by spectrophotometry, and the isoenzyme MB of creatine kinase (CK-MB) level was determined by immunosuppression method to evaluate the effects of different low-dose insulin glargine intervention on blood glucose, cardiac and renal functions in scalded rats with delayed resuscitation. The oxidative and antioxidant indices [xanthine oxidase (XOD), myeloperoxidase (MPO), copper-zinc superoxide dismutase (CuZn-SOD), catalase (CAT), glutathione peroxidase (GSH-Px), and total antioxidant capacity (T-AOC)] from the heart and kidney tissues of rats were detected by spectrophotometry to analyze the antioxidant effects of different low-dose insulin glargine interventions. Compared with the sham group, the blood glucose of the rats in the delayed resuscitation control group was significantly increased, the heart and kidney functions were significantly reduced, the oxidation capacity was enhanced, and the antioxidant indicators were significantly reduced. After the intervention of insulin glargine, with the increase of insulin glargine dose, the blood glucose, myocardial enzyme and renal function indicators of rats showed a gradual downward trend, the oxidation indicators continued to decrease, and the antioxidant indicators showed a gradual upward trend. When the dose was 2.0 U×kg-1×d-1, the blood glucose, LDH, CK, CK-MB, α-HBDH, AST, BUN, SCr, XOD and MPO were significantly lower than those in the delayed resuscitation control group [blood glucose (mmol/L): 5.91±0.25 vs. 11.76±0.36, LDH (U/L): 3 332.12±51.61 vs. 5 008.94±490.12, CK (kU/L): 0.49±0.03 vs. 0.85±0.04, CK-MB (U/L): 125.40±12.19 vs. 267.52±11.63, α-HBDH (U/L): 122.99±5.37 vs. 240.85±13.99, AST (U/L): 11.95±1.81 vs. 17.87±1.57, BUN (mmol/L): 4.72±0.15 vs. 7.16±0.34, SCr (μmol/L): 87.11±6.51 vs. 137.50±11.36, XOD (U/g): 166.29±3.27 vs. 204.90±4.82 in heart tissue, 63.51±1.46 vs. 79.69±1.75 in kidney tissue, MPO (U/g): 1.05±0.02 vs. 1.55±0.06 in heart tissue, 1.04±0.04 vs. 1.87±0.01 in kidney tissue, all P < 0.05], and CuZn-SOD, CAT, GSH-Px and T-AOC were significantly higher than those in the delayed resuscitation control group [CuZn-SOD (kU/g): 82.95±2.69 vs. 56.52±2.26 in heart tissue, 94.50±2.73 vs. 62.02±1.66 in kidney tissue, CAT (U/g): 36.07±2.01 vs. 15.15±2.22 in heart tissue, 184.49±4.53 vs. 156.02±3.96 in kidney tissue, GSH-Px (kU/g): 231.93±8.03 vs. 179.48±3.15 in heart tissue, 239.63±7.30 vs. 172.20±2.09 in kidney tissue, T-AOC (kU/g): 4.85±0.23 vs. 2.71±0.11 in heart tissue, 5.51±0.08 vs. 3.50±0.07 in kidney tissue, all P < 0.05]. Different low-dose of insulin glargine (≤ 2.0 U×kg-1×d-1) could exert antioxidant protection on the heart and kidney of rats with delayed resuscitation after burns, with a dose-dependent manner.

Multifunctional effects of wound dressing based on chitosan-coordinated argentum with resistant bacterial penetration

Third-degree scald, causing serious tissue destruction with continuous pain, easily leads to microbial infections and delayed wound healing. Therefore, a multifunctional treatment is attractive for seriously damaged tissue. Herein, carboxymethyl chitosan-coordinated argentum (Ag-CMC) was synthesized via a complexation method, and then the Ag+ release, antibacterial activity, biocompatibility, pain relief and wound healing properties of Ag-CMC were investigated in vitro and in vivo. The results revealed that Ag+ had interacted with carboxymethyl chitosan, containing approximately 1.2% of silver. The Ag-CMC (50–200 μg/mL) with Ag+ sustained release exhibited significant antibacterial activity against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Candida albicans, drug-resistant E. coli, PA, MRSA and good biocompatibility with L929 cells. Furthermore, antibacterial and wound healing experiments demonstrated that Ag-CMC achieved an effective contraction rate of 90% after 28 days by accelerating re-epithelialization, regulating inflammation response, relieving pain and infections. Therefore, Ag-CMC is a safe multifunctional treatment for wound healing and infections.

Obstruction of the formation of granulation tissue leads to delayed wound healing after scald burn injury in mice.

BackgroundDelayed wound healing remains a common but challenging problem in patients with acute or chronic wound following accidental scald burn injury. However, the systematic and detailed evaluation of the scald burn injury, including second-degree deep scald (SDDS) and third-degree scald (TDS), is still unclear. The present study aims to analyze the wound-healing speed, the formation of granulation tissue, and the healing quality after cutaneous damage.MethodsIn order to assess SDDS and TDS, the models of SDDS and TDS were established using a scald instrument in C57BL/6 mice. Furthermore, an excisional wound was administered on the dorsal surface in mice (Cut group). The wound-healing rate was first analyzed at days 0, 3, 5, 7, 15 and 27, with the Cut group as a control. Then, on the full-thickness wounds, hematoxylin and eosin (H&E) staining, Masson staining, Sirius red staining, Victoria blue staining and immunohistochemistry were performed to examine re-epithelialization, the formation of granulation tissue, vascularization, inflammatory infiltration and the healing quality at different time points in the Cut, SDDS and TDS groups.ResultsThe presented data revealed that the wound-healing rate was higher in the Cut group, when compared with the SDDS and TDS groups. H&E staining showed that re-epithelialization, formation of granulation tissue and inflammatory infiltration were greater in the Cut group, when compared with the SDDS and TDS groups. Immunohistochemistry revealed that the number of CD31, vascular endothelial growth factor A, transforming growth factor-β and α-smooth muscle actin reached preferential peak in the Cut group, when compared with other groups. In addition, Masson staining, Sirius red staining, Victoria blue staining, Gordon-Sweets staining and stress analysis indicated that the ratio of collagen I to III, reticular fibers, failure stress, Young’s modulus and failure length in the SDDS group were similar to those in the normal group, suggesting that healing quality was better in the SDDS group, when compared with the Cut and TDS groups.ConclusionOverall, the investigators first administered a comprehensive analysis in the Cut, SDDS and TDS groups through in vivo experiments, which further proved that the obstacle of the formation of granulation tissue leads to delayed wound healing after scald burn injury in mice.

Relationship between inflammatory responses and autophagy in lung tissues after scald and NOD2 signaling pathway in septic rats

Objective To evaluate the relationship between inflammatory responses and autophagy in lung tissues after scald and nucleotide-binding oligomerization domain-containing protein 2(NOD2)signaling pathway in septic rats. Methods Twenty SPF healthy male Sprague-Dawley rats, weighing 200-250 g, were divided into control group(group C, n=10) and sepsis after scald group(group SS, n=10)using a random number table.The rats were subjected to a third-degree scald burn covering 20% of total body surface area(body surface was shaved and then exposed to 99-100 ℃ water for 12 s), and 24 h later muramyldipeptide 5 mg/kg was intravenously injected to induce sepsis.The rats were only exposed to 20 ℃ water, and 24 h later normal saline 1 ml was given instead in group C. At 6 h after muramyldipeptide injection in group SS and at 6 h after normal saline injection in group C, arterial blood samples were collected for determination of serum tumor necrosis factor-α and interleukin-6 concentrations by enzyme-linked immunosorbent assay.Then rats were sacrificed and lungs were removed for measurement of activity of myeloperoxidase, NOD2 mRNA expression(using real-time polymerase chain reaction)and expression of receptor interacting protein 2, nuclear factor kappa Bp65 and microtubule-associated protein 1 light chain 3Ⅰ(LC3Ⅰ)and LC3Ⅱ in lung tissues(by Western blot). The LC3Ⅱ/Ⅰratio was calculated. Results Compared with group C, the expression of NOD2 mRNA, receptor interacting protein 2 and nuclear factor kappa Bp65 was significantly up-regulated, and the LC3Ⅱ/Ⅰratio and serum tumor necrosis factor-α and interleukin-6 concentrations were increased in group SS(P<0.05). Conclusion The mechanism underlying enhanced inflammatory responses and autophagy in lung tissues during sepsis after scald may be related to activation of NOD2 signaling pathway in rats. Key words: Burns; Sepsis; Nod2 signaling adaptor protein; Autophagy; Inflammation; Lung

Survey of the Statewide Impact of Payer Source on Referral of Small Burns to Burn Centers.

It is generally agreed that patients with large burns will be referred to organized burn centers, however, the referral of patients with smaller burns is less certain. A two-part survey was conducted to identify referral patterns for burn patients that meet American Burn Association referral criteria, and any effect insurance type might have on the referral patterns. The emergency departments of our state hospital association's member hospitals were contacted seeking a referral for a fictitious patient with a third-degree scald of the dominant hand. The referral sites were contacted twice, first stating that the patient had commercial insurance, next stating that the patient had Medicaid. Data collected included wait time for an appointment or reasons for denial of an appointment. Of 218 hospitals, 46 were excluded because they did not offer emergency care, and eight because they were listed as burn centers on the American Burn Association website. Of the remaining 164, 119 (73%) would refer to a burn center, 21 (13%) to a plastic surgeon, 10 (6%) to a hand surgeon, 7 (4%) to a wound center, 7 (4%) to another nonburn physician resource. There was no difference in wait time to the first available appointment with regards to insurance type (6.56 ± 4.68 vs 6.53 ± 5.05 days). Our state's referral pattern gives us insight into the regional referral pattern. This information will be used to guide a focused education and communication program to provide better service for the burn victims of our state.

The effect of electroacupuncture at ST36 on severe thermal injury-induced remote acute lung injury in rats

ObjectiveAcupuncture at ST36 can produce anti-inflammatory effects, which might be associated with vagus nerve activity. This study explored the effects of electroacupuncture (EA) at ST36 on severe thermal injury-induced remote acute lung injury in rats. InterventionsForty male Sprague-Dawley (SD) rats were randomly divided into five groups: (1) the sham (S) group, (2) the thermal injury (TEM) group subjected to 30% total body surface area (30% TBSA) third-degree scald, (3) the EA at ST36 group subjected to EA stimulation at ST36 (3V, 2ms, and 3Hz) after 30% TBSA scald, (4) the EA at non-acupoint group subjected to EA stimulation at non-acupoint after 30% TBSA scald, and (5) the α-bungarotoxin (α7 nicotinic acetylcholine receptor subunit antagonist) group administered 1.0μgkg−1 α-bungarotoxin before EA at ST36. Measurements and main resultsThermal injury of 30% TBSA induced leukocytosis in the alveolar space, interstitial edema, and the pro-inflammatory cytokines interleukin (IL)-1β, IL-6, and high-mobility group box 1 (HMGB-1); the expression of both HMGB-1 messenger RNA (mRNA) and protein in lung tissue was significantly enhanced. EA at ST36 significantly downregulated the levels of inflammatory cytokines and improved lung tissue injury. However, pretreatment with α-bungarotoxin reversed the effects of electrical stimulation of ST36. ConclusionsEA at ST36 might have a potential protective effect on severe thermal injury-induced remote acute lung injury via limitation of inflammatory responses in rats.

Estrogen-provided cardiac protection following burn trauma is mediated through a reduction in mitochondria-derived DAMPs

Mitochondria-derived danger-associated molecular patterns (DAMPs) play important roles in sterile inflammation after acute injuries. This study was designed to test the hypothesis that 17β-estradiol protects the heart via suppressing myocardial mitochondrial DAMPs after burn injury using an animal model. Sprague-Dawley rats were given a third-degree scald burn comprising 40% total body surface area (TBSA). 17β-Estradiol, 0.5 mg/kg, or control vehicle was administered subcutaneously 15 min following burn. The heart was harvested 24 h postburn. Estradiol showed significant inhibition on the productivity of H2O2 and oxidation of lipid molecules in the mitochondria. Estradiol increased mitochondrial antioxidant defense via enhancing the activities and expression of superoxide dismutase (SOD) and glutathione peroxidase (GPx). Estradiol also protected mitochondrial respiratory function and structural integrity. In parallel, estradiol remarkably decreased burn-induced release of mitochondrial cytochrome c and mitochondrial DNA (mtDNA) into cytoplasm. Further, estradiol inhibited myocardial apoptosis, shown by its suppression on DNA laddering and downregulation of caspase 1 and caspase 3. Estradiol's anti-inflammatory effect was demonstrated by reduction in systemic and cardiac cytokines (TNF-α, IL-1β, and IL-6), decrease in NF-κB activation, and attenuation of the expression of inflammasome component ASC in the heart of burned rats. Estradiol-provided cardiac protection was shown by reduction in myocardial injury marker troponin-I, amendment of heart morphology, and improvement of cardiac contractility after burn injury. Together, these data suggest that postburn administration of 17β-estradiol protects the heart via an effective control over the generation of mitochondrial DAMPs (mtROS, cytochrome c, and mtDNA) that incite cardiac apoptosis and inflammation.

Hyperglycemia Exacerbates Burn-Induced Liver Inflammation via Noncanonical Nuclear Factor-κB Pathway Activation

Hyperglycemia and inflammation are hallmarks of burn injury. In this study, we used a rat model of hyperglycemia and burn injury to investigate the effects of hyperglycemia on inflammatory responses in the liver. Hyperglycemia was induced in male Sprague-Dawley rats with streptozotocin (STZ) (35-40 mg/kg), followed by a 60% third-degree scald burn injury. Cytokine levels (by multiplex, in cytosolic liver extracts), hormones (by enzyme-linked immunosorbent assay [ELISA], in serum), nuclear factor (NF)-κB protein deoxyribonucleic acid (DNA) binding (by ELISA, in nuclear liver extracts) and liver functional panel (using VetScan, in serum) were measured at different time points up to 7 d after burn injury. Blood glucose significantly increased after burn injury in both groups with different temporal patterns. Hyperglycemic rats were capable of endogenous insulin secretion, which was enhanced significantly versus controls 12 h after burn injury. DNA binding data of liver nuclear extracts showed a robust and significant activation of the noncanonical NF-κB pathway in the hyperglycemic versus control burn animals, including increased NF-κB-inducing kinase expression (p < 0.05). Liver acute-phase proteins and cytokine expression were increased, whereas secretion of constitutive proteins was decreased after burn injury in hyperglycemic versus control animals (p < 0.05). These results indicate that burn injury to the skin rapidly activated canonical and noncanonical NF-κB pathways in the liver. Robust activation of the NF-κB noncanonical pathway was associated with increased expression of inflammatory markers and acute-phase proteins, and impaired glucose metabolism. Hyperglycemia is detrimental to burn outcome by augmenting inflammation mediated by hepatic noncanonical NF-κB pathway activation.

Effects of early excision and grafting on cytokines and insulin resistance in burned rats

Burn wound excision and grafting is a common clinical practice that decreases patient morbidity and mortality. It is not known, however, if the salutary effects of this procedure are related to effects on interleukin 6 (IL-6) and tumor necrosis factor (TNF-) α, and to reducing insulin resistance after burn. Sprague–Dawley rats were randomly divided into three groups: control, burn, burn ± excision groups. Rats in burn group were given a third-degree scald burn covering 30% total body surface area (TBSA) and no wound excision. Rats in burn ± excision group were subjected to a 30% third-degree burn followed by complete excision and allografting of the injury site within 15 min after burn. The rats in control group were treated in the same manner as the burn group, except that they were immersed in a room-temperature water. Glucose tolerance tests (GTT) were observed at 3 days after burn, euglycemic–hyperinsulinemic glucose clamps were performed at 4 days after burn and interleukin 6 (IL-6) and tumor necrosis factor (TNF-) α were determined after euglycemic–hyperinsulinemic glucose clamps. The levels of IL-6 and TNF-α increased after burn. Significant differences in GTT were observed between control and burn groups, and the rate of glucose infused measured in burned rats was significantly decreased compared with that in control at 4 days after burn. Early excision and grafting significantly decreased levels of IL-6 and TNF-α, and further reduced insulin resistance following thermal injury compared with burn group. Conclusion: Early excision and grafting appeared to have an effect on inflammatory mediators and further reduced insulin resistance induced by major burns.

Effect of sphingosine 1-phosphate on morphological and functional responses in endothelia and venules after scalding injury

Objective The uncontrolled increase of vascular permeability is the major obstacle in treatment of severe burns. Sphingosine 1-phosphate (S1P) has emerged as an important modulator of EC barrier function. This study was designed to explore the effect of S1P on morphological alteration in cultured endothelial cells (ECs) after burned plasma stimulation, and second to investigate the hyper-permeability response in intact vessels after scalding injury. Methods The distribution of VE-cadherin and F-actin was observed by double staining in primary cultured human umbilical vein endothelial cells (HUVECs) with immunofluorescence and fluorescent probes; respectively. Permeability changes were measured by a fluorescence ratio technique in isolated venules from rat skin. Burned plasma was obtained from a third-degree scald covering 30% of the total body surface area. Results The intervention with burned plasma on injured rats cultured HUVECs caused a significant disruption of intercellular adherens junction labeled by VE-cadherin staining, accompanied by the formation of F-actin stress fibers in the cells. S1P prevented or reversed these burned plasma-induced morphological alterations in cultured endothelial cells. The inhibition of S1P synthesis with N,N-dimethylsphingosine (DMS) mimicked the burned plasma-evoked redistribution of VE-cadherin and reorganization of F-actin. Venules isolated from burned rats demonstrated similar endothelial cytoskeleton changes with cultured cells under the influence of S1P or DMS. Both pre- and post-burn application of S1P attenuated increased permeability in isolated and perfused skin venules after burned plasma stimulation. Conclusion Our results indicate that S1P plays a role in maintaining basal vascular barrier function and could be protective in burn injury by enhancing the endothelial barrier function.

A ROLE FOR LONG CHAIN MYOSIN LIGHT CHAIN KINASE (MLCK-210) IN MICROVASCULAR HYPERPERMEABILITY DURING SEVERE BURNS

Microvascular leakage has been implicated in the pathogenesis of multiple organ dysfunction during trauma. Previous studies suggest the involvement of myosin light chain (MLC) phosphorylation-triggered endothelial contraction in the development of microvascular hyperpermeability. Myosin light chain kinase (MLCK) plays a key role in the control of MLC-phosphorylation status; thus, it is thought to modulate barrier function through its regulation of intracellular contractile machinery. The aim of this study was to further investigate the endothelial mechanism of MLC-dependent barrier injury in burns, focusing on the long isoform of MLCK (MLCK-210) that has recently been identified as the predominant isoform expressed in vascular endothelial cells. An MLCK-210 knockout mouse model was subjected to third-degree scald burn covering 25% total body surface area. The mesenteric microcirculation was observed using intravital microscopy, and the microvascular permeability was assessed by measuring the transvenular flux of fluorescein isothiocyanate-albumin. In a separate experiment, in vivo mesenteric hydraulic conductivity (Lp) was measured using the modified Landis technique. The injury caused a profound microvascular leakage, as indicated by a 2-fold increase in albumin flux and 4-fold increase in Lp at the early stages, which was associated with a high mortality within the 24-h period. Compared with wild-type control, the MLCK-210-deficient mice displayed a significantly improved survival with a greatly attenuated microvascular hyperpermeability response to albumin and fluid. These results provide direct evidence for a role of MLCK-210 in mediating burn-induced microvascular barrier injury and validate MLCK-210 as a potential therapeutic target in the treatment of burn edema.

Mesenteric lymph from burned rats induces endothelial cell injury and activates neutrophils.

Our previous studies indicated that mesenteric lymph duct ligation prevented burn-induced lung injury. Thus, the goal of the present study was to begin to investigate potential mechanisms of this protective effect. Prospective animal study with concurrent control. Small animal laboratory. Adult male Sprague-Dawley rats. Mesenteric lymph and portal vein plasma were collected from male rats subjected to a 40% third-degree scald burn or sham burn. The biological effects of these lymph and plasma samples were tested for their ability to kill human umbilical vein endothelial cells (HUVECs), increase HUVEC monolayer permeability, and activate polymorphonuclear leukocytes (PMNs), as reflected in CD11b adhesion molecule expression and superoxide production. Additionally, ileal specimens were harvested at the end of the experiment (6 hrs postburn) for histologic analysis. Postburn mesenteric lymph produced during the first 2 hrs after burn injury and tested at a 5% concentration, but not sham-burn lymph or portal plasma from burned rats, was toxic for HUVECs resulting in cell death after an 18-hr incubation period. Similarly, only postburn lymph increased HUVEC monolayer permeability. Postburn lymph activated both rat and human PMNs as reflected in increased CD11b expression and augmentation of the phorbol myristate acetate-induced superoxide response. Neither sham-burn lymph nor postburn portal vein plasma activated PMNs. Both the burn and sham-burn lymph samples were sterile, indicating that the effects of burn lymph on the HUVECs or PMNs were not due to translocating bacteria. Last, an association was found between burn-induced gut injury and the production of toxic burn lymph. Burn-induced gut injury results in the production of biologically active factors that are carried in the mesenteric lymph, but not the portal plasma, which injure endothelial cells and activate PMNs and thus could contribute to distant organ injury.

Myosin light chain kinase-dependent microvascular hyperpermeability in thermal injury.

Although the critical role of systemic inflammatory edema in the development of multiple organ failure in patients with massive burns has been fully recognized, the precise mechanisms responsible for the accumulation of blood fluid and proteins in tissues remote from the burn wound are poorly understood. The aim of this study was to test the hypothesis that circulating factors released during thermal injury cause microvascular leakage by triggering endothelial cell contraction and barrier dysfunction. A third-degree scald burn was induced in rats on the dorsal skin covering 25% total body surface area. The microcirculation and transvascular flux of albumin were observed in the rat mesentery using intravital fluorescence microscopy. The direct effect of circulating factors on microvascular barrier function was assessed by measuring the apparent permeability coefficient of albumin in isolated rat mesenteric venules during perfusion of plasma freshly withdrawn from burned rats. The in vivo study showed that the transvenular flux of albumin was significantly increased over a 6-h period with a maximal response seen at 3 h postburn. Importantly, perfusion of noninjured venules with burn plasma induced a time-dependent increase in albumin permeability. Pharmacological inhibition of protein kinase C, Src tyrosine kinases, or mast cell activation did not significantly affect the hyperpermeability response; however, blockage of myosin light chain phosphorylation with the myosin light chain kinase inhibitor ML-7 greatly attenuated the burn-induced increase in venular permeability in a dose-related pattern. The results support a role for endogenous circulating factors in microvascular leakage during burns. Myosin light chain phosphorylation-dependent endothelial contractile response may serve as an end-point effector leading to microvascular barrier dysfunction.

Gender-related differences in myocardial inflammatory and contractile responses to major burn trauma.

Gender-related differences in immune responses to hemorrhage and sepsis have been described. However, most trauma studies continue to limit experimental models to males to avoid the variable responses associated with hormonal fluctuation in proestrus/estrus females. In the present study, male and female (either diestrus or proestrus/estrus) Sprague-Dawley rats (250-325 g) were given a third-degree scald burn over 40% total body surface area and fluid resuscitated (4 ml/kg per %burn of lactated Ringer solution); sham burn males and diestrus as well as sham burn proestrus/estrus female rats were included to provide controls. Twenty-four hours postburn, hearts were either perfused to examine mechanical function (Langendorff, n = 8 to 9 hearts/group) or to prepare cardiomyocytes (collagenase digestion, n = 4 to 5 hearts/group). Left ventricular developed pressure and the positive and negative first derivative of left ventricular pressure responses to increases in preload were significantly lower in burned males compared with responses measured in either burned proestrus/estrus or burned diestrus females; burn trauma increased cardiomyocyte secretion of tumor necrosis factor-alpha, interleukin-1beta, and nitric oxide to a lesser extent in proestrus/estrus females than levels secreted by either diestrus females or males. Similarly, myocytes from proestrus/estrus females accumulated significantly less sodium/calcium compared with values measured in males (P < 0.05). Our data confirm gender-related differences in myocardial function and myocardial inflammatory responses to burn injury.

I kappa B overexpression in cardiomyocytes prevents NF-kappa B translocation and provides cardioprotection in trauma.

This study examined the effects of either IkappaBalpha overexpression (transgenic mice) or N-acetyl-leucinyl-leucinyl-norleucinal (ALLN) administration (proteosome inhibitor in wild-type mice) on cardiomyocyte secretion of tumor necrosis factor-alpha (TNF-alpha) and on cardiac performance after burn trauma. Transgenic mice were divided into four experimental groups. IkappaBalpha overexpressing mice were given a third-degree scald burn over 40% of the total body surface area or wild-type littermates were given either a scald or sham burn to provide appropriate controls. Pharmacological studies included ALLN (20 mg/kg) administration in either burned wild-type mice or wild-type shams. Burn trauma in wild-type mice promoted nuclear factor-kappaB (NF-kappaB) nuclear translocation, cardiomyocyte secretion of TNF-alpha, and impaired cardiac performance. IkappaBalpha overexpression or ALLN treatment of burn trauma prevented NF-kappaB activation in cardiac tissue, prevented cardiomyocyte secretion of TNF-alpha, and ablated burn-mediated cardiac contractile dysfunction. These data suggest that NF-kappaB activation and inflammatory cytokine secretion play a significant role in postburn myocardial abnormalities.

Effect of 2,4-diamino-6-hydroxy-pyrimidine on postburn Staphylococcus aureus sepsis in rats

Guanosine triphosphate-cyclohydrolase I (GTP-CHI) is the first and rate-limiting enzyme for the de novo biosynthesis of biopterin. The objective of present study was to observe the effect of 2,4-diamino-6-hydroxy-pyrimidine (DAHP), an inhibitor of GTP-CHI, on the development of postburn Staphylococcus aureus sepsis. A prospective, controlled animal study. A research laboratory in a hospital. Male Wistar rats. Fifty-six male Wistar rats were randomly divided into four groups as follows: normal control group (n = 10), scald control group (n = 10), postburn sepsis group (n = 20), and DAHP treatment group (n = 16). In the scald control group, rats were subjected to a 20% total body surface area third-degree scald injury and then were killed at 24 hrs. In the postburn sepsis group (n = 20), rats were inflicted with 20% total body surface area third-degree scald followed by Staphylococcus aureus challenge, and they were further divided into 2- and 6-hr groups. In the DAHP treatment group (n = 16), animals were intraperitoneally injected with a dose of 1 g/kg DAHP before Staphylococcus aureus challenge and then were further divided into 2- and 6-hr groups. Tissue samples from liver, kidneys, lungs, and heart were collected to determine GTP-CHI, inducible nitric oxide synthase, and tumor necrosis factor-alpha messenger RNA expression. Meanwhile, biopterin and nitric oxide concentrations in these tissues were also measured. After the scald injury followed by Staphylococcus aureus challenge, GTP-CHI messenger RNA expression and biopterin concentrations were significantly elevated in various tissues such as liver, heart, kidneys, and lungs, as were the values of inducible nitric oxide synthase messenger RNA expression and nitric oxide formation (p <.01). Pretreatment with DAHP significantly reduced GTP-CHI/biopterin induction (p <.05-.01), and the up-regulation of inducible nitric oxide synthase/nitric oxide was also suppressed. Furthermore, DAHP administration inhibited the gene expression of tumor necrosis factor-alpha. Two hours after septic challenge, tumor necrosis factor-alpha messenger RNA expression in liver, kidneys, and lungs in the DAHP-treated group was 35.7%, 37.3%, and 33.0% of that in the postburn septic group, respectively. Additionally, in animals without DAHP treatment, the 6-hr mortality rate was 55.6% (20 of 36), whereas it was only 25.0% in DAHP-treated animals (4 of 16, p =.08). Early treatment with DAHP might be a potential strategy to prevent the development of postburn Staphylococcal sepsis, which appears to be associated with down-regulation of biopterin and nitric oxide formation by DAHP.

IGF-I/BP-3 administration preserves hepatic homeostasis after thermal injury which is associated with increases in no and hepatic NF-kappa B.

After a severe trauma, such as a cutaneous thermal injury, an increase in hepatocyte apoptosis has been associated with hepatocyte damage and impairment in hepatic function. Insulinlike growth factor-I (IGF-I) exerts antiapoptotic effects in several organs, thus improving organ homeostasis. The purpose of the present study was to determine whether IGF-I in combination with its principle binding protein-3 (BP-3) attenuates liver damage after a burn and whether this attenuation is through signals of the apoptotic-proliferative axis of hepatocytes. Sprague-Dawley rats (56 males) received a 60% total body surface area third-degree scald burn and were randomly divided to receive either rhlGF-I/BP3 (10 mg/kg/day s.c.) or saline (control). Serum aspartate transaminase (AST) and nitric oxide (NO), and hepatocyte proliferation and apoptosis, were measured on postburn days 1, 2, 5, and 7. Hepatic interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) mRNA and hepatic nuclear-factor kappa B (NF-kappa B) were determined at 1 and 2 days postburn. IGF-I/BP-3 decreased serum AST and increased serum NO at 1, 2, and 5 days after burn when compared with controls (P < 0.05). IGF-I/BP-3 increased hepatocyte proliferation on the first day after burn and decreased hepatocyte apoptosis at day 7 postburn when compared with controls (P < 0.05). IGF-I/BP-3 decreased hepatic IL-1 beta and TNF-alpha mRNA 1 day after burn (P < 0.05). IGF-I/BP-3 further increased hepatic NF-kappa B concentration 1 and 2 days postburn when compared with controls (P < 0.05). Recombinant hIGF-I in combination with its principle binding protein conserves hepatic homeostasis, which is associated with a transient increase in hepatocyte proliferation and decrease in hepatocyte apoptosis possibly through NO and hepatic NF-kappa B.

Resistance of macrophages to the suppressive effect of interleukin-10 following thermal injury.

The activation of a macrophage (Mphi)-dependent proinflammatory cascade following thermal injury plays an important role in the development of immunosuppression and increased susceptibility to subsequent sepsis in burn patients. In contrast, although interleukin (IL)-10, an anti-inflammatory cytokine that can downregulate M phi activity, has also been implicated in postburn immune dysfunction, its role in the regulation of M phi function postburn remains unclear. To study this, C57BL/6 female mice were subjected to a 25% total body surface area third-degree scald burn, and splenic Mphis were isolated 7 days later. Lipopolysaccharide (LPS)-stimulated IL-10, IL-6, tumor necrosis factor (TNF)-alpha, and nitric oxide (NO) production were significantly increased in the burn group compared with shams. Blockade of endogenous IL-10 activity enhanced IL-6 and TNF-alpha release, but not NO release, in both groups. The addition of exogenous IL-10 to the M phi cultures dose dependently suppressed production of these inflammatory mediators in both groups. The timing of IL-10 addition to the cultures in relation to LPS stimulation, however, was critical. The suppressive effect of exogenous IL-10 was attenuated in both groups when the cells were exposed to IL-10 at 4-6 h after LPS stimulation; however, Mphis from injured mice were significantly better able to maintain inflammatory mediator-productive capacity. The resistance of Mphis from injured mice to IL-10-mediated suppression correlated with decreased IL-10 receptor (IL-10R) expression and increased CD11b expression. These findings suggest that Mphis, following thermal injury, display resistance to suppression by IL-10 due in part to downregulation of IL-10R expression.

Effect of Varying Burn Sizes and Ambient Temperature on the Hypermetabolic Rate in Thermally Injured Rats

Background. Small animals with scald covering 50% of their total body surface area (TBSA) have been used to study the hypermetabolic burn response. In the 50% TBSA burn rat model, the area of normal skin that is available for animal instrumentation is restricted and the mortality rate has been high. The purpose of this study was to determine whether a smaller burn size can induce a similar hypermetabolic response with mortality rates lower than those of the 50% TBSA model.Methods. Rats were randomly divided into four groups to receive a 0% (sham nonburned), 30%, 40%, or 50% TBSA third-degree scald burn. The hypermetabolic response was determined by measuring changes in body weight and oxygen consumption at ambient temperatures of 21, 26, and 31°C for each burn size. Weight measurements were made daily while oxygen consumption was measured 7, 11, and 14 days after thermal injury.Results. All thermally injured rats lost body weight; however, there were no significant differences between the 30, 40, and 50% TBSA burn groups. Burn induced a hypermetabolic response as indicated by an increase in oxygen consumption from 130 to 200% that of sham nonburned rats. No significant difference in oxygen consumption could be shown over the study period between the three burn sizes at different ambient temperatures. Mortality was 0% in the sham and 30% group, 10% for the 40% group, and 50% for the 50% TBSA burn group.Conclusions. From our study we conclude that a burn size covering 30% of the TBSA induces the same hypermetabolic response as a 50% TBSA burn.

Does burn wound excision after thermal injury attenuate subsequent macrophage hyperactivity and immunosuppression?

Studies have shown that cell mediated immunity is suppressed markedly following thermal injury. Macrophages and the activation of an inflammatory cascade that includes interleukin (IL)-1, IL-6, tumor necrosis factor-alpha (TNFalpha) and PGE2 have been implicated as causative factors. Burn wound excision and grafting is a common clinical practice that decreases patient morbidity and mortality. It is not known, however, if the salutary effects of this procedure are related to modulation of macrophage activity post-burn. Therefore, C57BL/6 female mice were subjected to a third-degree scald burn covering 25% of their total body surface area followed by complete excision and allografting of the injury site at 8, 24, or 72 h post-burn. Splenic macrophage function was assessed 7 days post-burn. Thermal injury without burn excision and grafting significantly increased macrophage TNFalpha, IL-6, nitric oxide, and PGE2 production in response to lipopolysaccharide stimulation, whereas IL-1beta production was not increased. Burn wound excision and grafting normalized TNFalpha production to sham levels, independent of when post-burn the procedure was conducted. In contrast, the elevated production of other inflammatory mediators (IL-1beta, IL-6, nitric oxide, PGE2) post-burn was unaffected by burn wound excision and grafting. Moreover, splenic T-lymphocyte proliferation was also suppressed at 7 days post-burn and was not improved by burn wound excision and grafting. These results, therefore, suggest that the beneficial effects of burn wound excision and grafting are likely to be related to the normalization of macrophage TNFalpha production as well as the maintenance of skin barrier function.