Pathophysiology of burn injury

The plasma of thermally injured rats showed dramatic increases in levels of xanthine oxidase activity, with peak values appearing as early as 15 minutes after thermal injury. Excision of the burned skin immediately after the thermal injury significantly diminished the increase in plasma xanthine oxidase activity [51, 53]. The skin permeability changes were attenuated by treating the animals with antioxidants (catalase, SOD, dimethyl sulfoxide, dimethylthiourea) or an iron chelator (DFO), thus supporting the role of oxygen radicals in the development of vascular injury as defined by increased vascular permeability [53]. Allopurinol, a xanthine oxidase inhibitor, markedly reduced both burn lymph flow and levels of circulating lipid peroxides, and further prevented all pulmonary lipid peroxidation and inflammation. This suggests that the release of oxidants from burned tissue was in part responsible for local burn edema, as well as distant inflammation and oxidant release [73]. The failure of neutrophil depletion to protect against the vascular permeability changes and the protective effects of the xanthine oxidase inhibitors (allopurinol and lodoxamide tromethamine) suggests that plasma xanthine oxidase is the more likely source of the oxygen radicals involved in the formation of burn edema. These oxygen radicals can increase vascular permeability by damaging microvascular endothelial cells [51, 53]. the use of antioxidants has been extensively investigated in animals, and some clinical trials suggest benefit. Antioxidants (vitamin C and E) are routinely administered to patients at many burn centers. High doses of antioxidant ascorbic acid (vitamin-C) have been found to be efficacious in reducing fluid needs in burn injured experimental animals when administered postburn [80–82]. The use of high doses (10–20g per day) of vitamin C was shown to be effective in one clinical trial, but ineffective in another [83, 84]. High dose vitamin C has not received wide clinical usage.

Platelet aggregation factor: Platelet aggregation (or activating) factor (PAF) can increase capillary permeability and is released after burn injury [66, 85]. Ono et al. showed in scald-injured rabbits that TCV-309 (Takeda Pharmaceutical Co Ltd., Japan), a PAF antagonist, infused soon after burn injury blocked edema formation in the wound and significantly inhibited PAF increase in the damaged tissue in a dose-dependent manner. In contrast, the

superoxide dismutase content in the group treated with TCV-309 was significantly higher than that of the control group [85]. These findings suggest that the administration of large doses of a PAF antagonist immediately after injury may reduce burn wound edema and the subsequent degree of burn shock by suppressing PAF and superoxide radical formation.

Angiotensin II and vasopressin: Angiotensin II and vasopressin or antidiuretic hormone (ADH), are two hormones that participate in the normal regulation of extracellular fluid volume by controlling sodium balance and osmolality through renal function and thirst [50]. However, during burn shock where sympathetic tone is high and volume receptors are stimulated, both hormones can be found in supranormal levels in the blood. Both are potent vasoconstrictors of terminal arterioles will little affect on the venules. Angiotensin II may be responsible for the selective gut and mucosal ischemia, which can cause translocation of endotoxins and bacteria and the development of sepsis and even multi-organ failure [86, 87]. In severely burn-injured patients angiotensin II levels were elevated two to eight times normal in the first 1 to 5 days after burn injury with peak levels occurring on day three [88]. Vasopressin had peak levels of 50 times normal upon admission and declined towards normal over the first five days after burn injury. Vasopressin, along with catecholamines may be largely responsible for increased system vascular resistance and left heart afterload, which can occur in resuscitated burn shock. Sun and others used vaso- pressin-receptor antagonist in rats with burn shock to improve hemodynamics and survival time, while vasopressin infusion exacerbated burn shock [89].

Corticotrophin-releasing factor: Cortico- trophin-releasing factor (CRF) has proven to be efficacious in reducing protein extravasation and edema in burned rat paw. CRF may be a powerful natural inhibitory mediator of the acute inflammatory response of the skin to thermal injury [90].

Hemodynamic consequences of acute burns

The cause of reduced cardiac output (CO) during the resuscitative phase of burn injury has been the subject of considerable debate. There is an immediate

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depression of cardiac output before any detectable reduction in plasma volume. The rapidity of this response suggests a neurogenic response to receptors in the thermally injured skin or increased circulating vasoconstrictor mediators. Soon after injury a developing hypovolemia and reduced venous return undeniably contribute to the reduced cardiac output. The subsequent persistence of reduced CO after apparently adequate fluid therapy, as evidenced by a reduction in heart rate and restoration of both arterial blood pressure and urinary output, has been attributed to circulating myocardial depressant factor(s), which possibly originates from the burn wound [21, 22]. Demling and collegues showed a 15% reduction in CO despite an aggressive volume replacement protocol after a 40% scald burn in sheep [28]. However, there are also sustained increases in catecholamine secretion and elevated systemic vascular resistance for up to five days after burn injury [70, 88]. Michie and others measured CO and SVR in anesthetized dogs resuscitated after burn injury [91]. They found that CO fell shortly after injury and then returned toward normal, however, reduced CO did not parallel the blood volume deficit. They concluded that the depression of CO resulted not only from decreased blood volume and venous return, but also from an increased SVR and from the presence of a circulating myocardial depressant substance. Thus, there are multiple factors that can significantly reduce CO after burn injury. However, resuscitated patients suffering major burn injury can also have supranormal CO from 2 to 6 days post-injury. This is secondary to the establishment of a hypermetabolic state.

Hypermetabolic response to burn injury

Marked and sustained increases in catecholamine, glucocorticoid, glucagon and dopamine secretion are thought to initiate the cascade of events leading to the acute hypermetabolic response with its ensuing catabolic state [92–100]. The cause of this complex response is not well understood. However, interleukins 1 and 6, platelet-activating factor, tumor necrosis factor (TNF), endotoxin, neutrophil-adher- ence complexes, reactive oxygen species, nitric oxide and coagulation as well as complement cascades

have also been implicated in regulating this response to burn injury [101]. Once these cascades are initiated, their mediators and by-products appear to stimulate the persistent and increased metabolic rate associated with altered glucose metabolism seen after severe burn injury [102].

Several studies have indicated that these metabolic phenomena post-burn occur in a timely manner, suggesting two distinct pattern of metabolic regulation following injury [103]. The first phase occurs within the first 48 hours of injury and has classically been called the “ebb phase” [103, 104], characterized by decreases in cardiac output, oxygen consumption, and metabolic rate as well as impaired glucose tolerance associated with its hyperglycemic state. These metabolic variables gradually increase within the first five days post-injury to a plateau phase (called the “flow” phase), characteristically associated with hyperdynamic circulation and the above mentioned hypermetabolic state. Insulin release during this time period was found to be twice that of controls in response to glucose load [105, 106] and plasma glucose levels are markedly elevated, indicating the development of an insulin-resistance [106, 107]. Current understanding has been that these metabolic alterations resolve soon after complete wound closure. However, recent studies found that the hypermetabolic response to burn injury may last for more than 12 months after the initial event [92, 93, 100, 108]. We found in recent studies that sustained hypermetabolic alterations post-burn, indicated by persistent elevations of total urine cortisol levels, serum cytokines, catecholamines and basal energy requirements, were accompanied by impaired glucose metabolism and insulin sensitivity that persisted for up to three years after the initial burn injury [109].

A 10 to 50-fold elevation of plasma catecholamines and corticosteroid levels occur in major burns which persist up to three years post-injury [49, 109–112]. Cytokine levels peak immediately after burn, approaching normal levels only after one month post injury. Constitutive and acute phase proteins are altered beginning 5–7 days post-burn, and remain abnormal throughout acute hospital stay. Serum IGF-I, IGFBP-3, parathyroid hormone, and Osteocalcin drop immediately after the injury 10 fold, and remain significantly decreased up to

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6 months post-burn compared to normal levels [111]. Sex hormones and endogenous growth hormone levels decrease around 3 weeks post-burn [111].

For severely burned patients, the resting metabolic rate at thermal neutral temperature (30°C) exceeds 140% of normal at admission, reduces to 130% once the wounds are fully healed, then to 120% at 6 months after injury, and 110% at 12 months postburn [92, 111]. Increases in catabolism result in loss of total body protein, decreased immune defenses, and decreased wound healing [92].

Immediately post-burn patients have low cardiac output characteristic of early shock [113]. However, three to four days post-burn, cardiac outputs are greater than 1.5 times that of non-burned, healthy volunteers [111]. Heart rates of pediatric burn patients’ approach 1.6 times that of non-burned, healthy volunteers [114]. Post-burn, patients have increased cardiac work [110, 115]. Myocardial oxygen consumption surpasses that of marathon runners and is sustained well into rehabilitative period [115, 116].

There is profound hepatomegaly after injury. The liver increases its size by 225% of normal by two weeks post-burn and remains enlarged at discharge by 200% of normal [111].

Post-burn, muscle protein is degraded much faster than it is synthesized [111, 114]. Net protein loss leads to loss of lean body mass and severe muscle wasting leading to decreased strength and failure to fully rehabilitate [117, 118]. Significant decreases in lean body mass related to chronic illness or hypermetabolism can have dire consequences. A 10% loss of lean body mass is associated with immune dysfunction. A 20% loss of lean body mass positively correlates with decreased wound healing. A loss of 30% of lean body mass leads to increased risk for pneumonia and pressure sores. A 40% loss of lean body mass can lead to death [119]. Uncomplicated severely burned patients can lose up to 25% of total body mass after acute burn injury [120]. Protein degradation persists up to nearly one year post severe burn injury resulting in significant negative wholebody and cross-leg nitrogen balance [110, 118, 121]. Protein catabolism has a positive correlation with increases in metabolic rates [118]. Severely burned patients have a daily nitrogen loss of 20–25 grams per

meter squared of burned skin [110, 122]. At this rate, a lethal cachexia can be reached in less than one month [122]. Burned pediatric patients’ protein loss leads to significant growth retardation for up to 24 months post injury [123].

Elevated circulating levels of catecholamines, glucagon, cortisol after severe thermal injury stimulate free fatty acids and glycerol from fat, glucose production by the liver, and amino acids from muscle [103, 124, 125]. Specifically, glycolytic-gluconeo- gengenic cycling is increased 250% during the postburn hypermetabolic response coupled with an increase of 450% in triglyceride-fatty acid cycling. [126]. These changes lead to hyperglycemia and impaired insulin sensitivity related to post-receptor insulin resistance demonstrated by elevated levels of insulin, fasting glucose, and significant reductions in glucose clearance [127–130].

Glucose metabolism

Glucose metabolism in healthy subjects is tightly regulated: under normal circumstances, a postprandial increase in blood glucose concentration stimulates release of insulin from pancreatic -cells. Insulin mediates peripheral glucose uptake into skeletal muscle and adipose tissue and suppresses hepatic gluconeogenesis, thereby maintaining blood glucose homeostasis [131, 132]. In critical illness, however metabolic alterations can cause significant changes in energy substrate metabolism. In order to provide glucose, a major fuel source to vital organs, release of the above mentioned stress mediators oppose the anabolic actions of insulin [133]. By enhancing adipose tissue lipolysis [125] and skeletal muscle proteolysis [134], they increase gluconeogenic substrates, including glycerol, alanine and lactate, thus augmenting hepatic glucose production in burned patients (Fig. 1) [131, 132, 135]. Hyperglycemia fails to suppress hepatic glucose release during this time [136] and the suppressive effect of insulin on hepatic glucose release is attenuated, significantly contributing to post-trauma hyperglycemia [137]. Catecho- lamine-mediated enhancement of hepatic glycogenolysis, as well as direct sympathetic stimulation of glycogen breakdown, can further aggravate the hyperglycemia in response to stress [132]. Cat-

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echolamines have also been shown to impair glucose disposal via alterations of the insulin signaling pathway and GLUT-4 translocation muscle and adipose tissue, resulting in peripheral insulin resistance (Fig. 1) [131, 138]. Cree et al. [137] showed an impaired activation of Insulin Receptor Substrate-1 at its tyrosine binding site and an inhibition of AKT in muscle biopsies of children at seven days post-burn. Work of Wolfe et al. indicates links between impaired liver and muscle mitochondrial oxidative function, altered rates of lipolysis, and impaired insulin signaling post-burn attenuating both the suppressive actions of insulin on hepatic glucose production and on the stimulation of muscle glucose uptake [106, 125, 136, 137]. Another counter-regulatory hormone of interest during stress of the critically ill is glucagon. Glucagon, like epinephrine, leads to increased glucose production through both gluconeogenesis and glycogenolysis [139]. The action of glucagons alone is not maintained over time; however, its action on gluconeogenesis is sustained in an additive manner with the presence of epinephrine, cortisol, and growth hormone [133, 139]. Likewise, epinephrine and glucagon have an additive effect on glycogenolysis [139]. Recent studies found that proinflammatory cytokines contribute indirectly to post-burn hyperglycemia via enhancing the release of the above mentioned stress hormones [140–142]. Other groups showed that inflammatory cytokines, including tumor necrosis factor (TNF), interleukin (IL) -6 and monocyte chemotactic protein (MCP) -1 also act via direct effects on the insulin signal transduction pathway through modification of signaling properties of insulin receptor substrates, contributing to post-burn hyperglycemia via liver and skeletal muscle insulin resistance [143–145]. Alterations in metabolic pathways as well as pro-inflammatory cytokines, such as TNF, have also been implicated in significantly contributing to lean muscle protein breakdown, both during the acute and convalescent phases in response to burn injury [121, 146]. In contrast to starvation, in which lipolysis and ketosis provide energy and protect muscle reserves, burn injury considerably reduces the ability of the body to utilize fat as an energy source.

Skeletal muscle is thus the major source of fuel in the burned patient, which leads to marked wasting of lean body mass (LBM) within days after injury

[110, 147]. This muscle breakdown has been demonstrated with whole body and cross leg nitrogen balance studies in which pronounced negative nitrogen balances persisted for 6 and 9 months after injury [118]. Since skeletal muscle has been shown to be responsible for 70–80% of whole body insulin-stimu- lated glucose uptake, decreases in muscle mass may significantly contribute to this persistent insulin resistance post-burn [148]. The correlation between hyperglycemia and muscle protein catabolism has been also supported by Flakoll et al. [149] in which an isotopic tracer of leucine was utilized to index whole-body protein flux in normal volunteers. The group showed a significant increase in proteolysis rates occurring without any alteration in either leucine oxidation or non-oxidative disposal (an estimate of protein synthesis), suggesting an hyperglycemia induced increase in protein breakdown. Flakoll et al. [149] further demonstrated that elevations of plasma glucose levels resulted in a marked stimulation of whole body proteolysis during hyperinsulinemia. A 10–15% loss in lean body mass has been shown to be associated with significant increases in infection rate and marked delays in wound healing [150]. The resultant muscle weakness was further shown to prolong mechanical ventilatory requirements, inhibit sufficient cough reflexes and delay mobilization in protein-malnourished patients, thus markedly contributing to the incidence of mortality in these patients [151]. Persistent protein catabolism may also account for delay in growth frequently observed in our pediatric patient population for up to 2 years post-burn [123].

Septic patients have a particularly profound increase in metabolic rates and protein catabolism up to 40% more compared to those with like-size burns that do not develop sepsis [92, 152, 153]. A vicious cycle develops, as patients that are catabolic are more susceptible to sepsis due to changes in immune function and immune response. The emergence of multi-drug resistant organisms have led to increases in sepsis, catabolism and mortality [153–155]. Modulation of the hypermetabolic, hypercatabolic response, thus preventing secondary injury is paramount in the restoration of structure and function of severely burned patients.

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