Atiyeh B.S., Al-Amm C.A.

Division of Plastic and Reconstructive Surgery, American University of Beirut Medical Center, Beirut, Lebanon

SUMMARY. Major injury due to trauma and burns and injury causing massive haemorrhage have been demonstrated to increase susceptibility to infectious complications and related multiple organ failure primarily because of a suppressed immune system. This is an interesting phenomenon that is still incompletely understood. Why should it be that just when an individual is injured and most needs to be able to resist infection, there is immunosuppression? Various components of the complex immune system have been incriminated, including neutrophil dysfunction, abnormality in opsonic activity, depletion of the complement cascade, helper cell dysfunction, macrophage dysfunction, production of soluble immunosuppressive substances, stress-associated hormones, and an increase in the prostaglandin E2 (PGE2) level. The most recent evidence suggests that the immunosuppression is the result of T-cell dysfunction with failure of interleukin-2 production. This may be only an endpoint of macrophage hyperactivity and activation of a pro-inflammatory cascade including prostaglandins and reactive nitrogen intermediates with the aberrant production of what is called “garbage” non-specific immunoglobulin G1.

Introduction

Despite recent advances in burn wound management, sepsis remains the main cause of morbidity and mortality in patients resuscitated after major thermal injury.1-3 Infectious complications account for as many as 75% of late deaths among seriously injured patients who survive resuscitation. Increased susceptibility to infection has been related to impaired humoral and cell-mediated immune response2,4,5 not only in burn victims but also following major trauma or haemorrhage.6 It seems that the normal immune defence mechanisms start to become suppressed in burn injuries of around 25% TBSA and in traumatic conditions with an Injury Score Scale of 25.7

Why it should be that there is immunosuppression whenever an individual is seriously injured and most needs to be able to resist infection remains unclear.8 Even though decreased immune responses have long been observed in severely burned patients (as evidenced by decreased delayed-type skin hypersensitivity allowing the advantageous use of allografts, xenografts, and other biological dressings without the rapid onset of an acute rejection phenomenon), the immune paradox of burns is still poorly understood (Fig. 1). It has been attributed to loss of barrier functions, tissue ischaemia and destruction, foreign body introduction, and inoculation with micro-organisms.6 The factors incriminated also include neutrophil dysfunction, abnormality in opsonic activity, depletion of the complement cascade, helper-cell dysfunction, macrophage dysfunction, production of soluble immunosuppressive substances, stress-associated hormones, and an increase in the PGE2 level.9 Several mechanisms have been proposed to explain the wide variety of abnormalities that exist, but as yet no explanation has proved wholly satisfactory.

Discussion

Clonal expansion

As a response to injury or bacterial invasion, it is now recognized that many immunological reactions require the rapid proliferation of a small number of cells into several million. This phenomenon has been termed clonal expansion (Fig. 2).10 It is largely controlled by interleukin-2 (IL-2), a cytokine produced primarily by peripheral blood mononuclear cells1 (PBMC) and T-helper (Th1) lymphocytes.3 In addition to its clonal expansion effect, IL-2 seems to be involved in T-cell blastogenesis and the promotion of both specific T-cell cytotoxicity and non-specific natural killer cytotoxicity.1,11 IL-2 production was found to be clearly reduced in burn patients. The reduction is significantly correlated to the extent of the burn. IL-2 levels return to normal only late during the recovery phase.1 The cause of IL-2 deficiency is by no means clear. It was initially thought to be caused by suppressor T-cells of the T8 subset activated by an increase in PGE2 released from monocytes.12,13 Recent data report other subsets of T-lymphocytes as being responsible.1,3,6,11,17

Complement cascade

Burn injury is characterized by three concentric zones. Centrally there is a zone of coagulation necrosis surrounded by an area of marginally viable tissues that may still be salvageable, surrounded by an area of intense inflammation. Devitalized tissue has exposed C3b binding sites as well as self-antigens. It is a powerful activator of the alternate complement system. Moreover, bacteria that may colonize the necrotic tissues are also powerful activators of the complement system.8 In major burns, actual complement depletion, in particular of the alternate complement factors, has been observed. Activation of the complement cascade leads to the diffusion of chemotactic factors in the surrounding blood stream (Fig. 3). Complement split factors, in turn, activate the neutrophils, leading to regional endothelial cell adhesion and migration.14 At the same time, lymphokines originally stored in the tissues or subsequently produced by invading cells are released in the wound itself. This stimulates monocyte invasion and potentiates their maturation into tissue macrophages, which are the central cells responsible for wound clearing of devitalized tissues, bacteria, and large amounts of self-antigens by the process of phagocytosis. This process is further enhanced by the opsonizing properties of the complement factors. Oxygen free radicals, lysosomes, and inflammatory cytokines are released into the wound as a result of phagocytosis. In small burns, these mediators remain confined locally and are rapidly cleared by neutralizing enzymes in surrounding tissues and by macrophages. In major burns, these mediators spill over into the systemic circulation. Such mediators are normally cleared by the lungs. Aggregates of macrophages, as also cytokines in the circulation reaching the lungs, stimulate the abundant lung macrophages. When exaggerated, this stimulation leads to lung injury. In cases of massive lung injury, and whenever lung capacity is overwhelmed by the massive influx of haematogenous mediators, systemic diffusion of these mediators occurs, leading to stimulation of macrophages and other immune competent cells in distant organs, causing injury and ultimately multiple organ failure.2,15-18

Th1 and Th2 lymphocytes

In addition to its phagocytic activity, the macrophage is responsible for antigen presentation to other immune competent cells, namely the lymphocytes (Fig. 4). Along with the major histocompatibility complex antigens, antigen presentation at the surface of the macrophages stimulates several T-lymphocyte CD4+ subtypes, including the progenitor helper cells Th0 which can be induced to mature into either Th1 or Th2 cells. The Th1 phenotype promotes cellular and humoral opsonizing immunity, whereas the Th2 phenotype inhibits it, though stimulating the less efficient non-opsonizing humoral immunity (IgE and IgG2) responsible for anaphylactic immune reactions.3,6,9 Interleukin IL-12 is mainly responsible for Th0 to Th1 transformation, thus promoting the immune response, while I-L4 and IL-10 have the opposite effect, facilitating the expression of the Th2 phenotype and thus inhibiting the development of an efficient immune response. IL-4, -5, -6, -10, and -13 expressed in the Th2 pathway predominate following major burns or massive trauma at the expense of IL-12, exerting a negative feedback on the macrophages and leading to a depressed immune state.1,3,6,11,17

Macrophage hyperstimulation

In major burns there is an abundant release of pro-inflammatory mediators. Soluble complement factors, tumour necrosis factor ·, interferon Á, plus a plethora of antigens, all contribute to macrophage hyperstimulation (Fig. 5).13,15,16,18-20 The stimulated macrophage, which is probably necessary in the acute phase of burns, becomes an “inhibitory macrophage” after several days (7-10 days in the mouse model).11 Although it still produces IL-1 and IL-6, it also changes its co-signalling, such that lymphocytes respond differently. IL-2 and IL-12 receptors become downregulated and the global immune proliferative and cytokine response become depressed. The stimulated macrophage initiates the production of IL-10, which potentiates the Th2 cascade (Figs. 6,7). IL-10 is a global immunosuppressant. It inhibits the Th1 cascade and exerts a depressant feedback effect on the macrophage. The macrophage also secretes PGE2, which is a modulator of the immune response. Reactive nitrogen intermediates (RNI) (normally synthesized by the macrophage and retained in intracellular granules which are usually utilized to fight specific intracellular pathogens such as Listeria, Legionella, Brucella, Mycobacterium, Toxoplasma, and other types of parasites) are released extracellularly following major burn injury.13 RNI suppress lymphocyte proliferation and specifically reduce the Th1 response. PGE2 and RNI stimulate aberrant production of what is called “garbage” non-specific immunoglobulin G1 (IgG1) (Fig. 8).

Gamma-delta lymphocytes

A special subset of T lymphocytes with Á‰ surface receptors, present in high concentrations in the skin, gut, and uterus and constituting an essential part of the innate immune system, seems to be involved in immune monitoring and epithelial repair (Fig. 9). Experimentally, mice lacking this cell type have a high initial mortality following burn injury. These lymphocytes are probably the first to be involved in the initiation of the immune reaction leading to increased macrophage stimulation. In major burns, these cells have been detected in the spleen and they have also been found to cause further systemic macrophage stimulation. Restoration of a normal immune response several weeks after the initial burn injury is associated with the emergence of a CD4+ contra-suppressor cell separate from the CD4+ Th1 or Th2 cells.21

The concept of a “suppressor” T-lymphocyte, which was abandoned in the 1980s, has recently seemed to be regaining ground (Fig. 10). Apparently, two suppressor cells, not just one, may be active. Th2 is a very likely suppressor in addition to the Á‰ lymphocyte. In promoting a pro-inflammatory macrophage, this cell participates in the suppression of the acquired immune response.19

Neutrophil action

The overwhelming role of the macrophage in immune response modulation following massive burns or injury must not overshadow the role of other cell types, namely the neutrophil. In regular situations, local chemoattractants cause neutrophil adhesion to endothelial cells. Actin polymerization/depolymerization allows the changing of cellular configuration, which is essential for cellular translocation into the zone of injury. Once in place, the release of neutrophil granules will initiate control of local infection in addition to clearing tissue necrosis. In situations of intense injury, neutrophil hyperstimulation causes irreversible actin polymerization with subsequent rigidity in cell wall structure (Fig. 11). This leads to incapacity of the neutrophil to translocate, causing blockage of the microcirculation and haematogenous release of neutrophil granules, which are distributed systemically.

Deitch’s two-hit theory

The recently advanced two-hit theory (Figs. 12a,b) greatly helps our understanding of the macrophage overstimulation which is central in the immune dysfunction observed in successfully resuscitated patients following major burns or massive trauma.16,18 An initial insult usually results in an adequate, well-orchestrated response of the immune cascade. Following the insult, the macrophage becomes primed, so that a secondary insult will result in a disproportionate response. Macrophage overstimulation leads to global downgrading of the immune response. In major burns, the second hit is most likely related to the gastrointestinal tract, which invariably loses its barrier function, leading to bacterial and endotoxin translocation. This is normally cleared by the liver, which has the highest macrophage concentration in the Kupffer cells. Overstimulation of these cells will lead to further systemic release of pro-inflammatory mediators reaching the lungs, which may contribute to the lung injury. On the other hand, Kupffer cell failure observed in cases of liver failure leads to uncontrolled delivery of translocated bacteria and endotoxins to the lungs and, when bypassed, to the systemic circulation, which adds to the secondary insult.

Alcohol and burns

It is also worth noting that alcohol-exposed burn patients were roughly three to six times more likely to die after thermal injury,15 primarily because of increased septic complications and decreased host defences. The significance of this observation is enhanced by the fact that nearly 50% of all adult burn patients admitted to hospitals in the USA are found to have had alcohol in their bloodstream at the time of their injury. The defects observed in cellular immune responses in ethanol-exposed and thermally injured subjects were largely mediated by increased production of IL-6 both systemically and by splenic macrophages. In high concentrations IL-6 inhibits immune function.

Strategies

Corticosteroids

Glucocorticoids are among the most potent regulators of IL-6 gene expression.15 Alcohol-associated suppression of cellular immune responsiveness in burn patients can be significantly, though partially, reversed by exogenous cortisone administration. This reversal is accompanied by attenuation of both circulating levels and macrophage-derived IL-6 production. Physiological doses of corticosteroids normally have a potentiating effect on the immune response. Pharmacological doses have variable and often contradictory effects on cytokine expression, receptor modulation, and proliferative activity. At high doses, corticosteroids have proved to be disappointing in clinical trials in subjects suffering from major trauma or burns.

Nonsteroidal anti-inflammatory drugs (NSAID)

Attempts to block the rise of PGE2 levels observed following major burn injuries or to avert its action in order to improve the immune response have been disappointing. NSAID have been partially successful in restoring the lymphocyte proliferative response. The restorative effect of NSAID is independent of their capacity to inhibit the cyclogenase pathway and to affect PGE2 levels. The most effective dosage is lower than the anti-inflammatory dosage. Weaker anti-inflammatory drugs such as salicylic acid and 5-aminosalicylic acid are more effective. Alternate pathways other than PGE2 level suppression may be involved. PGE2 is almost completely degraded by the lungs at the first passage. It may have systemic effects only in cases of extreme lung damage. Even then, its antagonism at best only partially restores the proliferative immune response.

Cytokine modulation

Owing to “cytokine networking” and the complexity of the multi-level regulatory mechanisms of the cytokine cascades, cytokine modulation has so far proved to be too complicated and only partially successful. Surprisingly, antagonizing IL-10, which is the main immunosuppressant, has resulted in a paradoxical increase in mortality. It seems that the generalized immune depression observed in successfully resuscitated severely injured patients might be a protective response against hyperstimulated pro-inflammatory macrophages preventing immune self-consumption and depletion. Furthermore, following major burn injuries, self-antigens become expressed. Immunosuppression may be instrumental in averting an auto-immune reaction. Administration of IL-12 to potentiate the Th1 immune cascade has been associated at least in one patient with an auto-immune reaction. It is believed that if the immune response is to be modulated it will have to be effected at several levels by selective use of both stimulatory and inhibito ry agents. At present, in the absence of a reliable test to determine the immunostimulation/immunosuppression balance, and in view of the narrow therapeutic range of available agents, immunomodulation may be too dangerous to attempt.

RESUME. Il a été démontré que les lésions les plus importantes causées par les traumatismes et les brûlures, comme aussi les lésions qui provoquent les hémorragies massives, augmentent sensiblement la susceptibilité aux complications infectives et à l’insuffisance organique multiple associée, causée principalement par la suppression du système immun. Ce phénomène intéressant n’a pas été bien compris. Quelle est la cause de la manifestation de l’immunosuppression qui se vérifie précisément quand une personne a subi une lésion et donc a plus besoin de résister à l’infection? On a pensé à divers aspects du complexe système immun: une dysfonction des neutrophiles, une abnormalité de l’activité opsonique, la déplétion de la cascade du complément, une dysfonction des cellules helper, une dysfonction des macrophages, la production de substances immunosuppressives solubles, les hormones associés des prostaglandines E2. L’évidence la plus récente indiquerait que l’immunosuppression est causée par une dysfonction des cellules T, avec l’insuffisance de la production de l’interleukine-2. Ceci est peut-être seulement le point final de l’hyperactivité des macrophages et de l’activation d’une cascade pro-inflammatoire, y compris les prostaglandines et les intermédiares d’azote réactifs, avec la production aberrante de ce qu’on appelle l’immunoglobuline non-spécifique “ordure” G1 (Ig G1).

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