William H. Alarcon

Can the clearance of tumor necrosis factor alpha and interleukin 6 be enhanced using an albumin dialysate hemodiafiltration system

Patients with acute hepatic failure (AHF) have elevated levels of inflammatory cytokines such as tumor necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6). Recently, we have shown selective hemodiafiltration with albumin dialysis, as an extracorporeal liver support device (ECLVS), to be effective in the clearance of multiple toxins that are elevated in AHF. Our objective was to evaluate whether ECLVS would be effective in the clearance of TNF-alpha and IL-6. An in vitro continuous hemodiafiltration circuit was used with single pass counter-current dialysis. A known amount of recombinant rat TNF-alpha and IL-6 was added to heparinized bovine blood and filtered across a polyalkyl sulfone hemofilter using matched filtration and dialysate flow rates. During 4 hours, the serial TNF-alpha and IL-6 concentrations were measured in the circulating blood, and the content of each cytokine was calculated using mass balance. For each cytokine, clearance was determined for two dialysate groups at constant temperature and pH (group 1: dialysate = 0.9 normal saline, n = 5; group 2: dialysate = albumin 2 gm/dl, n = 5). Analysis of data was performed using ANOVA and Students t-test. There was improved clearance of TNF-alpha and IL-6 when albumin was used in the dialysate (81+/-0.09% of the initial TNF-alpha and 77+/-0.04% of the IL-6 quantities) compared with when 0.9 normal saline was used as the dialysate (58+/-0.14% of the initial TNF-alpha and 56+/-0.18% of the IL-6 quantities); p < 0.03. An ECLVS utilizing hemodiafiltration with albumin dialysis is more effective than conventional hemofiltration in the clearance of TNF-alpha and IL-6 and, therefore, may benefit patients with acute hepatic failure.

Thermal injury induces expression of CD14 in human skin

BACKGROUND Skin is equipped with an array of immune mediators aimed at fighting invading microbes. CD14 has been shown to play a key role in modulating the activation of cells by LPS. Since LPS levels within burn wounds are often found to be elevated, we sought to examine the expression of CD14 within human skin following thermal injury. METHODS Patients who sustained partial thickness burns, were recruited into the study (n=57). Total RNA was isolated from both burn and normal (control) skin. Northern blot analysis and TaqMan RT-PCR were used to determine skin CD14 mRNA levels. Immunohistochemistry was used to localize CD14 expression in burned and normal skin. RESULTS Quantitative PCR showed significantly increased CD14 expression levels in the immediate post-burn period (P<0.05 burn versus non-burn). Immunohistochemistry revealed more pronounced CD14 staining 24 h after the injury, reaching normal levels approximately 5-7 days post-burn. CONCLUSION CD14 expression peaks within the first week post-burn before declining, reaching normal levels after 14 days. This loss of supranormal CD14 expression locally within the wound may contribute to a weakened host defense response 5-6 days after injury, when patients become especially vulnerable to infection.

Skin lipopolysaccharide-binding protein and IL-1β production after thermal injury

In response to a burn injury, skin can have an inflammatory response characterized by the production of inflammatory cytokines, recruitment of immune cells, containment of invading organisms, and clearance of noxious substances from the wound. Lipopolysaccharide-binding protein (LBP) is a molecule that is capable of coordinating all 4 functions; we previously found evidence that suggested that LBP is produced within surgical wounds. Because of the central role of LBP in the response to bacterial infection, as well as in the high rate of infection after burn injuries, we sought to determine whether a thermal injury could affect wound LBP production and thereby affect host responses against bacterial infection. Rats were given either a burn or a sham burn and were killed 24, 48, and 72 hours after the injuries. Wound specimens were assayed for bacterial counts and for the presence of LBP, messenger (m)RNA, and interleukin (IL)-1beta mRNA. Wound LBP mRNA was significantly upregulated at 24 hours in the group with burn injuries (P < .05; burn vs sham burn); this was followed by decreases at 48 and 72 hours. Immunohistochemistry showed LBP protein in the epidermis of animals with burns. Bacterial counts increased in the group with burn injuries (P < .05; burn vs sham burn) and continued to rise for 72 hours. IL-1beta mRNA levels were elevated at all time points in the group with burn injuries (P < .05). These results suggest an inverse correlation between burn wound LBP expression and bacterial wound counts. This failure to maintain local LBP production after severe thermal injury despite localized inflammation shown by high IL-1beta levels may predispose local wounds to bacterial invasion.