Endotoxemia and release of tumor necrosis factor and interleukin la in acute heatstroke ABDERREZAK BOUCHAMA, RANJIT S. PARHAR, ADNAN EL-YAZIGI, KIRKITANT SHETH, AND SULTAN AL-SEDAIRY Departments of Medicine, Biological and Medical Research, and Pathology and Laboratory King Faisal Specialist Hospital and Research Centre, Riyadh 11211, Saudi Arabia BOWCHAMA, ABI)ERREZAK,RANJITS.PARHAR,ADNAN ELYAZIGI,KIRKITANT SHETH, AND SULTAN AL-SEDAIRY~~~Otoxemia and release of tumor necrosis factor and interleukin 1a in acute heatstroke. J. Appl. Physiol. 70(6): 2640-2644,1991.-To

determine whether endotoxemia and release of tumor necrosis factor (TNF-a) and/or interleukin la! (IL-la) are involved in the pathogenesis of heatstroke, 17 adult patients with a mean rectal temperature of 42.1 t 0.2OC were studied. Blood samples were taken on admission and after cooling was completed. TNF-CY and IL-la levels were measured by enzyme-linked immunosorbent assay, and lipopolysaccharide (LPS) content was measured by the chromogenic substrate modification of the Limule amebocyte lysate. TNF-ar, IL&, and LPS were elevated in all patients [ 199 * 25 (SE) pg/ml, 480.5 t 68.3 pg/ml, and 8,60 & 1.19 rig/ml, respectively, compared with normal control values of 31.4 * 8.4 pg/ml, 53.7 t 5.32 pg/ml, and ~9 pg/ml]. There was no significant correlation between temperature and the circulating concentration of TNF-cu, IL-la, and LPS. Postcooling TNF-cu, IL&x, and LPS concentrations were significantly decreased but still above normal control values. The findings suggest that these mediators may have a role in the pathogenesis of heatstroke that could change the strategy of management. hyperthermia;

heat stress; cooling;

sepsis

HEATSTROKE causes a syndrome

similar to that of sepsis or endotoxemia: shock with elevated cardiac output and reduced systemic vascular resistance (27)) lactic acidosis (23, 29), and adult respiratory distress syndrome (14), culminating in multiorgan system failure and death (1, 9). The cause of this metabolic disorder and tissue injury is not well understood but is attributed to the direct cellular toxicity of heat (4, 26). Recent reports in animals suggest that endotoxin may have a role in the pathophysiology of heatstroke (3, 12, 16, 17). It has been demonstrated in primates subjected to heat stress that endotoxin enters the circulation at 40°C and its concentration increases with the temperature (17). Early reports implicated plasma endotoxin as a cause of circulatory collapse, coagulation disorders, and death in heatstroke victims (7, 20). Gram-negative bacterial endotoxin is the most potent known stimulator of the release of tumor necrosis factor (TNF-cu) (28). TNF-cw and interleukin la! (IL-la) are endogenous pyrogens that produce fever and are essential mediators of shock and tissue injury during endotoxemia (11, 32). 2640

0161-7567/91

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Medicine,

This study was undertaken to determine whether heatstroke is associated with endotoxemia and the release of TNF-ac and/or IL-la. MATERIALS AND METHODS

The study was conducted at the heatstroke center of King Faisal Hospital, Makkah, Saudi Arabia, during the 1989 pilgrimage season (July 12-15). The patients included in the study had classic (nonexertional) heatstroke, diagnosed by a rectal temperature >4O.l “C associated with neurological abnormalities such as delirium, convulsions, and coma (1). The time between onset of heatstroke and admission to the heatstroke center could not be determined precisely, but the average time was lO pg/ml. Ten plasma samples from healthy individuals were included in the test for determination of reference values, and the mean was 31.4 t 8.4 pglml. IL-la! was measured by an enzyme-linked immunosorbent assay technique by use of a commercial kit (Endogen, Boston, MA). This method detects IL-la! levels >30 pg/ml. Reference values for IL-la! were determined as for TNF-a; the mean was 53.7 t 5.32 pg/ml. To exclude the possibility that lipids, rheumatoid factor, and acute-phase proteins (6,13) had interfered with the assay results, additional experiments were performed in 5 of the 17 heatstroke patients with low-tohigh cytokine levels. Cytokine measurements in serial

1991 the American

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Society

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ENDOTOXIN

TABLE 1. Patient

&F,

AND CYTOKINES

data on admission

and leukocyte count are given in Table 1. Metabolic acidosis was noted in some patients with a high anion gap, which has been shown to be due to lactate in heatstroke (23, 29). Hyperglycemia and leukocytosis were also noted. No predisposing disease was found. One patient had a history of coronary artery disease, and no patient was taking any medication. On admission, the mean plasma concentrations of LPS, TNF-cu, and IL-la in the 17 patients were 8.60 t 1.19 rig/ml, 199 t 25 pg/ml, and 480.5 t 68.3 pg/ml, respectively. The relationship between temperature and LPS, TNF-cu, and IL-la! plasma concentration is shown in Fig. 1. There was no significant correlation between temperature and LPS (r = 0.36, P = 0.X), TNF-a (r = 0.24, P = 0.35), or IL-la (r = 0.39, P = 0.13). The correlation between LPS, TNF-cu, and IL-la was also analyzed (Fig. 2). There was no significant correlation between LPS and TNF-cu (r = 0.46, P = 0.06), LPS and IL-la (r = 0.47, P = 0.057), or TNF-cr and IL-la (r = 0.06, P = 0.82). Paired plasma samples were obtained from 7 of the 17 patients pre- and postcooling. The mean plasma concentrations of LPS (12.26 t 1.86 rig/ml), TNF-cw (247.4 t 41.2 pg/ml), and IL-la! (579.3 t 60.7 pg/ml) decreased significantly after cooling was completed to 6.12 t 1.18 rig/ml, 133 t 27.9 pg/ml, and 285 t 48 pg/ml, respectively (P < 0.05, Fig. 3). In two of the seven patients, the plasma levels of TNF-a! increased slightly. In one patient, this was associated with sustained endotoxemia.

5522.2

Yr

No. of males/no. of females Rectal temperature, “C PH Arterial Pco~, Torr Arterial PO,, Torr HCO,‘, mM Anion gap Blood glucose, mM Leukocyte count, X10*/1

1017 42.1t0.2 7.38kO.06 28k6 105k28 l&4

19*5 13&4 12.2k9.4

Values are means t SE of 17 patients. Data for pH, arterial PCO~ and PO,, and HCO, were obtained from 12 patients, and those for blood glucose and leukocyte count were obtained from 6 patients. Anion gap was calculated according to the formula Na+ - (Cl- + HCO,).

dilutions of patients’ plasma samples showed a dilution pattern parallel to the standard curve. After addition of recombinant cytokines, 91-99% recovery was achieved. Rheumatoid factor interference was excluded by adding 10 ~1 of 0.2 M dithiothreitol to plasma samples. The recovery of cytokine was unchanged. The chromogene substrate modification of the limulus amebocyte lysate technique (Kabi Diagnostica, Sweden) was used for determination of lipopolysaccharide (LPS) concentration in the samples. This technique has been described in detail elsewhere (10). The detection limit of the assay was 9 pg/ml. Statistical analysis. The data were analyzed on a Tandon AT-compatible microcomputer with the use of a standard statistical package (Statgraphics, STSC, Rockville, MD). Paired t tests were used to compare results pre- and postcooling, and linear regression was used to determine correlation coefficients. Differences were considered significant when P < 0.05. All values are expressed as means t SE.

DISCUSSION

In this study it was found that plasma concentrations of tumor TNF-ar, IL-la, and LPS were elevated in all patients with classical heatstroke. The concentrations decreased significantly after cooling but remained above the control values. It has been reported that TNF-a! is released in human volunteers after endotoxin injection (25) and in various diseases (18, 22) with marked variation in the peak circulatory levels and clinical response. In the present series of classical heatstroke, the plasma TNF-cu levels were between those observed in adult vol-

RESULTS

This study comprised 17 consecutive adult patients with a mean rectal temperature of 42.1 -t- 0.2OC (range 40.6-43‘3°C). Age, sex, arterial blood gas [corrected for body core temperature (30)], anion gap, blood glucose, 1300

r

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0’37 38 ’ 39‘ 40’ 41’ Um(SEU)blbdvJu



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TEMPERATURE,

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= 31.4(8.4)



43

FIG. 1. Correlation of lipopolysaccharide ature in heatstroke.



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40



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TEMPERATURE,



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IL-1 = 53.7(5.32)pg/ml

(LPS), tumor necrosis factor (TNF), and interleukin

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TEMPERATURE, LPS

Endotoxemia and release of tumor necrosis factor and interleukin 1 alpha in acute heatstroke.

To determine whether endotoxemia and release of tumor necrosis factor (TNF-alpha) and/or interleukin 1 alpha (IL-1 alpha) are involved in the pathogen...
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