- Email: [email protected]
Circulating levels of granulocyte macrophage colony-stimulating factor in patients with the systemic inflammatory response syndrome
Journal of Infection (2003) 47, 296–299
www.elsevierhealth.com/journals/jinf
Circulating levels of granulocyte macrophage colony-stimulating factor in patients with the systemic inflammatory response syndrome Donato Torrea,*, Roberto Tambinia, Mariangela Manfredib, Valerio Manganic, Paola Livid, Viviana Maldifassie, Maurizio Chiarandae, Paolo Campib, Filippo Speranzaa a
Section of Pediatric Infectious Diseases, Regional Hospital, Viale Borri 57, 21100 Varese, Italy Department of Clinical and Laboratory Immunology, San Giovanni di Dio Hospital, Florence, Italy c Intensive Care Unit, San Giovanni di Dio Hospital, Florence, Italy d Department of Anaesthesia and Intensive Care, Careggi Hospital, Florence, Italy e Department of Anaesthesia and Intensive Care, Regional Hospital, Varese, Italy b
Accepted 13 April 2003
KEYWORDS Systemic inflammatory response syndrome; Granulocytemacrophage; Colony stimulating factor
Summary Objectives. Granulocyte-macrophage colony stimulating factor (GM-CSF) is a key regulator cytokine that modulates the proliferation and maturation of polymorphonuclear and mononuclear progenitors. This study was designed to investigate and clarify the role of GM-CSF in 52 critically ill patients with systemic inflammatory response syndrome (SIRS). Methods. Serum levels of GM-CSF were detected by an immunoenzyme assay. Results. Our results clearly show that the serum concentrations of GM-CSF were significantly elevated in patients with infectious and noninfectious SIRS (33.2 ^ 45.7 pg/ml, controls: 17.2 ^ 9.8 pg/ml; p ¼ 0:0303). In addition, GM-CSF levels significantly decreased in patients with SIRS, particularly in patients with infectious SIRS, 5 and 7 days later. There was a clear tendency toward higher levels of GM-CSF in patients with poor, as compared with those having a good outcome of the disease. Conclusion. These results show that GM-CSF may play an important role in patients with infectious and noninfectious SIRS, and that GM-CSF levels progressively and significantly decrease in patients with infectious SIRS. Q 2003 The British Infection Society. Published by Elsevier Ltd. All rights reserved.
Introduction The systemic inflammatory response syndrome (SIRS), a massive pro-inflammatory immune state, is associated with a number of clinical infective and noninfective conditions.1 SIRS can develop inde*Corresponding author. Tel.: þ 39-0332-278446; fax: þ 390332-278580. E-mail address: [email protected]
pendently of any infection, including pancreatitis, polytrauma, or immune complex disease. Infectious as well as noninfectious events may activate cytokines and nitric oxide,2,3 and in particular increased levels of pro-inflammatory cytokines, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1 and interleukin (IL)-8, have been reported in septic patients, often correlating with severity of the disease.4 Neutrophils are considered to be important players in the host defense system during
0163-4453/$30.00 Q 2003 The British Infection Society. Published by Elsevier Ltd. All rights reserved. doi:10.1016/S0163-4453(03)00065-3
GM-CSF levels in patients with SIRS
the acute inflammatory response; their motility to infection or inflammatory sites is mediated through systemic release of chemotactic substances, and various cytokines, including granulocyte-colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF).5 The role of GM-CSF in patients with infectious or noninfectious SIRS is unclear and only two studies on this cytokine have been published. Preesneill et al.6 have demonstrated in the sera of 82 critically ill patients with sepsis and septic shock a significant increase of granulocyte-colony stimulating factor, whereas serum levels of GM-CSF were not elevated in these patients. In contrast, Fanning et al.7 showed increased levels of circulating GM-CSF in six patients with SIRS. In the present study, we determined circulating levels of GM-CSF in 52 critically ill patients with infectious or noninfectious SIRS to better clarify its role during the acute inflammatory response.
297
Cytokine assay To detect serum levels of GM-CSF, samples were taken from patients on admission and/or at the time of diagnosis, and after 1, 5 and 7 days. Healthy subjects served as the control group. Serum for cytokine detection was prepared from blood, centrifuged, and stored in 1 ml coded aliquots at 2 20 8C. Serum levels of GM-CSF were determined by commercially available solid-phase sandwich ELISA kit (R & D Systems, Minneapolis, USA), using the protocol recommended by the manufacture. The sensitivity of this ELISA kit was less than 3 pg/ml.
Statistical analysis
Materials and methods
Results are demonstrated as mean and standard deviation (^ 1 SD). Mean values were compared using unpaired Student’s t test with Welch’s correction. Analysis of variance with the Kruskal – Wallis test was used to assess results of cytokine levels at various time. p , 0:05 was considered statistically significant.
Patients
Results
In this prospective study, the patient population consisted of 52 adult critically ill patients, admitted to four intensive care units (Careggi and San Giovanni di Dio Hospital in Florence, Varese and Busto Arsizio) during a 9-month period. Patients were considered to have SIRS as defined in the guidelines of the America College of Chest Physician/Society of Critical Care Medicine.1 These criteria include two or more of the following conditions: temperature . 38 8C or , 36 8C, a heart rate . 90 beats/min, a respiratory rate . 20 breaths/min or PaCO2 , 32 Torr, a withe blood cell count . 12,000 cells ml or , 4000 cells/ml, or . 10% immature band forms. Sepsis was defined as a systemic response to infection including the criteria for SIRS. Patients with severe sepsis had to fulfill the criteria for sepsis including inadequate organ perfusion as indicated by hypoxemia, and/or oliguria, or hypotension, as manifested by a systolic blood pressure of , 90 mm Hg or a reduction of . 40 mm Hg from baseline without other causes for hypotension. SIRS patients had no signs of focal infection, and no microorganisms were isolated from blood cultures. The etiology of these patients includes trauma, cardiogenic shock, surgical interventions, pulmonary embolia, acute pulmonary edema, and intracranial hemorrhage.
The demographic characteristics of our patients with SIRS are illustrated in Table 1. As shown, we evaluated 52 patients, 31 of whom with infectious SIRS, whereas 21 patients suffered from noninfectious SIRS. Fifteen out of 31 patients had sepsis,
Table 1 Demographic characteristics of 52 patients with SIRS. Variables Gender (male/female) Age (years)
36/16 62.7 ^ 18.7
Infectious SIRS ðn ¼ 31Þ Sepsis Severe sepsis Septic shock Multiple organ dysfunction
15 9 7 17
Noninfectious SIRS ðn ¼ 21Þ Surgical interventions Trauma Pulmonary embolism Intracranial hemorrhage Acute pulmonary edema Cardiogenic shock
6 4 3 2 1 5
Mortality SIRS Infectious SIRS Noninfectious SIRS
15/52 (28.8%) 10/31 (32.2%) 5/21 (23.8%)
298
D. Torre et al.
Table 2 Serum levels of GM-CSF in 52 patients with SIRS at various days. Patients
SIRS, overall ðn ¼ 52Þ Infectious SIRS ðn ¼ 31Þ Noninfectious SIRS ðn ¼ 21Þ
GM-CSF (pg/ml) 0
1
5
7 days
33.2 ^ 45.7 30.6 ^ 39.7 37.1 ^ 54.2
38.4 ^ 64.3 40.6 ^ 65.7 34.5 ^ 64.1
22.3 ^ 26.2 21.9 ^ 23.2 22.6 ^ 30.1
18.6 ^ 21.6* 12.7 ^ 7.6** 26.8 ^ 31.0
*p ¼ 0:0148, and **p ¼ 0:0248 (Kruskal–Wallis test).
nine patients had severe sepsis, seven patients had septic shock, and 17 patients had multiple organ dysfunction. SIRS patients had a mortality of 28.8%, whereas mortality of patients with infectious SIRS and noninfectious SIRS was 32.2 and 23.8%, respectively. Results of serum levels of GM-CSF in SIRS patients and controls are shown in Fig. 1. A significant increase of GM-CSF (33.2 ^ 45.7 pg/ml, median: 16.20 pg/ml; controls: 17.2 ^ 9.8 pg/ml, median 10.5 pg/ml; p ¼ 0:0303) levels was observed in patients with SIRS, and in particular in patients with infectious SIRS (30.6 ^ 39.7 pg/ml, median: 18.0 pg/ml; p ¼ 0:0466) at the time of diagnosis, whereas GM-CSF levels of patients with noninfectious SIRS were not significantly increased (37.1 ^ 54.2 pg/ml, median 12.5 pg/ml). Furthermore, serum levels of GM-CSF were determined in SIRS patients at the time of diagnosis, and 1, 5 and 7 days later (Table 2). As shown, a significant decrease of GM-CSF levels was noted in the patients, in particular in patients with infectious SIRS, 5 and 7 days later, whereas patients with noninfectious SIRS had progressively, but nonsignificant, lower GM-CSF levels, especially after 5 and 7 days. Table 3 shows GM-CSF levels in SIRS patients with a good or poor outcome. As can be seen, GM-CSF
Figure 1 Serum levels of GM-CSF in 52 patients with SIRS, of whom 31 patients with infectious SIRS and 21 patients with noninfectious SIRS on admission. Horizontal bar indicates mean ^ SD levels of GM-CSF.
levels nearly reached statistical significance ðp ¼ 0:0537Þ in predicting poor outcome in patients with SIRS, whereas GM-CSF levels of patients with infectious or noninfectious SIRS separately were not predictive of poor outcome.
Discussion The results of this study clearly show increased levels of GM-CSF in patients with infectious and noninfectious SIRS. Among several mediators operating in SIRS, proinflammatory cytokines TNF-alpha, IL-1 and interleukin-6 are considered to play a key role, especially in patients with sepsis or septic shock.8 However, further studies have demonstrated a proinflammatory role of several other cytokines in SIRS, such as IL-8, interferon gamma and G-CSF.5,8,9 Kuhns et al.9 have shown increased levels of GCSF after intravenous administration of endotoxin in healthy volunteers. It is interesting to note that IL-1 is a strong inducer of GM-CSF and macrophagecolony stimulating factor.8 However, the role of GM-CSF in SIRS and in sepsis syndrome has not been fully investigated. Increased, but not significant, levels of GM-CSF were observed in four of nine healthy volunteers, who were treated with endotoxin at various dosages.9 Presneill et al.6 who studied 82 critically ill patients with sepsis, shock without sepsis, and septic shock, found that GM-CSF levels were rarely elevated in the first hours to days in many critically ill patients, particularly those with sepsis and septic shock. It should be noted that detection limit of GM-CSF levels of the immunoassay used by Presneill et al.6 was much higher (100 pg/ml) than that used in our immunoassay (3 pg/ml); furthermore, we cannot rule out that coagulation of blood of SIRS patients may interfere with GM-CSF production, inasmuch as plasma samples could influence GM-CSF levels in SIRS patients than serum samples. Furthermore, circulating GM-CSF was detectable in some patients with meningococcal septic shock.10
GM-CSF levels in patients with SIRS
299
Table 3 Predictive value of GM-CSF levels of outcome in patients with SIRS. Patients
SIRS, overall ðn ¼ 52Þ Infectious SIRS ðn ¼ 31Þ Noninfectious SIRS ðn ¼ 21Þ
GM-CSF (pg/ml) Good outcome
Poor outcome
15.0 ^ 18.0 ðn ¼ 37Þ 14.5 ^ 20.1 ðn ¼ 21Þ 15.6 ^ 15.4 ðn ¼ 16Þ
56.0 ^ 74.5* ðn ¼ 15Þ 44.8 ^ 62.5 ðn ¼ 10Þ 78.5 ^ 98.5 ðn ¼ 5Þ
*p ¼ 0:0537 by unpaired t test with Welch’s correction.
The failure to detect GM-CSF in the sera of patients with sepsis and septic shock may be misleading. In fact, Metcalf et al.11 suggest that in transgenic mice circulating GM-CSF is rapidly consumed by cells bearing GM-CSF receptors, such that free GM-CSF may be undetectable in serum by available assays. However, Fanning et al.7 showed that GM-CSF was consistently and significantly elevated in six patients with SIRS, and that GMCSF appears to be a major factor inhibiting neutrophil apoptosis in the systemic circulation of patients with SIRS. GM-CSF may also play a dominant role in inhibiting neutrophil apoptosis in the lungs of patients with adult respiratory distress syndrome.12 In fact, human neutrophils, in response to stimulation by lipopolysaccharide, in vitro secrete, several cytokines, including IL-1 ra, IL-4, and GM-CSF.13 It should be noted that other cytokines, including TNF alpha, IL-1, IL-6 and IL-8 could predict factors of fatal outcome in the infectious and noninfectious SIRS. However, several studies have shown that plasma levels of IL-1, TNF alpha, IL-6 fluctuate in patients with SIRS, and display a poor correlation with mortality,14,15 whereas in patients with septic shock, TNF alpha, IL-6 and IL-8 significantly correlate with fatal outcome.16,17 In conclusion, these findings of elevated levels of GM-CSF in patients with infectious and noninfectious SIRS suggest that GM-CSF, may stimulate the immune system in response to microbial and inflammatory stimuli may contribute to the pathogenesis in SIRS.
References 1. Bone RC, Balk RA, Cerra FB, et al. ACCP/SCCM consensus conference. Definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis. Chest 1992;101:1644—1655. 2. Casey L, Balk RA, Bone RC. Plasma cytokine and endotoxin levels correlate with survival in patients with sepsis syndrome. Ann Intern Med 1993;119:771—778. 3. Kilbourn R, Griffith O. Overproduction of nitric oxide in cytokine-mediated and septic shock. J Natl Cancer Inst 1992; 84:827—831.
4. Biliau A, Vandekerckhove F. Cytokines and their interactions with other inflammatory mediators in the pathogenesis of sepsis and septic shock. Eur J Clin Invest 1991;21:559—573. 5. Colotta F, Re F, Polentarutti N, Sozzani S, Mantovani A. Modulation of granulocyte survival and programmed cell death by cytokines and bacterial products. Blood 1997;89: 2012—2020. 6. Presneill JJ, Waring PM, Layton JE, et al. Plasma granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor levels in critical illness including sepsis and septic shock: relation to disease severity, multiple organ dysfunction, and mortality. Crit Care Med 2000;28: 2344—2354. 7. Fanning NF, Kell MR, Shorten GD, Kirwan WO, Bouchier-Hayes D, Cotter T, Redmond HP. Circulating granulocyte macrophage colony-stimulating factor in plasma of patients with the systemic inflammatory response syndrome delays neutrophil apoptosis through inhibition of spontaneous reactive oxygen species generation. Shock 1999;11:167—174. 8. Davies MG, Hagen PO. Systemic inflammatory response syndrome. Br J Surg 1997;84:920—935. 9. Kuhns DB, Alvord WG, Gallin JI. Increased circulating cytokine antagonists, and E-selectin after intravenous administration of endotoxin in humans. J Infect Dis 1995; 171:145—152. 10. Waring PM, Presneill JJ, Maher DW, et al. Differential alterations in plasma colony-stimulating factor concentrations in meningococcaemia. Clin Exp Immunol 1995; 102:501—506. 11. Metcalf D, Rasko JEJ. Leukemic transformation of immortalized FDC-P1 cells engrafted in GM-CSF transgenic mice. Leukemia 1993;7:878—886. 12. Matute-Bello G, Liles WC, Radella F, et al. Neutrophil apoptosis in the acute respiratory distress syndrome. Am J Respir Crit Care Med 1997;156:1969—1977. 13. Re F, Mengozzi M, Muzio M, Dinarello CA, Mantovani A, Colotta F. Expression of interleukin-1 receptor antagonist (IL-1 ra) by human circulating polymorphonuclear cells. Eur J Immunol 1993;23:570—573. 14. Friedland JS, Porter JC, Daranani S, et al. Plasma proinflammatory cytokine concentrations, acute physiology and chronic health evaluation (APACHE) III scores and survival in patients in an intensive care unit. Crit Care Med 1996;24: 1775—1781. 15. Presterl E, Staudinger T, Pettermann M, et al. Cytokine profile and correlation to the APACHE III and MPM II scores in patients with sepsis. Am J Respir Crit Care Med 1997;156:825—832. 16. Martin C, Boisson C, Haccoun M, Thomachot L, Mege JL. Patterns of cytokine evolution (tumor necrosis factor-alpha and interleukin-6) after septic shock, hemorrhagic shock, and severe trauma. Crit Care Med 1997;25:1771—1773. 17. Lin KJ, Lin J, Hanasawa K, Tani T, Kodama M. Interleukin-8 as a predictor of the severity of bacteremia and infectious disease. Shock 2000;14:95—100.
www.elsevierhealth.com/journals/jinf
Circulating levels of granulocyte macrophage colony-stimulating factor in patients with the systemic inflammatory response syndrome Donato Torrea,*, Roberto Tambinia, Mariangela Manfredib, Valerio Manganic, Paola Livid, Viviana Maldifassie, Maurizio Chiarandae, Paolo Campib, Filippo Speranzaa a
Section of Pediatric Infectious Diseases, Regional Hospital, Viale Borri 57, 21100 Varese, Italy Department of Clinical and Laboratory Immunology, San Giovanni di Dio Hospital, Florence, Italy c Intensive Care Unit, San Giovanni di Dio Hospital, Florence, Italy d Department of Anaesthesia and Intensive Care, Careggi Hospital, Florence, Italy e Department of Anaesthesia and Intensive Care, Regional Hospital, Varese, Italy b
Accepted 13 April 2003
KEYWORDS Systemic inflammatory response syndrome; Granulocytemacrophage; Colony stimulating factor
Summary Objectives. Granulocyte-macrophage colony stimulating factor (GM-CSF) is a key regulator cytokine that modulates the proliferation and maturation of polymorphonuclear and mononuclear progenitors. This study was designed to investigate and clarify the role of GM-CSF in 52 critically ill patients with systemic inflammatory response syndrome (SIRS). Methods. Serum levels of GM-CSF were detected by an immunoenzyme assay. Results. Our results clearly show that the serum concentrations of GM-CSF were significantly elevated in patients with infectious and noninfectious SIRS (33.2 ^ 45.7 pg/ml, controls: 17.2 ^ 9.8 pg/ml; p ¼ 0:0303). In addition, GM-CSF levels significantly decreased in patients with SIRS, particularly in patients with infectious SIRS, 5 and 7 days later. There was a clear tendency toward higher levels of GM-CSF in patients with poor, as compared with those having a good outcome of the disease. Conclusion. These results show that GM-CSF may play an important role in patients with infectious and noninfectious SIRS, and that GM-CSF levels progressively and significantly decrease in patients with infectious SIRS. Q 2003 The British Infection Society. Published by Elsevier Ltd. All rights reserved.
Introduction The systemic inflammatory response syndrome (SIRS), a massive pro-inflammatory immune state, is associated with a number of clinical infective and noninfective conditions.1 SIRS can develop inde*Corresponding author. Tel.: þ 39-0332-278446; fax: þ 390332-278580. E-mail address: [email protected]
pendently of any infection, including pancreatitis, polytrauma, or immune complex disease. Infectious as well as noninfectious events may activate cytokines and nitric oxide,2,3 and in particular increased levels of pro-inflammatory cytokines, tumor necrosis factor (TNF)-alpha, interleukin (IL)-1 and interleukin (IL)-8, have been reported in septic patients, often correlating with severity of the disease.4 Neutrophils are considered to be important players in the host defense system during
0163-4453/$30.00 Q 2003 The British Infection Society. Published by Elsevier Ltd. All rights reserved. doi:10.1016/S0163-4453(03)00065-3
GM-CSF levels in patients with SIRS
the acute inflammatory response; their motility to infection or inflammatory sites is mediated through systemic release of chemotactic substances, and various cytokines, including granulocyte-colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF).5 The role of GM-CSF in patients with infectious or noninfectious SIRS is unclear and only two studies on this cytokine have been published. Preesneill et al.6 have demonstrated in the sera of 82 critically ill patients with sepsis and septic shock a significant increase of granulocyte-colony stimulating factor, whereas serum levels of GM-CSF were not elevated in these patients. In contrast, Fanning et al.7 showed increased levels of circulating GM-CSF in six patients with SIRS. In the present study, we determined circulating levels of GM-CSF in 52 critically ill patients with infectious or noninfectious SIRS to better clarify its role during the acute inflammatory response.
297
Cytokine assay To detect serum levels of GM-CSF, samples were taken from patients on admission and/or at the time of diagnosis, and after 1, 5 and 7 days. Healthy subjects served as the control group. Serum for cytokine detection was prepared from blood, centrifuged, and stored in 1 ml coded aliquots at 2 20 8C. Serum levels of GM-CSF were determined by commercially available solid-phase sandwich ELISA kit (R & D Systems, Minneapolis, USA), using the protocol recommended by the manufacture. The sensitivity of this ELISA kit was less than 3 pg/ml.
Statistical analysis
Materials and methods
Results are demonstrated as mean and standard deviation (^ 1 SD). Mean values were compared using unpaired Student’s t test with Welch’s correction. Analysis of variance with the Kruskal – Wallis test was used to assess results of cytokine levels at various time. p , 0:05 was considered statistically significant.
Patients
Results
In this prospective study, the patient population consisted of 52 adult critically ill patients, admitted to four intensive care units (Careggi and San Giovanni di Dio Hospital in Florence, Varese and Busto Arsizio) during a 9-month period. Patients were considered to have SIRS as defined in the guidelines of the America College of Chest Physician/Society of Critical Care Medicine.1 These criteria include two or more of the following conditions: temperature . 38 8C or , 36 8C, a heart rate . 90 beats/min, a respiratory rate . 20 breaths/min or PaCO2 , 32 Torr, a withe blood cell count . 12,000 cells ml or , 4000 cells/ml, or . 10% immature band forms. Sepsis was defined as a systemic response to infection including the criteria for SIRS. Patients with severe sepsis had to fulfill the criteria for sepsis including inadequate organ perfusion as indicated by hypoxemia, and/or oliguria, or hypotension, as manifested by a systolic blood pressure of , 90 mm Hg or a reduction of . 40 mm Hg from baseline without other causes for hypotension. SIRS patients had no signs of focal infection, and no microorganisms were isolated from blood cultures. The etiology of these patients includes trauma, cardiogenic shock, surgical interventions, pulmonary embolia, acute pulmonary edema, and intracranial hemorrhage.
The demographic characteristics of our patients with SIRS are illustrated in Table 1. As shown, we evaluated 52 patients, 31 of whom with infectious SIRS, whereas 21 patients suffered from noninfectious SIRS. Fifteen out of 31 patients had sepsis,
Table 1 Demographic characteristics of 52 patients with SIRS. Variables Gender (male/female) Age (years)
36/16 62.7 ^ 18.7
Infectious SIRS ðn ¼ 31Þ Sepsis Severe sepsis Septic shock Multiple organ dysfunction
15 9 7 17
Noninfectious SIRS ðn ¼ 21Þ Surgical interventions Trauma Pulmonary embolism Intracranial hemorrhage Acute pulmonary edema Cardiogenic shock
6 4 3 2 1 5
Mortality SIRS Infectious SIRS Noninfectious SIRS
15/52 (28.8%) 10/31 (32.2%) 5/21 (23.8%)
298
D. Torre et al.
Table 2 Serum levels of GM-CSF in 52 patients with SIRS at various days. Patients
SIRS, overall ðn ¼ 52Þ Infectious SIRS ðn ¼ 31Þ Noninfectious SIRS ðn ¼ 21Þ
GM-CSF (pg/ml) 0
1
5
7 days
33.2 ^ 45.7 30.6 ^ 39.7 37.1 ^ 54.2
38.4 ^ 64.3 40.6 ^ 65.7 34.5 ^ 64.1
22.3 ^ 26.2 21.9 ^ 23.2 22.6 ^ 30.1
18.6 ^ 21.6* 12.7 ^ 7.6** 26.8 ^ 31.0
*p ¼ 0:0148, and **p ¼ 0:0248 (Kruskal–Wallis test).
nine patients had severe sepsis, seven patients had septic shock, and 17 patients had multiple organ dysfunction. SIRS patients had a mortality of 28.8%, whereas mortality of patients with infectious SIRS and noninfectious SIRS was 32.2 and 23.8%, respectively. Results of serum levels of GM-CSF in SIRS patients and controls are shown in Fig. 1. A significant increase of GM-CSF (33.2 ^ 45.7 pg/ml, median: 16.20 pg/ml; controls: 17.2 ^ 9.8 pg/ml, median 10.5 pg/ml; p ¼ 0:0303) levels was observed in patients with SIRS, and in particular in patients with infectious SIRS (30.6 ^ 39.7 pg/ml, median: 18.0 pg/ml; p ¼ 0:0466) at the time of diagnosis, whereas GM-CSF levels of patients with noninfectious SIRS were not significantly increased (37.1 ^ 54.2 pg/ml, median 12.5 pg/ml). Furthermore, serum levels of GM-CSF were determined in SIRS patients at the time of diagnosis, and 1, 5 and 7 days later (Table 2). As shown, a significant decrease of GM-CSF levels was noted in the patients, in particular in patients with infectious SIRS, 5 and 7 days later, whereas patients with noninfectious SIRS had progressively, but nonsignificant, lower GM-CSF levels, especially after 5 and 7 days. Table 3 shows GM-CSF levels in SIRS patients with a good or poor outcome. As can be seen, GM-CSF
Figure 1 Serum levels of GM-CSF in 52 patients with SIRS, of whom 31 patients with infectious SIRS and 21 patients with noninfectious SIRS on admission. Horizontal bar indicates mean ^ SD levels of GM-CSF.
levels nearly reached statistical significance ðp ¼ 0:0537Þ in predicting poor outcome in patients with SIRS, whereas GM-CSF levels of patients with infectious or noninfectious SIRS separately were not predictive of poor outcome.
Discussion The results of this study clearly show increased levels of GM-CSF in patients with infectious and noninfectious SIRS. Among several mediators operating in SIRS, proinflammatory cytokines TNF-alpha, IL-1 and interleukin-6 are considered to play a key role, especially in patients with sepsis or septic shock.8 However, further studies have demonstrated a proinflammatory role of several other cytokines in SIRS, such as IL-8, interferon gamma and G-CSF.5,8,9 Kuhns et al.9 have shown increased levels of GCSF after intravenous administration of endotoxin in healthy volunteers. It is interesting to note that IL-1 is a strong inducer of GM-CSF and macrophagecolony stimulating factor.8 However, the role of GM-CSF in SIRS and in sepsis syndrome has not been fully investigated. Increased, but not significant, levels of GM-CSF were observed in four of nine healthy volunteers, who were treated with endotoxin at various dosages.9 Presneill et al.6 who studied 82 critically ill patients with sepsis, shock without sepsis, and septic shock, found that GM-CSF levels were rarely elevated in the first hours to days in many critically ill patients, particularly those with sepsis and septic shock. It should be noted that detection limit of GM-CSF levels of the immunoassay used by Presneill et al.6 was much higher (100 pg/ml) than that used in our immunoassay (3 pg/ml); furthermore, we cannot rule out that coagulation of blood of SIRS patients may interfere with GM-CSF production, inasmuch as plasma samples could influence GM-CSF levels in SIRS patients than serum samples. Furthermore, circulating GM-CSF was detectable in some patients with meningococcal septic shock.10
GM-CSF levels in patients with SIRS
299
Table 3 Predictive value of GM-CSF levels of outcome in patients with SIRS. Patients
SIRS, overall ðn ¼ 52Þ Infectious SIRS ðn ¼ 31Þ Noninfectious SIRS ðn ¼ 21Þ
GM-CSF (pg/ml) Good outcome
Poor outcome
15.0 ^ 18.0 ðn ¼ 37Þ 14.5 ^ 20.1 ðn ¼ 21Þ 15.6 ^ 15.4 ðn ¼ 16Þ
56.0 ^ 74.5* ðn ¼ 15Þ 44.8 ^ 62.5 ðn ¼ 10Þ 78.5 ^ 98.5 ðn ¼ 5Þ
*p ¼ 0:0537 by unpaired t test with Welch’s correction.
The failure to detect GM-CSF in the sera of patients with sepsis and septic shock may be misleading. In fact, Metcalf et al.11 suggest that in transgenic mice circulating GM-CSF is rapidly consumed by cells bearing GM-CSF receptors, such that free GM-CSF may be undetectable in serum by available assays. However, Fanning et al.7 showed that GM-CSF was consistently and significantly elevated in six patients with SIRS, and that GMCSF appears to be a major factor inhibiting neutrophil apoptosis in the systemic circulation of patients with SIRS. GM-CSF may also play a dominant role in inhibiting neutrophil apoptosis in the lungs of patients with adult respiratory distress syndrome.12 In fact, human neutrophils, in response to stimulation by lipopolysaccharide, in vitro secrete, several cytokines, including IL-1 ra, IL-4, and GM-CSF.13 It should be noted that other cytokines, including TNF alpha, IL-1, IL-6 and IL-8 could predict factors of fatal outcome in the infectious and noninfectious SIRS. However, several studies have shown that plasma levels of IL-1, TNF alpha, IL-6 fluctuate in patients with SIRS, and display a poor correlation with mortality,14,15 whereas in patients with septic shock, TNF alpha, IL-6 and IL-8 significantly correlate with fatal outcome.16,17 In conclusion, these findings of elevated levels of GM-CSF in patients with infectious and noninfectious SIRS suggest that GM-CSF, may stimulate the immune system in response to microbial and inflammatory stimuli may contribute to the pathogenesis in SIRS.
References 1. Bone RC, Balk RA, Cerra FB, et al. ACCP/SCCM consensus conference. Definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis. Chest 1992;101:1644—1655. 2. Casey L, Balk RA, Bone RC. Plasma cytokine and endotoxin levels correlate with survival in patients with sepsis syndrome. Ann Intern Med 1993;119:771—778. 3. Kilbourn R, Griffith O. Overproduction of nitric oxide in cytokine-mediated and septic shock. J Natl Cancer Inst 1992; 84:827—831.
4. Biliau A, Vandekerckhove F. Cytokines and their interactions with other inflammatory mediators in the pathogenesis of sepsis and septic shock. Eur J Clin Invest 1991;21:559—573. 5. Colotta F, Re F, Polentarutti N, Sozzani S, Mantovani A. Modulation of granulocyte survival and programmed cell death by cytokines and bacterial products. Blood 1997;89: 2012—2020. 6. Presneill JJ, Waring PM, Layton JE, et al. Plasma granulocyte colony-stimulating factor and granulocyte-macrophage colony-stimulating factor levels in critical illness including sepsis and septic shock: relation to disease severity, multiple organ dysfunction, and mortality. Crit Care Med 2000;28: 2344—2354. 7. Fanning NF, Kell MR, Shorten GD, Kirwan WO, Bouchier-Hayes D, Cotter T, Redmond HP. Circulating granulocyte macrophage colony-stimulating factor in plasma of patients with the systemic inflammatory response syndrome delays neutrophil apoptosis through inhibition of spontaneous reactive oxygen species generation. Shock 1999;11:167—174. 8. Davies MG, Hagen PO. Systemic inflammatory response syndrome. Br J Surg 1997;84:920—935. 9. Kuhns DB, Alvord WG, Gallin JI. Increased circulating cytokine antagonists, and E-selectin after intravenous administration of endotoxin in humans. J Infect Dis 1995; 171:145—152. 10. Waring PM, Presneill JJ, Maher DW, et al. Differential alterations in plasma colony-stimulating factor concentrations in meningococcaemia. Clin Exp Immunol 1995; 102:501—506. 11. Metcalf D, Rasko JEJ. Leukemic transformation of immortalized FDC-P1 cells engrafted in GM-CSF transgenic mice. Leukemia 1993;7:878—886. 12. Matute-Bello G, Liles WC, Radella F, et al. Neutrophil apoptosis in the acute respiratory distress syndrome. Am J Respir Crit Care Med 1997;156:1969—1977. 13. Re F, Mengozzi M, Muzio M, Dinarello CA, Mantovani A, Colotta F. Expression of interleukin-1 receptor antagonist (IL-1 ra) by human circulating polymorphonuclear cells. Eur J Immunol 1993;23:570—573. 14. Friedland JS, Porter JC, Daranani S, et al. Plasma proinflammatory cytokine concentrations, acute physiology and chronic health evaluation (APACHE) III scores and survival in patients in an intensive care unit. Crit Care Med 1996;24: 1775—1781. 15. Presterl E, Staudinger T, Pettermann M, et al. Cytokine profile and correlation to the APACHE III and MPM II scores in patients with sepsis. Am J Respir Crit Care Med 1997;156:825—832. 16. Martin C, Boisson C, Haccoun M, Thomachot L, Mege JL. Patterns of cytokine evolution (tumor necrosis factor-alpha and interleukin-6) after septic shock, hemorrhagic shock, and severe trauma. Crit Care Med 1997;25:1771—1773. 17. Lin KJ, Lin J, Hanasawa K, Tani T, Kodama M. Interleukin-8 as a predictor of the severity of bacteremia and infectious disease. Shock 2000;14:95—100.