Newly Developed Endotoxin Measurement Method (the Endotoxin Activity Assay) May Reflect the Severity of Sepsis

Kiyohide Ishihata, Yasuyuki Kakihana, Tomotsugu Yasuda, Tohru Imabayashi, Norifumi Nakamura

Department of Anesthesiology, Faculty of Medicine, Kagoshima University, Kagoshima, Japan.
Department of Emergency and Intensive Care Medicine, Faculty of Medicine, Kagoshima University, Kagoshima, Japan.
Department of Oral and Maxillofacial Surgery, Field of Oral and Maxillofacial Rehabilitation, Advanced Therapeutic Course, Ka-goshima University Graduate School of Medical and Dental Sciences, Kagoshima, Japan.

DOI: 10.4236/ojpathology.2013.31001                

Abstract

Objectives: Endotoxin (ET) is a structural molecule of the Gram-negative bacilli extracellular membrance, which activates targeted cells including macrophages and neutrophils, and causes septic shock. But it is known that the conventional ET measurement method has many problems; for example, a discrepancy between plasma ET concentration and clinical manifestation in the septic patients has been reported. The purpose of this study was to evaluate the usefulness of a newly developed method (Endotoxin Activity Assay (EAA)) to measure the ET activity (EA) in patients under sepsis compared with the prior method of the limulus amebocyte lysate (LAL) assay and explore the association between EA levels and patients’ severity. Method: We measured the EA levels in 40 patients (aged 63.5 +/－ 17.7 years) admitted to the ICU. EA level was measured using a chemiluminometer (Autolumat LB953; EG & Berthold). Patients were divided into 5 groups: 1) control group; 2) systemic inflammatory response syndrome (SIRS) group; 3) sepsis (SIRS and infection) group; 4) severe sepsis group and 5) septic shock group. We then compared the EA level between each group and control group. We made the statistical evaluation by unpaired t test and significant difference was p < 0.05. Results: The EA levels were significantly increased as sepsis severity rises. The measured EA levels were (0.18 +/－ 0.09), (0.33 +/－ 0.19), (0.39 +/－ 0.16), (0.65 +/－ 0.25) and (0.78 +/－ 0.34) in control, SIRS, sepsis, severe sepsis and septic shock groups, respectively. In the EA level measured by EAA, severe patients had a tendency to exceed the cutoff value. Conclusion: The EA levels were significantly correlated as patients’ severity rise. Measuring EA levels on admission to ICU may provide a mechanism to identify and target severe septic patients.

Keywords

Endotoxin Activity Assay; Endotoxin; Sepsis

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	J. C. Marshall, D. Foster, J.-L. Vincent, D. J. Cook, J. Cohen, R. P. Dellinger, S. Opal, E. Abraham, S. J. Brett, T. Smith, S. Mehta, A. Derzko, A. Romaschin, MEDIC Study, “Diagnostic and Prognostic Implications of Endotoxemia in Critical Illness: Results of the MEDIC Study,” Journal of Infectious Diseases, Vol. 190, No. 3, 2004, pp. 527-534. doi:10.1086/422254
[2]	F. Valenza, L. Fagnani, S. Coppola, S. Froio, F. Sacconi, C. Tedesco, M. Maffioletti, M. Pizzocri, V. Salice, M. L. Ranzi, C. Marenghi and L. Gattinoni, “Prevalence of Endotoxemia after Surgery and Its Association with ICU Length of Stay,” Critical Care, Vol. 13, No. 3, 2009, p. R102. doi:10.1186/cc7934
[3]	D. A. Rocke, S. L. Gaffin, M. T. Wells, Y. Koen and J. G. Brock-Utine, “Endotoxemia Associated with Cardiopulmonary Bypass,” Journal of Thoracic Cardiovascular Surgery, Vol. 93, No. 6, 1987, pp. 832-837.
[4]	D. J. Klein, F. Briet, R. Nisenbaum, A. D. Romaschin and C. D. Mazer, “Endotoxemia Related to Cardiopulmonary Bypass Is Associated with Increased Risk of Infection after Cardiac Surgery: A Prospective Observational Study,” Critical Care, Vol. 15, No. 1, 2011, p. R69. doi:10.1186/cc10051
[5]	J. C. Marshall, P. M. Walker, D. M. Foster, D. Harris, M. Ribeiro, J. Paice, A. D. Romaschin and A. N. Derzko, “Measurement of Endotoxin Activity in Critically Ill Patients Using Whole Blood Neutrophil Dependent Chemiluminescence,” Critical Care, Vol. 6, No. 4, 2002, pp. 342-348. doi:10.1186/cc1522
[6]	R. Danner, R. Elin, J. Hosseini, R. Wesley, J. Reilly and J. Parillo, “Endotoxemia in Human Septic Shock,” Chest, Vol. 99, No. 1, 1991, pp. 169-175. doi:10.1378/chest.99.1.169
[7]	L. C. Casey, R. A. Balk and R. C. Bone, “Plasma Cytokine and Endotoxin Levels Correlate with Survival in Patients with the Sepsis Syndrome,” Annals Internal Medicine, Vol. 119, No. 8, 1993, pp. 771-778.
[8]	C. H. Wortel, M. A. von der Mohlen, S. J. van Deventer, C. L. Sprung, M. Jastremski, M. J. Lubbers, C. R. Smith, I. E. Allen and J. W. ten Cate, “Effectiveness of a Human Monoclonal Anti-Endotoxin Antibody (HA-1A) in Gram-Negative Sepsis: Relationship to Endotoxin and Cytokine Levels,” Journal of Infectious Diseases, Vol. 166, No. 6, 1992, pp. 1367-1374. doi:10.1093/infdis/166.6.1367
[9]	J. Bion, I. Badger and H. Crosby, “Selective Decontamination of the Digestive Tract Reduces Gram-Negative Pulmonary Colonization but Not Systemic Endotoxemia in Patients Undergoing Elective Liver Transplantation,” Critical Care Medicine, Vol. 22, No. 1, 1994, pp. 40-49.
[10]	G. B. Gaeta, P. Perna, L. E. Adinolfi, R. Utili and G. Ruggiero, “Endotoxemia in a Series of 104 Patients with Chronic Liver Diseases: Prevalence and Significance,” Digestion, Vol. 23, No. 4, 1982, pp. 239-244.
[11]	R. Roumen, J. Frieling and H. Van Tits, “Endotoxemia after Major Vascular Operations,” Journal of Vascular Surgery, Vol. 18, No. 5, 1993, pp. 853-857. doi:10.1016/0741-5214(93)90341-I
[12]	S. Dholakia, D. Inwald, H. Betts and S. Nadel, “Endotoxemia in Pediatric Critical Illness—A Pilot Study,” Critical Care, Vol. 15, No. 3, 2011, p. R141. doi:10.1186/cc10264
[13]	P. C. Fink, L. Lehr, R. M. Urbaschek and J. Kozak, “Limulus Amebocyte Lysate Test for Endotoxemia: Investigations with a Femtogram Sensitive Spectrophotometric Assay,” Klinische Wochenschrift, Vol. 59, No. 5, 1981, pp. 213-218. doi:10.1007/BF01476578
[14]	M. Sakata, T. Inoue, M. Todokoro and M. Kunitake, “Limulus Amebocyte Lysate Assay for Endotoxins by an Adsorption Method with Polycation-Immobilized Cellulose Beads,” Analytical Sciences, Vol. 26, No. 3, 2010, pp. 291-296. doi:10.2116/analsci.26.291
[15]	Y. Sanada, K. Mizuta, T. Urahashi, Y. Ihara, T. Wakiya, N. Okada, N. Yamada, K. Ushijima, S. Otomo, K. Sakamoto and Y. Yasuda, “Impact of Hepatic Clearance of Endotoxin Using Endotoxin Activity Assay,” Hepatology International, Vol. 6, 2011, pp. 489-493. doi:10.1007/s12072-011-9289-4
[16]	R. C. Bone, R. A. Balk, F. B. Cerra, R. P. Dellinger, A. M. Fein, W. A. Knaus, R. M. Schein and W. J. Sibbald, “Definitions for Sepsis and Organ Failure and Guidelines for the Use of Innovative Therapies in Sepsis. The ACCP/ SCCM Consensus Conference Committee. American College of Chest Physicians/Society of Critical Care Medicine,” Chest, Vol. 101, No. 6, 1992, pp. 1644-1655. doi:10.1378/chest.101.6.1644
[17]	A. D. Romaschin, D. M. Harris, M. B. Ribeiro, J. Paice, D. M. Foster, P. M. Walker and J. C. Marshall, “A rapid Assay of Endotoxin in Whole Blood Using Autologous Neutrophil Dependent Chemiluminescence,” Journal of Immunological Methods, Vol. 212, No. 2, 1998, pp. 169-185. doi:10.1016/S0022-1759(98)00003-9
[18]	I. Hilmi, J. A. Kellum, R. Planinsic, D. Foster, A. Abdullah, D. Damian, S. Gligor, D. Klein and K. Abu-Elmagd, “Endotoxemia is Common Following Abdominal Organ Transplantation and Is Associated with Reperfusion and Rejection,” Journal of Organ Dysfunction, Vol. 5, No. 4, 2009, pp. 254-260. doi:10.3109/17471060902859113
[19]	P. G. Jansen, H. Te Velthuis, H. M. Oudemans-Van Straaten, E. R. Bulder, S. J. van Deventer, A. Sturk, L. Eijsman and C. R. Wildevuur, “Perfusion-Related Factors of Endotoxin Release during Cardiopulmonary Bypass,” European Journal Cardiothoracic Surgery, Vol. 8, No. 3, 1994, pp. 125-129. doi:10.1016/1010-7940(94)90167-8
[20]	R. J. Elin, R. A. Robinson, A. S. Levine and S. M. Wolff, “Lack of Clinical Usefulness of the Limulus Test in the Diagnosis of Endotoxemia,” New England Journal of Medicine, Vol. 293, No. 11, 1975, pp. 521-524. doi:10.1056/NEJM197509112931102
[21]	D. W. Bates, J. Parsonnet, P. A. Ketchum, E. B. Miller, T. J. Novitsky, K. Sands, P. L. Hibberd, P. S. Graman, P. N. Lanken, J. S. Schwartz, K. Kahn, D. R. Snydman, R. Moore, E. Black and R. Platt, “Limulus Amebocyte Lysate Assay for Detection of Endotoxin in Patients with Sepsis Syndrome,” Clinical Infectious Diseases, Vol. 27, No. 3, 1998, pp. 582-591. doi:10.1086/514713
[22]	M. Kjelgaard-Hansen, B. Wiinberg, B. Aalbaek, L. Olsen, D. Harris, A. Romaschin, A. T. Kristensen and A. L. Jensen, “Endotoxin Activity in Whole Blood Measured by Neutrophil Chemiluminescence Is Applicable to Canine Whole Blood,” Comparative Immunology, Microbiology and Infectious Diseases, Vol. 31, No. 6, 2008, pp. 477-485. doi:10.1016/j.cimid.2007.08.004
[23]	E. Abdala, C. E. S. Baía, S. Mies, P. C. B. Massarollo, N. de Paula Cavalheiro, V. R. M. Baía, C. A. F. Inácio, H. C. Sef and A. A. Barone, “Bacterial Translocation during Liver Transplantation: A Randomized Trial Comparing Conventional with Venovenous Bypass vs. Piggyback Methods,” Liver Transplantation, Vol. 13, No. 4, 2007, pp. 488-496. doi:10.1002/lt.21085
[24]	G. Behre, I. Schedel, B. Nentwig, B. W?rmann, M. Essink and W. Hiddemann, “Endotoxin Concentration in Neutropenic Patients with Suspected Gram-Negative Sepsis: Correlation with Clinical Outcome and Determination of Anti-Endotoxin Core Antibodies during Therapy with Polyclonal Immunoglobulin M-Enriched Immunoglobulins,” Antimicrobial Agents and Chemotherapy, Vol. 36, No. 10, 1992, pp. 2139-2146. doi:10.1128/AAC.36.10.2139
[25]	P. Brandtzaeg, P. Kierulf, P. Gaustad, A. Skulberg, J. N. Bruun, S. Halvorsen and E. Sorensen, “Plasma Endotoxin as a Predictor of Multiple Organ Failure and Death in Systemic Meningococcal Disease,” Journal of Infectious Diseases, Vol. 159, No. 2, 1989, pp. 195-204. doi:10.1093/infdis/159.2.195
[26]	N. B. Aydin, H. Gercekoglu, B. Aksu, V. Ozkul, T. Sener, I. Kiygil, T. Turkoglu, S. Cimen, F. Babacan and M. Demirtas, “Endotoxemia in Coronary Artery Bypass Surgery: A Comparison of the Off-Pump Technique and Conventional Cardiopulmonary Bypass,” Journal of Thoracic and Cardiovascular Surgery, Vol. 125, No. 4, 2003, pp. 843-848. doi:10.1067/mtc.2003.323