Nucleated Red Blood Cells Are Associated with a Higher Mortality Rate in Patients with Surgical Sepsis

Abstract

Background:

Nucleated red blood cells (NRBCs) are present in certain non-oncologic disease states and are associated with a poor prognosis. The purpose of this study was to evaluate NRBCs as an early prognostic marker for death in patients with surgical sepsis.

Methods:

Retrospective evaluation of data collected prospectively from 275 patients from our Investigational Review Board-approved surgical sepsis database over a 27-mo period. The NRBC values were correlated with patient outcomes. The χ² test was used for testing of categorical variables and the Mann-Whitney U was used for testing of continuous variables. The level of significance was set at 0.05.

Results:

At sepsis recognition, 48 patients (17.5%) were NRBC-positive. The mortality rate was greater in patients who were NRBC positive while in the intensive care unit (ICU); (27% vs. 12%; p=0.007) and during the hospital stay (35.4% vs. 15%; p=0.001). When NRBC-values at all time points are considered, 116 patients (42.2%) were NRBC-positive. The mortality rate was greater in patients who were NRBC-positive in both the ICU (23.3% vs. 8.2%; p<0.001) and during the hospital stay (31% vs. 9.4%; p<0.001). In-hospital and ICU mortality rates increased with increasing NRBC-concentration. For the 153 patients with severe sepsis, NRBC positivity at any time was associated with a higher ICU mortality rate (20% vs. 3.2%; p=0.001). Significant mortality differences did not occur between NRBC-positive and NRBC-negative patients with sepsis (n=48) or septic shock (n=74).

Conclusions:

Surgical sepsis patients with detectable NRBCs are at higher risk of ICU and in-hospital death than those with non-detectable NRBCs. The mortality difference is underscored in surgical patients with severe sepsis. This study suggests NRBCs may be a biomarker of outcomes in patients with surgical sepsis.

Sepsis among surgical patients continues to be a common and deadly problem. Identifying biomarkers of sepsis severity can aid in early recognition and help risk-stratify patients. Ideally, these biomarkers would be available readily through routine laboratory testing and could be measured repeatedly throughout the treatment course. Previously, baseline elevation of serum lactate, β-type natriuretic peptide (BNP), international normalized ratios (INR), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and total bilirubin concentration were identified as discriminators of non-survivors in patients with surgical sepsis [1]. However, these markers can be elevated by various other disease states also, so the identification of additional, more specific, biomarkers for sepsis prognostication would be advantageous.

Nucleated red blood cells (NRBCs) are immature erythrocytes, also known as erythroblasts. Under normal physiologic circumstances, the peripheral blood of adults is free of NRBCs. However, certain disease states and critical illness may lead to the appearance of these cells in the blood stream. The mechanism by which these cells are generated remains unclear. It may be a result of increased red blood cell production and differentiation via the upregulation of erythropoietin, interleukin (IL)-3, and IL-6 [2]. Another proposed mechanism is via dysregulation of the bone marrow and its normal screening process to preclude NRBCs from entering the circulation [3]. Other potential causes are extramedullary hematopoiesis and splenectomy [4].

Regardless of the mechanism, the presence of NRBCs in the blood is a marker of both disease severity and increased risk of death in certain patient populations [2 –11]. Such cells have been associated with a variety of disease states, including cancer [5,6], congestive heart failure [7,8], acute and chronic anemia [9], hypoxia [9], and hematologic diseases [4,10]. Recent work by Stachon et al. showed that the appearance of NRBCs in the circulating blood is associated with a higher risk of death in intensive care unit (ICU) patients [11 –13].

To date, the importance of NRBCs in patients with surgical sepsis has not been evaluated. The purpose of this study was to determine the early prognostic significance of NRBCs in ICU patients with surgical sepsis. We hypothesized that the presence of NRBCs under these conditions is a biomarker of disease severity and in-hospital death.

Patients and Methods

Study site and patients

This study was conducted in the surgical intensive care unit (SICU) at The Methodist Hospital, a 948-bed, academic tertiary referral hospital located in The Texas Medical Center in Houston, Texas. The SICU is a 27-bed non-cardiac unit that serves a diverse group of surgical patients, including critically ill general, vascular, oncologic, transplant, thoracic, orthopedic, plastic, urologic, and head and neck surgical patients. All patients are screened for sepsis using our validated physiologic screening tool [14]. This tool has a sensitivity of 96.5%, a specificity of 96.7%, a positive predictive value (PPV) of 80.2%, and a negative predictive value (NPV) of 99.5% [14]. Once identified, these patients are managed by an evidence-based protocol that includes goal-directed fluid resuscitation with time-appropriate interventions based on clinical consensus and evidence-based guidelines for the management of sepsis [15]. With informed consent, all SICU patients managed with this protocol have data acquired for our sepsis research database. Data collection and entry are performed prospectively by a research nurse and informaticist, and all cases are reviewed and maintained by the surgical sepsis research team. Data reports are coded and password protected to ensure patient confidentiality. The sepsis research database is maintained with approval of The Methodist Hospital Research Institute Institutional Review Board.

Surgical sepsis definitions

All study patients were categorized as having sepsis, severe sepsis, or septic shock according to our modification of the American College of Chest Physician/Society of Critical Care Medicine Consensus Conference definitions [16]. We define surgical sepsis as systemic inflammatory response syndrome (SIRS) plus an infection requiring surgical intervention for source control or SIRS plus an infection within 14 d of a major surgical procedure. Such a procedure is defined as any requiring general anesthesia for >1 h. Severe sepsis is defined as SIRS plus infection plus acute organ dysfunction. The types of acute organ dysfunction are defined as follows:

1. Neurologic: Glasgow Coma Scale (GCS) score <13 at recognition of sepsis or deterioration of the GCS to <13 during the first 24 h;

2. Pulmonary: PaO₂:F_iO₂ (partial pressure of oxygen/inspired fraction of oxygen) ratio<250 (<200 if lung is primary site of infection) and pulmonary capillary wedge pressure (PCWP, if available) not suggestive of fluid overload;

3. Renal (one of the following): Urine output <0.5 mL/kg for ≥1 h despite adequate volume resuscitation, increase in serum creatinine concentration ≥0.5 mg/dL from baseline (measured within 24 h of start of sepsis resuscitation) despite adequate volume resuscitation or increase in serum creatinine ≥0.5 mg/dL during the first 24 h of sepsis management despite adequate volume resuscitation. Adequate volume resuscitation is defined as a minimum intravenous fluid infusion of 20 mL/kg/ideal body weight (IBW), or central venous pressure (CVP)≥8 mm Hg or PCWP≥12 mm Hg;

4. Coagulation (one of the following): International Normalized Ratio (INR)>1.5, platelet count<80,000 or ≥50% decrease compared with 24 h before instituting sepsis resuscitation or in the 24 h after starting sepsis resuscitation in the absence of chronic liver disease;

Hypoperfusion: Serum lactate concentration >4 mmol/L.

Septic shock was defined as SIRS plus infection plus acute cardiac dysfunction or hypotension, which is defined by both persistent hypotension despite intravenous fluid challenge ≥20 mL/kg/IBW of isotonic crystalloid infusion or CVP≥8 mm Hg or PCWP≥12 mm Hg and requirement for vasopressors to increase mean arterial blood pressure (MAP)≥65 mm Hg.

Study design and data collection

We evaluated our SICU sepsis database from September 1, 2007 through December 31, 2009 for all patients with a diagnosis of surgical sepsis. The database was queried for patient demographics, baseline biomarkers of organ dysfunction (including serum lactate concentration, white blood cell count, platelet count, BNP concentration, AST, ALT, alkaline phosphatase, total bilirubin concentration, INR, NRBC count), source of infection, Acute Physiology and Chronic Health Evaluation (APACHE) II score [17], Sequential Organ Failure Assessment (SOFA) score [18], and in-hospital death. Patients were grouped into the categories of sepsis, severe sepsis, and septic shock according to the definitions given above.

Sources of the infection were grouped into the following categories: Abdomen, pulmonary/thoracic, wound/soft tissue, vascular access/blood stream, urinary tract, or other. Abdominal sources included any intra-abdominal organ except the bladder, which, as noted, was classified under urinary tract. Thoracic/pulmonary sources of infection included the lung parenchyma and the thoracic esophagus. Wound/soft tissue sources included superficial surgical site infections, cellulitis, necrotizing soft tissue infections, and other abscess that did not involve the abdominal or thoracic cavities. Vascular access/blood stream sources included infected temporary and permanent vascular access devices and bacteremia. Urinary tract sources included pyelonephritis and urinary tract infections. The other category included infections secondary to endocarditis, infection of the central nervous system, and sinus infections. Intensive care unit-free days were calculated by subtracting the number of days a patient was in the ICU from 28. Patients who died in the ICU were assigned an ICU-free day value of zero.

Nucleated red blood cell values were collected at any time after a diagnosis of sepsis was made, with a subset of values (initial) gathered within 24 h of sepsis protocol initiation. Nucleated red blood cell values above zero were defined as positive, whereas undetectable counts were classified as negative. The peak NRBC value was used for all other analyses unless otherwise specified. The NRBC value was reported on all complete blood counts (CBCs) performed. Thus, each NRBC value corresponded to a white blood cell count (WBC), hemoglobin concentration (Hgb), hematocrit, and platelet count. Nucleated red blood cell values were reported by the laboratory Sysmex XE-2100 Automated Hematology System® Sysmex America, Inc., Mundelin, IL) to the electronic medical record. The Sysmex XE-2100 provides an accurate estimate of NRBCs, as judged by results garnered using a microscopic determination according to the National Committee for Clinical Laboratory Standards protocol. The correlation between the Sysmex XE-2100 and the manual microscopic NRBC count was 0.97 to 0.99 in these studies [19].

Statistical analysis

Data are presented as the median and interquartile range. Data were analyzed using MINITAB software version 14 for Windows (MINITAB Inc., State College, PA). The χ² test was used for categorical variables and the Mann-Whitney U test for continuous variables. Significance was set at 0.05.

Results

Two hundred seventy-five patients were included in the study. At sepsis protocol initiation, 48 patients (17.5%) were NRBC-positive. Baseline demographics for the groups are displayed in Table 1. When all NRBC values are considered, 116 patients (42.2%) were positive at some point. There was no difference in sex or age between the positive and negative groups. There was a significant difference in the APACHE II score (p<0.001), SOFA score (p=0.008), and severity of sepsis (p<0.001) between the NRBC-positive and -negative groups. Although the most common source of infection in both groups was abdominal, the NRBC-positive group had a higher percentage of patients with a pulmonary/thoracic source of infection. There also were some notable differences in peak laboratory values between NRBC-positive and NRBC-negative patients (Table 2).

Table 1.

Demographics

	Nucleated red blood cells
	Positive	Negative	p value
Male (%)^a	53 (46)	70 (44)	0.80
Age (years)^a	59 (48–71)	60 (49–72)	0.58
APACHE II score^a	30 (24–35)	22 (16–28)	<0.001
SOFA score^a	10 (7–13)	8 (6–11)	0.008
No. (%) having emergency surgery	85 (73)	109 (69)	0.42
Classification (%)
Septic shock	44 (38)	30 (19)	<0.001
Severe sepsis	60 (52)	93 (58)	<0.001
Sepsis	12 (10)	36 (23)	<0.001
Source of infection (%)
Abdomen	68 (59)	118 (74)	0.23
Vascular/blood stream	4 (3)	11 (7)	0.24
Pulmonary/thoracic	20 (17)	8 (5)	0.003
Wound/soft tissue	17 (15)	12 (8)	0.09
Urinary tract	5 (4)	6 (4)	0.83
Other	2 (2)	4 (3)	0.66

Median (interquartile range).

APACHE=Acute Physiology and Chronic Health Evaluation; SOFA=Sepsis-related Organ Failure Assessment.

Table 2.

Peak Laboratory Values for Patients with and without Nucleated Red Blood Cells after Initiation of Sepsis Protocol ^a

	Nucleated red blood cells
	Positive (n=116)	Negative (n=159)	P value
β-type natriurietic peptide (pg/mL)	99 (30–361)	76 (26–221)	0.04
Partial pressure of oxygen: inspired fraction of oxygen (mm Hg)	203 (134–288)	244 (168–362)	0.012
Serum creatinine concentration (mg/dL)	1.5 (0.8–2.2)	1.0 (0.7–1.5)	0.0014
International Normalized Ratio	1.5 (1.3–1.8)	1.4 (1.3–1.6)	0.016
White blood cells (1,000/mcL)	15 (10–22)	13 (8–19)	0.02
Hemoglobin concentration (g/dL)	11 (10–12)	11 (10–12)	0.2
Platelet count (1,000/mcL)	285 (126–424)	276 (167–422)	0.64
Serum lactate concentration (mmol/L)	3.6 (1.9–6.5)	1.7 (1.2–2.7)	<0.001
Serum alkaline phosphatase (U/L)	136 (89–236)	110 (74–175)	0.01
Aspartate aminotransferase (U/L)	71 (39–132)	38 (26–72)	<0.001
Alanine aminotransferase (U/L)	39 (20–80)	24 (15–44)	<0.001

Values are median (interquartile range).

The overall hospital mortality rate for all study patients was 19% (51/275). Death was more likely among patients who were initially NRBC-positive than in those who were initially -negative for both ICU (27% vs. 12%; p=0.007) and hospital (35.4% vs. 15%; p=0.001) stay. Likewise, the mortality rate was greater in patients who were NRBC-positive than in those who were NRBC-negative for both ICU (23.3% vs. 8.2%; p<0.001) and hospital stay (31% vs. 9.4%; p<0.001). For patients who were NRBC-positive at any point in the ICU stay, the number of ICU-free days at day 28 was 11 less than in patients who were NRBC-negative (p<0.001). These results are summarized in Table 3. Sixty-six of the 116 patients (57%) who were NRBC-positive transitioned to NRBC-negative. For those patients who remained positive, the mortality rate was higher for both ICU (38% vs. 12%; p=0.001) and hospital (52% vs. 15%; p<0.001) stay.

Table 3.

Patient Outcomes ^a,b

	Nucleated red blood cells
	Positive (n=116)	Negative (n=159)
Intensive care unit death (%)	27 (23)	13 (8)
In-hospital death (%)	36 (31)	15 (9)
Intensive care unit-free days at day 28 (interquartile range)	13 (0–20)	24 (18–25)
Hospital stay (days)(interquartile range)	23.7 (12–42)	16 (10–25)

All values are expressed as medians.

The p value of all differences is <0.001.

Intensive care unit and hospital mortality rates increased with increasing NRBC concentration. As shown in Figure 1, the ICU mortality rate among patients who were NRBC-positive was higher than in those patients who had undetectable NRBCs either at sepsis protocol initiation or at any time after the start of the protocol. A peak NRBC count >500/mcL either at sepsis protocol initiation or when all NRBC values are considered was associated with a 50% or greater ICU mortality rate. Of note, only 1/9 patients (11%) survived the hospital stay when the peak NRBC count was >2,000/mcL.

FIG. 1.

Mortality rate in the intensive care unit according to nucleated red blood cell (NRBC) count. iNRBC=initial NRBC count.

The peak NRBC also was analyzed according to sepsis severity. The median peak count for patients with sepsis was 69/mcL, for those with severe sepsis 74/mcL, and for those with septic shock 162/mcL. In addition, NRBC exposure was calculated for each patient by adding the NRBC values from each day a patient had a detectable value. As evidenced in Figure 2, the ICU mortality rate generally increased with higher counts.

FIG. 2.

Deaths in intensive care unit based on exposure to nucleated red blood cells (NRBC).

For ICU death, detectable NRBCs at sepsis protocol initiation had a sensitivity of 33%, a specificity of 85%, a PPV of 27%, and an NPV of 88%. When all NRBC values are considered, the sensitivity increased to 68% and the specificity decreased to 62% for ICU death; the PPV decreased to 23%, and the NPV increased to 92%.

Discussion

Sepsis continues to present a substantial burden in hospitalized patients. A recent report from the U.S. Centers for Disease Control and Prevention stated that sepsis rates more than doubled in the United States between 2000 and 2008 [20]. It is estimated that there were 1.4 million cases of sepsis in the United States in 2008 at an estimated cost of $14.6 billion [20]. Early aggressive treatment remains the mainstay of care; however, this action is contingent on early recognition. The use of a sepsis screening program in conjunction with early evidence-based therapy improves dramatically the sepsis-related mortality rate [14,15,21]. Identifying biomarkers available routinely could aid clinicians in the early recognition of sepsis. Biomarkers that provide prognostic data could help delineate care further.

As the use of mechanical blood analyzers has become more routine, the incidence and importance of NRBCs in hospitalized patients has received increased attention. Multiple prior studies have shown NRBCs to be associated with a poor prognosis in a general population of ICU patients [2,11,12,22,23]. Further study within an SICU population revealed a larger proportion of NRBC-positive than NRBC-negative patients were afflicted with sepsis or infection (18.4% vs. 9.2%) [11]. On the basis of these findings, we considered the use of NRBC positivity as a biomarker of outcomes for patients with sepsis. To our knowledge, the current study is the first to evaluate the prognostic significance of erythroblastemia specifically in ICU patients with surgical sepsis.

The current study revealed an NRBC-positive rate of 42.2% among SICU patients with surgical sepsis. This incidence is slightly higher than the overall NRBC rate of 32% reported in two other cohorts of SICU patients by Stachon et al. [11,24]. The higher incidence of detectable NRBCs in our study could be attributable to more frequent monitoring, which enables a higher rate to be detected if NRBCs appear transiently and subsequently dissipate. Our focus on SICU patients with surgical sepsis also could account for the differences.

Although the proportion of NRBC-positive patients in our study was greater than in previous studies conducted in similar patient populations, the hospital mortality rate among the NRBC-positive patients in our SICU was significantly lower. Thirty-one percent of NRBC-positive patients did not survive to hospital discharge in our study population versus 44%–52% in previous studies conducted in the SICU [11,13]. A recent analysis of the National Surgical Quality Improvement Program data set reported a 34% mortality rate for surgical patients with severe sepsis/septic shock [25]. The overall mortality rate for this cohort of patients with surgical sepsis, regardless of NRBC status, was only 19%. This discrepancy likely is secondary to our use of an aggressive sepsis screening program and a computerized clinical decision support (CCDS) protocol to implement early evidence-based care [15]. Within our SICU, our severe sepsis/septic shock-related mortality rate decreased from a baseline of 34% to 14% after implementation of sepsis screening and a CCDS sepsis management protocol [15]. Our overall lower sepsis-related mortality rate may explain the lower mortality rate of NRBC-positive patients in this cohort.

Intensive care unit and hospital mortality rates were higher in patients with detectable NRBCs. Furthermore, a higher peak NRBC count was associated with a higher ICU mortality rate. Patients in the NRBC-positive cohort also demonstrated statistically higher baseline values for other biomarkers, including BNP, creatinine concentration, INR, WBC counts, and serum lactic acid, AST, ALT, and alkaline phosphate concentrations. Baseline elevation of BNP and lactic acid correlate with both sepsis severity and death [1,26 –28]. Prior work by Stachon et al. documented differences in admission creatinine and ALT concentrations but no difference in admission WBC counts in NRBC-positive versus -negative patients [24]. In this study, we found that NRBC-positive patients had higher baseline WBC counts than NRBC-negative patients. Patients in the NRBC-positive group also had significantly higher APACHE II and SOFA scores. The fact that NRBCs are present in the same patients who have higher APACHE II and SOFA scores, as well as baseline elevation of known biomarkers of sepsis severity and death, supports the view that the presence of NRBCs is an accurate marker of sepsis severity and higher risk of death.

There are several limitations to our study, including its retrospective nature. The number of patients having a splenectomy was not recorded, nor was the number of patients receiving granulocyte and macrophage colony-stimulating factors, which may induce release of NRBCs into the peripheral blood. Therefore, the NRBC value in these patients may have been elevated falsely. Also, because NRBC counts were recorded from sepsis protocol initiation to SICU discharge, some abnormal values may have appeared when the patients were still in the SICU but no longer experiencing sepsis.

This study shows patients with surgical sepsis and erythroblastemia are at greater risk of death than patients with undetectable NRBCs. Because the presence of NRBCs may be an early biomarker of inflammation and hypoxia, we expected patients with surgical sepsis and detectable NRBCs to have worse outcomes, an idea that was substantiated by our study. An NRBC count exceeding 500/mcL at sepsis protocol initiation or a peak count >500/mcL after protocol initiation was associated with a 50% or greater ICU mortality rate. Because the ICU mortality rate is greater in NRBC-positive patients, the next logical step is to determine whether early detection and aggressive management (i.e., antibiotics) of NRBC-positive patients changes outcomes.

Author Disclosure Statement

No competing financial interests exist.

References

Moore

, McKinley

, Turner

et al. The epidemiology of sepsis in general surgery patients. J Trauma, 2011; 70:672–680.

Stachon

, Bolulu

, Holland-Letz

, Krieg

. Association between nucleated red blood cells in blood and the levels of erythropoietin, interleukin 3, interleukin 6, and interleukin 12p70. Shock, 2005; 24:34–39.

Lehnhardt

, Katzy

, Langer

et al. Prognostic significance of erythroblasts in burns. Plast Reconstr Surg, 2005; 115:120–127.

Marmont

. Nuclear extrusion by erythroblasts in the peripheral blood of a splenectomized patient. Haematologica, 1998; 83:943.

Delsol

, Guiu-Godfrin

, Guiu

et al. Leukoerythroblastosis and cancer frequency, prognosis, and physiopathologic significance. Cancer, 1979; 44:1009–1013.

West

, Ley

, Pearson

. Myelophthisic anemia in cancer of the breast. Am J Med, 1955; 18:923–931.

Groen

, Godfried

. The occurrence of normoblasts in the peripheral blood in congestive heart failure: An indication of unfavorable prognosis. Blood, 1948; 3:1445–1452.

Frumin

, Mendell

, Mintz

et al. Nucleated red blood cells in congestive heart failure. Circulation, 1959; 20:367–370.

Schwartz

, Stansbury

. Significance of nucleated red blood cells in peripheral blood: Analysis of 1,496 cases. JAMA, 1954; 154:1339–1340.

10.

Yokose

, Ogata

, Dan

, Nomura

. Aplastic anemia with circulating erythroblasts. Int J Hematol, 1994; 60:111–117.

11.

Stachon

, Becker

, Holland-Letz

et al. Estimation of the mortality risk of surgical intensive care patients based on routine laboratory parameters. Eur Surg Res, 2008; 40:263–272.

12.

Stachon

, Holland-Letz

, Krieg

. High in-hospital mortality of intensive care patients with nucleated red blood cells in blood. Clin Chem Lab Med, 2004; 42:933–938.

13.

Stachon

, Kempf

, Holland-Letz

et al. Daily monitoring of nucleated red blood cells in the blood of surgical intensive care patients. Clin Chim Acta, 2006; 366:329–335.

14.

Moore

, Jones

, Kreiner

et al. Validation of a screening tool for the early identification of sepsis. J Trauma, 2009; 66:1539–1546.

15.

McKinley

, Moore

, Sucher

et al. Computer protocol facilitates evidence-based care of sepsis in the surgical intensive care unit. J Trauma, 2011; 70:1153–1167.

16.

Bone

, Balk

, Cerra

et al. 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, 1992; 101:1644–1655.

17.

Knaus

, Zimmerman

, Wagner

et al. APACHE—Acute Physiology and Chronic Health Evaluation: A physiologically based classification system. Crit Care Med, 1981; 9:591–597.

18.

Vincent

, Moreno

, Takala

et al. on behalf of the Working Group on Sepsis-Related Problems of the European Society of Intensive Care Medicine. The SOFA (Sepsis-related Organ Failure Assessment) score to describe organ dysfunction/failure. Intensive Care Med, 1996; 22:707–710.

19.

Gulati

, Behling

, Kocher

, Schwarting

. An evaluation of the performance of Sysmex XE-2100 in enumerating nucleated red cells in peripheral blood. Arch Pathol Lab Med, 2007; 131:1077–1083.

20.

Hall

, Williams

, DeFrances

, Golosinskiy

. Inpatient care for septicemia or sepsis: A challenge for patients and hospitals. Hyattsville, MD: National Center for Health Statistics, 2011.

21.

Moore

, Turner

, Todd

et al. Computerized clinical decision support improves mortality in intra abdominal surgical sepsis. Am J Surg, 2010; 200:839–843.

22.

Stachon

, Holland-Letz

, Kempf

et al. Poor prognosis indicated by nucleated red blood cells in peripheral blood is not associated with organ failure of the liver or kidney. Clin Chem Lab Med, 2006; 44:955–961.

23.

Stachon

, Segbers

, Hering

et al. A laboratory-based risk score for medical intensive care patients. Clin Chem Lab Med, 2008; 46:855–862.

24.

Stachon

, Becker

, Kempf

et al. Re-evaluation of established risk scores by measurement of nucleated red blood cells in blood of surgical intensive care patients. J Trauma, 2008; 65:666–673.

25.

Moore

, Moore

, Todd

et al. Sepsis in general surgery: The 2005–2007 National Surgical Quality Improvement Program perspective. Arch Surg, 2010; 145:695–700.

26.

Shapiro

, Howell

, Talmor

et al. Serum lactate as a predictor of mortality in emergency department patients with infection. Ann Emerg Med, 2005; 45:52–4528.

27.

Shapiro

, Trzeciak

, Hollander

et al. A prospective, multicenter derivation of a biomarker panel to assess risk of organ dysfunction, shock, and death in emergency department patients with suspected sepsis. Crit Care Med, 2009; 37:96–104.

28.

Vorwerk

, Loryman

, Coats

et al. Prediction of mortality in adult emergency department patients with sepsis. Emerg Med J, 2009; 26:254–258.