Mathias Oberhoffer

Procalcitonin expression in human peripheral blood mononuclear cells and its modulation by lipopolysaccharides and sepsis-related cytokines in vitro.

Procalcitonin (PCT), the precursor of calcitonin, was recently put forward as a diagnostic marker of systemic bacterial infection and sepsis. The major PCT production site in sepsis still remains unclear. Because of a certain association between increased levels of PCT and leukocyte-derived cytokines during sepsis, we assessed the possible expression of PCT in human peripheral blood mononuclear cells (PBMCs) and the modulation of PCT by lipopolysaccharides (LPS) and various sepsis-related cytokines by reverse transcriptase-polymerase chain reaction (RT-PCR) by using a novel primer set and flow cytometric analysis with intracellular staining with antibodies to the PCT components calcitonin and katacalcin. RT-PCR and flow cytometric analysis demonstrated that PBMCs express PCT both on mRNA and on protein levels. LPS and various proinflammatory cytokines (interleukin-1beta (IL-1beta), IL-6, tumor necrosis factor-alpha (TNF-alpha), IL-2) had pronounced stimulatory effects on the expression of PCT mRNA. Under identical experimental conditions the anti-inflammatory cytokine IL-10 had no effect on the expression of mRNA for PCT. Flow cytometric analysis demonstrated increased intracellular amounts of PCT components after LPS stimulation. Thus we demonstrate for the first time that PCT is expressed in PBMCs. This expression is modulated by bacterial LPS and sepsis-related cytokines. Therefore PBMCs may be among the sources of elevated PCT levels in patients with sepsis.

Procalcitonin — A new indicator of the systemic response to severe infections

Severe infection and sepsis with consecutive multiple organ dysfunction or failure (MODS) are major causes of morbidity and mortality in modern intensive care units [1]. Recent advances in our understanding of the pathogenesis of sepsis have made it clear that uncontrolled infections, whether clinically manifest or occult, are not the only cause of systemic inflammation and organ dysfunction. Other stimuli such as pancreatitis [2], major trauma [3], and burns [4] can also trigger an excessive inflammatory response and lead to MODS. As a result, it is now generally accepted that not all patients displaying the clinical signs of sepsis have an underlying infection. It is therefore frequently difficult to distinguish patients with systemic infection from those who appear septic but have no bacteriological or clinical evidence of infection. Common signs of systemic inflammation such as changes in body temperature, leukocytosis, and tachycardia may have an infectious or noninfectious etiology and are neither ,specific nor sensitive for sepsis. Thus, of 61 patients with sepsis (documented infection and at least two signs of remote organ failure), 35 (57%) presented with leukocytosis, 18 (29%) had leukopenia, and eight (14%) had normal leukocyte counts (unpublished data). Likewise, patients with sepsis may present with fever, hypothermia, or normal body temperature. Bacteriological evidence of infection, although considered a gold standard, may have some drawbacks. Negative blood cultures do not exclude sepsis and multiple samples may be required over extended periods before positive cultures emerge [5]. Furthermore, positive or negative cultures may not develop concurrently with clinical signs of sepsis. Because these common clinical and laboratory parameters lack sensitivity and specificity, others are needed to provide an early marker of the infectious etiology of a generalized inflammatory response and thus allow early diagnosis and the application of more specific therapeutic interventions. Furthermore, new parameters also may help identify subgroups of septic patients who may benefit from proor antiinflammatory therapies. One such parameter, procalcitonin, has recently attracted attention as a possible marker of the systemic inflammatory response to infection.

Outcome prediction by traditional and new markers of inflammation in patients with sepsis.

Abstract Patients (n = 242) admitted to intensive care unit for longer than 48 hours were categorised for sepsis according to American College of Chest Physicians/Society of Critical Care Medicine (ACCP/SCCM) Consensus Conference criteria. Body temperature, leukocyte count, C-reactive protein (CRP) and procalcitonin (PCT) as well as tumor necrosis factor-α (TNF-α), interleukin (IL)-6, IL-8, IL-10 and HLA-DR expression on monocytes were determined. Data of one randomly choosen day per patient entered analysis. Immunologic mediators contributing significantly to outcome were determined by logistic regression analysis. Area under the curves (AUC) of receiver operating characteristic curves of clinical markers of inflammation predicting prognosis were compared with AUC of relevant immunologic mediators. TNF-α, IL-6 and HLA-DR expression on monocytes were significantly associated with outcome; the AUCs were 0.835, 0.844 and 0.761 respectively. AUCs for clinical markers were 0.878, 0.811, 0.620 and 0.614 for PCT, CRP, leukocyte count and body temperature respectively. PCT had the highest AUC compared to other clinical markers. These data indicate that PCT might be a better marker than the classic criteria of inflammation, CRP, leukocyte count, and body temperature to identify patients endangered by severe infection or sepsis.

Sensitivity and specificity of various markers of inflammation for the prediction of tumor necrosis factor-α and interleukin-6 in patients with sepsis

OBJECTIVES To determine correlations and predictive strength of surrogate markers (body temperature, leukocyte count, C-reactive protein [CRP], and procalcitonin [PCT]) with elevated levels of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) in septic patients. DESIGN Prospective consecutive case series. SETTING Surgical intensive care unit (ICU) of a university hospital. PATIENTS A total of 175 patients experiencing intensive care unit stays >48 hrs categorized for sepsis according to ACCP/ SCCM Consensus Conference criteria. MEASUREMENTS AND MAIN RESULTS CRP and PCT were both significantly correlated with TNF-alpha and IL-6. Based on the area-under-the-curve of the receiver operating characteristics curves, predicting capability was highest for PCT (0.814 for TNF-alpha >40 pg/mL and 0.794 for IL-6 >500 pg/mL), moderate with CRP (0.732 and 0.716, respectively), and lowest for leukocyte count (0.493 and 0.483, respectively) and body temperature (0.587 and 0.589, respectively). Sensitivity, specificity, positive, and negative predictive values and test effectiveness all followed this same pattern of being highest for PCT followed by CRP, with leukocyte count and body temperature being lowest. CONCLUSION PCT may be an early and better marker of elevated cytokines than the more classic criteria of inflammation.

Molecular Aspects and Natural Source of Procalcitonin

Abstract The search for sensitive and specific markers of systemic infection has shown that procalcitonin levels are increased in sepsis, and, consequently, this plasma protein has come into the focus of clinical research. Human procalcitonin is encoded by the Calc-I gene, which gives rise to two alternatively spliced transcripts. Despite systemic investigation of the Calc-I gene and mechanisms of the tissue-specific regulation and alternative splicing, little is known about the biology of procalcitonin and the cells which express this protein during inflammation. Here we focus on the molecular and biochemical properties of the molecule and summarize the known biological functions of procalcitonin. We report on the structure of the Calc-I gene, the amino acid conservation of procalcitonin in different species, and the consensus sequences of the protein with regard to sites relevant for posttranslational modification, spatial distribution, and homologies to other cytokines. We discuss aspects of intracellular location of procalcitonin and demonstrate that it has the characteristics of a secreted protein.

Total plasma antioxidant capacity is not always decreased in sepsis

OBJECTIVE To compare total plasma antioxidant capacity and selected individual antioxidants in patients with varying degrees of severity of sepsis. DESIGN A prospective, observational, consecutive case study. SETTING A 16-bed intensive care unit (ICU) in a university teaching hospital. INTERVENTIONS None. PATIENTS Forty-six healthy controls, ten ICU patients, nine patients with systemic inflammatory response syndrome (SIRS), 11 septic patients, and 14 septic shock patients. Plasma was obtained in healthy patients scheduled for minor surgery immediately before anesthesia and in ICU patients within 24 hrs of admittance to the unit or diagnosis of SIRS, sepsis, or septic shock. MEASUREMENTS AND MAIN RESULTS Using the total peroxyl radical trapping method, we found plasma antioxidant capacity to be lower in septic patients but higher in septic shock patients, as compared with controls. Bilirubin was the greatest contributor to the increase with shock, followed by uric acid. Neopterin also correlated with the peroxyl radical trapping antioxidant parameter values. CONCLUSION Although total plasma antioxidant capacity is decreased from normal levels in septic patients, an increase in some oxidants contributes to an increased total antioxidant capacity in septic shock patients.

Katacalcin and calcitonin immunoreactivity in different types of leukocytes indicate intracellular procalcitonin content

PURPOSE Procalcitonin is a new marker of severity of nonviral, in particular, bacterial infections. In respect of sepsis its site of production remains unknown. This study was carried out to determine whether subsets of human leukocytes contain procalcitonin. MATERIALS AND METHODS Blood samples of 17 patients who had demonstrated various degrees of serum procalcitonin levels on the day before evaluation were analyzed for serum procalcitonin by immunoluminometry and for intracellular reaction of monocytes, granulocytes, B-, and T-lymphocytes against katacalcin- or calcitonin-sensitive antibodies. Katacalcin and calcitonin are part of the procalcitonin molecule. Associations of these reactions with serum procalcitonin levels as well as differences between groups with a normal or elevated serum level were analyzed. RESULTS Intracellular antibody reaction against katacalcin was demonstrated in all cell types. We also found a lower rate of intracellular antibody reaction against calcitonin. Associations of serum procalcitonin with the two antibody reactions were demonstrated. Differences in intracellular reactions in the group with elevated serum procalcitonin were seen with both antibodies compared with a normal control. CONCLUSION Intracellular antibody reaction against katacalcin supports the notion that various types of leukocytes contain procalcitonin.

ACCP/SCCM Consensus Conference definitions correlate better with procalcitonin than tumor-necrosis-factor-α and interleukin-6

ConclusionIn contrary to TNF-α and IL-6, PCT differs in all stages of systemic inflammation proportional to severity in accordance with the consensus conference definitions. PCT values are helpful in newly and severely infected patients, primarily of bacterial cause. After endotoxin injection PCT increases shortly after TNF-α and before IL-6 (1). Hence PCT might serve as an early and dependable indicator in patients suffering from severe infection/systemic inflammation. The results have yet to be confirmed by greater sample sizes.

Procalcitonin Ein neuer diagnostischer Parameter bei schweren Infektionen und Sepsis

ZusammenfassungProcalcitonin (PCT), ein aus 116 Aminosäuren bestehendes Glykoprotein, wurde als neuer Parameter der schweren Infektion inauguriert. Sein Syntheseort in diesem Zusammenhang ist nicht bekannt. Mittels immunoluminometrischen Assays kann die PCT-Konzentration innerhalb von ca. 2 h aus 40 µl Serum oder Plasma bestimmt werden. PCT-Erhöhungen sind u.a. mit Lipopolysaccharid und mit den Zytokinen TNF-α und IL-6 assoziiert. Bakterielle, parasitäre und pilzbedingte generalisierte Infektionen weisen im Gegensatz zu lokal begrenzten Infekten häufig einen Wert über 2 ng/ml auf, wobei die Spezifität des Parameters mit der Höhe des Werts steigt. Therapeutische Maßnahmen, die zur lokalen Begrenzung der Infektion führen, resultieren in einem Abfall von PCT. In den ersten Tagen nach großen Operationen, Polytrauma, bei wenigen malignen Erkrankungen, Hitzschlag wie auch im Rahmen spezieller Behandlungen hämatologischer Krankheiten kann der Parameter erhöht sein, ohne daß eine Sepsis oder schwere Infektion vorliegt. Bisherige Studien legen nahe, daß PCT bei Patienten mit Sepsis im Vergleich zu traditionellen Entzündungs- bzw. Sepsisparametern Vorteile besitzt, wobei an erster Stelle die Eigenschaft, eine generalisierte Infektion anzuzeigen, zu nennen ist.AbstractProcalcitonin (PCT), a glycoprotein consisting of 116 amino acids, has been proposed as a new marker of severe infection. The site of production under this condition remains unknown. The serum PCT concentration is determined by an immunoluminometric assay of 40 µl serum or plasma requiring approximately two hours. Elevations of PCT are for instance associated with levels of lipopolysaccharide and the cytokines TNF-α and IL-6. Bacterial, parasitic or fungal infections developing septic complications in contrast to local infections, often show values ex- ceeding 2 ng/ml. The specificity of the parameter in this context increases with its concentrations. Therapeutic actions that confine the infection locally are reflected by a decrease of the PCT value. PCT may be elevated within the first days after extended surgery or polytrauma, in some malignancies, heatstroke and during treatment of some hematologic diseases without an existing sepsis or severe infection. Previous studies indicate certain benefits of PCT compared to traditional markers of inflammation or sepsis, where the ability to indicate a generalized infection is the primary advantage.