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Predictive Identification of High Risk Sepsis and Shock Patients in Intensive Care Medicine |
1. Introduction:
Intensive care patients are in life threatening conditions when
affected by sepsis or non infectious shock. Granulocytes
represent a major defense barrier against these affections.
They phagocytose microorganisms or tissue breakdown products,
but they also release enzymes or pharmacologically active mediators.
In this way they can represent a danger for the organism if
their potent functionalities escape inhibitory control mechanisms.
- It is clinically of high importance to recognize the danger of sepsis or
shock as early as possible. Unfortunately, this is not readily
possible by the determination of humoral biochemical markers in
the vascular or other body compartments.
2. Goal:
The concept of this work was to determine cell function parameters
of granulo- and monocytes at a molecular level to obtain predictive
(prognostic) indicators of imminent danger to patients.
- Our earlier flow cytometric work (
1,
2)
using
bacterial phagocytosis (
6,
7),
ADB
intracellular pH and esterase (
1,
2)
measurements as well as
acridine orange
as indicator of cellular and bacterial RNA and DNA(
7)
had shown for the first time that the prediction of imminent
danger of sepsis and non infectious shock in intensive care
(IC) patients was already possible two to three days
in advance to the appearence of clinical symptoms
(CL1).
These findings provide a significantly increased
therapeutic lead time for the clinician.
- Although conceptually promising, the use of bacteria in a phagocytosis
assay is comparatively complicated e.g. in automatically operated
flow cytometers. The concept was therefore to facilitate the practical
approach to cell function assays. This was achieved by:
1. the use of humoral stimulators like e.g. cytokines and
2. the development of the sensitive oxidative
burst indicator dye
dihydrorhodamine123 (DHR) (
8,
10,
11,
14)
and of the specific
rhodamine110 substrates
for the determination of
protease activity (
12,
13,
17-22,
24)
in vital cells. These developments have substantially simplified the
determination of blood cell functions in infection, sepsis or non infectious
shock.
3. CLASSIF1
Data Pattern Analysis
Flow cytometric data of such measurements are typically collected
as list mode files. They are then evaluated in a
standardized and automated way by the CLASSIF1
(CL1,CL2)
multiparameter data classification program.
- The analysis of the entire data set in this way
reveiled that the incubation of ex-vivo leukocyte preparations:
- alone (ex-vivo)
- with physiological stimulators such as: suboptimal concentrations
of FMLP (formyl-methionyl-leucyl-phenylalanyl bacterial peptide),
TNF-alpha (tumor necrosis factor-alpha),
FMLP+TNF-alpha and
- with phorbol ester (PMA, phorbol-myristate-acetate) as
maximum stimulator
provides a sufficient amount of predictive information
(CL1)
for the early risk determination in septically admitted IC patients
e.g. already on the day of admission, similarly as the cytometric
determination of proteolytic enzyme activities like:
- cysteine proteinases or
- serine proteinases
- The optimization of the classification process
for the same group of septically admitted IC patients showed
(CL1)
that the most discriminatory predictive information was contained in
the FMLP and TNF-alpha stimulated oxidative burst (DHR123)
assays.
- As a practical consequence of the CLASSIF1
multiparameter data analysis, only two out of the seven
performed assays were really required for survival prediction in this
group of septically admitted IC patients (CL1).
4. Conclusions:
A. Functional parameters of granulocytes and monocytes provide
predictive information for individualized risk assessment in
septically admitted IC patients already on the day of admission
B. Automated operation with assay preparation,
flow cytometric measurement, on-line data analysis in combination with
standardized result classification is possible in principle either with
laboratory flow cytometer + cell staining robot station
but conceptually also with larger hematology analyzers equipped
with fluorescence channels such as increasingly available for
clinical routine laboratories in the context of
medical cytomics or clinical cytomics.
Literature References:
CL1.
G.Valet, G.Roth, W.Kellermann: Risk assessment for intensive care
patients by automated classification of flow cytometric oxidative burst,
serine and cysteine proteinase measurements using CLASSIF1 triple matrix
analysis, in: Cytometric Cellular Analysis, Eds: J.P.Robinson, G.Babcock,
Wiley-Liss, New York 1998, p.289-306
CL2.
G.Rothe, W.Kellermann, J.Briegel, B.Schaerer, G.Valet:
Activation of neutrophils by tumor necrosis factor-alpha during sepsis, in:
Immune Consequences of Trauma, Shock and Sepsis Vol.II, Ed: E.Faist, J.Ninnemann,
D.Green, Springer Verlag, Berlin 1993, p.727-733
CL3.
G.Rothe, W.Kellermann, G.Valet: Flow cytometric parameters
of neutrophil function as early indicators of sepsis or trauma-related
pulmonary or cardiovascular organ failure.
J.Lab.Clin.Invest.115:52-61(1990)
| © 2012 G.Valet |