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S1P Distribution in Plasma of Septic Patients: Characterization Using Flow Induced Dispersion Analysis (FIDA)
Isabelle Seidita 1, Anke C. Ziegler, Markus H. Gräler2,3,4
1 Department of Experimental and Clinical Biomedical Sciences Mario Serio, University of Florence, 50121 Firenze, Italy
2 Department of Anesthesiology and Intensive Care Medicine, Jena University Hospital, Jena, Germany.
Sepsis is defined as a life-threatening organ dysfunction caused by a dysregulated host response to infection. Various studies have shown an involvement of sphingosine-1-phosphate (S1P) in the condition, with a decrease of serum and plasma S1P levels in sepsis patients that correlates with disease severity. S1P is a bioactive sphingolipid that regulates several processes involved in the pathophysiology of sepsis, such as endothelial barrier protection, lymphocyte trafficking and cytokine production. In circulation, S1P is present bound to apolipoprotein M (ApoM) in high densitiy lipoproteins (HDL) or associated with human serum albumin (HSA), which were identified as the two main carriers of S1P in plasma. Several studies have suggested different signalling properties for HDL-S1P and HSA-S1P: HDL-S1P, but not HSA-S1P, was able to reduce vascular inflammation and induce endothelial barrier stabilization. A recent study by Winkler et al. used liquid chromatography coupled to triple-quadrupole mass spectrometry to analyze the levels of HSA- and HDL-associated S1P in the plasma of intensive care unit patients with surgical trauma, sepsis or septic shock and healthy controls. The results showed significantly decreased plasma S1P levels with sepsis severity, while in surgical trauma patients the total S1P levels in the plasma were unaltered. Interestingly, however, in all the patient groups, a redistribution of S1P among its carriers was observed, with decreased HSA-S1P and increased HDL-S1P compared to healthy controls: in the less severe cases, total plasma S1P levels were maintained via increased HDL-S1P levels.
Flow induced dispersion analysis (FIDA) is a technology that relies on the analysis of a parabolic hydrodynamic flow profile for measuring the apparent hydrodynamic radius of a selective ligand (indicator) as it binds to an analyte of interest. The indicator is injected into a capillary together with the analyte, and its flow profile is detected via a fluorescent detector: large molecules or complexes diffuse slowly, while smaller molecules diffuse faster. The apparent hydrodynamic radius (size) depends on the apparent diffusivity, which in turn is dependant on the fractions of free and complexed indicator in the sample.
In this study, we established a method to use the FIDA technology to characterize the distribution of S1P among its carriers in plasma samples in a fast and easy way, using FITC-labelled S1P as the indicator and patient plasma as the analyte. We compared healthy controls with surgical trauma, sepsis and septic shock patients, and we were able to confirm the shift from HAS-S1P to HDL-S1P in the patients compared to the controls found by Winkler et al. Moreover, in the more severe cases, we found significant correlations between the apparent hydrodynamic radius measured in plasma samples taken on day 1 after the diagnosis and important clinical outcomes several days later.
Thus, this preliminary study shows promising results for the development of a fast and simple way of characterizing the distribution of S1P in patient plasma samples, that might also serve as a predictive factor for positive clinical outcome in the more severe cases.
Speakers
Isabelle Seidita