Sepsis Research Today is a free monthly online journal that collates and summarizes the latest research about Sepsis, including details on septicemia, diagnosis, symptoms, treatment. | ||||||||
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Use of multiple fluorophores for evaluating microvascular permeability in control rats and rats with sepsis.Assaly RA, Habib RH, Azizi M, Shapiro JI, Dignam JD Department of Medicine, University of Toledo College of Medicine, Toledo, OH 43614, USA. ragheb.assaly@utoledo.edu Capillary leak accompanying systemic inflammatory response conditions is a significant clinical problem. In the present study, we describe and verify a method for studying capillary leak that is based on the injection of proteins that differ significantly in size and have spectrally distinguishable fluorophores. Control (n=11) and post-CLP (caecal ligation and puncture; n=14) Sprague-Dawley rats were injected with tracer amounts of albumin and PEG-Alb [albumin covalently linked to methoxy-poly(ethylene glycol)] labelled with fluorescein and Texas Red. Blood samples were withdrawn between 5 min and 144 h, and the fluorescence of the labelled proteins was determined. The relative retention of the PEG-Alb and albumin was assessed via measurement of the TER (transcapillary escape rate; in %/h) and the t(50%) estimate, defined as the time when the actual concentration reached 50% of its baseline. The concentration-time trends for both albumin and PEG-Alb tracers exhibited two-compartmental behaviour and were analysed using bi-exponential modelling. Retention times were significantly greater for PEG-Alb in both control and CLP rats. TER(PEG-Alb) was significantly lower than TER(albumin) for both control (8.1+/-5.6 compared with 14.8+/-7.1 %/h respectively; P<0.01) and CLP (14.8+/-6.6 compared with 22.5+/-7.3 %/h respectively; P<0.001) rats. The t(50%[PEG-Alb]) was substantially greater than the corresponding t(50%[albumin]) for both control (29.8+/-9.8 compared with 7.2+/-2.0 h respectively; P<0.001) and CLP (12.9+/-5.6 compared with 5.1+/-1.6 h respectively; P<0.001) rats. The result was similar irrespective of the fluorophore-protein combination, validating the multifluorophore technique. In conclusion, the double-fluorophore approach described in the present study may provide the future basis for a method to quantify capillary leak in disease. Published 7 December 2007 in Clin Sci (Lond), 114(2): 123-30.
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