In Vivo Measurement of Site-Specific Peritoneal Solute Transport Using a Fiber-Optic-based Fluorescence Photobleaching Technique

Abstract

Fluorescence recovery after photobleaching (FRAP) is a well-established method commonly used to measure the diffusion of fluorescent solutes and biomolecules in living cells or tissues. Here a fiber-optic-based FRAP (f-FRAP) system was developed, and validated using macromolecules in water and agarose gels of different concentrations. We applied f-FRAP to measure the site-specific diffusion of fluorescein (NaFluo) in peritoneal membranes (PMs) on the liver, cecum, and kidney of a living rat during peritoneal dialysis. Diffusion of fluorescein in PM varied in a time-dependent manner according to the type of organ (DPM on Liver/DNaFluo = 0.199 ± 0.085, DPM on Cecum/DNaFluo = 0.292 ± 0.151, DPM on Kidney/DNaFluo = 0.218 ± 0.110). The proposed method allows direct quantitative measurement of the three-dimensional diffusion in local PM in vivo, which was previously inaccessible by peritoneal function test methods such as peritoneal equilibration test (PET) and standardized PM assessment (SPA). f-FRAP is promising for local and dynamic assessments of peritoneal pathophysiology and the mass transport properties of PMs, presumed to be affected by variation of tissue structures over different organs and functional changes of the PM with years of peritoneal dialysis.