Biomedical subjects
Publications and source records attributed to Luciana L Almada.
This study provides evidence of a new treatment approach for colorectal peritoneal metastases, a condition associated with a worse prognosis than other types of metastases.
Ischemia-reperfusion injury is a major cause of early graft dysfunction after liver transplantation. The bile flow has been suggested as an index of ischemic damage, and severely impaired bile flow seems to be predictive of poor survival in experimental studies. Looking for injury markers, biliary inorganic phosphate has the potential of being a useful endogenous marker of diminished hepatobiliary function because this anion is excreted in the bile by a paracellular pathway and it can detect changes in permeability. The goal of this study was to evaluate the effects of cold preservation-reperfusion of the liver on bile flow and bile inorganic phosphate and their relationship with storage-related graft failure. The isolated and perfused rat liver was used to evaluate the injury for ischemia-reperfusion. The intrahepatic resistance, lactate dehydrogenase release, and potassium and biliary inorganic phosphate concentration were used to estimate viability and function of freshly isolated or cold-preserved livers. The intrahepatic resistance and the bile flow were consistent and significantly decreased throughout the perfusion time in relation to the increment in storage. Inorganic phosphate is more concentrated in bile from preserved livers, showing an alteration in paracellular pathway, confirmed by the biliary excretion of horseradish peroxidase. After preservation, concentration and excretion of the paracellular marker were increased during the first peak. The second peak appears earlier in preserved livers (10 minutes) with a different shape but without changes in concentration. In conclusion, inorganic phosphate in bile shows changes in paracellular permeability as occurs in livers after 48 hours of cold preservation.
The ammonium removal efficiency (ARE) and cell viability was investigated in freshly isolated rat hepatocytes exposed to increasing ammonium loads (0.1-2.0 mM). No difference was observed in both ARE and cell viability at the different ammonium concentrations tested. Storage of hepatocytes at 0 degrees C for 72 hours inhibited ammonium removal and urea synthesis. Rewarming of cells at 37 degrees C for 120 min was followed by an ARE fully comparable to freshly isolated hepatocytes. These data indicated that cold preservation of rat hepatocytes for 3 days in UW followed by a rewarming is associated with normal ammonium detoxification efficiency.
Livers cold preserved in University of Wisconsin (UW) solution followed by reperfusion suffer ischemia/reperfusion injuries. Microcirculation is the primary target of damage, characterized by sinusoidal perfusion failure due, mainly, to morphological changes of sinusoidal endothelial cells. Here, we demonstrated that the addition of S-nitrosoglutathione (GSNO) to the UW solution before cold storage, as a nitric oxide (NO) donor, attenuated hepatic injuries.Wistar adult rat livers were stored in UW solution (0 degrees C-48 hs) and then reperfused during 60 minutes using the Isolated Perfused Rat Model (IPRL). We assayed four GSNO concentration (50, 100, 250 and 500 mM). NO concentration was estimated calculating the amount of nitrite (NO2-) generated in the UW solution. Injuries during cold preservation were established measuring lactate dehydrogenase (LDH) released to the UW solution. Meanwhile, intrahepatic resistance (IR), LDH released to the perfusate, the effluent/perfusate ratio for K+, bile flow, liver glycogen content and sinusoidal endothelial cell morphology were studied after 1 hour of reperfusion in the IPRL system. In cold preserved livers without GSNO, glycogen content was dramatically reduced, IR increased markedly, LDH released was high, bile flow diminished and sinusoidal endothelial cells appeared rounded and detached from perisinusoidal matrix after reperfusion. The presence of 100 mM GSNO prevented the IR rise and LDH release, improved bile production and partially reduced endothelial cells damages. In conclusion, the addition of 100 mM GSNO to UW solution improved hemodynamic and function capacity of cold preserved/reperfused livers.
A rapid and simple assay (6 min, two steps) is described for determination of cell viability of hepatocytes subjected to cold preservation protocols. In this method, cells are incubated with the fluorescent marker propidium iodide (PI) and the fluorescence intensity is measured before (direct fluorescence--Fd) and after (total fluorescence--Ft) addition of digitonin, which allows the dye to enter the hepatocytes. The Fd originated from non-viable cells that have membrane damage and taken up PI. The Ft originated from all cells in the sample. The ratio between the two fluorescence values is used as an indicator of cell viability. The assay was challenged versus two classical viability tests: LDH retention and Trypan Blue exclusion. Our assay shows good correlation only with Trypan Blue test. In addition, a fluorescence confocal microscopy protocol was used to evaluate the possible toxicity of PI in hepatocyte suspensions.