Role of platelets in hepatic allograft preservation injury in the rat

Abstract

Cold preservation of liver allografts injuries hepatic sinusoidal lining cells. This injury is exacerbated on reperfusion, in part because of adhesion of leukocytes. Platelets also adhere to activated endothelial surfaces. In this study we examined the role of platelets in preservation injury. Our specific aim was to determine whether the degree of platelet adhesion on reperfusion of preserved rat livers was related to duration of cold or warm ischemia and whether platelet adhesion resulted in injury to allografts. We also examined the effect of prior activation of platelets on adhesion and injury. Rat livers were preserved at 1 degree C for different time periods in University of Wisconsin solution and then reperfused for 3 hr on the isolated perfused rat liver system with Krebs-Henseleit solution to which unactivated isolated rat platelets were added. Other livers were rewarmed before reperfusion or reperfused with activated platelets. Platelets were lost from the circulation in all studies; the percentage reduction of circulating platelets was dependent on the length of preservation. The initial platelet concentration did not affect the rate of reduction of platelets in the circuit. Rewarming before reperfusion increased platelet adherence, and prior activation also increased adherence. With electron microscopy we determined that platelets adhered in small aggregates to endothelial cells or endothelial cell remnants. Adherent platelets appeared more activated and contained fewer granules than did unperfused platelets. Liver injury as measured by release of transaminases into perfusate was worsened by longer periods of cold preservation and by addition of rewarming to the protocol. The presence of platelets under these circumstances aggravated injury. Prior activation of platelets also increased the extent of injury. These studies show that platelets have an important role in cold preservation-reperfusion injury.

Evidence that actin disassembly is a requirement for matrix metalloproteinase secretion by sinusoidal endothelial cells during cold preservation in the rat

“…These remnants are of interest because on reperfusion of the liver with unactivated platelets, main site of adherence of platelets is to these remnants rather than to exposed matrix or the rounded SECs, as we have previously shown. 7 This suggests that in the process of retraction and rounding of SECs, adhesive sites for platelets become exposed on the cell remnants. How MMP2 and MMP9 figure in these more complex events is at present unclear, however they may be great importance because both effective liver preservation solutions, UW solution and HTK solution, retard these changes, and the common property of these solutions is that they contain cryptic inhibitors of MMPs.…”

“…This appearance in cell culture of retraction of cell processes into the cell body, while leaving behind small cell remnants attached to matrix, is very similar to what we have described using transmission electron microscopy in whole rat livers that have been cold preserved. 4,7 Pretreatment of SECs with phalloidin of SECs, which were then exposed to 8 hours of cold preservation, resulted in a remarkable maintenance of normal cell shape (Fig. 4C).…”

“…4,6 After preservation, at the moment of reperfusion, recipient leukocytes, platelets, and clotting factors come into contact with the exposed donor connective tissue matrix and hepatocytes, as well as the altered sinusoidal endothelial cells (SECs), in the hepatic sinusoid. This leads to adherence and activation of the platelets 7 and leukocytes 8,9 as well as thrombosis 10 with resultant damage to the allograft. The degree to which these events occur on reperfusion is directly related to the duration of cold preservation in every model.…”

Decreased survival in rat liver transplantation with extended cold preservation: Role of portal vein clamping time

“…It is currently hypothesized that Kupffer cell activation plays a causal role in cold ischemiareperfusion injury of the liver, precipitating sinusoidal endothelial cell (SEC) death, and leading to microcirculatory disturbances. [1][2][3][4][5][6][7] This hypothesis is based on experimental findings using mainly the orthotopic rat liver transplantation (ORLT) model, first described by Lee et al 8 and subsequently modified by Kamada and Calne. 9 However, recent experimental reports have shown that, after ORLT following extended cold ischemia, the mortality of animals could not be related to the severity of SEC dysfunction.…”

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“…It is currently hypothesized that sinusoidal endothelial cell impairment, [9][10][11] activation of Kupffer cells, 12,13 microcirculatory disturbances, and sinusoidal accumulation of leukocytes [14][15][16] and platelets 17,18 are major causes of preservation-related graft failure. This hypothesis is strongly supported by observations that endothelial cells lose their function with increasing time of cold preservation 19,20 and die during a brief period of reperfusion.…”