Protein Variation Related to Sporidesmin Induced Liver Damage and to Genetic Selection For or Against Sporidesmin Induced Damage

Abstract

Quantitative two-dimensional polyacrylamide gel electrophoresis was used to study protein variation related to liver damage caused by the fungal toxin sporidesmin, This toxin causes a photosensitising disease, called facial eczema, in livestock. Photosensitisation is secondary to liver damage. Protein variation was measured in C57BL/6 mice and Romney sheep. Changes in liver proteins which were caused by sporidesmin dosing were measured in both species, and changes in gall bladder, bile and serum proteins were also measured in the mice. A further study of protein variation was made between flocks of sheep which had been selected for resistance or susceptibility to sporidesmin induced liver damage. The goal of these studies was to detect changes which might be related to the mechanisms of sporidesmin toxicity, and to detect proteins which might be used as the basis of tests for detecting livestock which were resistant to the effects of the toxin. Significant differences (P<0.05), in the amounts of 19 out of 30 proteins which were measured, were detected in the liver homogenates of C57BL/6 mice 1 or 4 days after dosing with sporidesmin. Up to two fold losses or increases in protein amount occurred after dosing. There were also differences in the in vitro translation of mRNA which had been extracted from the livers of affected and control mice. Substantial histological damage and protein change also occurred in the walls of the gall bladders of dosed mice. Approximately 90% of the measured gall bladder proteins showed up to 10-20 fold changes in abundance after dosing. Protein change appeared to precede histological injury. The protein profile of bile completely changed after dosing with sporidesmin. Serum proteins appeared to dominate in bile of dosed animals. Changes in abundance of several serum proteins were also detected after dosing. This included increases in the acute phase reactants α2 HS glycoprotein and haptoglobin β. Liver protein variation was also measured in flocks of Romneys which had been selected for either resistance or susceptibility to sporidesmin induced liver damage. There were 7 significant (P<0.05) differences in abundance among 62 proteins which were compared between ewes from a resistant flock and a susceptible flock (5 animals per flock). Quantitative data was also obtained for 78 proteins for groups of 4 rams from two resistant flocks, a susceptible flock and a control unselected flock. There were 37 significant (P<0.05) flock related differences, including a protein which was more abundant in the two resistant flocks than in the susceptible and control flocks, and another protein which had also differed between the ewe flocks. When an analysis was made of changes in protein abundance in the liver of ewes from one of the resistant flocks (n=8) which had been experimentally dosed with sporidesmin, 0.1-3 mq/kg body weight, 25 of the 79 proteins whose amounts were measured showed significant change (P<0.05). Nine of these proteins showed more than 2 fold change (5 gains and 4 losses) following the dosing. There was also a change in abundance of one of the proteins which was previously shown to vary between resistant and susceptible ewes. This approximately 25 kDa, pI 5.6 unidentified protein showed a mean 35% decrease in abundance following sporidesmin dosing. This protein had a 3.9 fold greater abundance in undosed ewes from the susceptible flock compared with the resistant flock and 1.4 - 1.5 fold greater abundance in rams from the two resistant flocks compared with rams from the susceptible flock. In conclusion, dosing with sporidesmin produced many changes in abundance of liver proteins in mice and sheep. Greater changes occured in the gall bladder than in the liver of C57BL/6 mice. There was also some variation in the amounts of individual liver proteins among flocks of Romneys which had been selected for resistance or susceptibility to the toxic effects of sporidesmin on the liver. Some of these differences may be related to specific actions of sporidesmin, or to less specific responses of the liver to toxic challenge.