Medical Hypotheses 4: 353-361, 1978.
LIPID PEROXIDATION IN THE HEMOLYTIC UREMIC SYNDROME S. O'Regan* and J.S.C. Fong. The Department of Nephrology, University of Montreal, Hbpital Ste-Justine - Institute of Research*, 3175 CGte Ste-Catherine Road, Montreal H3T lC5, Quebec, Canada, Department of Nephrology, McGill University, Montreal Childrens Hospital Research Institute, Montreal, P.Q. Canada. ABSTRACT Based on recent evidence of a genetic influence on prognosis (1) and the existence of red cell membrane phospholipid depletion with low or absent serum a-tocopherol (2) levels in three children with the Hemolytic Uremic Syndrome (H.U.S.), we wish to suggest the existence of an inborn error of antioxident capacity as the basic pathogenetic mechanism in the development of the hemolytic uremic syndrome (H.U.S.). Key Words:
Hemolytic Uremic Syndrome, Lipid Metabolism
353
INTRODUCTION Since the initial description of the HUS by Gasser et al., (3) this symptom complex has been reported with increasing frequency. (4,5) Marked variations in the geographical, (6) age, (7) and racial (6) distribution must be reconciled in any attempts to propose a pathogenetic mechanism. A unifying concept is required to explain its association with such diverse conditions as viral (overt (8) and vaccination (9), bacterial infection, (10) oral contraceptive therapy, (11) and post renal transplantation. (12) A role for injested peroxidized lipids has previously been proposed by Wolman. (13) This however would be expected to produce larger numbers of cases in clusters, i.e., whole families. We wish to extend the peroxidized lipid hypothesis and propose an endogenous source of lipid precipitating a pathologic cascade to account for the major features of the disease, i.e., haemolytic anemia with schizocytosis, thrombocytopenia, and renal failure. We recently reported (2) the association of depressed c-tocopherol activity and red cell membrane phospholipid (phosphatidyl ethanolamine) depletion in the HUS in 3 children. One parent with 3 affected children also had an abnormal erythrocyte membrane phosphdlipid pattern associated with undetectable tocopherol activity in her serum. A marked genetic influence on prognosis has been noted from examination of the outcome of the disease in 25 sibships. (1) These two features help substantiate our proposal of the existence of an inborn error in lipid metabolism, i.e., decreased antioxident capacity of intracellular enzyme systems normally active in maintaining the integrity of the cell membrane. Marked alterations in lipid metabolism are a common feature of gram negative septicemia. (14) Disturbances in lipid metabolism have also been reported in association with oral contraceptive therapy (15) and renal transplantations. (16) Marked rises in serum triglyceride and cholesterol occur in all these conditions as in the HUS. (17) Vit. E as an Antioxident In the absence of severe protein calorie malnutrition or fat malabsorption deficiency of u-tocopherol is exceedingly rare, only one condition to date demonstrating depressed serum levels because of increased utilization. (18) The major role of vitamin E is considered to be that of an antioxident. It acts as a free radical scavenger (20) and thus prevents the peroxidation of polyunsaturated fatty acids. It may also be active in the synthesis of heme containing proteins active in the scavenging of peroxides (21). Thus an important action of this compound is in the maintenance of cellular and intracellular membrane integrity by possible interactions with polyunsaturated lipids in membranes. (22) This importance of a-tocopherol for membrance stability has been studied on erythrocytes of rats fed on vitamin E deficient diet. (23) The primary
354
defect noted in the membrane lipid was an almost complete depletion of phosphatidyl-ethenolamine (P.E.), demonstrated following in vitro exposure of cells to H202. (Peroxide fragility test). Reappearance of P.E. was dramatic when vitamin E sufficiency was maintained. An additional finding was the inability of the mature erythrocyte to repair losses of lipid components of the cell membranes. Thus the effect of the antioxident capacity of vitamin E playing an in vivo role in protecting against red cell membrane lipid degradation was to slow the rate of peroxidation and not to stimulate regeneration of lipids in the mature cell. Evolution of the Syndrome. When a free radical-derived from the breakdown of H202, or any other source, (e.g. peroxidized lipid), reaches a lipid double bond in the presence of 02 unstable lipoperoxides are produced. These can then spontaneously degenerate to form more free radicals which attack other double bonds, to produce more lipoperoxides. This peroxide challenge without effective antioxident capacity may propagate an autocatalytic chain reaction. The process might be accelerated by a state in which rapid production of more peroxidized lipid is continuing from endogenous sources and is being released into the circulation. Such a release phenomenon would have its initial and major effects on membranes of circulating cells, i.e., RBC's, platelets - they being rapidly subjected to massive exposure to peroxidized lipids. Due to the necessity of diffusion from the circulation, etc., - effects would be slower on "fixed" tissues. The susceptibility to massive membrane peroxidation would depend on: a.
Rapid utilization of available tocopherol due to continuous release of peroxidizing lipids with ultimate saturation of antioxident (extracellular) capacity.
b.
Failure of intracellular mechanisms of antioxidation after complete utilization of extracellular capacity.
Other involved tissues would also demonstrate effects of peroxidation, i.e., phospholipid content, especially if subject to hypoxic stress with consequent depression of intracellular enzyme mechanisms. A feature of the process of P.E. depletion of red cell membranes is the progressive loss of plasticity of the cell with increased rigidity of the cell membrane. (24) Such cells are susceptible to removal by the spleen due to their failure to negotiate the splenic sinusoids (25) and tranverse its barriers due to their loss of plasticity. The same is true for all circulating cells, including platelets, with damaged membranes.
355
High serum levels of phospholipid will cause 'acanthocyte' formation due to progressive loss of red cell membrane cholesterol formation (in rats).(26) The elevation of serum lipids in the HUS is similar to that seen in Type II, Hyperlipoproteinemia in which there is an increased tendency to platelet aggregation. (27) Aggregation has been shown to alter the lipid configuration of platelets (28) and therefore presumably make them more susceptible to splenic sequestration if they do not partake of clot formation. Animal correlates of the disease. Wolman noted the similarity of certain animal models of renal cortical necrosis and 'eclampsia' and the HUS. A fatal 'eclampsia' type syndrome (29) may be produced in pregnant rats by feeding them a high fat diet deficient in vitamin E, or by feeding oxidized cod liver oil. (30) Damage is extensive with lesions in the placenta, spleen, and kidney.(31) Red blood cells show an increased perocide fragility an indicator of red cell membrane susceptibility to peroxidation. (31) This is usually seen in states associated with decreased serum tocopherol levels. Rats produce a similar lesion when fed a choline deficient diet. (32) This disease may be prevented by antioxident administration. Rats fed oxidized polyunsaturated fatty acids develop a similar symptom complex with depression of phospholipid content of the kidney and other organs. (33) The renal lesion in the rat model bears many similarities to that seen in the HUS with glomerular endothelial cell swelling with fibrin thrombi in the capillaries and arterioles. (31) Fibrin is also demonstrated in the arterioles in both. (31) Marked lipid deposition in the kidneys is a feature of both (31,34) as is depression of renal phospholipid content. (31) Platelet sequestration takes place in the kidney in the eclamptic rat. (31) The same process may occur in the kidney in the early states of the HUS.(35) Katz -et al. have demonstrated platelet sequestration in the spleen in the established phase of the disease. (35) CONCLUSION Proposed mechanism. The following cascade of events is proposed to explain the development of the HUS. Infection (viral, bacterial, vaccination) oral contraceptive therapy or renal transplantation produce marked alterations in lipid metabolism. A patient with an inborn defect in intracellular antioxident capacity may not be able to cope with a continuous endogenous peroxidized lipid challenge should the extracellular antioxident mechanism, i.e., tocopherol become depleted. Massive exposure to such peroxidizing
356
lipids would first occur to circulating red blood cells, platelets, and white cells. With failure of intracellular antioxident mechanisms P.E. depletion would occur in red cell membranes with subsequent haemolysis, especially in older cells, or loss of cell plasticity and increase in membrane rigidity. Cholesterol depletion secondary to increased serum phospholipid levels would occur with consequent acanthrocyte (or schistocyte) formation. Due to severe rapid haemolysis of the older cells who already have P.E. depletion associated with age, a marked drop in hematocrit would occur with coagulation secondary to hemolysis. The high glomerular perfusion pressure (36) combined with fibrin formation would result in very bizarre shaped erythrocytes due to mechanical trauma sustained in traversing damaged capillaries and vessels. (38) While thrombocytopenia might be accounted for through sequestration, platelet aggregation may be expected to play a role. It has been reported that the addition of exogenous Hz02 induces platelets to aggregate. (39) Furthermore this Hz02 effect may be abolished by the addition of Vitamin E. Further stimulation to induce platelet aggregation might be provided through damaged endothelial cells that release factor VIII and expose their basement membranes. (40)
357
Peroxidation
Antioxident Vitamin E
LIPID PEROXIDATION IN THE HEMOLYTIC UREMIC SYNDROME S. O'Regan* and J.S.C. Fong. The Department of Nephrology, University of Montreal, Hbpital Ste-Justine - Institute of Research*, 3175 CGte Ste-Catherine Road, Montreal H3T lC5, Quebec, Canada, Department of Nephrology, McGill University, Montreal Childrens Hospital Research Institute, Montreal, P.Q. Canada. ABSTRACT Based on recent evidence of a genetic influence on prognosis (1) and the existence of red cell membrane phospholipid depletion with low or absent serum a-tocopherol (2) levels in three children with the Hemolytic Uremic Syndrome (H.U.S.), we wish to suggest the existence of an inborn error of antioxident capacity as the basic pathogenetic mechanism in the development of the hemolytic uremic syndrome (H.U.S.). Key Words:
Hemolytic Uremic Syndrome, Lipid Metabolism
353
INTRODUCTION Since the initial description of the HUS by Gasser et al., (3) this symptom complex has been reported with increasing frequency. (4,5) Marked variations in the geographical, (6) age, (7) and racial (6) distribution must be reconciled in any attempts to propose a pathogenetic mechanism. A unifying concept is required to explain its association with such diverse conditions as viral (overt (8) and vaccination (9), bacterial infection, (10) oral contraceptive therapy, (11) and post renal transplantation. (12) A role for injested peroxidized lipids has previously been proposed by Wolman. (13) This however would be expected to produce larger numbers of cases in clusters, i.e., whole families. We wish to extend the peroxidized lipid hypothesis and propose an endogenous source of lipid precipitating a pathologic cascade to account for the major features of the disease, i.e., haemolytic anemia with schizocytosis, thrombocytopenia, and renal failure. We recently reported (2) the association of depressed c-tocopherol activity and red cell membrane phospholipid (phosphatidyl ethanolamine) depletion in the HUS in 3 children. One parent with 3 affected children also had an abnormal erythrocyte membrane phosphdlipid pattern associated with undetectable tocopherol activity in her serum. A marked genetic influence on prognosis has been noted from examination of the outcome of the disease in 25 sibships. (1) These two features help substantiate our proposal of the existence of an inborn error in lipid metabolism, i.e., decreased antioxident capacity of intracellular enzyme systems normally active in maintaining the integrity of the cell membrane. Marked alterations in lipid metabolism are a common feature of gram negative septicemia. (14) Disturbances in lipid metabolism have also been reported in association with oral contraceptive therapy (15) and renal transplantations. (16) Marked rises in serum triglyceride and cholesterol occur in all these conditions as in the HUS. (17) Vit. E as an Antioxident In the absence of severe protein calorie malnutrition or fat malabsorption deficiency of u-tocopherol is exceedingly rare, only one condition to date demonstrating depressed serum levels because of increased utilization. (18) The major role of vitamin E is considered to be that of an antioxident. It acts as a free radical scavenger (20) and thus prevents the peroxidation of polyunsaturated fatty acids. It may also be active in the synthesis of heme containing proteins active in the scavenging of peroxides (21). Thus an important action of this compound is in the maintenance of cellular and intracellular membrane integrity by possible interactions with polyunsaturated lipids in membranes. (22) This importance of a-tocopherol for membrance stability has been studied on erythrocytes of rats fed on vitamin E deficient diet. (23) The primary
354
defect noted in the membrane lipid was an almost complete depletion of phosphatidyl-ethenolamine (P.E.), demonstrated following in vitro exposure of cells to H202. (Peroxide fragility test). Reappearance of P.E. was dramatic when vitamin E sufficiency was maintained. An additional finding was the inability of the mature erythrocyte to repair losses of lipid components of the cell membranes. Thus the effect of the antioxident capacity of vitamin E playing an in vivo role in protecting against red cell membrane lipid degradation was to slow the rate of peroxidation and not to stimulate regeneration of lipids in the mature cell. Evolution of the Syndrome. When a free radical-derived from the breakdown of H202, or any other source, (e.g. peroxidized lipid), reaches a lipid double bond in the presence of 02 unstable lipoperoxides are produced. These can then spontaneously degenerate to form more free radicals which attack other double bonds, to produce more lipoperoxides. This peroxide challenge without effective antioxident capacity may propagate an autocatalytic chain reaction. The process might be accelerated by a state in which rapid production of more peroxidized lipid is continuing from endogenous sources and is being released into the circulation. Such a release phenomenon would have its initial and major effects on membranes of circulating cells, i.e., RBC's, platelets - they being rapidly subjected to massive exposure to peroxidized lipids. Due to the necessity of diffusion from the circulation, etc., - effects would be slower on "fixed" tissues. The susceptibility to massive membrane peroxidation would depend on: a.
Rapid utilization of available tocopherol due to continuous release of peroxidizing lipids with ultimate saturation of antioxident (extracellular) capacity.
b.
Failure of intracellular mechanisms of antioxidation after complete utilization of extracellular capacity.
Other involved tissues would also demonstrate effects of peroxidation, i.e., phospholipid content, especially if subject to hypoxic stress with consequent depression of intracellular enzyme mechanisms. A feature of the process of P.E. depletion of red cell membranes is the progressive loss of plasticity of the cell with increased rigidity of the cell membrane. (24) Such cells are susceptible to removal by the spleen due to their failure to negotiate the splenic sinusoids (25) and tranverse its barriers due to their loss of plasticity. The same is true for all circulating cells, including platelets, with damaged membranes.
355
High serum levels of phospholipid will cause 'acanthocyte' formation due to progressive loss of red cell membrane cholesterol formation (in rats).(26) The elevation of serum lipids in the HUS is similar to that seen in Type II, Hyperlipoproteinemia in which there is an increased tendency to platelet aggregation. (27) Aggregation has been shown to alter the lipid configuration of platelets (28) and therefore presumably make them more susceptible to splenic sequestration if they do not partake of clot formation. Animal correlates of the disease. Wolman noted the similarity of certain animal models of renal cortical necrosis and 'eclampsia' and the HUS. A fatal 'eclampsia' type syndrome (29) may be produced in pregnant rats by feeding them a high fat diet deficient in vitamin E, or by feeding oxidized cod liver oil. (30) Damage is extensive with lesions in the placenta, spleen, and kidney.(31) Red blood cells show an increased perocide fragility an indicator of red cell membrane susceptibility to peroxidation. (31) This is usually seen in states associated with decreased serum tocopherol levels. Rats produce a similar lesion when fed a choline deficient diet. (32) This disease may be prevented by antioxident administration. Rats fed oxidized polyunsaturated fatty acids develop a similar symptom complex with depression of phospholipid content of the kidney and other organs. (33) The renal lesion in the rat model bears many similarities to that seen in the HUS with glomerular endothelial cell swelling with fibrin thrombi in the capillaries and arterioles. (31) Fibrin is also demonstrated in the arterioles in both. (31) Marked lipid deposition in the kidneys is a feature of both (31,34) as is depression of renal phospholipid content. (31) Platelet sequestration takes place in the kidney in the eclamptic rat. (31) The same process may occur in the kidney in the early states of the HUS.(35) Katz -et al. have demonstrated platelet sequestration in the spleen in the established phase of the disease. (35) CONCLUSION Proposed mechanism. The following cascade of events is proposed to explain the development of the HUS. Infection (viral, bacterial, vaccination) oral contraceptive therapy or renal transplantation produce marked alterations in lipid metabolism. A patient with an inborn defect in intracellular antioxident capacity may not be able to cope with a continuous endogenous peroxidized lipid challenge should the extracellular antioxident mechanism, i.e., tocopherol become depleted. Massive exposure to such peroxidizing
356
lipids would first occur to circulating red blood cells, platelets, and white cells. With failure of intracellular antioxident mechanisms P.E. depletion would occur in red cell membranes with subsequent haemolysis, especially in older cells, or loss of cell plasticity and increase in membrane rigidity. Cholesterol depletion secondary to increased serum phospholipid levels would occur with consequent acanthrocyte (or schistocyte) formation. Due to severe rapid haemolysis of the older cells who already have P.E. depletion associated with age, a marked drop in hematocrit would occur with coagulation secondary to hemolysis. The high glomerular perfusion pressure (36) combined with fibrin formation would result in very bizarre shaped erythrocytes due to mechanical trauma sustained in traversing damaged capillaries and vessels. (38) While thrombocytopenia might be accounted for through sequestration, platelet aggregation may be expected to play a role. It has been reported that the addition of exogenous Hz02 induces platelets to aggregate. (39) Furthermore this Hz02 effect may be abolished by the addition of Vitamin E. Further stimulation to induce platelet aggregation might be provided through damaged endothelial cells that release factor VIII and expose their basement membranes. (40)
357
Peroxidation
Antioxident Vitamin E
