Erythrocyte Pathophysiology of Riboflavin deficiency in rats

Pathophysiology of erythrocytes from riboflavin-deficient rats has been investigated. Riboflav in-deficiency was produced by feeding a riboflavin-deficient diet ad-libitum to weanling male Wistar specified pathogen-free, albino rats (40-SOg) housed individually in wire-bottomed cages. Characteristic signs of severe deficiency were produced in most animals,and included weight loss, hair discolouration and skin lesions. Biochemical status of riboflavin deficiency was assessed enzymatically by measuring the erythrocyte glutathione reductase activity coefficient (EGR-AC) and the association of this index with all other changes which occurred, was investigated. Control animals were fed the riboflavin-deficient diet plus 22mg of riboflavin/kg of diet. Pair-fed animals were given the average amount of food which was eaten by the deficient rats the previous day. The results reported in this thesis showed that riboflavin-deficiency is directly correlated with red blood cell fragility as measured by either in-vitro exposure of these cells to a H2O2-generating system and/or hypotonic saline solutions. The degree of haemolysis obtained was found to be negatively correlated with concentrations of reduced glutathione (GSH) of the riboflavin-·defiicient blood, It was found also that thyroid hormone prevented the in-vitro haemolysis of red cells from riboflavin-deficient rats expoecd to the above system and a preliminary experiment suggested that prior administration of thyroxine in-vivo had a similar effect on red cell integrity when tested subsequently irt-vitro. When erythrocytes from riboflavin-deficient rats were separated into fractions of different mean age, haemoglobin concentrations in the red cell fractions suuggested that there was a progressive reduction in the number of young cells as the severity of the riboflavin-deficiency increased and a corresponding increase in the proportion of old cells. There was also a progressive induction in the number of reticulocytes and plasma iron and an increase in the non-haem and ferritin iron in liver tissue as biochemical riboflavin-deficiency increased. Red cells from the deficient animals were significantly more fragile in all fractions than the comparable fractions from control animals. Other changes which accompanied riboflavin-deficiency were increases in the concentration of erythrocyte peroxides, methaemoglobin and the activities of glutathione peroxidase, NADH-methaemoglobin reductase and NADPH-methacmoglobin reductase. Riboflavin-deficiency causes a reduction in the activity of glutathione reductase and it is suggested that the increased fragility of red cells from riboflavin-deficient rats may be a consequence of the inability to maintain concentrations of GSH. The reduced concentrations of GSH may also be inadequate to supply glutathione peroxidase with adequate substrate causing increased concentrations of peroxides to accumulate and possibly exert damaging effects on lipid structures, for example, cell membranes. The increase in erythrocyte methaemoglobin may be further evidence of the defence mechanisms against oxidant moieties and the increase in methaeomoglobin reductase and glutathione peroxidase, compensatory effects to counteract the oxidant stress. Many of the changes found in erythrocytes of riboflavin-deficient rats occurred also, to a variable degree, in erythrocytes front iodine-deficient rats. In addition it was confirmed that concentration of plasma thyroxine fell in blood from riboflavin-deficient rats. The interaction of thyroid status and riboflavin-deficiency is causing the changes found in erythrocytes is discussed.