Overview

Pyruvate kinase deficiency (PKD) is an extremely rare inherited metabolic disorder due to mutations in the PKLR gene, which result in a deficiency of pyruvate kinase, an enzyme that produces adenosine triphosphate (ATP) in red blood cells (RBCs). Since mature RBCs rely on glycolysis only to obtain energy due to the lack of mitochondria, impaired ATP means dehydration, stiffness, and increased destruction in the spleen, resulting in haemolytic anaemia and chronic haemolysis. This causes jaundice, pallor, weakness, splenomegaly, and gallstone predisposition due to excess bilirubin. For ongoing RBC breakdown, erythropoiesis is increased in the bone marrow in compensation, elevating the demand for folic acid needed for DNA synthesis and RBC formation. In the lack of sufficient folic acid, defective maturation of erythroid cells can lead to megaloblastic anaemia, which is an additional complication of the already compromised RBC formation in PKD. Therefore, having proper levels of folic acid is essential to promote erythropoiesis and neutralise the effects of chronic haemolysis in PKD patients.¹

Pyruvate kinase deficiency and erythropoiesis

Pathophysiology of PKD

Disturbance of energy metabolism in RBCs

PKD results in impaired ATP production, which impacts several ATP-dependent processes crucial for RBC stability and function:²

• Ion homeostasis and RBC dehydration: ATP energises ion transporters such as Na⁺/K⁺-ATPase and Ca²⁺-ATPase, which regulate intracellular ion concentrations. Loss of ATP causes disturbed ion homeostasis, resulting in RBC dehydration, shrinkage, and stiffness. Dehydrated RBCs are prone to premature destruction in the spleen³
• Membrane structure and elasticity: ATP is needed to sustain cytoskeletal proteins (actin and spectrin) for the deformability of RBC. In the absence of ATP, RBCs become unable to retain their biconcave disc shape, becoming fragile and incapable of passing through microvasculature, and hence cause splenic sequestration and haemolysis⁴
• Resistance to oxidative stress: RBCs are continually subjected to oxidative injury by reactive oxygen species (ROS). Normally, glutathione (GSH), the main antioxidant, is reduced at the expense of ATP and thus conserved. In PKD, reduced levels of ATP cause impaired antioxidant defence, hence exposing RBCs to severe oxidative damage, speeding up their destruction even more⁵

Premature red cell destruction and anaemia

As RBCs lose water, they become rigid and prone to oxidative stress, causing them to undergo chronic extravascular haemolysis, where macrophages in the spleen lyse them prematurely. This leads to:⁶

• Haemolytic anaemia: Reduced RBC lifespan causes chronic anaemia, impairing oxygen delivery to tissues
• Jaundice and hyperbilirubinemia: Excess bilirubin from RBC breakdown accumulates in the blood, causing yellow discolouration of the skin and eyes
• Splenomegaly: The spleen enlarges due to increased RBC clearance
• Gallstone formation: Predisposition to pigment gallstones due to increased bilirubin results in cholecystitis and biliary disease
• Chronic fatigue and weakness: Decreased oxygen-carrying capacity results in chronic fatigue, pallor, and intolerance to exercise

Compensatory erythropoiesis in response to haemolysis

To offset the ongoing haemolysis, the body accelerates RBC production through compensatory erythropoiesis, under the control of erythropoietin (EPO). The resultant increased erythropoiesis is very expensive in terms of nutrients needed, particularly folic acid, to sustain productive red cell manufacture.⁷

Bone marrow response to haemolysis⁷

• The kidneys release additional EPO in reaction to chronic RBC destruction, stimulating the bone marrow to elevate erythropoiesis
• The number of erythroid precursors rises to compensate for the continuing loss of RBCs, requiring increased input of required nutrients
• Iron and folic acid are needed for DNA synthesis, cell division, and normal erythroid progenitor maturation. Under folate deficiency, the bone marrow is unable to sustain increased RBC production, leading to ineffective erythropoiesis and exacerbation of anaemia

Increased reticulocyte production and folic acid utilisation⁷

• To replace lost RBCs, the bone marrow releases reticulocytes (young RBCs) into the circulation. In PKD, reticulocytosis is an associated compensatory change
• Due to their active proliferation and differentiation, reticulocytes also have a greater metabolic demand for folic acid to serve as a cofactor for DNA synthesis, nucleotide synthesis, and RNA metabolism
• Folic acid deficiency may lead to megaloblastic change in erythroid precursors, leading to impaired RBC production and aggravation of anaemia

Clinical significance

With constant RBC turnover and reticulocytosis in PKD, it is important to maintain adequate folic acid to:

• Prevent megaloblastic anaemia, which can result from folate deficiency
• Permit normal erythropoiesis, which can maintain continuous replacement of RBCs
• Improve clinical outcomes, reducing symptoms such as fatigue and worsening anaemia

Role of folic acid in supporting erythropoiesis

Folic acid plays a crucial role in the upkeep of erythropoiesis, particularly in conditions like PKD, where RBCs are continuously being destroyed. Folic acid as a coenzyme for nucleotide biosynthesis guarantees the production of thymidine, a critical precursor of DNA synthesis, ensuring proper DNA replication and erythroid precursor cell proliferation within the bone marrow for effective RBC maturation. In the absence of sufficient folic acid, megaloblastic transformation occurs in the erythroid precursors, which yields big, functionless RBCs and increases anaemia. Moreover, given the accelerated RBC turnover in PKD, folic acid has a key function in generating rapid red cells through sustaining new RBC formation and thus preventing anaemia related to folate insufficiency.⁸

Clinical implications of folic acid supplementation in PKD

Prevention of folate deficiency and megaloblastic anaemia

In PKD, chronic haemolysis of RBCs leads to excessive erythropoiesis and a high demand for folic acid to support the production of new RBCs. Without adequate folate, a risk of folate deficiency and megaloblastic anaemia exists, with erythroid precursors in the bone marrow failing to mature, resulting in the production of large, ineffective RBCs. This worsens the inherent anaemia of PKD patients. Folic acid supplementation guarantees the bone marrow sufficient resources to manufacture RBCs under normal circumstances, preventing the complications of folate deficiency and maximising erythropoiesis.²

Dietary intake and supplementation guidelines

The proposed dietary folic acid requirement for patients with PKD may be dependent on age, sex, and severity of disease. The general recommended daily allowance for the population is approximately 400 micrograms per day, and in states of increased cell turnover, such as in PKD. Supplementation with folic acid is necessary in PKD patients to meet the increased demand. Folic acid supplementation is typically given in the form of over-the-counter vitamin B9 supplements, and the dosage is typically 1 to 5 milligrams daily, depending on the severity of anaemia and the degree of folate deficiency. Dosage will be decided by medical professionals according to clinical symptoms, laboratory findings, and response to treatment.

Evidence from clinical studies and case reports

Clinical reports and case studies consistently suggest folic acid supplement benefits for patients with PKD.⁹ Folate supplementation has proven to lead to red blood cell production normalisation and prevention of megaloblastic anaemia, thus improving haematologic function in individuals with PKD. In one of the clinical trials of PKD patients, folic acid-supplemented patients were found to have augmented RBC counts, reduced reticulocyte counts, and reduced incidence of megaloblastic abnormalities compared to control patients not supplemented with folic acid. These findings attest to the role of folic acid in the management of PKD and the beneficial effect of preventive supplementation on such patients' clinical outcomes.

Summary

Folic acid plays a crucial role in supporting erythropoiesis in individuals with PKD by aiding DNA synthesis, preventing megaloblastic anaemia, and supporting the rapid turnover of RBCs due to chronic haemolysis. Clinical recommendations emphasise the importance of folic acid supplementation to prevent folate deficiency and support optimal red blood cell production in PKD patients. Given the critical role of folate in managing PKD-related anaemia, future research should focus on optimising nutritional interventions, exploring personalised dosing regimens, and investigating the potential of other micronutrients in managing haemolytic anaemias to enhance patient outcomes and better address the underlying mechanisms of RBC destruction in PKD.