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Original Article
Red Cell Pyruvate Kinase Deficiency in Neonatal Jaundice Cases in India Prabhakar S. Kedar, Prashant Warang, Roshan B. Colah and Dipika Mohanty Institute of Immunohaematology, Indian Council of Medical Research, K.E.M. Hospital Campus, Parel, Mumbai, India
ABSTRACT Objective. Pyruvate Kinase (PK) deficiency is the most common enzymopathy of the glycolytic pathway in erythrocytes. It constitutes one of the common causes of hereditary non-spherocytic hemolytic anemia. The aim of this study was to screen newborns in India for pyruvate kinase (PK) deficiency in relation to unconjugated hyperbilirubinemia. Methods. Laboratory investigations done included complete blood counts, reticulocyte counts, direct and indirect bilirubin, assay of G6PD and PK activity, ATP and 2,3 DPG levels. All variables were studied in 50-cord blood samples from normal deliveries and 218 neonates with hyperbilirubinemia. Results. 7 of the 218 cases of neonatal jaundice were PK deficient with 30-40 % reduction in PK activity. These cases also had a 3-4-fold increase in 2,3 DPG:ATP ratios, which is one of the additional indicators for PK deficiency. Six of the 7 infants had a severe clinical course. Conclusion. This study shows that the prevalence of PK deficiency in Indian neonatal jaundice cases is 3.21%, which is relatively high. This emphasizes the need for screening neonatal hyperbilirubinemia cases in India for PK deficiency. [Indian J Pediatr 2006; 73(11) : 985-988] E-mail :
[email protected]
Key words : Pyruvate Kinase deficiency; Hereditary non-spherocytic hemolytic anemia; Neonatal jaundice; Hyperbilirubinemia
Red cell pyruvate kinase (PK.EC.2.7.1.40) deficiency is the most common enzyme abnormality in the Embden Meyerhoff pathway of glycolysis in humans. PK deficiency is associated with hereditary non-spherocytic hemolytic anemia (HNSHA) and is transmitted as an autosomal recessive disorder, with both the sexes being equally affected. The clinical severity of this disorder varies widely, ranging from a mildly compensated anemia to severe anemia of childhood, though in some cases it does not manifest itself until adulthood. Affected newborns usually present with unconjugated hyperbilirubinemia and may require exchange transfusion. PK deficiency was first described by Valentine et al in 1961 and since then, approximately 450 cases have been described in the literature1-2. Most of these are of North European origin while occasional cases have been reported from the Middle East, Japan, China, Spain and Saudi Arabia.3-6
Correspondence and Reprint requests : Dr. Roshan B. Colah, Ph.D., Deputy Director, Institute of Immunohaematology (Indian Council of Medical Research), 13th Floor, New Multistoried Building, K.E.M. Hospital, Parel, Mumbai-400012 (India); Fax: 9122-24138521
Indian Journal of Pediatrics, Volume 73—November, 2006
The common causes for inherited hemolytic anemia in the Indian Subcontinent include thalassemia, sickle cell anemia, other hemoglobinopathies and glucose-6 phosphate dehydrogenase deficiency (G6PD). The distribution of these hereditary defects in this population have been described earlier. 7 There is no data on the prevalence of PK deficiency in the Indian population. This study was thus undertaken to determine the incidence of PK deficiency in Indian newborns presenting with neonatal jaundice and to evaluate the severity of the disease in these cases. MATERIALS AND METHODS 50-cord blood samples from clinically and hematologically normal deliveries were collected in EDTA and analyzed within 24 hr (Normal controls). 218 neonates (103 males and 115 females) presenting with unconjugated hyperbilirubinemia were included in this study. Hematological indices were measured on an automated cell counter. (Sysmex- K-1000, Japan) Peripheral smear examination and reticulocyte counts were done using standard techniques. Direct and indirect 985
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Prabhakar S. Kedar et al antiglobulin test was done to exclude immune causes of hemolysis. Total and indirect bilirubin levels were determined by standard methods. Depletion of reticulocytes was done by differential centrifugation and confirmation was done by reticulocyte staining with new methylene blue.8 The red cells were prepared for enzyme assay by passing the whole blood through a column containing a mixture of α-cellulose and microcrystalline cellulose (Sigma, USA) to remove leucocytes.9 Glucose-6 phosphate dehydrogenase and pyruvate kinase activities were assayed by the method described by Beutler.10 2,3 Diphospho glycerate (2-3 DPG) level was determined by a modification of the chromotropic acid method of Bartlett as described by Eaton.11 ATP was estimated on perchloric acid extracts using the G6PD: Hexokinase double enzyme method.12
hematologically normal deliveries and it ranged from 7.4 - 12.5 IU/g Hb. This is lower than the adult levels studied by us earlier (12.5- 17.5 IU/g Hb). 7 of the 218 cases of neonatal jaundice were PK deficient with a 30-40 % reduction in PK activity. The prevalence of PK deficiency in newborns with hyperbilirubinemia was thus 3.21 %. The hematological and biochemical data of the PK deficient cases with neonatal jaundice at presentation and follow-up are presented in table 1 and 2. All these cases presented with unconjugated hyperbilirubinemia with reticulocytosis in the first 2 weeks of postnatal life. All of them needed phototherapy and three of them required exchange blood transfusions. (Table 1). All the cases were followed up between one and nine-months of age to reconfirm PK deficiency. Their hemoglobin levels on follow-up ranged from 4.3 to 11.0 g/dl and they had marked reticulocytosis (6 to 30%) (Table 2). Other causes of intracorpuscular hemolysis like presence of unstable hemoglobins, thalassemia, hereditary spherocytosis and G6PD deficiency were ruled out. The presence of auto antibodies and cold antibodies were also excluded. These cases also had a 3-4-fold increase in 2-3 DPG:ATP ratios,
RESULTS The normal PK activity in newborns was established using 50-cord blood samples from clinically and
TABLE 1. Hematological and Biochemical Parameters in 7 PK Deficient Cases at First Presentation Parameter
Case 1
Case 2
Case 3
Case 4
Case 5
Case 6
Case 7
Caste Origin Age (days) Sex Hb (g/dl) Retic (%) RBC 106 X mm3 MCV (fl) MCH (pg) MCHC (%) Exchange Transfusion Total bilirubin (mg/dl) Indirect Bilirubin (mg/dl) Pyruvate Kinase (IU/g Hb)
Koli MH 7 Male 14.0 7.0 3.98 107.8 35.2 32.6 Yes 21.4 20.6 4.24
Patel GJ 9 Female 15.9 9.0 4.31 113.0 36.9 32.6 No 9.9 9.2 4.34
Hindu GJ 9 Male 14.7 3.0 4.00 111.5 36.7 33.0 No 13.4 11.8 4.92
Muslim Khan UP 4 Male 13.4 14.0 3.76 105.8 35.0 32.0 Yes 18.6 16.2 3.54
Gupta UP 7 Female 15.8 17.1 4.32 110.6 36.5 32.4 No 9.8 7.8 4.55
Patel GJ 2 Male 12.2 5.0 3.82 109.5 36.0 31.8 Yes 19.8 17.2 4.53
Maratha MH 8 Male 13.9 9.0 3.84 108.3 36.2 32.4 No 3.2 2.3 5.12
MH: Maharashtra, GJ: Gujrat, UP: Utter Pradesh TABLE 2. Hematological and Biochemical Data in PK Deficient Cases at Follow Up. Parameter
Age (Months) on Follow-up Hb (g/dl) Retic (%) Blood Transfusion (Units) Total bilirubin (mg/dl) Indirect Bilirubin (mg/dl) Pyruvate Kinase (IU/g Hb) G6PD (IU/mlRBC/min) 2-3DPG (µmoles/gHb) ATP (µmoles/gHb) 2-3 DPG: ATP Ratio
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50 Normal Newborns Mean +SD 19.13+3.4 < 2.0%
