Study of glycolytic intermediates in hereditary elliptocytosis with thalassemiaRoshan S Pavri1, AJ Baxi1, Shobha Grover2, RA Parande3
1 Blood Group Reference Centre, Seth G. S. Medical College, Parel, Bombay-400 012., India
2 Department of Pathology, Government Medical College, Nagpur., India
3 Clinical Investigation Laboratory, Mahal, Nagpur-2., India
Correspondence Address: Source of Support: None, Conflict of Interest: None PMID: 615267
Source of Support: None, Conflict of Interest: None
Glycolytic intermediates like ATP, DPG and GSH have been studied in a family with. hereditary elliptocytosis and thalassemia. Results indicate a fall in ATP with a concomitant rise in DPG in the Patient. Findings are discussed in relation to other data.
Hereditary elliptocytosis (HE) is a rare hematological disorder which sometimes occurs with other genetic abnormalities like sickle cell anemia, Hb C, beta thalassemia and others. ,,,,, A close linkage of Rhesus antigen with HE has been shown by a few investigators. ,, Biochemical investigations in HE suggest abnormalities in glycolytic metabolisms. ,, Marked GSH instability in two families of HE with and without hemolysis was observed by us earlier.  We present below biochemical changes observed in glycolytic (Embden-Meyerhof) and pentose phosphate pathways in a family of HE interacting with thalassemia trait.
Hematological investigations were carried out according to standard methods.  Adenosine triphosphate (ATP) was determined by double enzyme method.  2,3 -diphospho-glycerate was estimated by the modified chromotropic acid method of Bartlett.  Glucose-6-phosphate dehydrogenase (G-6-PD) and pyruvate kinase (PK) were assayed as described previously.  Glutathione (GSH) and GSH stability were determined by Beutler's procedure. , Hemoglobin electrophoresis including Hb A 2 was performed as described by Yunis.  Hemoglobin F was determined by the method of Singer. 
DK, a 23 year old Maharashtrian male, presented the following features on initial examination.
The patient (propositus) had recurrent episodes of mild jaundice, low grade fever and pain in the left hypochondrium usually lasting for 1-2 weeks during the last three years. He had two bouts of haematemesis during the last two months. Past history did not reveal any attack of jaundice or having taken drugs, alcohol etc. There was no history of joint swelling, leg ulcer or peptic ulcer in the past. His family consisted of three brothers and mother. Father died of lymphosarcoma at the age of 56 years. Mother was not available for investigation.
On examination, the patient was found to be of an average build with mild jaundice and moderate pallor. No sternal tenderness, glands or purpura was noticeable. Hepato-splenomegaly was present, liver being 5 cm firm and nontender and spleen 7 cm firm and palpable below the costal margin. Other systemic examination was normal.
Hematological examination : Hb. was 9.75 gm/ dl. total WBC was 10,000/µl; differential count: P-70, L-28, M-1 and E-1. Oval cells (elliptocytes) were greater than 50%; hypcchromia and macrocytosis were present; evidence of hemolysis and regeneration was also present; reticulocyte count was 8%; Hb F (Singer) was 5%; Hb A, was 4.3%. No abnormal Hb was demonstrated. Osmotic fragility was decreased. Heinz bodies were not seen; isopropyl test was negative; heat stability was 0.5%; serum bilirubin level was 1 mg.%; urobilinogen was present but bile was absent. Bone marrow presented hypoplastic-normoblastic picture. Iron staining was negative but tissue iron was abundantly present. Sideroblasts count was more than 60%. No ring sideroblast was seen. Roentgenologic examination was normal.
Diagnosis of HE with thalassemia trait was arrived at after the family study [Figure 1].
Hematological findings of the patient (DK) and other brothers are presented in [Table 1]. The patient and another brother (MK) showed both abnormal traits i.e. elliptocytosis and thalassemia while the third brother (KK) showed the presence of elliptocytosis. The remaining brother (MRK) was normal. The family tree showing inheritance of both the genetic traits is shown in [Figure 1]. Though the patient and one brother (MK) are both having double genetic disease, the incidence of hemolytic anemia was obtained only in the former.
Biochemical findings on ATP and DPG in the patient (DK) showed reduction and increase respectively. Levels of these two phosphorous intermediates were normal in other three brothers studied. G-6-PD level was moderately raised in all four brothers while PK activity was high (p < 0.05) in the patient only, but was diminished in other three brothers [Table 2].
Normal GSH with greater stability was demonstrable in all four brothers. Methemoglobin level was slightly raised in the patient.
Aberrent glycolytic metabolism in hereditary elliptocytosis has been shown by De Gruchy et al.  Cross sectional view of the anaerobic (E.M.) pathway and pentose phosphate pathway in HE-thalassemia appears to differ from HE alone,  or thalassemia alone.  De Gruchy et al,  have pointed out that the levels of ATP and DPG are normal in fresh cells but fall on incubation. These results differ from our own findings wherein we found reduction in ATP level by about 65% and increase in DPG by almost the same margin (65%) in the patient in fresh red cells.
The difference in results of the patient from his brother (MK) who also demonstrated both the genetic abnormalities raises the question as to what causes the loss of ATP with concomitant rise in DPG? Low level of ATP has been demonstrated in diverse types of hemolytic anemias by us previously.  High level of PK indicates that the glycolytic metabolism is accelerated in the patient but is normal in the other brother (MK). Thus, taking into account overall glycolytic changes, we feel that ATP utilization is of importance in hereditary elliptocytosis with thalassemia. Variable expression of the disease may be responsible for difference in susceptibility to hemolysis in the patient and his brother. Further studies will be needed to elucidate this point.
[Table 1], [Table 2]