Objective: Megaloblastosis (i.e., megaloblastic transformation of erythroid precursor cells in the bone marrow) is the cytomorphological hallmark of megaloblastic anemia resulting from vitamin B12 and folate deficiency. It is characterized by a finely stippled lacy pattern of nuclear chromatin, which is believed to be an expression of deranged cellular DNA synthesis. However, the molecular basis of these cytomorphological aberrations still remains obscure. The current presentation describes the results of our studies on some molecular events associated with the development of megaloblastosis. Methods: Transmission electron microscopy was used to study megaloblasts as well as DNA fibers extracted from megaloblastic and normoblastic bone marrows with and without treatment with proteinase K during the extraction procedure; cellular DNA synthesis in bone marrow cultures was studied by incorporation of 3H-thymidine and deoxyuridine suppression test, while histone biosynthesis in bone marrow cells was studied by in vitro incorporation of 3H-tryptophan, 3H-lysine and 3H-arginine into histones. Results: Derangement of DNA synthesis occurred due to an impaired de novo pathway of thymidylate synthesis in both vitamin-B12- and folate-deficient human megaloblastic bone marrows as well as in the bone marrows of rhesus monkeys and rats with experimentally induced folate deficiency. Interestingly, folate-deficient monkeys developed frank megaloblastic bone marrows, but folate-deficient rats did not. On the other hand, megaloblastic changes in the bone marrow of human patients with myelodysplastic syndrome and erythroleukemia were not associated with this DNA synthetic abnormality. Biosynthesis of predominantly arginine-rich histones in megaloblastic bone marrows was markedly reduced as compared to normoblastic bone marrows, which was consistently associated with elongation and despiralization of chromosomes and finely stippled nuclear chromatin in megaloblasts. Conclusion: The impaired biosynthesis of predominantly arginine-rich nuclear histones appeared to be a common molecular event (a denominator) underlying the development of megaloblastosis with or without abnormal DNA synthesis.
© 2005 S. Karger AG, Basel
- Electron microscopy
- Chromosomal despiralization
- 3H-thymidine incorporation
- DNA synthesis
- Deoxyuridine suppression test
- Histone biosynthesis
- Chanarin I: The Megaloblastic Anaemia, ed 3. Oxford, Blackwell Scientific Publications, 1990, pp 315–331.
- Wills L: Treatment of pernicious anaemia of pregnancy and ‘tropical anaemia’ with special reference to yeast extract as a curative agent. Br Med J 1931;1:1059–1064.
- Wills L: Treatment of pernicious anaemia, nutritional macrocytic anaemia and tropical sprue. Blood 1948;3:36–56.
- Chatterjea JB: Nutritional megaloblastic anaemia in tropical zones. Proc 11th Congr Int Soc Hematol, Sidney, 1966, pp 120–132.
- Das KC: Nutritional megaloblastic anaemia. Proc Symp Nutr Anemias, Plenary Sess 2, Meet Asian Pacific Div, Int Soc Hematol, Melbourne, 1971, pp 11–13.
- Herbert V: The five possible causes of all nutrient deficiency. Aust NZ J Med 1972;1:69–72.
- Das KC, Herbert V: Vitamin B12-folate interrelations. Clin Haematol 1976;5:997–725.
- Herbert V: The Megaloblastic Anemias, ed 1. New York, Grune and Stratton, 1959, p 1.
- Lee RG: Megaloblastic anemias: Disorders of impaired DNA synthesis; in Lee RG, Foester J, Lukens J, Parakevas JP, Rodgers GM (eds): Wintrobe’s Clinical Hematology, ed 10. Baltimore, Williams & Wilkins, 1999, pp 941–978.
- Das KC, Hoffbrand AV: Lymphocyte transformation in megaloblastic anaemias. Morphology and DNA synthesis. Br J Haematol 1970;19:459–468.
- Das KC, Herbert V: The lymphocyte as a marker of past nutritional status. Persistence of abnormal lymphocyte deoxyuridine (dU) suppression test and chromosomes in patients with past deficiency of folate and vitamin B12. Br J Haematol 1978;38:219–223.
- Metz J, Kelly A, Swett VC, Waxman S, Herbert V: Deranged DNA synthesis by bone marrow from vitamin B12-deficient humans. Br J Haematol 1968;14:575–592.
- Mohanty D, Das KC: Effect of folate deficiency on the reproductive organs of female Rhesus monkeys: A cytomorphological and cytokinetic study. J Nutr 1982;112:1505–1576.
- Johnson EM, Nelson MM, Monie IN: The composition of folate-deficient diet for rats. Anat Rec 1963;146:215.
- Dacie JV, Lewis SM: Practical Haematology, ed 8. London, Churchill Livingstone, 1995, pp 49–143.
- Das KC, Herbert V: In vitro DNA synthesis by megaloblastic bone marrow: Effect of folates and cobalamins on thymidine incorporation and de novo thymidylate synthesis. Am J Hematol 1989;31:11–20.
- Das KC, Manusselis C, Herbert V: In vitro DNA synthesis by bone marrow cells and PHA-stimulated lymphocytes. Suppression by non radioactive thymidine of the incorporation of 3H-deoxyuridine into DNA: Enhancement of incorporation when inadequate vitamin B12 and folate is corrected. Br J Haematol 1980;44:51–63.
- Das KC, Garewal G, Mohanty D: Derangement of DNA synthesis in erythroleukemia. Normal deoxyuridine suppression and impaired thymidine incorporation in bone marrow culture. Acta Haematol 1980;64:121–134.
- Das KC, Mohanty D, Garewal G: Cytogenetics in nutritional megaloblastic anaemia: Prolonged persistence of chromosomal abnormalities in lymphocytes after remission. Acta Haematol 1986;76:146–154.
- Sambrook J, Fritsch EF, Maniatis J: Molecular Cloning: A Laboratory Manual. New York, Cold Spring Harbor Laboratory Press, 1989, pp 1–25.
- Murray K: The acid extraction of histones from calf thymus deoxyribonucleoprotein. J Mol Biol 1966;15:409–419.
- Das KC, Kaufman RP, Gay H: Autoradiographic evidence of synthesis of an arginine-rich histone during spermatogenesis in Drosophila melanogaster. Nature 1964;204:1008–1009.
- Tidwell T, Allfrey VF, Mirsky AE: The methylation of histone during regeneration of the liver. J Biol Chem 1968;243:707–715.
- Gershey FL, Halett GW, Vidali G, Allfrey VG: Chemical studies on histone methylation. Evidence for the occurrence of 3-methyl-histidine in avian erythrocyte histone fraction. J Biol Chem 1969;244:4871–4877.
- Longo DL, Herbert V: Radioassay for serum and red cell folate. J Lab Clin Med 1976;87:138–151.
- Das KC, Manusselis, Herbert V: Determination of vitamin B12 (cobalamin) in serum and erythrocytes by radioassay and of holo-transcobalamin II ( holo-TC-II) and holo-haptocorrin (holo-TC1 and III) in serum by adsorbing holo-TC-II on microfine silica. J Nutr Biochem 1991;2:455–463.
- Kiossoglou, KA, Mitus WJ, Dameshek W: Chromosomal aberrations in pernicious anemia. Study of three cases before and after therapy. Blood 1965;5:662–682.
- Heath CW: Cytogenetic observations in vitamin B12 and folate deficiency. Blood 1966;27:800–815.
- Das KC: Nutritional megaloblastosis. Symp Nutr Anaemias. Proc Second Congr Asian Pacific Div, Int Soc Haematol, New Delhi, 1976, pp 14–15.
- Hoffbrand AV, Pegg AE: Base composition of normal and megaloblastic bone marrow DNA. Nat New Biol 1972;235:187–188.
- Khobta A, Carlo-Stella C, Capranico G: Specific histone patterns and acetylase/deacetylase activity at the breakpoint-cluster region of the human MLL gene. Cancer Res 2004;64:2656–2662.
- Zhao J: Coordination of DNA synthesis and histone gene expression during normal cell cycle progression and after DNA damage. Cell Cycle 2004;3:695–697.
- Norbury CJ, Zhivolovsky B: DNA damage- induced apoptosis. Oncogene 2004;23:2797–2808.
- Wang C, Fu M, Pestell RG: Histone acetylation/deacetylation as a regulator of cell cycle gene expression. Methods Mol Biol 2004;241:207–216.
- McLaughlin F, La Thangue NB: Histone deacetylase inhibitors open new doors in cancer therapy. Biochem Pharmacol 2004;68:1139–1144.
- Ingram CF, Davidoff AN, Marais F, Sherman GG, Mendelow BV: Evaluation of DNA analysis for evidence of apoptosis in megaloblastic anaemia. Br J Haematol 1997;96:576–583.
- Koury MJ, Price JO, Hicks GG: Apoptosis in megaloblastic anemia occurs during DNA synthesis p53-independent, nucleoside-reversible mechanism. Blood 2000;96:3249–3255.
- Gu L, Wu J, Qiu L, Jennings CD, Li GM: Involvement of DNA mismatch repair infolate deficiency-induced apoptosis small star, filled. J Nutr Biochem 2002;13:355–363.
- Li GM, Presnell SR, Gu I: Folate deficiency, mismatch repair-independent apoptosis and human disease. J Nutr Biochem 2003;14:568–575.
- Das KC, Easow SK, Jadaon M, Das M, Mohanty D: The development of megaloblastosis is a species-restricted phenomenon. 9th Annu Health Sci Poster Conf, Kuwait, April 19–21, 2004, p 284.
Dr. K.C. Das
Hematology Unit, Department of Pathology, Faculty of Medicine, Kuwait University
PO Box 24923, 13110 Safat (Kuwait)
Tel. +965 531 9476, Fax +965 533 8905
E-Mail firstname.lastname@example.org, email@example.com
Received: August 15, 2004
Revised: October 25, 2004
Published online: July 09, 2008
Number of Print Pages : 13
Number of Figures : 9, Number of Tables : 6, Number of References : 40
Medical Principles and Practice (International Journal of the Kuwait University Health Sciences Centre)
Vol. 14, No. Suppl. 1, Year 2005 (Cover Date: Released July 2005)
Journal Editor: Al Awadi, F. (Kuwait)
ISSN: 1011–7571 (Print), eISSN: 1423–0151 (Online)
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