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Decrease in Apoptosis and Increase in Polyploidization of Megakaryocytes by Stem Cell Factor During Ex Vivo Expansion of Human Cord Blood CD34⁺ Cells Using Thrombopoietin

^a Department of Pathology,
^d Internal Medicine,
^e Clinical Pathology, College of Medicine, Yonsei University, Seoul, Korea;
^b Department of Pathology, National Health Insurance Cooperation Ilsan Hospital, Koyang, Korea;
^c Department of Forensic Pathology, National Institute of Scientific Investigation, Seoul, Korea;
^f Department of Pediatrics, Seoul National University College of Medicine, Seoul, Korea;
^l Department of Microbiology,
^g Hematology-Oncology,
^h Biochemistry,
ⁱ Pediatrics,
^j Clinical Pathology,
^k Parasitology, Medical Research Center, College of Medicine, Ewha Womans University, Seoul, Korea

Key Words. Cord blood • Megakaryocyte • Thrombopoietin • Stem cell factor • Ex vivo expansion • Apoptosis

Ju-Young Seoh, M.D., Ph.D., Department of Microbiology, College of Medicine, Ewha Womans University, Mok-6-Dong 911-1, Yangchon-Gu, Seoul 158-710, Korea. Telephone: 822-650-5738; Fax: 822-653-8891; e-mail: jyseoh{at}mm.ewha.ac.kr

Thrombopoietin (TPO) is widely used for ex vivo expansion of hematopoietic stem cells. Previously, we have reported that TPO induces a characteristic pattern of apoptosis, and the TPO-induced apoptosis is closely associated with megakaryocyte (MK) differentiation. In the present study, several cytokines, flt3-ligand, stem cell factor (SCF), interleukin-3 (IL-3), IL-6, IL-11, leukemia inhibitory factor, G-CSF, and erythropoietin, which are known to affect megakaryocytopoiesis, have been evaluated to elucidate their effects on the TPO-induced apoptosis.

Measurement of apoptosis by flow cytometry revealed that only SCF absolutely reduced the TPO-induced apoptosis in MK fractions, particularly in the late phase of ex vivo expansion. Platelet production was demonstrated by electron microscopy in a later phase when SCF was added. Simultaneous measurement of DNA contents with immunophenotyping demonstrated a significant increase in polyploidization in the CD41⁺ cell fraction when cultured with SCF. These results suggested that SCF not only inhibited premature senescence but also enhanced maturation of the differentiating cells of MK lineage during ex vivo expansion using TPO.