EXPRESSION OF 5-HT RECEPTORS AND NEURONAL MARKER MAP 2, GFAP AND
TAU ON HUMAN MEGAKARYOCYTES AND MEGAKARYOCYTIC CELL LINES
Yang M, Li K, Yuen PMP, Pong NH, Wong R, Chik KW, Li
CK, Shing MK, Fok TF Department of Paediatrics, The Prince of Wales Hospital,
The Chinese University of Hong Kong, Hong Kong
Objective: Serotonin (5-Hydroxytryptamine, 5-HT) is a monoamine
with a molecular weight of 176.2 dalton can affect the functions of many
cells including platelets. It has been reported that serotonin has mitogenic
effects on smooth muscle cells and fibroblasts. Recently, serotonin has
been recognized not only as a neurotransmitter and vasoactive agent, but
also as a growth factor representative of a class of molecules promoting
cell growth by interacting with G protein coupled receptors. Our previous
studies have shown that serotonin significantly stimulates megakaryocyte
colony formation with maximum stimulation at 100 nM and ketanserin, a 5-HT2
receptor antagonist, blocks the mitogenic effect of serotonin on
megakaryocytopoiesis.
Methods: In this study, we investigated the expression of
5-HT2A, 2B and 2C receptors, and other neuronal marker on megakaryocytic
lineage and the effect of serotonin on anti-apoptosis in megakaryocytes
using RT-PCR, flowcytometry, immunofluorescence staining, annexin V and
caspase-3 assay.
Results: We found that: (1) Using RT-PCR method, 5-HT2A
receptors were identified on the four megakaryocytic cell lines: Meg-01,
Dami, CHRF-288-11 and M-07e. 5-HT2B receptors were also detected on Meg-01,
Dami and M-07e but not on CHRF-288-11 cells; (2) Using flow cytometry with
monoclonal antibodies, 5-HT2A, 2B and 2C receptors were identified on all
four megakaryocytic cell lines. It was not detected on the control cells
K562; (3) The expression of the three receptors were further confirmed on
human bone marrow megakaryocytes by using immunofluorescence staining; (4)
Furthermore, the anti-apoptotic effect of serotonin was studied in M-07e
cells by annexin V and active caspase-3 assay using flow cytometry. Our
preliminary results demonstrated the apoptotic effect of thrombospondin-1
(TSP-1). (2.5 mg/ml)
was abolished by serotonin (100 nM) as well as TPO (50 ng/ml) in the
culture of M-07e cells. And (5) The neuronal marker MAP2, GFAP and Tau were
also identified on human bone marrow megakaryocytes and human
megakaryocytic cell lines (Meg-01, Dami and M-07e cells).
Conclusions: This
study demonstrated that megakaryocytic lineage expresses 5-HT2A, 2B and 2C
receptors and serotonin has anti-apoptotic effect on megakaryocytic cell
line in vitro.