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919 The Role of 5-HT on Proplatelet Formation and Thrombopoietin Production

Program: Oral and Poster Abstracts
Session: 502. Hematopoiesis: Regulation of Gene Transcription, Cytokines, Signal Transduction, Apoptosis, and Cell Cycle Regulation: Poster I
Hematology Disease Topics & Pathways:
Diseases, Bleeding and Clotting, ITP, Thrombocytopenias
Saturday, December 5, 2020, 7:00 AM-3:30 PM

Mo Yang1*, Enyu Liang, PhD2*, Jieyu Ye, PhD3*, Beng H Chong, MBBS, PhD4,5 and Liang Li, PhD1*

1The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, China
2Department of Laboratory Medicine, the Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, China
3Department of Hematology, Nanfang Hospital, Southern Medical University, Guangzhou, China
4Medicine, University of New South Wales, Kogarah, Australia
5St George Hospital, Sydney, Australia

Background: Our previous work confirmed that serotonin(5-HT)promotes the proliferation of hemopoietic stem cells and megakaryocytes (Yang M et al, Stem Cells, 2007; 2014). However, the mechanisms remain indefinite.

Methods: Q-PCR, Flow Cytometry, Western Blot, or Immunofluorescence microscope were used in the receptor and TPO study. MTT/CCK-8, Proplatelet assay, and Flow Cytometry were also used in cell proliferation and apoptosis study. The relationship between 5-HT and TPO was studied in a traumatic stress mice model.

Results: In-vitro study, there was a stimulating effect of 5-HT on proplatelet formation in human bone marrow megakaryocytes. Human BM MK progenitors cultured in serum-free medium with either 5-HT (200nM) or TPO (100 ng/ml) had more proplatelet bearing MKs than the control group (5-HT (12.3 ± 5.0)% vs. Control (6.2 ± 3.5)%, P=0.025; TPO (15.6 ± 2.5)% vs. Control, P=0.04; n=4). The 5-HT treatment group showed more mature and more in the final stage MK cells as compared to the TPO group. 5-HT2A, 2B, 2C receptors were detected in the surface of megakaryocytes. The effect of 5-HT on proplatelet formation in MK cells was via 5-HT2 receptors and this effect was reduced by 5-HT2 receptor inhibitor ketanserin. 5-HT acted on cytoskeleton reorganization in MKs via 5-HT2 receptors and ERK1/2 pathway. Using an immunofluorescence microscope with F-actin specific binder rhodamine-phalloidin staining, the polymerized actin level was lower in the control group than the 5-HT group and actin distributed diffusely throughout the cytoplasm. In contrast, the polymerization actin level was higher in the 5-HT group. Adding ketanserin and ERK1/2 inhibitor PD98059 to 5-HT treatment, the fluorescence intensity was correspondingly reduced. Our data also demonstrated that ERK1/2 was activated in MKs treated with 5-HT for 30 minutes. In a traumatic stress mice model, both of 5-HT and TPO were increased, but the increasing of TPO is posterior to 5-HT. After added LX1606, the synthesis inhibitor of 5-HT, 5-HT was reduced markedly, as well as TPO. The expression of TPO mRNA and the production of TPO protein were increased as compared with the control in this model.

Conclusions: This study suggests that 5-HT promotes thrombopoiesis from two aspects: one is the direct effect on megakaryocytes. 5-HT could promote the proplatelet formation from megakaryocytes. The second is the indirect effect by promoting the production of TPO, which is a paracrine secretion to influence thrombopoiesis.

Disclosures: No relevant conflicts of interest to declare.

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