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ATRA CAN CORRECT DEFECTIVE HIF-1Α/S1P AXIS-MEDIATED CYTOSKELETAL REORGANIZATION IN PROPLATELET FORMATION OF ITP
Author(s): ,
Qiu-Sha Huang
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Jing Xue
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Feng-Qi Liu
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Qi Chen
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Gao-Chao Zhang
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Xue-Yan Sun
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Chen-Cong Wang
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Li-Ping Yang
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
,
Yue-Ying Li
Affiliations:
Chinese Academy of Sciences (CAS) Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, CAS, Beijing, China,Beijing,Chine;Chinese Academy of Sciences (CAS) Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, CAS, Beijing,
,
Qian-Fei Wang
Affiliations:
Chinese Academy of Sciences (CAS) Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, CAS, Beijing, China,Beijing,Chine;Chinese Academy of Sciences (CAS) Key Laboratory of Genomic and Precision Medicine, Collaborative Innovation Center of Genetics and Development, Beijing Institute of Genomics, CAS, Beijing,
,
Jun Peng
Affiliations:
Department of Hematology, Shangdong Key Laboratory of Immunochematology, and Shandong Provincial Clinical Medicine Research Center for Hematology, Qilu Hospital, Cheeloo College of Medicine, Shandong University,Jinan,Chine;Department of Hematology, Shangdong Key Laboratory of Immunochematology, and Shandong Provincial Clinical Medicine Research Center for Hematology, Qilu Hospital, Cheeloo College
,
Ming Hou
Affiliations:
Department of Hematology, Shangdong Key Laboratory of Immunochematology, and Shandong Provincial Clinical Medicine Research Center for Hematology, Qilu Hospital, Cheeloo College of Medicine, Shandong University,Jinan,Chine;Department of Hematology, Shangdong Key Laboratory of Immunochematology, and Shandong Provincial Clinical Medicine Research Center for Hematology, Qilu Hospital, Cheeloo College
,
Xiao-Jun Huang
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
Xiao-Hui Zhang
Affiliations:
Peking University People's Hospital,Beijing,Chine;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,Cina;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;Peking University People's Hospital,Beijing,China;P
(Abstract release date: 05/12/22) EHA Library. Huang Q. 06/11/22; 357154; S290
Qiu-Sha Huang
Qiu-Sha Huang
Contributions
Abstract
Presentation during EHA2022: All Oral presentations will be presented between Friday, June 10 and Sunday, June 12 and will be accessible for on-demand viewing from Monday, June 20 until Monday, August 15, 2022 on the Congress platform.

Abstract: S290

Type: Oral Presentation

Session title: Thrombocytopenia - Management insights in ITP and TTP

Background

Bone marrow physiological hypoxia plays a crucial role in haematopoietic stem cell homeostasis. Hypoxia inducible factor (HIF) is central to mediating the cellular response to hypoxia. HIF-1α expression was decreased in the bone marrow of patients with immune thrombocytopenia (ITP), and HIF-1α activation was shown to enhance megakaryopoiesis in mice. Recent studies suggest that "inside-out" signalling by S1P in megakaryocytes (MKs) plays a critical role in proplatelet formation (PPF) (Blood, 2013; J EXP MED, 2012). Our previous data indicated that impaired PPF contributed to the development of thrombocytopenia in ITP. To further explore the underlying mechanism of impaired PPF in ITP, we found that HIF-1α/S1P axis-mediated cytoskeletal reorganization was defective in the PPF of ITP. All-trans retinoic acid (ATRA), which has been shown to be a promising treatment option for ITP patients in our clinical studies (Blood, 2021; Lancet haematology, 2017; Lancet haematology, 2021), could restore cytoskeletal reorganization and correct impaired PPF.

Aims

 This study aimed to explore the role of hypoxia inducible factor-1α (HIF-1α) in proplatelet formation (PPF) and the underlying mechanisms of ATRA treatment in ITP patients.

Methods

Thirty consecutive patients with newly diagnosed ITP and 30 healthy donors were included in our study. MKs were isolated from bone marrow samples. Targeted and untargeted metabolomic profiling through metabolomic analysis was performed to explore the relationship between the metabolome and ITP. Confocal microscopy and transmission electron microscopy were used to observe the PPF and cytoskeleton structure of ITP MKs. An ITP mouse model was established to observe the therapeutic effects of ATRA in the PPF.

Results

In the present study, we observed that MKs displayed altered cytoskeletal reorganization and impaired proplatelet formation (PPF) in ITP patients. Targeted and untargeted metabolite profiling revealed a decreased sphingosine-1-phosphate (S1P) level in ITP. Downregulated sphingosine kinase 2 (SPHK2) expression in MKs accounted for the low level of S1P in ITP. S1P is essential for S1P receptor 1 (S1PR1) and Rac1 activation, Src family kinases (SFKs) activity, and subsequent cytoskeletal reorganization and PPF regulation. Moreover, we demonstrated that HIF-1α mediated SPHK2 activation and S1P production. Decreased HIF-1α levels were found in the MKs of patients with ITP, contributing to impaired PPF.

We then investigated the effect of ATRA on PPF in ITP patients. ATRA upregulated HIF-1α and SPHK2 expression, increased S1P production and corrected impaired PPF in vitro. In an ITP mouse model, ATRA alleviated thrombocytopenia and restored cytoskeletal reorganization. ATRA corrected impaired PPF by upregulating HIF-1α expression. The exposure of ITP MKs to selective RARα (AM580) or RARγ (BMS961) agonists did not change PPF. However, the treatment of ITP MKs with a selective RARβ agonist (CD2314) significantly increased PPF. Furthermore, we found that the effect of ATRA on enhancing PPF in ITP MKs was reversed in the presence of an RARβ antagonist (CD2665). These data suggest that impaired PPF in ITP MKs is corrected by ATRA in a RARβ-dependent manner.

Conclusion

Together, our data show that the HIF-1α/S1P axis mediates altered cytoskeletal reorganization and impaired PPF in ITP and suggest that ATRA correction of impaired PPF is a potential mechanistic explanation for the clinical efficacy of ATRA in ITP.

Keyword(s): Cytoskeleton, Immune thrombocytopenia (ITP), Proplatelet, Treatment

Presentation during EHA2022: All Oral presentations will be presented between Friday, June 10 and Sunday, June 12 and will be accessible for on-demand viewing from Monday, June 20 until Monday, August 15, 2022 on the Congress platform.

Abstract: S290

Type: Oral Presentation

Session title: Thrombocytopenia - Management insights in ITP and TTP

Background

Bone marrow physiological hypoxia plays a crucial role in haematopoietic stem cell homeostasis. Hypoxia inducible factor (HIF) is central to mediating the cellular response to hypoxia. HIF-1α expression was decreased in the bone marrow of patients with immune thrombocytopenia (ITP), and HIF-1α activation was shown to enhance megakaryopoiesis in mice. Recent studies suggest that "inside-out" signalling by S1P in megakaryocytes (MKs) plays a critical role in proplatelet formation (PPF) (Blood, 2013; J EXP MED, 2012). Our previous data indicated that impaired PPF contributed to the development of thrombocytopenia in ITP. To further explore the underlying mechanism of impaired PPF in ITP, we found that HIF-1α/S1P axis-mediated cytoskeletal reorganization was defective in the PPF of ITP. All-trans retinoic acid (ATRA), which has been shown to be a promising treatment option for ITP patients in our clinical studies (Blood, 2021; Lancet haematology, 2017; Lancet haematology, 2021), could restore cytoskeletal reorganization and correct impaired PPF.

Aims

 This study aimed to explore the role of hypoxia inducible factor-1α (HIF-1α) in proplatelet formation (PPF) and the underlying mechanisms of ATRA treatment in ITP patients.

Methods

Thirty consecutive patients with newly diagnosed ITP and 30 healthy donors were included in our study. MKs were isolated from bone marrow samples. Targeted and untargeted metabolomic profiling through metabolomic analysis was performed to explore the relationship between the metabolome and ITP. Confocal microscopy and transmission electron microscopy were used to observe the PPF and cytoskeleton structure of ITP MKs. An ITP mouse model was established to observe the therapeutic effects of ATRA in the PPF.

Results

In the present study, we observed that MKs displayed altered cytoskeletal reorganization and impaired proplatelet formation (PPF) in ITP patients. Targeted and untargeted metabolite profiling revealed a decreased sphingosine-1-phosphate (S1P) level in ITP. Downregulated sphingosine kinase 2 (SPHK2) expression in MKs accounted for the low level of S1P in ITP. S1P is essential for S1P receptor 1 (S1PR1) and Rac1 activation, Src family kinases (SFKs) activity, and subsequent cytoskeletal reorganization and PPF regulation. Moreover, we demonstrated that HIF-1α mediated SPHK2 activation and S1P production. Decreased HIF-1α levels were found in the MKs of patients with ITP, contributing to impaired PPF.

We then investigated the effect of ATRA on PPF in ITP patients. ATRA upregulated HIF-1α and SPHK2 expression, increased S1P production and corrected impaired PPF in vitro. In an ITP mouse model, ATRA alleviated thrombocytopenia and restored cytoskeletal reorganization. ATRA corrected impaired PPF by upregulating HIF-1α expression. The exposure of ITP MKs to selective RARα (AM580) or RARγ (BMS961) agonists did not change PPF. However, the treatment of ITP MKs with a selective RARβ agonist (CD2314) significantly increased PPF. Furthermore, we found that the effect of ATRA on enhancing PPF in ITP MKs was reversed in the presence of an RARβ antagonist (CD2665). These data suggest that impaired PPF in ITP MKs is corrected by ATRA in a RARβ-dependent manner.

Conclusion

Together, our data show that the HIF-1α/S1P axis mediates altered cytoskeletal reorganization and impaired PPF in ITP and suggest that ATRA correction of impaired PPF is a potential mechanistic explanation for the clinical efficacy of ATRA in ITP.

Keyword(s): Cytoskeleton, Immune thrombocytopenia (ITP), Proplatelet, Treatment

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