Page 122 - 2021_03-Haematologica-web

X. Ma et al.
megakaryocytes, and quantitative reverse transcription PCR revealed no difference in Mpl transcripts between WT and Plt Slc35a1–/– megakaryocytes, suggesting reduced Mpl expression in Slc35a1–/– megakaryocytesis is likely caused by impaired protein stability. Deficiency of Mpl may contribute to the defective platelet production as the percentage of proplatelet-forming megakaryocytes and absolute count of reticulated platelets were significantly decreased in Plt Slc35a1–/– mice, although we did not repro- ducibly detect impaired Mpl-mediated signaling in Slc35a1–/– megakaryocytes. Desialylation of other glyco- proteins, such as GPIba, on megakaryocytes and platelets may also contribute to reduced platelet production. Previous studies have shown that refrigeration-mediated desialylation in platelets results in GPIba shedding in vitro.44,45 In our study, while both Slc35a1–/– megakaryocytes and platelets exhibited significant reductions of sialyla- tion, GPIba level was only decreased on Slc35a1–/– platelets, while it remained unchanged on Slc35a1–/– megakaryocytes. These results support that reduced sialy- lation causes shedding of GPIba on circulating platelets, likely due to the action of plasma proteases. However, as the level of GPIba on Slc35a1–/– megakaryocytes is unchanged, it less likely to contribute to reduced platelet generation. Nevertheless, whether desialylation directly affects the function of these surface glycoproteins needs further study.
Genetic mutations of the SLC35A1 gene cause a rare form of a congenital disorder of glycosylation, SLC35A1- CDG. So far, only three human cases associated with SLC35A1 mutations have been reported. A recent study of such a patient showed that CST is not required for pro- platelet formation,23 a finding that differs from our mouse results which show that Slc35a1–/– megakaryocytes have defective proplatelet formation. There are a few differ- ences that may contribute to some of the inconsistencies between the study of the human patient and our mouse model, including: (i) the SLC35A1-CDG patient has a point mutation in the transmembrane domain, which might still have some residual CMP-SA transporting activ- ities; (ii) in vitro-differentiated megakaryocytes from isolat- ed CD34+ cells from the patient’s peripheral blood were used in the human study while bone marrow megakary- ocytes were used in our study; (iii) the SLC35A1-CDG
patient has reduced sialylation in all cell types while our mouse model has deletion of Slc35a1 specifically in megakaryocytes and platelets, and (iv) potential differ- ences between human and mouse megakaryocytes. Nevertheless, we believe our mouse model, which is the first mouse model of Slc35a1 deficiency in megakaryo- cytes and platelets, will be valuable for determining the role of sialylation in platelet biology and helping to dissect the molecular and cellular pathogenesis of thrombocy- topenia, a primary clinical presentation of patients with SLC35A1-CDG. The study of patients and the mouse model are complementary.
In addition to SLC35A1-CDG, desialylation contributes to many acquired platelet disorders, such as refractory immune thrombocytopenia and prolonged isolated thrombocytopenia after hematopoietic stem cell trans- plantation.45,46 Our current and future studies using the Plt Slc35a1–/– mice as a tool may reveal more insights into the pathogenesis of these platelet problems, which may in turn lead to new diagnoses and therapies.
Disclosures
This study is a collaboration between Dr. Changgeng Ruan's laboratory at Jiangsu Institute of Hematology (JIH), Soochow University, China, and the Oklahoma Medical Research Foundation (OMRF), USA. The study was funded by JIH and OMRF. There are no intellectual conflicts of interests.
Contributions
XM, YL, YK, JH, SAA and PA designed and conducted experiments, analyzed data, and drafted the manuscript. HS, YK, YJ, and XB contributed to manuscript preparation. JF, CR and LX designed the study, analyzed data, and wrote the paper.
Funding
This study was supported by grants from the National Natural Science Foundation of China (81520108005,81470825, 30928010, 81370617, 81800114), Jiangsu Provincial Key Medical Center (ZX201102), Jiangsu Provincial Special Program of Medical Science (BL2012005), and the Priority Academic Program Development (PAPD) of Jiangsu Higher Education Institutions (YXZXA2016002), National Institutes of Health (IS10OD018530, P41GM10349010), and a research grant from Oklahoma Medical Research Foundation.
References
1.de Gaetano G. Historical overview of the role of platelets in hemostasis and thrombo- sis. Haematologica. 2001;86(4):349-356.
2. Bryckaert M, Rosa JP, Denis CV, Lenting PJ. Of von Willebrand factor and platelets. Cell Mol Life Sci. 2015;72(2):307-326.
3. Herzog BH, Fu J, Wilson SJ, et al. Podoplanin maintains high endothelial venule integrity by interacting with platelet CLEC-2. Nature. 2013;502(7469):105-109.
4. Holinstat M. Normal platelet function. Cancer Metastasis Rev. 2017;36(2):195-198.
5. Mauler M, Bode C, Duerschmied D. Platelet serotonin modulates immune functions. Hamostaseologie. 2016;36(1):11-16.
6. Nurden AT. Platelet membrane glycopro- teins: a historical review. Semin Thromb Hemost. 2014;40(5):577-584.
7. Li Y, Fu J, Ling Y, et al. Sialylation on O-gly-
cans protects platelets from clearance by liver Kupffer cells. Proc Natl Acad Sci U S A. 2017;114(31):8360-8365.
8.Schauer R. Sialic acids as regulators of molecular and cellular interactions. Curr Opin Struct Biol. 2009;19(5):507-514.
9. Chen X, Varki A. Advances in the biology and chemistry of sialic acids. ACS Chem Biol. 2010;5(2):163-176.
10. Chen W, Druzak SA, Wang Y, et al. Refrigeration-induced binding of von Willebrand factor facilitates fast clearance of refrigerated platelets. Arterioscler Thromb Vasc Biol. 2017;37(12):2271-2279.
11. Grozovsky R, Giannini S, Falet H, Hoffmeister KM. Regulating billions of blood platelets: glycans and beyond. Blood. 2015;126(16):1877-1884.
12. Grewal PK, Uchiyama S, Ditto D, et al. The Ashwell receptor mitigates the lethal coagu- lopathy of sepsis. Nat Med. 2008;14(6):648- 655.
13. Rumjantseva V, Grewal PK, Wandall HH, et al. Dual roles for hepatic lectin receptors in the clearance of chilled platelets. Nat Med. 2009;15(11):1273-1280.
14. Grozovsky R, Begonja AJ, Liu K, et al. The Ashwell-Morell receptor regulates hepatic thrombopoietin production via JAK2-STAT3 signaling. Nat Med. 2015;21(1):47-54.
15.
16.
17.
Wang Y, Jobe SM, Ding X, et al. Platelet bio- genesis and functions require correct protein O-glycosylation. Proc Natl Acad Sci U S A. 2012;109(40):16143-16148.
Kudo T, Sato T, Hagiwara K, et al. C1galt1- deficient mice exhibit thrombocytopenia due to abnormal terminal differentiation of megakaryocytes. Blood. 2013;122(9):1649- 1657.
Sorensen AL, Rumjantseva V, Nayeb- Hashemi S, et al. Role of sialic acid for platelet life span: exposure of beta-galactose results in the rapid clearance of platelets from the circulation by asialoglycoprotein
768
haematologica | 2021; 106(3)