Autophagosome-rich platelets are increased in immune thrombocytopenia

Immune thrombocytopenia (ITP) is characterized by increased platelet destruction and reduced platelet production, mediated by antiplatelet autoantibodies. Owing to the absence of definitive tests, diagnosis of ITP mainly involves exclusion of other thrombocytopenic disorders. Accumulating evidence h...

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Published inJournal of Iwate Medical Assiociation Vol. 72; no. 3; pp. 103 - 113
Main Authors Hitomi, Jiro, Izumita, Wataru, Miyajima, Shinri, Seki, Yuki, Ito, Shigeki, Otsu, Akihiro, Kowata, Shugo, Ishida, Yoji, Murai, Kazunori, Kiyohara, Kazuki
Format Journal Article
LanguageEnglish
Published Iwate Medical Association 01.08.2020
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ISSN0021-3284
2434-0855
DOI10.24750/iwateishi.72.3_103

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Abstract Immune thrombocytopenia (ITP) is characterized by increased platelet destruction and reduced platelet production, mediated by antiplatelet autoantibodies. Owing to the absence of definitive tests, diagnosis of ITP mainly involves exclusion of other thrombocytopenic disorders. Accumulating evidence has implicated enhanced autophagy in thrombopoiesis and ITP. However, it remains unknown whether platelet autophagy is enhanced in patients with ITP and has diagnostic potential. To determine this, we measured the proportion of autophagosome-rich platelets (ARPs%) in patients with thrombocytopenia, including ITP. We examined 34 patients with ITP and 52 patients with other types of thrombocytopenic disorders. To detect and quantify autophagosomes within platelets, we used flow cytometry with a cationic fluorescence dye, Cyto-ID, which specifically labels autophagic compartments. Among thrombocytopenic patients, ARPs% was significantly elevated only in ITP patients (p ‹ 0.01) and was found to be significantly correlated with the proportion of reticulated platelets (RPs%) (r = 0.58) and mean platelet volume (r = 0.55). ARPs% showed a greater sensitivity but reduced specificity (88.2% and 71.2%, respectively) compared with those of RPs% (85.3% and 90.4%, respectively). These data suggest that enhanced autophagy of platelets is a characteristic finding in patients with ITP and shows diagnostic potential in thrombocytopenia.
AbstractList Immune thrombocytopenia (ITP) is characterized by increased platelet destruction and reduced platelet production, mediated by antiplatelet autoantibodies. Owing to the absence of definitive tests, diagnosis of ITP mainly involves exclusion of other thrombocytopenic disorders. Accumulating evidence has implicated enhanced autophagy in thrombopoiesis and ITP. However, it remains unknown whether platelet autophagy is enhanced in patients with ITP and has diagnostic potential. To determine this, we measured the proportion of autophagosome-rich platelets (ARPs%) in patients with thrombocytopenia, including ITP. We examined 34 patients with ITP and 52 patients with other types of thrombocytopenic disorders. To detect and quantify autophagosomes within platelets, we used flow cytometry with a cationic fluorescence dye, Cyto-ID, which specifically labels autophagic compartments. Among thrombocytopenic patients, ARPs% was significantly elevated only in ITP patients (p ‹ 0.01) and was found to be significantly correlated with the proportion of reticulated platelets (RPs%) (r = 0.58) and mean platelet volume (r = 0.55). ARPs% showed a greater sensitivity but reduced specificity (88.2% and 71.2%, respectively) compared with those of RPs% (85.3% and 90.4%, respectively). These data suggest that enhanced autophagy of platelets is a characteristic finding in patients with ITP and shows diagnostic potential in thrombocytopenia.
Author Ishida, Yoji
Izumita, Wataru
Seki, Yuki
Miyajima, Shinri
Otsu, Akihiro
Kiyohara, Kazuki
Ito, Shigeki
Murai, Kazunori
Kowata, Shugo
Hitomi, Jiro
Author_xml – sequence: 1
  fullname: Hitomi, Jiro
  organization: Division of Human Embryology, Department of Anatomy, School of Medicine, Iwate Medical University, Yahaba, Japan
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  fullname: Izumita, Wataru
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
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  fullname: Miyajima, Shinri
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
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  fullname: Seki, Yuki
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
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  fullname: Ito, Shigeki
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
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  fullname: Otsu, Akihiro
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
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  fullname: Kowata, Shugo
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
– sequence: 1
  fullname: Ishida, Yoji
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
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  fullname: Murai, Kazunori
  organization: Division of Hematology, Iwate Prefecture Central Hospital, Morioka, Japan
– sequence: 1
  fullname: Kiyohara, Kazuki
  organization: Division of Hematology and Oncology, Department of Internal Medicine, School of Medicine, Iwate Medical University, Yahaba, Japan
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Shintani T and Klionsky DJ: Autophagy in health and disease: a double-edged sword. Science 306, 990-995, 2004.
Liu Z and Mei T: Immune thrombocytopenia induces autophagy and suppresses apoptosis in megakaryocytes. Mol Med Rep 18, 4016-4022, 2018.
Arnal C, Piette JC, Leone J, et al.: Treatment of severe immune thrombocytopenia associated with systemic lupus erythematosus: 59 cases. J Rheumatol 29, 75-83, 2002.
Provan D, Stasi R, Newland AC, et al.: International consensus report on the investigation and management of primary immune thrombocytopenia. Blood 115, 168-186, 2010.
Khellaf M, Chabrol A, Mahevas M, et al.: Hydroxychloroquine is a good second-line treatment for adults with immune thrombocytopenia and positive antinuclear antibodies. Am J Hematol 89, 194-198, 2014.
Barsam SJ, Psaila B, Forestier M, et al.: Platelet production and platelet destruction: assessing mechanisms of treatment effect in immune thrombocytopenia. Blood 117, 5723-5732, 2011.
Guo S, Liang Y, Murphy SF, et al.: A rapid and high content assay that measures Cyto-ID-stained autophagic compartments and estimates autophagy flux with potential clinical applications. Autophagy 11, 560-572, 2015.
Kanda Y: Investigation of the freely available easy-to-use software 'EZR' for medical statistics. Bone Marrow Transplant 48, 452-458, 2013.
Kienast J and Schmitz G: Flow cytometric analysis of thiazole orange uptake by platelets: a diagnostic aid in the evaluation of thrombocytopenic disorders. Blood 75, 116-121, 1990.
Mortensen M, Soilleux EJ, Djordjevic G, et al.: The autophagy protein Atg7 is essential for hematopoietic stem cell maintenance. J Exp Med 208, 455-467, 2011.
Ouseph MM, Huang Y, Banerjee M, et al.: Autophagy is induced upon platelet activation and is essential for hemostasis and thrombosis. Blood 126, 1224-1233, 2015.
Matic GB, Chapman ES, Zaiss M, et al.: Whole blood analysis of reticulated platelets: improvements of detection and assay stability. Cytometry 34, 229-234, 1998.
Lee SH, Du J, Stitham J, et al.: Inducing mitophagy in diabetic platelets protects against severe oxidative stress. EMBO Mol Med 8, 779-795, 2016.
Choi AM, Ryter SW, Levine B, et al.: Autophagy in human health and disease. N Engl J Med 368, 1845-1846, 2013.
Guo S, Pridham KJ, Sheng Z, et al.: Detecting autophagy and autophagy flux in chronic myeloid leukemia cells using a Cyto-ID fluorescence spectrophotometric assay. Methods Mol Biol 1465, 95-109, 2016.
Roche O, Net A, Rakotonjanahary J, et al.: Evaluation of the efficiency of hydroxychloroquine in treating children with immune thrombo-cytopenia. Am J Hematol 92, E79-81, 2017.
Brown AS and Martin JF: The megakaryocyte platelet system and vascular disease. Eur J Clin Invest 24, 9-15, 1995.
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Kashiwagi H and Tomiyama Y: Diagnosis in general. In "Autoimmune thrombocytopenia" eds by Ishida Y, et al., pp. 87-95. Springer, Singapore, 2017.
Houwerzijl EJ, Blom NR, van der Want JJ, et al.: Megakaryocytic dysfunction in myelodysplastic syndromes and idiopathic thrombocytopenic purpura is in part due to different forms of cell death. Leukemia 20, 1937-1942, 2006.
Sakuragi M, Hayashi S, Maruyama M, et al.: Clinical significance of IPF% or RP% measurement in distinguishing primary immune thrombocytopenia from aplastic thrombocytopenic disorders. Int J Hematol 101, 369-735. 2015.
Tomiyama Y: Differential diagnosis: Hypoplastic thrombocytopenia. In "Autoimmune thrombo-cytopenia" eds by Ishida Y, et al., pp. 107-114. Springer, Singapore, 2017.
Kashiwagi H and Tomiyama Y: ITP in adults. In "Autoimmune thrombocytopenia"eds by Ishida Y, et al., pp. 75-84. Springer, Singapore, 2017.
References_xml – reference: Togawa R and Kowata S: The basic approach to measure nascent platelets by autophagosome detection in blood sample. J Iwate Med Assoc 69, 209-217, 2017.
– reference: Mortensen M, Soilleux EJ, Djordjevic G, et al.: The autophagy protein Atg7 is essential for hematopoietic stem cell maintenance. J Exp Med 208, 455-467, 2011.
– reference: Roche O, Net A, Rakotonjanahary J, et al.: Evaluation of the efficiency of hydroxychloroquine in treating children with immune thrombo-cytopenia. Am J Hematol 92, E79-81, 2017.
– reference: Guo S, Liang Y, Murphy SF, et al.: A rapid and high content assay that measures Cyto-ID-stained autophagic compartments and estimates autophagy flux with potential clinical applications. Autophagy 11, 560-572, 2015.
– reference: Ouseph MM, Huang Y, Banerjee M, et al.: Autophagy is induced upon platelet activation and is essential for hemostasis and thrombosis. Blood 126, 1224-1233, 2015.
– reference: Houwerzijl EJ, Blom NR, van der Want JJ, et al.: Megakaryocytic dysfunction in myelodysplastic syndromes and idiopathic thrombocytopenic purpura is in part due to different forms of cell death. Leukemia 20, 1937-1942, 2006.
– reference: Arnal C, Piette JC, Leone J, et al.: Treatment of severe immune thrombocytopenia associated with systemic lupus erythematosus: 59 cases. J Rheumatol 29, 75-83, 2002.
– reference: Kanda Y: Investigation of the freely available easy-to-use software 'EZR' for medical statistics. Bone Marrow Transplant 48, 452-458, 2013.
– reference: Kowata S and Yoji I: Megakaryocytopoiesis and thrombopoiesis. In "Autoimmune thrombocytopenia" eds by Ishida Y, et al., pp. 9-19, Springer, Singapore, 2017.
– reference: Liu Z and Mei T: Immune thrombocytopenia induces autophagy and suppresses apoptosis in megakaryocytes. Mol Med Rep 18, 4016-4022, 2018.
– reference: Khellaf M, Chabrol A, Mahevas M, et al.: Hydroxychloroquine is a good second-line treatment for adults with immune thrombocytopenia and positive antinuclear antibodies. Am J Hematol 89, 194-198, 2014.
– reference: Kashiwagi H and Tomiyama Y: Diagnosis in general. In "Autoimmune thrombocytopenia" eds by Ishida Y, et al., pp. 87-95. Springer, Singapore, 2017.
– reference: Tomiyama Y: Differential diagnosis: Hypoplastic thrombocytopenia. In "Autoimmune thrombo-cytopenia" eds by Ishida Y, et al., pp. 107-114. Springer, Singapore, 2017.
– reference: Matic GB, Chapman ES, Zaiss M, et al.: Whole blood analysis of reticulated platelets: improvements of detection and assay stability. Cytometry 34, 229-234, 1998.
– reference: Provan D, Stasi R, Newland AC, et al.: International consensus report on the investigation and management of primary immune thrombocytopenia. Blood 115, 168-186, 2010.
– reference: Kienast J and Schmitz G: Flow cytometric analysis of thiazole orange uptake by platelets: a diagnostic aid in the evaluation of thrombocytopenic disorders. Blood 75, 116-121, 1990.
– reference: Choi AM, Ryter SW, Levine B, et al.: Autophagy in human health and disease. N Engl J Med 368, 1845-1846, 2013.
– reference: Sakuragi M, Hayashi S, Maruyama M, et al.: Clinical significance of IPF% or RP% measurement in distinguishing primary immune thrombocytopenia from aplastic thrombocytopenic disorders. Int J Hematol 101, 369-735. 2015.
– reference: Brown AS and Martin JF: The megakaryocyte platelet system and vascular disease. Eur J Clin Invest 24, 9-15, 1995.
– reference: Kashiwagi H and Tomiyama Y: ITP in adults. In "Autoimmune thrombocytopenia"eds by Ishida Y, et al., pp. 75-84. Springer, Singapore, 2017.
– reference: Kowata S, Isogai S, Murai K, et al.: Platelet demand modulates the type of intravascular protrusion of megakaryocytes in bone marrow. Thromb Haemost 112, 743-756, 2014.
– reference: Lee SH, Du J, Stitham J, et al.: Inducing mitophagy in diabetic platelets protects against severe oxidative stress. EMBO Mol Med 8, 779-795, 2016.
– reference: Shintani T and Klionsky DJ: Autophagy in health and disease: a double-edged sword. Science 306, 990-995, 2004.
– reference: Guo S, Pridham KJ, Sheng Z, et al.: Detecting autophagy and autophagy flux in chronic myeloid leukemia cells using a Cyto-ID fluorescence spectrophotometric assay. Methods Mol Biol 1465, 95-109, 2016.
– reference: Barsam SJ, Psaila B, Forestier M, et al.: Platelet production and platelet destruction: assessing mechanisms of treatment effect in immune thrombocytopenia. Blood 117, 5723-5732, 2011.
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Snippet Immune thrombocytopenia (ITP) is characterized by increased platelet destruction and reduced platelet production, mediated by antiplatelet autoantibodies....
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StartPage 103
SubjectTerms autophagy
Immune thrombocytopenia
platelet
Title Autophagosome-rich platelets are increased in immune thrombocytopenia
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