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Antibody-mediated glycophorin C coligation on K562 cells induces phosphatidylserine exposure and cell death in an atypical apoptotic process

Authors

  • Duncheng Wang,

    1. Immunohematology Reference Laboratory, BloodCenter of Wisconsin, Milwaukee, Wisconsin
    2. Pathology & Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada
    3. Laboratory Medicine & Pathobiology, University of Toronto, Toronto, Ontario, Canada
    4. Research & Development, Canadian Blood Services, Toronto, Ontario, Canada
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  • Eva Seto,

    1. Immunohematology Reference Laboratory, BloodCenter of Wisconsin, Milwaukee, Wisconsin
    2. Pathology & Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada
    3. Laboratory Medicine & Pathobiology, University of Toronto, Toronto, Ontario, Canada
    4. Research & Development, Canadian Blood Services, Toronto, Ontario, Canada
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  • Jenny Shu,

    1. Immunohematology Reference Laboratory, BloodCenter of Wisconsin, Milwaukee, Wisconsin
    2. Pathology & Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada
    3. Laboratory Medicine & Pathobiology, University of Toronto, Toronto, Ontario, Canada
    4. Research & Development, Canadian Blood Services, Toronto, Ontario, Canada
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    • These authors contributed equally to the work presented in this paper. This work was supported by a National Science and Engineering Research Council scholarship to JAM and by Canadian Blood Services infrastructure grant TO22656.
  • Jonathan A. Micieli,

    1. Immunohematology Reference Laboratory, BloodCenter of Wisconsin, Milwaukee, Wisconsin
    2. Pathology & Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada
    3. Laboratory Medicine & Pathobiology, University of Toronto, Toronto, Ontario, Canada
    4. Research & Development, Canadian Blood Services, Toronto, Ontario, Canada
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    • These authors contributed equally to the work presented in this paper. This work was supported by a National Science and Engineering Research Council scholarship to JAM and by Canadian Blood Services infrastructure grant TO22656.
  • Bernard J. Fernandes,

    1. Immunohematology Reference Laboratory, BloodCenter of Wisconsin, Milwaukee, Wisconsin
    2. Pathology & Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada
    3. Laboratory Medicine & Pathobiology, University of Toronto, Toronto, Ontario, Canada
    4. Research & Development, Canadian Blood Services, Toronto, Ontario, Canada
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  • Gregory A. Denomme

    Corresponding author
    1. Pathology & Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada
    2. Laboratory Medicine & Pathobiology, University of Toronto, Toronto, Ontario, Canada
    3. Research & Development, Canadian Blood Services, Toronto, Ontario, Canada
    • Immunohematology Reference Laboratory, BloodCenter of Wisconsin, Milwaukee, Wisconsin
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Address reprint requests to: Gregory A. Denomme, PhD, Immunohematology Reference Laboratory, BloodCenter of Wisconsin, 638 North 18 Street, PO Box 2178, Milwaukee, WI 53201-2178; e-mail: greg.denomme@bcw.edu

Abstract

Background

Glycophorin C (GPC) is necessary in the maintenance of red blood cell structure. Severe autoimmune hemolytic anemia and hemolytic disease of the fetus and newborn (HDFN) have been associated with Gerbich (Ge) blood group system antigens expressed on GPC. Previous in vitro studies with cord blood progenitor cells have shown that anti-Ge suppresses erythropoiesis.

Study Design and Methods

Here, we evaluated the K562 erythroleukemic cell line to study the cellular effects of a murine anti-GPC. Cell proliferation was evaluated after treatment with anti-GPC. Flow cytometry was used to evaluate exofacial phosphatidylserine (PS) expression and cell viability (propidium iodide binding). Cell morphology was evaluated under light microscopy with cytospin preparations stained with May-Grünwald Giemsa.

Results

Anti-GPC dramatically inhibited K562 proliferation and increased PS expression, consistent with cytoplasmic blebbing, suggesting evidence of apoptosis. Z-VAD-FMK, an inhibitor of classical apoptosis, was unable to reverse the suppressive effect of anti-GPC. However, hemin was able to attenuate growth suppression.

Conclusion

Together, the data suggest that anti-GPC suppresses erythroid proliferation through the induction of nonclassical apoptosis.

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