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Lymphoma

In vivo adhesion of malignant B cells to bone marrow microvasculature is regulated by α4β1 cytoplasmic-binding proteins

Leukemia volume 30, pages 861–872 (2016)Cite this article

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Abstract

Multiple myeloma (MM) and chronic lymphocytic leukemia (CLL) cells must attach to the bone marrow (BM) microvasculature before lodging in the BM microenvironment. Using intravital microscopy (IVM) of the BM calvariae we demonstrate that the α4β1 integrin is required for MM and CLL cell firm arrest onto the BM microvasculature, while endothelial P-selectin and E-selectin mediate cell rolling. Talin, kindlin-3 and ICAP-1 are β1-integrin-binding partners that regulate β1-mediated cell adhesion. We show that talin and kindlin-3 cooperatively stimulate high affinity and strength of α4β1-dependent MM and CLL cell attachment, whereas ICAP-1 negatively regulates this adhesion. A functional connection between talin/kindlin-3 and Rac1 was found to be required for MM cell attachment mediated by α4β1. Importantly, IVM analyses with talin- and kindlin-3-silenced MM cells indicate that these proteins are needed for cell arrest on the BM microvasculature. Instead, MM cell arrest is repressed by ICAP-1. Moreover, MM cells silenced for talin and kindlin-3, and cultured on α4β1 ligands showed higher susceptibility to bortezomib-mediated cell apoptosis. Our results highlight the requirement of α4β1 and selectins for the in vivo attachment of MM and CLL cells to the BM microvasculature, and indicate that talin, kindlin-3 and ICAP-1 differentially control physiological adhesion by regulating α4β1 activity.

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Acknowledgements

We thank the multiple myeloma and chronic lymphocytic leukemia patients who donated samples for this research. We thank Dr Pedro Lastres for his help in flow cytometry. We also thank JM Adrover for movie editing. This work was supported by the following grants from the Ministerio de Economía y Competitividad (Spain): SAF2011-24022 to JT; SAF2012-31613 to AG-P; SAF2012-31142 and SAF2013-49662-EXP to AH; PI13/01454 to PS-M; RD12/0036/0061 to JT and AG-P; RD12/0036/0058 to NCG. The work was also funded by grant P2010/BMD-2314 from the Comunidad de Madrid to AG-P, JT, PS-M and AH, and partially supported by a CRIS foundation grant to fight cancer to JM-L. NCG is also funded by a grant from the Asociación Española Contra el Cáncer (GCB120981SAN), and from the Gerencia Regional de Salud, Junta de Castilla y León (GRS-702/A/11). The CNIC is supported by the Pro-CNIC Foundation.

Author contributions

MM-M, ML and NA-M performed experiments and analyzed data. SS-M, SIV and NA-S performed some experiments, analyzed data and prepared valuable reagents. NG, RM, JM-L, IB and JAG-M collected and provided primary cells from MM and CLL patients. JT, AG-P, AH and PS-M designed the research and wrote the manuscript. All authors reviewed and approved the manuscript.

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Author notes

  1. M Martínez-Moreno and M Leiva: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Cellular and Molecular Medicine, Centro de Investigaciones Biológicas (CSIC), Madrid, Spain

    M Martínez-Moreno, N Aguilera-Montilla, S Sevilla-Movilla, S Isern de Val, N Arellano-Sánchez, A García-Pardo & J Teixidó

  2. Area of Cell and Developmental Biology, Fundación Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain

    M Leiva & A Hidalgo

  3. Department of Hematology, Hospital Universitario de Salamanca, Salamanca, Spain

    N C Gutiérrez

  4. Section of Hematology, Hospital Universitario 12 de Octubre, Madrid, Spain

    R Maldonado & J Martínez-López

  5. Section of Hematology, Hospital General Universitario Gregorio Marañón, Instituto de Investigación Sanitaria Gregorio Marañón, Madrid, Spain

    I Buño

  6. Hematology Unit, Hospital Universitario Puerta de Hierro, Madrid, Spain

    J A García-Marco

  7. Section of Immuno-Oncology, Hospital General Universitario Gregorio Marañón, Complutense University School of Medicine, Madrid, Spain

    P Sánchez-Mateos

  8. Institute for Cardiovascular Prevention, Ludwig-Maximilians-University, Munich, Germany

    A Hidalgo

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  1. M Martínez-Moreno
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Martínez-Moreno, M., Leiva, M., Aguilera-Montilla, N. et al. In vivo adhesion of malignant B cells to bone marrow microvasculature is regulated by α4β1 cytoplasmic-binding proteins. Leukemia 30, 861–872 (2016). https://doi.org/10.1038/leu.2015.332

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