Assessment of Proteolytic Activities in the Bone Marrow Microenvironment

  • Andreas MaurerEmail author
  • Gerd Klein
  • Nicole D. Staudt
Part of the Methods in Molecular Biology book series (MIMB, volume 2017)


During cytokine- or chemotherapy-induced hematopoietic stem cell (HSC) mobilization, a highly proteolytic microenvironment can be observed in the bone marrow that has a strong influence on adhesive and chemotactic interactions of HSC with their niches. The increase of proteases during mobilization goes along with a decrease of endogenous protease inhibitors. Prominent members of the proteases involved in HSC mobilization belong to the families of matrix metalloproteinases and cathepsins, which are able to degrade chemokines/cytokines, extracellular matrix components, and membrane-bound adhesion receptors. To determine the functional activity of different proteolytic enzymes, zymographic analyses with different substrates and pH conditions can be employed. An involvement of cysteine cathepsins can be determined by the “active site labeling” technique using a modified inhibitor irreversibly binding to the active center of the enzymes. Intact or degraded chemokines and cytokines, which fall into the range between 1000 and 20,000 Da, can readily be detected by MALDI-TOF analysis. These three methods can help to detect proteolytic activities directly involved in the mobilization process.

Key words

Matrix metalloproteinases Cathepsins Gelatin zymography Collagen zymography Active site labeling Matrix-assisted laser desorption and ionization time-of-flight mass spectrometry 


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Preclinical Imaging and Radiopharmacy, Werner Siemens Imaging CenterUniversity of TübingenTübingenGermany
  2. 2.Department of Internal Medicine II, Center for Medical ResearchUniversity of TübingenTübingenGermany
  3. 3.Pharmaceutical BiologyUniversity of TübingenTübingenGermany

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