Using Chemotherapeutic Drugs to Trigger Hematopoietic Stem Cell Activation and Determine Hematological Toxicity

Nathan Boles1

1 Neural Stem Cell Institute, Rensselaer, New York
Publication Name:  Current Protocols in Toxicology
Unit Number:  Unit 22.1
DOI:  10.1002/0471140856.tx2201s47
Online Posting Date:  February, 2011
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Hematopoietic stem cells (HSCs) replenish blood cells throughout the lifetime of an animal. Maintaining the HSC pool is of paramount importance to the survival of the organism. Thus, when treating a patient or an animal with chemotherapeutic agents, all care must be taken to ensure that the HSC pool is minimally damaged while simultaneously eradicating the target population. Discussed here is a method to assess the potency of chemotherapeutic drugs on the hematopoietic system, their ability to activate the HSC compartment, and the damage done to the HSC pool. Curr. Protoc. Toxicol. 47:22.1.1‐22.1.6. © 2011 by John Wiley & Sons, Inc.

Keywords: hematopoietic stem cells; activation; chemotherapy; FACS

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Table of Contents

  • Introduction
  • Basic Protocol 1: Proliferation of Hematopoietic Cells Following Treatment with Chemotherapeutic Agents
  • Commentary
  • Literature Cited
  • Figures
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Basic Protocol 1: Proliferation of Hematopoietic Cells Following Treatment with Chemotherapeutic Agents

  • Adult mice
  • 5‐Fluorouracil (5‐FU; Sigma) or other chemotherapeutic agent
  • Phosphate‐buffered saline (PBS; appendix 2A)
  • DMEM+: Dulbecco's modified Eagle medium (DMEM) with 2% fetal calf serum (FCS) and 10 mM HEPES
  • HBSS+: HBSS ( appendix 2A) with 2% FCS and 10 mM HEPES
  • 1 mg/ml Hoechst 33342 stock solution (200×)
  • Lineage‐PE antibodies (BD Biosciences, 1:100 dilution)
    • Gr‐1‐PE
    • Mac‐1‐PE
    • Ter‐119‐PE
    • CD4‐PE
    • CD8‐PE
    • B220‐PE
  • Additional antibodies (BD Biosciences, 1:100 dilution except as noted)
    • Gr‐1‐APC
    • B220‐APC
    • Sca‐1‐APC
    • FITC‐conjugated mouse anti–human Ki‐67 (1:10)
  • 200 µg/ml propidium iodide
  • BD Cytofix/Cytoperm Plus (BD Biosciences)
  • BD Perm/Wash Buffer (BD Biosciences)
  • 37°C water bath
  • Hemocytometer
  • 40‐µm nylon filters, sterile
  • Flow cytometer and cell sorter with a UV, blue, and red laser
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Literature Cited

Literature Cited
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