Stem cell transplantation in an in vitro simulated ischemia/reperfusion model

Attila Cselenyák, Zsolt Benko, Mónika Szepes, L. Kiss, Z. Lacza

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Stem cell transplantation protocols are finding their way into clinical practice. Getting better results, making the protocols more robust, and finding new sources for implantable cells are the focus of recent research. Investigating the effectiveness of cell therapies is not an easy task and new tools are needed to investigate the mechanisms involved in the treatment process. We designed an experimental protocol of ischemia/reperfusion in order to allow the observation of cellular connections and even subcellular mechanisms during ischemia/reperfusion injury and after stem cell transplantation and to evaluate the efficacy of cell therapy. H9c2 cardiomyoblast cells were placed onto cell culture plates. Ischemia was simulated with 150 minutes in a glucose free medium with oxygen level below 0.5%. Then, normal media and oxygen levels were reintroduced to simulate reperfusion. After oxygen glucose deprivation, the damaged cells were treated with transplantation of labeled human bone marrow derived mesenchymal stem cells by adding them to the culture. Mesenchymal stem cells are preferred in clinical trials because they are easily accessible with minimal invasive surgery, easily expandable and autologous. After 24 hours of co-cultivation, cells were stained with calcein and ethidium-homodimer to differentiate between live and dead cells. This setup allowed us to investigate the intercellular connections using confocal fluorescent microscopy and to quantify the survival rate of postischemic cells by flow cytometry. Confocal microscopy showed the interactions of the two cell populations such as cell fusion and formation of intercellular nanotubes. Flow cytometry analysis revealed 3 clusters of damaged cells which can be plotted on a graph and analyzed statistically. These populations can be investigated separately and conclusions can be drawn on these data on the effectiveness of the simulated therapeutical approach.

Original languageEnglish
Article numbere3575
JournalJournal of visualized experiments : JoVE
Issue number57
DOIs
Publication statusPublished - Nov 2011

Fingerprint

Stem Cell Transplantation
Stem cells
Reperfusion
Ischemia
Flow cytometry
Oxygen
Cell culture
Glucose
Cell- and Tissue-Based Therapy
Confocal microscopy
Mesenchymal Stromal Cells
Confocal Microscopy
Flow Cytometry
Surgery
Nanotubes
Microscopic examination
Bone
Fusion reactions
Cells
Cell Fusion

Keywords

  • Confocal microscopy
  • Flow cytometry
  • Ischemia/reperfusion model
  • Issue 57
  • Medicine
  • Stem cell transplantation

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Chemical Engineering(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)

Cite this

Stem cell transplantation in an in vitro simulated ischemia/reperfusion model. / Cselenyák, Attila; Benko, Zsolt; Szepes, Mónika; Kiss, L.; Lacza, Z.

In: Journal of visualized experiments : JoVE, No. 57, e3575, 11.2011.

Research output: Contribution to journalArticle

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