Chapter title |
Engineering Human Mesenchymal Stem Cells to Release Adenosine Using miRNA Technology
|
---|---|
Chapter number | 16 |
Book title |
RNAi and Small Regulatory RNAs in Stem Cells
|
Published in |
Methods in molecular biology, July 2017
|
DOI | 10.1007/978-1-4939-7108-4_16 |
Pubmed ID | |
Book ISBNs |
978-1-4939-7106-0, 978-1-4939-7108-4
|
Authors |
Ren, Gaoying, Boison, Detlev, Gaoying Ren M.D., Detlev Boison, Gaoying Ren |
Editors |
Baohong Zhang |
Abstract |
Adenosine is an important modulator of metabolic activity with powerful tissue and cell protective functions. Adenosine kinase (ADK), the major adenosine-regulating enzyme, is critical to adapt its intra- and extracellular levels in response to environmental changes. Lentiviral RNAi-mediated downregulation of ADK in human mesenchymal stem cells (hMSCs) has therefore been considered an effective tool for engineering therapeutically effective adenosine-releasing cell grafts that could constitute patient-identical autologous implants for clinical application. We constructed lentiviral vectors that co-express miRNA directed against ADK and an emerald green fluorescent protein (EmGFP) reporter gene. Following lentiviral transduction of hMSCs, we demonstrated up to 80% downregulation of ADK and 98% transduction efficiency. Transduced hMSCs continued to express EmGFP after four to six consecutive passages, and EmGFP-positive hMSC grafts survived in the hippocampal fissure of mouse brains and provided efficient adenosine-dependent neuroprotection in a mouse model of seizure-induced cell loss. |
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