PMID- 25590859
OWN - NLM
STAT- MEDLINE
DCOM- 20150306
LR  - 20181202
IS  - 1940-087X (Electronic)
IS  - 1940-087X (Linking)
IP  - 95
DP  - 2015 Jan 4
TI  - High throughput characterization of adult stem cells engineered for delivery of 
      therapeutic factors for neuroprotective strategies.
PG  - e52242
LID - 10.3791/52242 [doi]
LID - 52242
AB  - Mesenchymal stem cells (MSCs) derived from bone marrow are a powerful cellular 
      resource and have been used in numerous studies as potential candidates to 
      develop strategies for treating a variety of diseases. The purpose of this study 
      was to develop and characterize MSCs as cellular vehicles engineered for delivery 
      of therapeutic factors as part of a neuroprotective strategy for rescuing the 
      damaged or diseased nervous system. In this study we used mouse MSCs that were 
      genetically modified using lentiviral vectors, which encoded brain-derived 
      neurotrophic factor (BDNF) or glial cell-derived neurotrophic factor (GDNF), 
      together with green fluorescent protein (GFP). Before proceeding with in vivo 
      transplant studies it was important to characterize the engineered cells to 
      determine whether or not the genetic modification altered aspects of normal cell 
      behavior. Different culture substrates were examined for their ability to support 
      cell adhesion, proliferation, survival, and cell migration of the four 
      subpopulations of engineered MSCs. High content screening (HCS) was conducted and 
      image analysis performed. Substrates examined included: poly-L-lysine, 
      fibronectin, collagen type I, laminin, entactin-collagen IV-laminin (ECL). Ki67 
      immunolabeling was used to investigate cell proliferation and Propidium Iodide 
      staining was used to investigate cell viability. Time-lapse imaging was conducted 
      using a transmitted light/environmental chamber system on the high content 
      screening system. Our results demonstrated that the different subpopulations of 
      the genetically modified MSCs displayed similar behaviors that were in general 
      comparable to that of the original, non-modified MSCs. The influence of different 
      culture substrates on cell growth and cell migration was not dramatically 
      different between groups comparing the different MSC subtypes, as well as culture 
      substrates. This study provides an experimental strategy to rapidly characterize 
      engineered stem cells and their behaviors before their application in long-term 
      in vivo transplant studies for nervous system rescue and repair.
FAU - Sharma, Anup D
AU  - Sharma AD
AD  - Department of Chemical and Biological Engineering, Iowa State University.
FAU - Brodskiy, Pavel A
AU  - Brodskiy PA
AD  - Department of Chemical and Biological Engineering, Iowa State University; Biology 
      Program, Iowa State University.
FAU - Petersen, Emma M
AU  - Petersen EM
AD  - Department of Genetics, Development and Cell Biology, Iowa State University; 
      Biology Program, Iowa State University.
FAU - Dagdeviren, Melih
AU  - Dagdeviren M
AD  - Department of Genetics, Development and Cell Biology, Iowa State University.
FAU - Ye, Eun-Ah
AU  - Ye EA
AD  - Department of Genetics, Development and Cell Biology, Iowa State University.
FAU - Mallapragada, Surya K
AU  - Mallapragada SK
AD  - Department of Chemical and Biological Engineering, Iowa State University.
FAU - Sakaguchi, Donald
AU  - Sakaguchi D
AD  - Department of Genetics, Development and Cell Biology, Iowa State University; 
      Biology Program, Iowa State University; dssakagu@iastate.edu.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
PT  - Video-Audio Media
DEP - 20150104
PL  - United States
TA  - J Vis Exp
JT  - Journal of visualized experiments : JoVE
JID - 101313252
RN  - 0 (Brain-Derived Neurotrophic Factor)
RN  - 0 (Glial Cell Line-Derived Neurotrophic Factor)
RN  - 0 (Neuroprotective Agents)
RN  - 147336-22-9 (Green Fluorescent Proteins)
SB  - IM
MH  - Adult Stem Cells/cytology/*physiology
MH  - Animals
MH  - Brain-Derived Neurotrophic Factor/genetics
MH  - Cell Engineering/*methods
MH  - Genetic Vectors/genetics
MH  - Glial Cell Line-Derived Neurotrophic Factor/*genetics
MH  - Green Fluorescent Proteins/genetics
MH  - High-Throughput Screening Assays/*methods
MH  - Lentivirus/genetics
MH  - Mesenchymal Stem Cells/cytology/*physiology
MH  - Mice
MH  - Mice, Inbred C57BL
MH  - Neuroprotective Agents/*administration & dosage
PMC - PMC4354513
EDAT- 2015/01/16 06:00
MHDA- 2015/03/07 06:00
PMCR- 2017/01/04
CRDT- 2015/01/16 06:00
PHST- 2015/01/16 06:00 [entrez]
PHST- 2015/01/16 06:00 [pubmed]
PHST- 2015/03/07 06:00 [medline]
PHST- 2017/01/04 00:00 [pmc-release]
AID - 52242 [pii]
AID - 10.3791/52242 [doi]
PST - epublish
SO  - J Vis Exp. 2015 Jan 4;(95):e52242. doi: 10.3791/52242.