Proteomic analysis of reporter genes for molecular imaging of transplanted embryonic stem cells.
Study of stem cells may reveal promising treatment for diseases. The fate and function of transplanted stem cells remain poorly defined. Recent studies demonstrate that reporter genes can monitor real-time survival of transplanted stem cells in living subjects. We examined the effects of a novel and versatile triple fusion (TF) reporter gene construction on embryonic stem (ES) cell function by proteomic analysis. Murine ES cells were stably transduced with a self-inactivating lentiviral vector containing fluorescence (firefly luciferase; Fluc), bioluminescence (monomeric red fluorescence protein; mRFP), and positron emission tomography (herpes simplex virus type 1 truncated thymidine kinase; tTK) reporter genes. Fluorescence-activated cell sorting (FACS) analysis isolated stably transduced populations. TF reporter gene effects on cellular function were evaluated by quantitative proteomic profiling of control ES cells versus ES cells stably expressing the TF construct (ES-TF). Overall, no significant changes in protein quantity were observed. TF reporter gene expression had no effect on ES cell viability, proliferation, and differentiation capability. Molecular imaging studies tracked ES-TF cell survival and proliferation in living animals. In summary, this is the first proteomic study, demonstrating the unique potential of reporter gene imaging for tracking ES cell transplantation non-invasively, repetitively, and quantitatively.
Wu
Joseph C
JC
Department of Radiology and Molecular Imaging Program at Stanford, Stanford University School of Medicine, Stanford, CA 94305, USA. joewu@stanford.edu
Cao
Feng
F
Dutta
Sucharita
S
Xie
Xiaoyan
X
Kim
Elmer
E
Chungfat
Neil
N
Gambhir
Sanjiv
S
Mathewson
Sean
S
Connolly
Andrew J
AJ
Brown
Matthew
M
Wang
Evelyn W
EW
D000818:Animals; D019677:Artificial Gene Fusion; D053595:Embryonic Stem Cells / Q000378:metabolism* / Q000637:transplantation*; D020869:Gene Expression Profiling / Q000379:methods*; D017930:Genes, Reporter / Q000502:physiology*; D016086:Lentivirus / Q000235:genetics; D008163:Luminescent Measurements; D008164:Luminescent Proteins / Q000096:biosynthesis; D051379:Mice; D010103:Oxygen Isotopes; D040901:Proteomics; D014162:Transfection
D008164:Luminescent Proteins; D010103:Oxygen Isotopes