title: Development and characterization of enhanced green fluorescent protein and luciferase expressing cell line for non-destructive evaluation of tissue engineering constructs.
abstract: This study investigates the utility of genetically modified cells developed for the qualitative and quantitative non-destructive evaluation of cells on biomaterials. The Fisher rat fibroblastic cell line has been genetically modified to stably express the reporter genes enhanced green fluorescence protein (EGFP) and luciferase. These reporter genes provide two unique opportunities to evaluate cell growth on materials without destruction of the sample. Utilizing the fluorescence of EGFP expressed in the cells, we were able to demonstrate distribution of cells in a oligo(poly(ethylene glycol) fumarate) hydrogel material and on a titanium fiber mesh scaffold using an inverted fluorescent light microscope. In addition, we were able to utilize a molecular light imaging system to macroscopically image the cells on these materials both with fluorescence and luminescence, as well as quantify the signal from the samples. Quantification of cell growth on the titanium mesh material for a period of 28 days was accomplished using the molecular light imaging system. Imaging was extended in vivo to cells on the titanium mesh scaffolds subcutaneously implanted in Fisher rats for a period of 28 days. This study outlines a non-destructive method to evaluate cells growing on biomaterials in vitro and in vivo.
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, BCM505, Houston, TX 77030, USA.
authors.forename: Jeremy S
authors.initials: JS
authors.lastname: Blum
authors.forename: Johnna S
authors.initials: JS
authors.lastname: Temenoff
authors.forename: Hansoo
authors.initials: H
authors.lastname: Park
authors.forename: John A
authors.initials: JA
authors.lastname: Jansen
authors.forename: Antonios G
authors.initials: AG
authors.lastname: Mikos
authors.forename: Michael A
authors.initials: MA
authors.lastname: Barry
mesh_terms: D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry*; D001709:Biotechnology / Q000379:methods*; D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000378:metabolism; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000737:chemistry; D008156:Luciferases / Q000378:metabolism*; D008164:Luminescent Proteins / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D008954:Models, Biological; D010957:Plasmids / Q000378:metabolism; D051381:Rats; D011916:Rats, Inbred F344; D012190:Retroviridae / Q000235:genetics; D013997:Time Factors; D023822:Tissue Engineering / Q000379:methods*; D014025:Titanium / Q000737:chemistry; D014759:Viral Envelope Proteins / Q000378:metabolism
chemical_list: D001672:Biocompatible Materials; C028391:G protein, vesicular stomatitis virus; D008164:Luminescent Proteins; D008562:Membrane Glycoproteins; D014759:Viral Envelope Proteins; D049452:Green Fluorescent Proteins; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate; D014025:Titanium; D008156:Luciferases