Reference - Detail
|Author||Mitra M, Mohanty C, Harilal A, Maheswari UK, Sahoo SK, Krishnakumar S.|
|Title||A novel in vitro three-dimensional retinoblastoma model for evaluating chemotherapeutic drugs.|
PURPOSE:Novel strategies are being applied for creating better in vitro models that simulate in vivo conditions for testing the efficacy of anticancer drugs. In the present study we developed surface-engineered, large and porous, biodegradable, polymeric microparticles as a scaffold for three dimensional (3-D) growth of a Y79 retinoblastoma (RB) cell line. We evaluated the effect of three anticancer drugs in naïve and nanoparticle-loaded forms on a 3-D versus a two-dimensional (2-D) model. We also studied the influence of microparticles on extracellular matrix (ECM) synthesis and whole genome miRNA-gene expression profiling to identify 3D-responsive genes that are implicated in oncogenesis in RB cells.
METHODS:Poly(D,L)-lactide-co-glycolide (PLGA) microparticles were prepared by the solvent evaporation method. RB cell line Y79 was grown alone or with PLGA-gelatin microparticles. Antiproliferative activity, drug diffusion, and cellular uptake were studied by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, a yellow tetrazole (MTT) assay, fluorescent microscope, and flow cytometry. Extra cellular matrix (ECM) synthesis was observed by collagenase assay and whole genome miRNA-microarray profiling by using an Agilent chip.
RESULTS:With optimized composition of microparticles and cell culture conditions, an eightfold increase from the seeding density was achieved in 5 days of culture. The antiproliferative effect of the drugs in the 3-D model was significantly lower than in the 2-D suspension, which was evident from the 4.5 to 21.8 fold differences in their IC(50) values. Using doxorubicin, the flow cytometry data demonstrated a 4.4 fold lower drug accumulation in the cells grown in the 3-D model at 4 h. The collagen content of the cells grown in the 3-D model was 2.3 fold greater than that of the cells grown in the 2-D model, suggesting greater synthesis of the extracellular matrix in the 3-D model as the extracellular matrix acted as a barrier to drug diffusion. The microarray and miRNA analysis showed changes in several genes and miRNA expression in cells grown in the 3-D model, which could also influence the environment and drug effects.
CONCLUSIONS:Our 3-D retinoblastoma model could be used in developing effective drugs based on a better understanding of the role of chemical, biologic, and physical parameters in the process of drug diffusion through the tumor mass, drug retention, and therapeutic outcome.
|MeSH||Antineoplastic Agents / pharmacology* Biocompatible Materials / chemistry Carboplatin / pharmacology* Cell Culture Techniques / methods Cell Proliferation / drug effects Cell Survival / drug effects Diffusion Doxorubicin / pharmacology* Etoposide / pharmacology* Extracellular Matrix / drug effects Extracellular Matrix / metabolism Gene Expression Profiling Humans Inhibitory Concentration 50 Kinetics Lactic Acid / chemistry MicroRNAs / biosynthesis* Nanoparticles / chemistry Oligonucleotide Array Sequence Analysis Particle Size Polyglycolic Acid / chemistry Polylactic Acid-Polyglycolic Acid Copolymer Porosity Retinoblastoma / drug therapy Retinoblastoma / metabolism Retinoblastoma / pathology* Tissue Engineering / methods Tissue Scaffolds|
|WOS Category||OPHTHALMOLOGY BIOCHEMISTRY & MOLECULAR BIOLOGY|
|Human and Animal Cells|