title: On-line monitoring of adhesion and proliferation of cultured hepatoma cells using optical waveguide lightmode spectroscopy (OWLS).
abstract: Monitoring of cell adhesion, cell spreading, and cell proliferation opens attractive perspectives in the on-line control of monolayer cell cultures in toxicity tests, in bioreactors as used for the serial production of skin grafts, or in extracorporeal liver devices. In this study the hepatoma Hep G2 cell adhesion and proliferation was monitored using an integrated optical method, optical waveguide lightmode spectroscopy (OWLS). This method is based upon refractive index measurements within a 100-nm thin layer above a Si(Ti)O(2) surface on which the cells were cultured and exposed to cytotoxic and cytostatic agents. The OWLS signal was proportional to cell density during the spreading period (4 h), and in long-term experiments (46 h) the OWLS signal correlated on a logarithmic scale with cell density. After administration of the protein synthesis inhibitor cycloheximide (4 microg/mL) to fully spread hepatoma cells, cell growth was arrested and change of the OWLS signal became noticeable within 6 h after drug administration. For exposure to increasing concentrations of the anticancer drug cyclophosphamide (2.5-20 mM) a concentration-dependent reduction of the OWLS signal was found. For cycloheximide and cyclophospamide the OWLS signal was also confirmed by cell viability measurements using the neutral red assay, the thiazolylblue tetrazoliumbromide assay, total protein measurements, and cell morphology. It was demonstrated that the OWLS signal detects minor changes in cell adhesion, which serve as indicators of metabolic state and growth behavior. OWLS is thus a quantitative tool to characterize impaired cell growth mediated by culture medium, by extracellular matrix, or after exposure to a toxin.
authors.affiliation: Chemical Engineering Department, ETH (Swiss Federal Institute of Technology) Hönggerberg, HCI/F130, 8093 Zürich, Switzerland. thomassilvan@bluemail.ch
authors.forename: T S
authors.initials: TS
authors.lastname: Hug
authors.forename: J E
authors.initials: JE
authors.lastname: Prenosil
authors.forename: P
authors.initials: P
authors.lastname: Maier
authors.forename: M
authors.initials: M
authors.lastname: Morbidelli
mesh_terms: D019149:Bioreactors; D006528:Carcinoma, Hepatocellular / Q000473:pathology*; D002448:Cell Adhesion; D018929:Cell Culture Techniques / Q000379:methods; D002455:Cell Division; D002470:Cell Survival; D003513:Cycloheximide / Q000494:pharmacology; D003520:Cyclophosphamide / Q000494:pharmacology; D005336:Fiber Optic Technology; D006801:Humans; D009862:Online Systems; D013057:Spectrum Analysis / Q000295:instrumentation*; D023822:Tissue Engineering / Q000295:instrumentation*; D014407:Tumor Cells, Cultured
chemical_list: D003520:Cyclophosphamide; D003513:Cycloheximide