DNA IMAGE CYTOMETRY AND AG-NORS-STAINING APPLICATION IN BIOCOMPATIBILITY STUDIES ON HUMAN OSTEOBLAST CELLS IN-VITRO

We report here the study of the biocompatibility of a bone graft mater ial, the Pyrost(R), using a previously established in vitro model of h uman osteoblasts, The effect of this material on cell proliferation wa s evaluated by the MTS assay. Results indicated the absolute absence o f cytotoxic or cytostatic effect of Pyrost(R) on cultured osteoblasts. Viability rate was more than 90% in cells cultured with the material compared to the control. Morphological analysis, undertaken by scannin g electron microscopy showed a good adhesion and a spreading of osteob lasts in contact with the material that was colonized by cultured cell s. In the second part of this work, we have introduced two methods as complementary biocompatibility tests: DNA image cytometry and interpha se Ag-NORs quantification. DNA content was measured in cells cultured with or without Pyrost(R) for 3, 9, 15 and 30 days. The determination of DNA indicated that the majority of osteoblasts population was diplo id without aneuploidy. The DNA index and cell distribution profile in DNA histograms were similar in all cell populations. The Ag-NORs amoun t was used as a parameter for cell kinetic evaluation. We have measure d the Ag-NORs index like DNA quantification. The proliferation rate, e valuated by Ag-NORs counts in osteoblasts cultured with or without the material, was identical. However, a decrease in Ag-NORs index was obs erved from day 3 to day 15 of incubation. These results showed a satis factory biocompatibility of the Pyrost(R) in human osteoblasts culture , The material did not alter cell viability and had no inducing effect either on proliferation rate or on cell ploidy as demonstrated by DNA image cytometry and Ag-NORs proteins staining. (C) 1998 Published by Elsevier Science Ltd. All rights reserved.