Objectives: In this study we assessed the cytotoxic effect of nanohydroxyapatite (NHA) incorporated into resin modified and conventional glass ionomer cements (RMGICs and CGICs) on L929 murine fibroblasts. Materials and Methods: In this in vitro study, 0wt%, 1wt%, 2wt%, 5wt%, 7wt% and 10wt% concentrations of NHA were added to Fuji II LC RMGIC and Fuji IX CGIC powders. Eighteen samples (5×3mm) were fabricated from each type of glass ionomer, in six experimental groups (n=3): CG0, CG1, CG2, CG5, CG7, CG10, RMG0, RMG1, RMG2, RMG5, RMG7, and RMG10. Samples were incubated for 72h. The overlaying solution was removed and added to L929 fibroblasts. The methyl thiazolyl tetrazolium bromide (MTT) assay was performed at 24, 48 and 72h. The wavelength was read by a spectrophotometer. Data were analyzed by ANOVA and Tukey’s test. Results: There was no significant difference in cytotoxicity of the two types of glass ionomers, with and without NHA, except for CG0 and RMG0 groups after 72h. RMG0 group was significantly more cytotoxic than the CG0 group (P<0.05). In CG groups during the first 24h, the cytotoxicity of CG5 and CG7 groups was significantly higher than that of CG1; while, there was no significant difference between the RMG groups. Cytotoxicity significantly decreased in all groups after 24h (P<0.05). Conclusion: Incorporation of NHA into Fuji II LC RMGIC and Fuji IX CGIC did not affect their biocompatibility and therefore its addition to these materials can provide favorable biological properties, especially considering its beneficial effects on the other properties of GICs.