Calcifying nanoparticles (CNPs, previously called nanobacteria) are self-propagating, cultivable macromolecular complexes. Their extraordinary characteristic is that they can aggregate carbonate apatite on their envelope from soluble calcium and phosphorus at physiologic concentrations and display cytotoxic effects on murine and human fibroblast cell lines. The question arises whether CNPs contribute to the degeneration of pulp tissue and thus result in clinically significant human dental pulp stones as nidies. This study evaluates CNPs' effects upon human dental pulp cells (HDPCs, the host cells in pulp tissue). We observed the ultrastructural variation of HDPCs attacked by CNPs. The spatial relationship of HDPCs and CNPs after coculture was also identified by immunofluorescence staining. Furthermore, it was verified by MTT viability assay that CNPs isolated from dental pulp stones exerted cytotoxic effect on HDPCs. Therefore, it could be concluded that the existence of CNPs might interfere with the normal physiologic function of the cells, and that might lead to dental pulp calcification. Elucidation of the cytotoxic characteristics of CNPs may offer a new perspective for understanding the etiology of human dental pulp stones.