In vitro assessment of alendronate toxic and apoptotic effects on human dental pulp stem cells

Objective(s): Osteonecrosis of the jaw, as an exposed necrotic bone in the oral cavity, is one of the adverse effects of bisphosphonates, which have an affinity for bone minerals. This study investigates the cytotoxic effects of alendronate (ALN) as a nitrogen-containing bisphosphonate, on human dental pulp stem cells (hDPSCs). Materials and Methods: The mesenchymal stem cells (MSCs), obtained from third molar tooth pulps were characterized by immunophenotyping assay in order to identify surface markers to evaluate their expression. To detect multipotency hDPSCs, they were differentiate into osteocytes and adipocytes. Cell proliferation was measured by MTT assay. PI staining of DNA fragmentation by flowcytometry (sub-G۱ peak) was performed for determination of apoptotic cells and Bax, Bcl-۲, and cleaved caspase ۳ expressions. Protein expression was detected by Western blotting. Results: As the results revealed, ALN decreased viable cells (in ۰.۸–۱۰۰ µM) after ۷۲ hr and ۱۶۸ hr (P<۰.۰۰۱), significantly. ALN could lower cell proliferation in hDPSCs in a concentration and time-dependent manner. Sub-G۱ peak as an indicator of flowcytometry histogram of treated cells by ALN, showed apoptosis was involved in ALN-induced cytotoxicity. Expressions of cleaved caspase ۳ and Bax protein, as pro-apoptotic proteins, were increased and Bcl-۲ protein as anti-apoptotic protein was decreased in response to increases in the concentration of ALN (۰.۸–۲۵ µM).Conclusion: Long-term effects of ALN on cell proliferation and apoptosis in hDPSCs can result in either initiation or potentiation of ALN-induced osteonecrosis.

Keywords:

Alendronate , Apoptosis , Bisphosphonates , Human dental pulp stem cells , Proliferation