NAC prevents apoptosis through inverse regulation of NFkB and JNK

Objective: The mechanisms by which N-acetyl cysteine (NAC) inhibits cell death mediated by resin based materials have not been completely elucidated. The objective of our study is to demonstrate that NAC inhibits 2-hydroxyethyl methacrylate (HEMA) induced apoptotic cell death of oral keratinocytes and Dental Pulp Stromal Cells (DPSCs) via NF&KappaB dependent and independent and JNK dependent pathways thereby promoting subsequent pulpal regeneration.

Methods: DPSCs and two different NF&KappaB knockdown cell lines, an oral larygngeal carcinoma HEp2 and an oral keratinocyte cell line HOK-16B were used. Dual staining with FITC- Annexin V and Propidium iodide was used to assess cell death and viability in NF&KappaB knockdown cells when treated with HEMA. Whole cell extracts were used to measure the levels of JNK and nuclear extracts measured the levels of NF&KappaB in untreated and HEMA treated cells. Additionally the effects of a JNK inhibitor on cell death in the knockdown cells were assessed.

Results: HEMA induced decrease in mitochondrial membrane potential, increase in cleaved caspases, increase in cell death and loss of cell function were potently inhibited in the presence of NAC. The protective effect of NAC was related to its ability to partly induce NF&KappaB and promote differentiation in oral keratinocytes and DPSCs. In addition, NAC was capable of rescuing a small but reproducible fraction of NF&KappaB knock down cells from HEMA mediated cell death. Therefore, there is an NF&KappaB independent and JNK dependent pathway of protection from HEMA mediated cell death by NAC, since NAC is capable of decreasing JNK in NF&KappaB knock down cells.

Conclusion: Coordinated decrease in NF&KappaB and increase in JNK are the major mechanisms by which HEMA induces cell death of oral keratinocytes and DPSCs. These effects are significantly prevented by NAC which then subsequently promotes pulpal regeneration.

Supported by RO1- DE 10331:08 from NIH-NIDCR.