Volume 32 (2009) Issue 8 Pages 1328-1334
Nitric oxide (NO) and heme oxygenase-1 (HO-1) play important roles in the regulation of stem cell proliferation and differentiation. However, it has not been examined whether human periodontal ligament (PDL) cells can differentiate into osteoblast-like cells by NO activity mediated via HO-1. The objective of this study was to determine the effect of NO on proliferation and differentiation in human PDL cells, and to identify the underlying mechanism of its actions. Primary human PDL cells were cultured with NO donor sodium nitroprusside (SNP); cell proliferation and differentiation were measured. NO production, cell viability and cell proliferation were evaluated using the Griess reagent, MTT assay and BrdU incorporation, respectively. To analyze differentiation, we measured alkaline phosphatase (ALP) activity, osteocalcin (OC), osteonectin (ON) expression, and bone sialoprotein (BSP) by Western blotting. SNP-induced NO production is associated with inducible nitric oxide synthase induction in a time and dose-dependent manner. SNP resulted in decreased cell proliferation and increased expression of osteogenic differentiation markers such as ALP, OC, ON and BSP. Maximal HO-1 was reached with 0.05 mM SNP and gradually decreased with 1.0 mM. Treatment with an HO-1 inhibitor and selective inhibitors of extracellular regulated kinase 1/2 and nuclear factor-kappaB blocked the SNP-induced growth inhibition, as well as osteoblastic differentiation. These data suggest that NO-induced osteogenic differentiation through HO-1 may be an important mediator of periodontal regeneration or bone tissue engineering.