Nitric oxide (NO), synthesized from L-arginine by a family of constitutive and inducible NO synthases (iNOS), is a small, diffusible, diatomic, highly reactive molecule with a variety of cellular functions in all mammalian cells. 1) Recently, it was shown that NO plays an important role in stem cell proliferation and differentiation. It has been reported that, at physiological concentrations, NO functions as a negative regulator of stem/progenitor cell proliferation and is critical to the initiation of cell differentiation.2,3) NO also promotes bone and chondrocyte terminal differentiation ; stimulates preadipocyte differentiation in rat 6) ; and mediates osteoblastic differentiation.7) In addition, NO synthase inhibitors have been shown to arrest differentiation toward a cardiac phenotype in mouse embryonic stem cells; differentiation is rescued by NO donors. 8) In contrast, NO donor sodium nitroprusside (SNP) inhibits mineralization in the mouse chondrocyte-like ATDC5 cell line. 9) However, whether NO can affect the differentiation of periodontal ligament (PDL) cells is unknown.The PDL is a non-mineralized connective tissue which surrounds the tooth and exhibits osteoblast-like features, such as high alkaline phosphatase (ALP) activity and expression of osteonectin (ON), bone sialoprotein (BSP), osteocalcin (OC), and osteopontin (OPN). The cells that compose the PDL are capable of differentiating into osteoblasts or cementoblasts.10) Moreover, PDL cells have demonstrated stem cell properties such as self-renewal, clonogenicity, and multitissue differentiation potential.11) In addition, PDL cells can differentiate in response to a variety of extracellular stimuli, serving either to maintain homeostasis or to remodel, repair, and regenerate the surrounding hard tissue.12) Recently, increased NO production during osteogenic differentiation has been reported in human PDL cells, 13) but the role of NO in osteoblastic differentiation of these cells requires further investigation.Heme oxygenase (HO) is the rate-limiting enzyme involved in the catabolism of heme, yielding equimolar quantities of carbon monoxide (CO), free iron, and biliverdin; the latter two compounds are converted to ferritin and bilirubin, respectively. 14) One of three mammalian HO isoforms, HO-1 (also called heat shock protein 32), is a stress-responsive protein induced by various agents and is involved in a variety of regulatory and protective mechanisms in cells. 15,16) We previously reported that HO-1 is induced by pro-inflammatory cytokines, NO, substance P, hydrogen peroxide, and mechanical stress, [17][18][19][20][21][22][23] and that it may play a cytoprotective role in human pulp and PDL cells. However, HO-1-mediated NOinductive effects of osteoblastic differentiation in PDL cells has not yet been reported.Understanding the mechanisms that regulate osteogenic differentiation in human PDL cells will have important implications for the development of new therapeutic strategies in periodontal defects. This study aimed to investigate whether h...

No abstract

No abstract

Genomic DNA was isolated from buccal swabs by the QIAamp DNA Mini Kit (QIAGEN). The primers used for amplification and sequencing of exon 6 and 7 of ABO gene were suggested by Lee and Chang (1), and Ogasawara et al. (2), respectively. About 1-2 ng of genomic DNA was used for PCR in 25 μL reaction volume. PCR mixture contained 10 mM Tris-HCl pH 8.5, 50 mM KCl, 1.5 mM MgCl2, 200 μM of each dNTP, 0.25 μM of each primer, and 1 U of AmpliTaq Gold DNA Polymerase (Applied Biosystems). Thermal cycling was performed initially at 95°C for 10 min, then 35 cycles consisting of 95°C for 1 min, 60°C for 1 min for exon 6 and 63°C for 1 min for exon 7, 72°C for 1 min, followed by 10 min extension at 72°C in a GeneAmp PCR System 9600 (Perkin Elmer).