Exogenous Nitric Oxide Induced Early Mineralization in Rat Bone Marrow Mesenchymal Stem Cells via Activation of Alkaline Phosphatase

Abstract

Background:Since the low concentration and short-time treatment with sodium nitroprusside (SNP), a nitric oxide (NO)–donor, cause no harm to rat bone marrow mesenchymal stem cells (MSCs), we studied the impact of SNP on MSCs differentiation. Methods:MSCs were treated with 100 and 1000 µM of SNP for 1 hour in every 48 hours and after 5, 10, 15, and 21 days in osteogenic media. The viability and the level of mineralization were determined using MTT assay and alizarin red staining, respectively. Morphology of the cells was studied using fluorescent dye. Concentration of calcium and the activity of alanine transaminase (ALT), aspartate transaminase (AST), lactate dehydrogenase (LDH), and alkaline phosphatase (ALP) were evaluated by commercial kits. Results:SNP with the concentration of 1000 µM significantly reduced viability from day 5 to day 20, but 100 µM did not affect the viability until the day 15. The low concentration of SNP increased matrix deposition from day 10 and reached almost its maximum (4.40 ± 2.4) at the day 15. Also, increasing the activity of ALP (419 ± 2.2), due to low concentration of SNP, started at day 10 and continued till the day 20, while LDH (2026 ± 11) and AST (25.6 ± 0.4) elevations were observed from day 5 onwards. In case of ALT, we observed a significant decrease (36%) from day 5 till day 20. Conclusion:Based on our findings, low concentrations of SNP might be useful in the promotion of bone repair.

Off-Stoichiometric Reactions at the Cell–Substrate Biomolecular Interface of Biomaterials: In Situ and Ex Situ Monitoring of Cell Proliferation, Differentiation, and Bone Tissue Formation

“…Extensively used as a venous vasodilator, sodium nitroprusside serves to immediately decrease the blood pressure in critical clinical situations [69]. The efficacy of exogenous NO on the in vitro osteogenic differentiation of mesenchymal stem cells has been the object of a recent study by Abnosi and Pari [70]. These researchers discovered that low concentrations of exogenous sodium nitroprusside (in the order of <0.1 mM generating ~1.2 nM of NO) promoted the ability to proliferate and the capacity to differentiate into osteoblasts of rat bone marrow mesenchymal stem cells.…”

Exogenous Nitric Oxide Up-regulates the Runx2 Via Bmp7 Overexpression to Increase the Osteoblast Matrix Production In Vitro

“…Felka et al reported the osteogenic inhibitory effect of exogenous NO at high concentrations (1 mM) in the early stage of osteogenesis, but in our previous study, we showed that the 100 µM of SNP as the NO-releasing agent induced the early matrix formation of BMSCs at day 5 in vitro (19,20). Some studies reported that the matrix formation starts from day 7 when the activity of ALP begins as an early gene (23,24).…”

“…SNP for 1 hour every 48 hours elevated matrix formation from day 10 of the differentiation process up to the 20th day of incubation (20).…”

Therapeutic Potential of Reactive Oxygen Species: State of the Art and Recent Advances

“…This suggests that the mechanism of action was independent of hydrogen peroxide formation. 163 The addition of the nitric oxide (NO) donors ( Z )-1[ N -(2-aminoethyl)- N -(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA-NONOate) and sodium nitroprusside 110,169–171 was also used to stimulate NO production—which belongs to reactive nitrogen species yet acts similarly to ROS. Nevertheless, while simple and convenient, due to their instability, the exogenous administration of ROS appears infeasible for tissue regeneration purposes.…”