mahsa maleklou

Although exogenous nitric oxide (NO) is used as medicine, in the previous we showed its inhibitory effect on the proliferation ability of rat bone marrow mesenchymal stem cells (BMSCs). In the present investigation, the inhibitory role of exogenous NO on BMSCs cell cycle was studied.

BMSCs after the third passage were treated for one hour every 48 hours with 100μM of sodium nitroprusside as an NO donor. Then, after 5,10,15, and 20 days of treatment, the viability, proliferation, and cell cycle of the BMSCs was investigated. In addition, the expression of the Raf1, CDK2, CDK4, P53, and GAPDH genes was studied.

Cell treatment caused a significant reduction in viability and proliferation at 5,10,15, and 20 days. Also, the treatment caused cell cycle arrest at G1 after 20 days. In addition, it was found that the CDK2 and CDK4 expression were down-regulated whereas the P53 expression was up-regulated, but the expression of Raf1 as well as GAPDH remained the same.

This study showed that prolonged treatment with a NO donor arrest the BMSCs cell cycle due to overexpression of P53, which inhibits the expression of Cdk2 and Cdk4.

Nitric oxide (NO) is a signaling molecule that is required for the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). According to previous reports, high concentrations of sodium nitroprusside (SNP) inhibit the osteogenic differentiation of BMSCs, while its low concentration promotes this process.

The present investigation focused on evaluating the underlying mechanism of the osteogenic differentiation of BMSCs treated with low concentrations of SNP as an NO generating agent.

The BMSCs after the 3rd passage was differentiated to osteoblasts when treated with 100 µM for 1 hour every 48 hours until 5, 10, 15, and 20 days of incubation. Then, the matrix production was estimated by quantitative alizarin red assay and calcium determination. The expression of different genes involved in osteogenic differentiation was statistically determined using the reverse transcriptase polymerase chain reaction. Finally, alkaline phosphatase activity was measured by a commercial kit.

The exogenous NO caused a significant (P<0.05) increase in the matrix production of differentiated BMSCs from day 5 to 20. The results showed the elevation of alkaline phosphatase activity and the up-regulation of its gene. Eventually, an increase was observed in the expression of a cascade of other genes such as osteonectin, Bmp7, Smad1, Runx2, and Raf1 in treated BMSCs.

Overall, short-time treatment with a low concentration of exogenous NO increases the matrix production via gene up-regulation and protein production, which might open a new window in treating the low-density bone complication.