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L-NMMA delays apoptosis in the nucleus pulposus cells of intervertebral discs

Chun-Lei Liu, Xiang-Jiang Wang, Yi-He Hu, Bei-Ji Lu, Gui-Qing Wang

Apoptosis plays an important role in degeneration of Nucleus Pulposus (NP) cells. L-Monomethyl- Arginine (L-NMMA) is an analogue of L-arginine and can be inactivated irreversibly by induced NOS (iNOS). L-NMMA can also compete with L-arginine for binding sites on iNOS molecules. In addition, LNMMA has been reported to influence the apoptosis of hyaline cartilage cells in vitro. Thus, we hypothesized that L-NMMA would influence the apoptosis of NP cells in vitro. Rabbit primary NP cells were cultured and subjected to flow cytometry to determine the levels of apoptosis in each group at each time point. Western blotting was performed to detect the Bcl-2/Bax ratio, as well as protein levels of caspase-3 and caspase-9. A significant increase in apoptosis was observed between the NP cells cultured with 1% FBS medium with versus without L-NMMA (P<0.01). Likewise, the Bcl-2/Bax ratio was also increased in NP cells cultured in 1% FBS with L-NMMA, versus those cultured without L-NMMA (P<0.05). Meanwhile, the activity levels of both caspase-3 and caspase-9 were reduced in the presence of L-NMMA, compared with levels observed in the absence of L-NMMA (P<0.01). This study showed that L-NMMA treatment delayed the senescence and apoptosis of NP cells in vitro, raising the possibility that L-NMMA may be a novel treatment to delay intervertebral disc degeneration.