Altogether, these results clearly suggest that NOS is involved in cell differentiation ob served in PC12 cells selleckbio grown on Inhibitors,Modulators,Libraries ns TiO2 without NGF. In particular, Inhibitors,Modulators,Libraries since iNOS has been described as the enzyme predominantly involved in the production of NO preced ing the development of the differentiated phenotype in duced by NGF in PC12 cells grown on PLL glass, the results suggest that iNOS is involved in the differentiation process also in our experimental system. This is in keeping with the Inhibitors,Modulators,Libraries data of NOS expression reported in Figure 4 and confirms our hypothesis that nanotopography mimics the effect of NGF, promoting NOS expression and cytoskel etal protein nitration.
Effect of nanostructured TiO2 on the human neuroblastoma SH SY5Y cell line We then aimed at defining whether the effects produced Inhibitors,Modulators,Libraries by nanostructured TiO2 on neurite growth was specific for PC12 cells or was a generalized effect produced by the substrate on different neuronal like cell types. Therefore, we studied the behaviour on glass or ns TiO2 20 nm and 29 nm rms roughness of SH SY5Y human neuroblastoma cells which are considered as in vitro cell model of dopaminergic neurons and have been widely studied as cell model for Parkinsons disease. As shown in the case of PC12 cells, neuroblast oma cells grown on 20 or 29 nm rms ns TiO2 displayed longer neuritis with respect to cells grown on glass or on flat substrates, as revealed by bright field examination, as well as by the staining for the protein SNAP 25. The neurite length distri butions analysis showed an evident shift of the normal distribution toward higher length values.
No difference between different ns TiO2 roughnesses was observed. Inhibitors,Modulators,Libraries Western blot analysis by anti nitroTyr antibodies, shows that there is an increase in protein nitration triggered by the ns TiO2 as described above in PC12 cells suggesting that this behavior is common to different neuronal like cell types. Interestingly, in SY5Y cells evidence in literature indi cates that marked increases in the levels of nitrated pro teins induce apoptotic cell death. We show here that modest induction of protein nitration induces instead increased neuritogenesis in the same cell line. Involvement of ERK signaling cascade in nanostructured induced neuritogenesis The addition of NGF to PC12 cells causes neurite elon gation through a sustained activation of ERK, a mitogen activated protein kinase whose phosphorylation is essential to neuronal differentiation.
As reported by Yamazaki antiangiogenic et al.this activation occurs upon activation of NOS and can be obtained also by NO itself, in the absence of NGF, during NO induced neuritogenesis. These observa tions prompted us to check if the ERK signaling cascade may be also involved in the differentiation process trig gered by nanotopography.