CNTF up-regulation of TIMP-2 in neuroblastoma cells

The ciliary neurotrophic factor (CNTF) can regulate survival and differentiation of many types of developing and adult neurons; in metastatic SK-N-BE neuroblastoma cells, it promotes differentiation and neurite outgrowth. The expression of Gelatinase A (MMP-2) and its specific tissue inhibitor (TIMP-2), a degradative system whose balance is involved in matrix invasion and metastasis, was investigated in SK-N-BE cells cultured with and without CNTF or NGF. Zymographic analysis of conditioned media revealed that the cells constitutively secrete two gelatinases, mainly pro-MMP-2 but also traces of pro-MMP-9. In a time-course experiment in the presence of 25 ng/ml of CNTF, the MMP-2 mRNA expression showed no significant modulation, while TIMP-2 mRNA up-regulated to > 2-fold after 48 h and then fell dramatically. At the same concentrations, NGF showed no effect. TIMP-2 mRNA expression showed a dose-dependent increase of up to 8-fold from 1 to 250 ng/ml of CNTF and increased secretion of TIMP-2 was confirmed by Western blotting. MMP-2 was only slightly over-expressed under the same conditions, at either mRNA or protein level, with no correlation with neurocytokine concentration. These results suggest that boosting the expression of TIMP-2 by CNTF could restrain both matrix degradation following nervous system injury and neuroblastoma aggressiveness.