title: Protease Nexin-1 affects the migration and invasion of C6 glioma cells through the regulation of urokinase Plasminogen Activator and Matrix Metalloproteinase-9/2.
abstract: Protease Nexin-1 (PN-1) or Serpine2 is a physiological regulator of extracellular proteases as thrombin and urokinase (uPA) in the brain. Besides, PN-1 is also implicated in some human cancers and further identified as a substrate for Matrix Metalloproteinase (MMP)-9, a key enzyme in tumor invasiveness. Our aim was to study the role of PN-1 in the migration and invasive potential of glioma cells, using the rat C6 glioma cell line as stable clones transfected with pAVU6+27 vector expressing PN-1 short-hairpin RNA. We find that PN-1 knockdown enhanced the in vitro migration and invasiveness of C6 cells which also showed a strong gelatinolytic activity by in situ zymography. PN-1 silencing did not alter prothrombin whereas increased uPA, MMP-9 and MMP-2 expression levels and gelatinolytic activity in a conditioned medium from stable C6 cells. Selective inhibitors for MMP-9 (Inhibitor I), MMP-2 (Inhibitor III) or exogenous recombinant PN-1 added to the culture medium of C6 silenced cells restored either the migration and invasive ability or gelatinolytic activity thus validating the specificity of PN-1 silencing strategy. Phosphorylation levels of extracellular signal-related kinases (Erk1/2 and p38 MAPK) involved in MMP-9 and MMP-2 signaling were increased in PN-1 silenced cells. This study shows that PN-1 affects glioma cell migration and invasiveness through the regulation of uPA and MMP-9/2 expression levels which contribute to the degradation of extracellular matrix during tumor invasion.
authors: Dipartimento di Scienze della Salute, Università di Catanzaro "Magna Graecia", Viale Europa, Germaneto, 88100 Catanzaro, Italy.
authors: Valentina
authors: V
authors: Pagliara
authors: Dipartimento di Farmacia e Scienze della Salute e della Nutrizione, Università della Calabria, Via P. Bucci, Arcavacata di Rende, 87036 Cosenza, Italy.
authors: Annagrazia
authors: A
authors: Adornetto
authors: Dipartimento di Scienze della Salute, Università di Catanzaro "Magna Graecia", Viale Europa, Germaneto, 88100 Catanzaro, Italy.
authors: Maria
authors: M
authors: Mammì
authors: Dipartimento di Scienze Motorie e del Benessere, Università di Napoli "Parthenope", Via Medina 40, 80133 Napoli, Italy; CEINGE, Biotecnologie Avanzate, S.C. a R.L., Via G. Salvatore 486, 80145 Napoli, Italy.
authors: Mariorosario
authors: M
authors: Masullo
authors: CEINGE, Biotecnologie Avanzate, S.C. a R.L., Via G. Salvatore 486, 80145 Napoli, Italy; Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università di Napoli "Federico II", Via S. Pansini 5, 80131 Napoli, Italy.
authors: Daniela
authors: D
authors: Sarnataro
authors: Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Università di Napoli "Federico II", Via S. Pansini 5, 80131 Napoli, Italy.
authors: Concetta
authors: C
authors: Pietropaolo
authors: Dipartimento di Scienze Motorie e del Benessere, Università di Napoli "Parthenope", Via Medina 40, 80133 Napoli, Italy; CEINGE, Biotecnologie Avanzate, S.C. a R.L., Via G. Salvatore 486, 80145 Napoli, Italy. Electronic address: rosaria.arcone@uniparthenope.it.
authors: Rosaria
authors: R
authors: Arcone
keywords: C6 glioma cell; Cell invasion; Matrix Metalloproteinase-2 (MMP-2); Matrix Metalloproteinase-9 (MMP-9); Protease Nexin-1 (Serpine2); urokinase Plasminogen Activator (uPA)