Activation of calpain 1 and protein kinase C alpha promotes a constitutive expression and release of matrix metalloproteinase 9 in peripheral blood mononuclear cells from cystic fibrosis patients

Matrix metalloproteinase 9 (MMP9) is physiologically involved in remodeling the extracellular matrix components but its abnormal release has been observed in several human pathologies, including cystic fibrosis. We have studied if the alteration in intracellular Ca2+homeostasis, observed in peripheral blood mononuclear cells (PBMCs), isolated from cystic fibrosis (CF) patients homozygous for deletion of phenylalanine 508 in gene of cystic fibrosis transmembrane conductance regulator (F508del-CFTR), could be involved in the abnormal presence of MMP9 in the extracellular fluids of CF patients. PBMCs were isolated from 23 healthy donors and 26 CF patients homozygous for F508del-CFTR. MMP9 levels were evaluated following 2 h of cell incubation. Our investigations show that all CF PBMCs analyzed constitutively express and release high levels of MMP9; conversely, in PBMCs from healthy donors, expression and secretion of MMP9 are undetectable but both events can be evoked, after 12 h of cell culture, by a possible paracrine stimulation. We have demonstrated that in CF and 12 h-cultured control PBMCs MMP9 secretion is prevented by chelation of intracellular Ca2+ and mediated by the concomitant activation of calpain and protein kinase Ca (PKCa) and also that MMP9 expression is mediated by the sequential activation of PKC and extracellular signal-regulated protein kinases 1 and 2 (ERK1/2). Moreover, the rescue of active F508del-CFTR reduces the extent of MMP9 secretion, correlating the channel defect to the [Ca2+]i dysregulation of CF PBMCs. Our results indicate that the high level of intracellular Ca2+ concentration in CF PBMCs, promoting the aberrant activation of both calpain and PKCa, induces a constitutive release of MMP9. These data characterize new alterations in mononuclear leukocytes of CF patients that may be of primary importance in the progression of the disease and indicate that PBMCs may contribute to the accumulation of MMP9 in fluids of CF patients.