Tissue inhibitors of metalloproteinases (TIMP) are specific inhibitors of matrix metalloproteinases (MMPs) and thus participate in maintaining the balance between extracellular matrix deposition and degradation in several physio-pathological processes. Nevertheless, TIMP must be regarded as multifunctional proteins involved in cell growth, angiogenesis and apoptosis. The molecular mechanisms induced by TIMP remain largely unknown. In the present study, we provide evidence that TIMP-1 induces a significant anti-apoptotic effect in the human erythroleukaemic cell line UT-7 and in the murine myeloid cell line 32D. Using specific kinases inhibitors, we show that TIMP-1-mediated cell survival is dependent upon Janus kinase (JAK) 2 and phosphoinositide 3-kinase (PI 3-kinase) activities. By transient transfection of dominant-negative Akt in UT-7 cells, we demonstrate that this kinase is crucial for the TIMP-1 anti-apoptotic effect. Moreover, TIMP-1 enhances specific phosphorylation of both Akt and Bad (Bcl-2/Bcl-XL-antagonist, causing cell death) in a PI 3-kinase-dependent manner and, besides, controls the level of the anti-apoptotic protein Bcl-XL. We conclude that TIMP-1 induces haematopoietic cell survival via the JAK2/PI 3-kinase/Akt/Bad pathway.
Abbreviations used: Bad, Bcl-2/Bcl-XL-antagonist, causing cell death; (p)EGFP, (plasmid) enhanced green fluorescent protein; EPA, erythroid potentiating activity; Epo, erythropoietin; IL-3, interleukin-3; JAK, Janus kinase; MAP kinase, mitogen-activated protein kinase; MEM, minimal essential medium; MMP, matrix metalloproteinase; PI 3-kinase, phosphoinositide 3-kinase; TBS, Tris-buffered saline; TBST, TBS and 0.1% (v/v) Tween 20; TIMP, tissue inhibitor of metalloproteinases.