Concentration-dependent switch-on and -off mechanism of cell-growth promoting activity of TIMP-1

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It has been already reported that TIMP-1 shows a typical bell-shaped dose curve on its cell-growth promoting activity, suggesting the presence of some concentration-dependent regulation of the cell-growth promoting activity of TIMP-1(Hayakawa et al., J. Cell Sci. 107, 2373-2379, 1994; Yamashita et al., FEBS Lett. 396, 103-107, 1996). To confirm this point, human TIMP-1 gene-transfected [CGEMT1] and empty vector-transfected [CGEMEV] HTlO8O cells were first prepared, and compared their proliferation and TIMP-1 expression/secretion in fetal calf serum-free Dulbecco's modified Eagle's minimal essential medium(DMEM) were examined. CGEMT1 cells secreted TIMP-1 and proliferated after 2 days lag period. CGEMEV cells, however, neither secreted TIMP-1 nor proliferated at all. Proliferation of CGEMT1 cells were completely suppressed by the addition of a minute amount of anti-TIMP-t monoclonal antibody, clearly indicating that the proliferation is totally depend on the TIMP-1 produced by cells the … More mselves, that is, an autocrine mechanism of the cell-growth promoting activity of TIMP-1. Furthermore, cell proliferation leveled off at around 4 culture day, and TIMP-1 secretion ceased and got a plateau. TIMP-1 concentration in the culture medium was around 100ng/ml. Then, we kept CGEMT1 cells in D-MEM supplemented with 〜160 ng/ml human recombinant TIMP-1. The cells neither proliferated nor secreted TIMP-1, but started to secrete TIMP-1 and proliferate from 1 day after replacing the culture medium with DMEM alone. Concerning to the intracellular signal transduction in HT1080 cells, TIMP-1 activated both MEK and p238 MAPK maximally at the low concentration (〜25 ng/ml), but not activated at the higher concentration (>100 ng/ml). ERK1/2, however, were activated constantly by TIMP-1 at all the concentrations examined (25〜150 ng/ml). These results strongly support our hypothesis, i.e., the presence of a concentration-dependent switch-on and -off mechanism for the control of cell-growth promoting activity of TIMP-1. Less