Mitogenic effects of urokinase on melanoma cells are independent of high affinity binding to the urokinase receptor

The structural and functional properties of the urokinase-type plasminogen activator (u-PA) that are involved in the mitogenic effect of this proteolytic enzyme on human melanoma cells M14 and IF6 and the role of the u-PA receptor (u-PAR) in transducing this signal were analyzed. Native u-PA purified from urine induced a mitogenic response in quiescent IF6 and M14 cells that ranged from 25 to 40% of the mitogenic response obtained by fetal calf serum. The half-maximum response in M14 and IF6 cells was reached at u-PA concentrations of approximately 35 and 60 nM, respectively. Blocking the proteolytic activity of u-PA resulted in a 30% decrease of the mitogenic effect, whereas inhibition of plasmin activity did not alter the mitogenic effect. No mitogenic response was elicited by low molecular weight u-PA, lacking the growth factor domain and the kringle domain. The ATF domain of u-PA induced a mitogenic response that was similar to complete u-PA. Defucosylated ATF and recombinant u-PA purified from Escherichia coli lacking all post-translational modifications did not induce a mitogenic response. Blocking the interaction of u-PA with u-PAR, using a specific monoclonal antibody, did not alter the mitogenic effect induced by u-PA. The binding of radiolabeled u-PA to M14 and IF6 cells was characterized by high affinity binding mediated by u-PAR and low affinity binding to an unknown binding site. These results demonstrate that proteolytically inactive u-PA is able to induce a mitogenic response in quiescent melanoma cells in vitro by a mechanism that involves the ATF domain but is independent of high affinity binding to u-PAR. Furthermore, it suggests that u-PA is able to bind with low affinity to a hitherto unidentified membrane associated protein that could be involved in u-PA-induced signal transduction.     

Potential nitrate removal in a coastal freshwater sediment (Haringvliet Lake, The Netherlands) and response to salinization

Nitrogen transformations and their response to salinization were studied in bottom sediment of a coastal freshwater lake (Haringvliet Lake, The Netherlands). The lake was formed as the result of a river impoundment along the south-western coast of the Netherlands, and is currently targeted for restoration of estuarine conditions. Nitrate porewater profiles indicate complete removal of NO(3)(-) within the upper few millimeters of sediment. Rapid NO(3)(-) consumption is consistent with the high potential rates of nitrate reduction (up to 200 nmol N cm(-3) h(-1)) measured with flow-through reactors (FTRs) on intact sediment slices. Acetylene-block FTR experiments indicate that complete denitrification accounts for approximately half of the nitrate reducing activity. The remaining NO(3)(-) reduction is due to incomplete denitrification and alternative reaction pathways, most likely dissimilatory nitrate reduction to NH(4)(+) (DNRA). Results of FTR experiments further indicate that increasing bottom water salinity may lead to a transient release of NH(4)(+) and dissolved organic carbon from the sediment, and enhance the rates of nitrate reduction and nitrite production. Increased salinity may thus, at least temporarily, increase the efflux of NH(4)(+) from the sediment to the surface water. This work shows that salinity affects the relative importance of denitrification compared to alternative nitrate reduction pathways, limiting the ability of denitrification to remove bioavailable nitrogen from aquatic ecosystems.     

An effective shaking procedure for 2D and 3D strip packing problems

Strip packing intends to minimise the height required for placing a set of rectangular items into a strip with infinite height. The present paper introduces a fast, yet effective shaking algorithm for the two- and three-dimensional strip packing problems, which are both NP-hard. The proposed heuristic procedure starts from an ordered item list, from which it alternates between forward and backward construction phases. The algorithm builds upon the common (deepest) bottom-left-fill algorithm, but shows significantly better results. Improvements on the solution quality of more than 9% can be observed. Moreover, applying the shaking procedure as a post processing algorithm to existing high performance heuristics also leads to improvements.