RESUMO
It is shown that the release of matrix metalloproteinase-9 (gelatinase B) by THP-1 and U937 cells into conditioned media is increased under the action of recombinant single-chain urokinase. This effect is not accompanied by proteolytic activation of gelatinase B and is related to release of a pro-form of the enzyme. The action of urokinase on monocytes is time-dependent and becomes significant 12-24 h after the beginning of cell incubation. The dependence of the effect on the concentration of urokinase is characterized by half-maximum at about 20 nM and saturation at about 200 nM. The urokinase-induced gelatinase B release is not dependent on the action of plasmin because plasmin inhibitors aprotinin and alpha2-antiplasmin do not abolish this action. Additionally, tissue type plasminogen activator does not induce gelatinase B release by monocytes as observed under the action of urokinase. Nevertheless, the catalytic activity of urokinase participates in the development of the observed effect because it is significantly depressed by the natural urokinase inhibitor PAI-1. The effect of urokinase is completely abolished by actinomycin D and cycloheximide, indicating the participation of transcription and translation processes in its development.
Assuntos
Fibrinolisina/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Monócitos/metabolismo , Ativador de Plasminogênio Tipo Uroquinase/metabolismo , Aprotinina/farmacologia , Células Cultivadas , Humanos , Inibidores de Metaloproteinases de Matriz , Monócitos/efeitos dos fármacos , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Inibidor 1 de Ativador de Plasminogênio/farmacologia , Inibidores de Serina Proteinase/farmacologia , Ativador de Plasminogênio Tecidual/metabolismo , Ativador de Plasminogênio Tecidual/farmacologia , Ativador de Plasminogênio Tipo Uroquinase/farmacologia , alfa 2-Antiplasmina/farmacologiaRESUMO
In order to identify the optimal target sites for antisense oligonucleotides in the human multiple drug resistance mRNA, the secondary structure of the 5'-terminal part of this mRNA (nucleotides 1-678) was investigated. By using results of probing with ribonucleases T1, ONE and V1 and results of computer simulations, a model of the 5'-region of the PGY1/MDR1 mRNA was built. The molecule is formed by three major domains comprising several hairpins separated by single-stranded fragments. The predicted single-stranded regions of the PGY1/MDR1 mRNA efficiently bind complementary oligonucleotides.