Addition Of PAI-1 Research Articles

The Overall Haemostatic Potential (OHP) Assay Detects Abnormalities in Fibrinolytic Factors

No routine assays are available which quantify abnormalities in the fibrinolytic system. The Overall Haemostatic Potential (OHP) assay is a global assay of fibrin generation and fibrinolysis. Our aim in this study was to assess changes in fibrinolysis measured by the OHP assay in samples with variable levels of the different components of the fibrinolytic system: plasminogen, a2-antiplasmin, PAI-1 and TAFI. Plasmas deficient in these factors were obtained commercially. Samples were spiked with Lys-plasminogen, Glu-plasminogen, PAI-1, TAFI or pooled normal plasma to produce serial increases in these factors. Potato tuberose carboxypeptidase inhibitor (PTCI) was also used to inhibit TAFI in PNP and patient samples. OHP assays were performed utilizing either thrombin (0.03 IU/ml) or tissue factor (5 pM) to trigger fibrin generation, and rt-PA (350 ng/ml) to trigger fibrinolysis. Data analysis involved calculation of Spearman correlations for OHP assay parameters and concentrations of the various fibrinolytic factors tested. The OHP assay demonstrated markedly reduced fibrinolysis in plasminogen deficient plasmas, which normalised when plasminogen levels were increased to the normal range. At elevated Lys-plasminogen levels fibrin generation was impaired. Overall fibrinolytic potential was mildly increased in a2-antiplasmin deficiency. PAI-1 depleted plasma had mildly increased fibrinolysis compared with PNP but with addition of PAI-1, to concentrations of 300 ng/ml or higher, fibrinolysis was markedly impaired. TAFI deficient plasma had reduced fibrin generation but no change in fibrinolysis was seen in spiked samples, even with elevated TAFI levels. Representative hypofibrinolytic patient samples showed normalisation of fibrinolysis with TAFI inhibition by PTCI.The OHP assay shows appropriate changes in OFP with variations in the concentrations of activators and inhibitors of the fibrinolytic system. The assay provides a simple means of quantifying overall fibrinolysis and is sensitive to abnormalities in individual components of the fibrinolytic system. It should be useful for investigating fibrinolytic abnormalities in patients, where no routine assay is currently available.

Quorum sensing in Pseudomonas aeruginosa controls expression of catalase and superoxide dismutase genes and mediates biofilm susceptibility to hydrogen peroxide.

Quorum sensing (QS) governs the production of virulence factors and the architecture and sodium dodecyl sulphate (SDS) resistance of biofilm-grown Pseudomonas aeruginosa. P. aeruginosa QS requires two transcriptional activator proteins known as LasR and RhlR and their cognate autoinducers PAI-1 (N-(3-oxododecanoyl)-L-homoserine lactone) and PAI-2 (N-butyryl-L-homoserine lactone) respectively. This study provides evidence of QS control of genes essential for relieving oxidative stress. Mutants devoid of one or both autoinducers were more sensitive to hydrogen peroxide and phenazine methosulphate, and some PAI mutant strains also demonstrated decreased expression of two superoxide dismutases (SODs), Mn-SOD and Fe-SOD, and the major catalase, KatA. The expression of sodA (encoding Mn-SOD) was particularly dependent on PAI-1, whereas the influence of autoinducers on Fe-SOD and KatA levels was also apparent but not to the degree observed with Mn-SOD. beta-Galactosidase reporter fusion results were in agreement with these findings. Also, the addition of both PAIs to suspensions of the PAI-1/2-deficient double mutant partially restored KatA activity, while the addition of PAI-1 only was sufficient for full restoration of Mn-SOD activity. In biofilm studies, catalase activity in wild-type bacteria was significantly reduced relative to planktonic bacteria; catalase activity in the PAI mutants was reduced even further and consistent with relative differences observed between each strain grown planktonically. While wild-type and mutant biofilms contained less catalase activity, they were more resistant to hydrogen peroxide treatment than their respective planktonic counterparts. Also, while catalase was implicated as an important factor in biofilm resistance to hydrogen peroxide insult, other unknown factors seemed potentially important, as PAI mutant biofilm sensitivity appeared not to be incrementally correlated to catalase levels.