New compound targets bloodclots without increased bleeding risk ubcnews NatureComms
, MPI 8 demonstrates great potential as a therapeutic, by providing nontoxic thrombo-protection without the bleeding risk. The data presented support the hypothesis that minimized charge density paired with specifically smart binding groups result in targeted polyP inhibition with minimal interference with hemostasis, a significant improvement over current polyP inhibitors. These findings lay the groundwork for a safer and effective antithrombotic.
A thrombin generation assay was carried out at 37 °C by measuring the fluorescence intensity upon cleavage of the fluorogenic thrombin substrate, Z-Gly-Gly-Arg-AMC. Commercially available pooled normal platelet poor plasma from George King Bio-Medical, USA was mixed 1:1 with HBS . Phosphatidylcholine : phosphatidylserine liposomes were added to obtain a final concentration of 20 μM. Serial dilutions of MPI candidates and UHRA were prepared fresh for these experiments.
Thrombin calibrator was added to each well following the manufacturer’s instructions and the thrombin generation assay was initiated by the addition of fluorogenic substrate . Substrate hydrolysis was monitored on a Thrombinograph™ plate reader from Diagnostica Stago. The fluorescence intensity was recorded at 37 °C every 30 s over a period of 1.5 h and analyzed using Thrombinoscope™ software from Diagnostica Stago.
A mouse model was used to assess the effect of MPI on bleeding in the absence of added polyP or any anticoagulants. Heparin was used as a positive control and saline was used as a negative control. Eight-to-ten-week-old C57/BL6 mice were obtained from The Jackson Laboratories , and the experimental protocol was approved by the International Animal Care and Use Committee at the University of Michigan . Light cycles in the animal holding rooms are set for 12 h on and 12 h off.
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