Revisiting the effect of cholesteryl sulfate on clotting and fibrinolysis: Inhibition of human thrombin and other human blood proteases
Research article published in Heliyon (2024)
Abstract
Cholesteryl sulfate (CS) was quantitatively synthesized by microwave-assisted sulfonation of cholesterol followed by sodium exchange chromatography. In vitro effects of CS on human thrombin and other serine proteases of the coagulation and fibrinolysis processes were investigated using a series of biochemical and biophysical techniques. CS was found to inhibit thrombin with an IC50 value of 140.8 ± 21.8 μM at pH 7.4 and 25 ○C. Michaelis-Menten kinetics indicated that thrombin inhibition by CS is non-competitive (allosteric) in nature. Fluorescence-based binding studies indicated that CS binds to thrombin with a KD value of 180.9 ± 18.9 μM. Given the lack of competition with heparins and a hirudin peptide in competitive inhibition assays, it appears that CS does not bind to thrombin's exosites 1 or 2 and it rather recognizes a different allosteric exosite. CS was found to partially inhibit thrombin-mediated fibrinogen activation with an IC50 value of 175.5 ± 17.5 μM and efficacy of ∼26.0 ± 6.6%. Likewise, CS selectively doubled the activated partial thromboplastin time with EC2x of 521 μM. Interestingly, CS was found to also inhibit factors Xa and XIa as well as plasmin with IC50 values of ∼85-250 μM and efficacy of 94-100%. Nevertheless, CS most potently inhibited factor XIIa with an IC50 Value of ∼17 μM and efficacy of 60%. Surprisingly, CS did not inhibit factor IXa. These results encourage further in vitro and in vivo investigation of CS to better understand its (patho-) physiological roles in coagulation and hemostasis.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Zusammenfassung
Cholesteryl sulfate (CS) was quantitatively synthesized by microwave-assisted sulfonation of cholesterol followed by sodium exchange chromatography.
Warum dies für die Hirudotherapie relevant ist
Diese In-vitro-Studie synthetisierte Cholesterylsulfat (CS) und charakterisierte dessen hemmende Wirkungen auf menschliches Thrombin und andere Gerinnungs-/Fibrinolyseproteasen. CS hemmte Thrombin (IC50 ~140,8 μM) durch nicht-kompetitive (allosterische) Bindung, hemmte teilweise die Fibrinogenaktivierung und verlängerte die aPTT; es hemmte außerdem die Faktoren Xa, XIa, XIIa und Plasmin, jedoch nicht Faktor IXa. Kompetitive Hemmtests zeigten keine Konkurrenz mit Heparinen oder einem Hirudin-Peptid, was darauf hinweist, dass CS an eine distinkte allosterische Exosite auf Thrombin bindet. Für das ASH-Gebiet ist die Verbindung indirekt: Ein Hirudin-Peptid wird als Werkzeug verwendet, um die Bindung an die Thrombin-Exosite-1 zu kartieren, was bestätigt, dass CS an anderer Stelle wirkt, aber Hirudin ist nicht das untersuchte Agens. Die Studie betrifft ein synthetisches Sterolsulfat, nicht die Blutegeltherapie oder das Blutegelsekretom; die Relevanz beschränkt sich darauf, mechanistischen Kontext zur Thrombinhemmung jenseits des Hirudin-Wirkmechanismus zu liefern.
Zitation
Revisiting the effect of cholesteryl sulfate on clotting and fibrinolysis: Inhibition of human thrombin and other human blood proteases
Al-Horani RA · Heliyon, 2024
Verwandter klinischer Kontext
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