2. Antithrombin III

3. Protein C system Thrombin + thrombomodulin Protein C aProtein C + protein S Cleaves Factor Va and VIIIa

4. Thrombin + thrombomodulin Thrombin-activatable fibrinolysis inhibitor (TAFI) Removes the terminal lysine on the fibrin molecule, renders the clot more susceptible to lysis by plasmin

5. Degradation of the Fibrin Clot Natural part of hemostasis Fibrin fibrils in the clot are dense, dissolution occurs at “coiled coils” to form oligomers which are digested into smaller units Highly regulated in terms of gene expression Plasmin Tissue plasminogen activator (t-PA) urokinase streptokinase

6. Plasminogenesis

7. Plasmin Serine protease formed from plasminogen Plasminogen found in plasma and extracellular space and has high affinity for fibrin Inactive and requires activation to plasmin by urokinase and t-PA Formation of plasmin is highly regulated Cleaves fibrin into degradation products. Circulating plasmin is inhibited by α2-antiplasmin. Forms complex with plasmin and prevents it from binding to fibrin

8. Tissue Plasminogen Activator Serine protease released from ECs in response to injury (in response to thrombin and some vasoactive peptides) Chiefly responsible for conversion of plasminogen to plasmin Secretion highly regulated at transcriptional level Major binding is to fibrin clots and extracellular matrix Highly specific ( site of clot formation ) Inhibited by plasminogen activator inhibitor-1 (PAI-1) Recombinant protein used clinically to inhibit thrombosis

9. Urokinase Serine protease secreted as pro-urokinase Produced by endothelial cells, fibroblasts, and monocytes/macrophages Potent plasminogen activator but non-specific secreted as one chain: fragments in urine generated by plasmin Functions to degrade ECM, enabling cells to migrate Half life of only 7 minutes Inhibited by plasminogen activator-2 (PAI-2) synthesis tightly regulated by cytokines and inflammatory mediators Used clinically (e.g. Abbokinase)

10. Streptokinase Protein isolated from certain types of streptococci bacteria Potent plasminogen activator Less selective than t-PA Can result in circulating plasmin Can produce degradation of fibrinogen as well as fibrin May result in formation of plasmin in excess of that inhibited by α2-antiplasmin and result in bleeding

11. BECAUSE OF THE COMPLEX NATURE OF HEMOSTASIS, POTENTIAL INTERFERENCE IN THE PROCESS CAN OCCUR AT MANY LEVELS.

12. HEMOSTATIC DEFECTS CONGENITAL Coagulation Factor deficiency Hemophilia von Willebrand’s disease Factor XI,II, V, X, XIII deficiency Platelet function defects Glanzmann’s thrombastenia Bernard-Soulier syndrome Storage pool disease

13. HEMOSTATIC DEFECTS Major surface protein abnormality Glanzmann’s thrombastenia Glycoprotein IIb / IIIa Absence of platelet aggregation Bernard-Soulier syndrome Glycoprotein Ib / IX / V Absence of platelet adhesion

14. HEMOSTATIC DEFECTS ACQUIRED Platelet abnormalities quantitative defects failure of production Bone marrow disorder shortened survival ITP / DIC Sequestration hypersplenism

15. Conditions of Excess Bleeding Hemophilia ( inherited sex-linked rec. ) Hemophilia A (classic)- deficiency of Factor VIII (85%) Hemophilia B – deficiency of Factor IX (15%) Spontaneous bleeding Joints frequently, crippling arthropathies Retroperitoneal hematoma Gastrointestinal/ genitourinary NORMAL PLATELET FUNCTION

16. Conditions of Excess Bleeding von Willebrand’s disease Autosomal dominant Low level of vWF CARRIER FOR FACTOR VIII NORMAL PLATELET ADHESION ABNORMAL PLATELET FUNCTION Easy bruising and mucosal bleeding Menorrhagia is common

17. Conditions of Excess Bleeding Thrombocytopenia Most common abnormality of hemostasis in surgical patients Platelets fall from normal 150-400,000 to < 100,000/µl Massive blood loss Heparin induced Impaired platelet function Vit b12 / folic acid def ITP hypersplenism

18. Conditions of Excess Bleeding Coagulopathy of liver disease Vitamin K deficiency Required by liver for formation of prothrombin and Factors VII, IX, X and protein C Leads to serious bleeding tendencies

19. Conditions of Clotting Dysfunction Thromboembolic conditions Can result when endothelial surface is compromised or BF is slow Associated with atherosclerosis, infection, or trauma Femoral venous thrombosis When BF is slow (bed ridden, prolonged sitting) Clot grows and large piece can break off Can pass through right side of heart to lodge in pulmonary arteries (pulmonary embolism) If large enough can be fatal

20. COAGULANTS Thrombosis Inappropriate activation of hemostatic mechanism; Venous –associated with stasis of blood with small platelet component Arterial – associated with artherosclerosis with large platelet component

21. COAGULANTS Vitamin K Essential for the formation of clotting factors (II, VII, IX and X) Given orally or thru IV (Natural Vit K requires Bile-acid while Menadiol Na Phosphate does not but takes longer to act) USE – bleeding 2o to oral anticoagulants, babies, Vit. K deficiencies

22. COAGULANTS Antifibrinolytic Agent Tranexamic acid (Inhibit plasminogen activation),for conditions with bleeding or risk of bleeding,life-threatening bleeding following thrombolytic drug administration and hereditary angioedema. Aprotinin-inhibits proteolytic enzymes, used for hyperplasminemia due to fibrinolytic overdose & during cardiac surgery

23. Conditions of Clotting Dysfunction Disseminated intravascular coagulation Clotting activated in widespread areas Often from severe trauma or shock (endotoxin) Plugging of vessels limits oxygen delivery lethal in ~85% of the cases

24. ANTICOAGULATION AND BLEEDING

25. Prevention of Blood Clotting Critical for clots to dissolve or not form when not needed Endothelial cell surface- Most important factor – integrity prevents contact activation of intrinsic clotting system by collagen Charged EC glycocalyx repels platelets and clotting factors Thrombomodulin-EC membrane protein, binds thrombin to slow clotting process. Thrombomodulin-thrombin complex activates a protein C which inactivates Factors V and VIII.

26. Removal of Thrombin (blocks common pathway) Fibrin – absorbs 85-90% of thrombin by absorption into fibers Anti-thrombin III – binds remaining thrombin Prevention of Blood Clotting

27. Vitamin K cycle (post-translational carboxylation) Factors VII, IX, X and prothrombin

29. Anticoagulants for Clinical Use Heparin Coumarins Aspirin Cyclooxygenase inhibitor Prevents formation of thromboxane A2 and activation of platelets Calcium-deionizing agents Sodium, ammonium and potassium citrate combines with calcium in blood Several factors require calcium for activation Used in test tubes to prevent clotting

30. Antithrombin III

31. ANTICOAGULANT Heparin -binds to antithrombin III Combines with anti-thrombin III and increases activity 100-1000X Immediate effect ,half life of 60-90min Monitor by aPTT

32. ANTICOAGULANT Low molecular weight heparin- 4-6 kDa Selective Xa inhibitor More favorable antithrombotic effect,less bleeding,highly predictable bioavailability Longer half life

33. ANTICOAGULANTS Coumarin derivatives- warfarin,acenocoumarol,phenprocoumon Block coagulation factors(2,7,9,10) Warfarin decreases formation of Factors VII, IX and X by the liver Competes with vitamin K for reactive sites Restoration after coumarin tx- 3-5 days Prothrombin time INR- corrects the differences of the various thromboplastin activity. INR=1 no anticoagulation

34. ANTICOAGULATION Antiplatelet Drugs Aspirin Dipyridamole Thienopyridine Derivatives Glycoprotein IIB/IIIA receptor antagonists Other antiplatelet drugs

35. ANTICOAGULATION Antiplatelet Drugs Aspirin Inhibits cyclooxygenase irreversibly Inhibits TXA2 Main drug – acute MI, High-risk for MI, after coronary artery Bypass, after angioplasty, unstable coronary syndromes,TIA and AF if oral anticoagulant is contraindicated

36. ANTICOAGULATION Antiplatelet Drugs Dipyridamole Phosphodiesterase inhibitor Additive effect to aspirin Less effective than aspirin Headache but no excess risk of bleeding

37. ANTICOAGULATION Antiplatelet Drugs Thienopyridine Derivatives Inhibits ADP-dependent aggregation Orally given, additive with aspirin Ticlopidine slow onset, unwanted effects-blood dyscrasias (neutropenia) Clopidogrel – same as Ticlopidine except for neutropenia

38. ANTICOAGULATION Antiplatelet Drugs Glycoprotein IIB/IIIA Receptor Antagonists Abciximab – for angioplasty patient as adjunct to heparin and aspirin, reduces restenosis, Tirofiban – an oligopeptide Abciximab - Risk of bleeding & immunogenicity limits its use They inhibit diverse agonists (e.g. ADP, TXA2 etc. )

39. ANTICOAGULATION Thrombolytic agents- plasminogen activators. Recombinant endogenous plasminogen activators(recombinant tPA) Streptokinase- exogenous source bleeding

40. ANTICOAGULATION Fibrinolytic Drugs Streptokinase Protein extracted from cultures of streptococci Activates plasminogen, given IV Additive with aspirin Action blocked by antistreptococcal antibodies Allow 1 year before it can be used again

41. ANTICOAGULATION Fibrinolytic Drugs Alteplase, Duteplase and Reteplase Recombinant tPA (plasminogen activator) ‘Clot-selective’ – fibrin bound plasminogen Not antigenic-substitute for patients w/ ab for streptokinase Reteplase –longer half-life, bolus admn.simple

42. Tests of hemostasis Primary deficiency/ defect in one component Pharmacologic therapy Anticoagulant/ antiplatelet Comorbid condition Thrombocytopenia Sepsis/ hepatic disease

43. Tests of hemostasis Careful review of patient’s clinical history most important/ initial approach Drug use Basic laboratory tests Platelet count Prothrombin time ( PT ) or INR Activated partial thromboplastin time ( aPTT ) Bleeding time ( BT ) and clotting time ( CT )

44. Evaluation of surgical patient Patients history Questions to ask Prolonged bleeding or swelling after biting the lip or tongue Bruises without apparent injury Prolonged bleeding after dental extraction Excessive menstrual bleeding Bleeding problems associated with major and minor operations Medical problems receiving a physician’s attention within the past 5 years Medications including aspirin or remedies for headache taken within the past 10 days Relative with a bleeding problem

45. Four levels of concern Level I Hx(-); Sx (m) no test Level II Hx(-); Sx (M) platelet ct PTT Level III Hx (suggestive) PT, PTT, BT, CT, pltelet ct Level IV Hx(+) Hematologic consultation

46. Platelet dysfunction Normal 150,000 to 400,000 /ul Clinical signs of thrombocytopenia < 100,000 / ul Increased bleeding complications in major surgical procedures < 50,00 / ul Increased bleeding complications in minor surgical procedures < 20,000 / ul SPONTANEOUS HEMORRHAGE

47. Platelet dysfunction Bleeding time ( BT ) Evaluate platelet , vWD and vascular dysfunction Several methods Ivy test = 7mins Clotting time ( CT ) … + Aspirin use

48. Clotting dysfunction PT reagent contains thromboplastin and calcium + plasma = clot PT measures Factors I, II, V, X and VII Factor VII extrinsic pathway Abnormal coagulation due to Vit K deficiency warfarin / coumadin therapy

49. Clotting dysfunction Variations in thromboplastin activity from different sources PT value is adjusted = INR ISI ( international sensitivity index ) Value for each batch of thromboplastin Optimal reagent has as ISI of 1.3 to 1.5 INR = ( PTpatient / PTnormal )ISI INR- corrects the differences of the various thromboplastin activity. INR = 1.0 is NORMAL

50. Clotting dysfunction aPTT reagent contains Phospholipid substitutes, activator and calcium + plasma = clot aPTT measures Factors I, II and V of the common pathway Factors VIII, IX, X and XII intrinsic pathway Heparin therapy

51. HEMOSTASIS RENE PSA MENDOZA, MD, MHSA Associate Professor, Department of Surgery FEU-NRMF Institute of Medicine