Victor Gurewich

Risk of idiopathic cardiovascular death and rionfatal venous thromboembolism in women using oral contraceptives with differing progestagen components

Concern about the risks of cardiovascular illness in women using combined oral contraceptives (OC) containing the progestagens desogestrel and gestodene prompted two studies of data from the UK General Practice Research Database. We compared the risks of certain cardiovascular illnesses in otherwise healthy women exposed to one of three OCs containing < 35 micrograms oestrogen plus levonorgestrel, desogestrel, or gestodene. In the first study, based on some 470 general practices, there were 15 cases of unexpected idiopathic cardiovascular death among 303,470 women who were current users of one of the study OCs. The estimated incidence rates were 8/184,536 (4.3 per 100,000) woman-years at risk for users of combined OCs containing levonorgestrel, 2/135,567 (1.5 per 100,000) for desogestrel users, and 5/105,201 (4.8 per 100,000) for gestodene users. The relative risk (RR) estimates were 0.4 (95% CI 0.1-2.1) and 1.4 (CI 0.5-4.5) for desogestrel and gestodene, respectively, compared with levonorgestrel. In the second study, derived from some 370 general practices, there were 80 cases of nonfatal venous thromboembolism (VTE) in a cohort of 238,130 otherwise healthy women. The incidence rates of VTE per 100,000 woman-years at risk were 16.1 for levonorgestrel users, 29.3 for desogestrel, and 28.1 for gestodene. The adjusted RR estimates from the cohort analysis were 1.9 (1.1-3.2) and 1.8 (1.0-3.2) for desogestrel and gestodene users, respectively, compared with users of levonorgestrel. In a nested case-control analysis the adjusted matched RR estimates were 2.2 (1.1-4.4) and 2.1 (1.0-4.4) for desogestrel and gestodene users, respectively, compared with users of levonorgestrel. The excess risk for nonfatal VTE associated with the new generation of combined OCs containing low-dose oestrogen and the progestagens desogestrel or gestodene compared with levonorgestrel is estimated to be 16 per 100,000 woman-years.

Effective and fibrin-specific clot lysis by a zymogen precursor form of urokinase (pro-urokinase). A study in vitro and in two animal species.

A single-chain 55,000-mol wt form of urokinase (UK), similar to that previously isolated from urine, was purified from a transformed kidney cell culture medium and characterized; and its fibrinolytic properties were evaluated. The preparation immunoprecipitated with UK antiserum, had a low intrinsic amidolytic activity that was 0.1% of its active derivative, and resisted diisopropyl fluorophosphate treatment and inactivation by plasma inhibitors. The single-chain UK was therefore designated pro-UK. In the presence of plasmin and during clot lysis, activation by conversion to two-chain, 55,000-mol wt UK (TC-UK) was demonstrated. This did not occur during blood clotting nor on incubation with purified thrombin. Clot lysis in plasma consistently occurred in 2-5 h with 50-100 IU per ml of pro-UK, whereas comparable lysis was inconsistently achieved by 500-1,000 IU of UK. Pro-UK, in sharp contrast to UK, caused no fibrinogen degradation at fibrinolytic concentrations. In the absence of a clot, pro-UK in plasma was stable for more than 2 d. When a clot was added after incubation (37 degrees C) for 50 h, activation to full lytic activity took place. The findings in vivo were comparable but the rapid clearance of pro-UK required that it be given by a constant infusion despite its plasma stability. In rabbits, a UK-resistant species, pro-UK was significantly (P less than 0.001) more efficacious than TC-UK but neither induced significant fibrinogen degradation. In dogs, a more sensitive species, the high specificity of thrombolysis by pro-UK contrasted with the defibrinogenation and uncontrollable bleeding that accompanied thrombolysis by UK. It was concluded that clot lysis by pro-UK is more effective and specific than UK. The advantage of pro-UK is in the limitation of its activation to the site of a clot. This can be explained by an activation mechanism that is dependent, under physiological conditions, on fibrin-stabilized plasmin.

Thrombolytic Therapy in Thrombosis: A National Institutes of Health Consensus Development Conference

Excerpt For over three decades, the primary method of therapy used by almost all physicians for the management of acute deep-vein thrombosis and pulmonary embolism has been anticoagulation. This fo...

Purification and partial characterization of a single-chain high-molecular-weight form of urokinase from human urine

Affinity chromatography on fibrin/celite, a previously described technique for isolating plasminogen activators with a high fibrin affinity (S. Husain, B. Lipinski, and V. Gurewich, 1981, Proc. Nat. Acad. Sci. USA 78, 4265-4269), was performed on freshly voided urine. About 10-25% of the plasminogen activator activity present was adsorbed whereas less than 1% was adsorbed when urine was stored at room temperature for 12-24 h. The adsorbed activator was isolated by elution with arginine (0.1 M) and further purified by gel filtration. This plasminogen activator was similar in molecular weight (56,000) to the known high-molecular-weight form of urokinase but differed from it by its binding affinity for fibrin/celite, single-chain subunit structure, lower specific activity (approximately 20,000 IU/mg), and slower reaction with diisopropylfluorophosphate. The enzymatic activity of the purified activator was quenched by anti-urokinase antibody and therefore established it to be a new form of urokinase. The relationship of this high affinity, single-chain urokinase to the previously described forms of urokinase is unclear.

Complementary modes of action of tissue-type plasminogen activator and pro-urokinase by which their synergistic effect on clot lysis may be explained.

Tissue plasminogen activator (t-PA) and/or pro-urokinase (pro-UK) induced lysis of standard 125I-fibrin clots suspended in plasma was studied. Doses were kept below the concentration at which a nonspecific effect was seen, i.e., where fibrinogenolysis and major plasminogen consumption were observed. Small amounts of t-PA potentiated clot lysis by pro-UK by attenuating the lag phase characteristic of pro-UK, and causing a much earlier transition to the rapid phase of lysis. Similar promotion of the fibrinolytic effect of pro-UK was obtained when clots were pretreated with UK or with a little plasmin (less than 1% clot lysis). Promotion by plasmin was nullified by a subsequent treatment of the clot with carboxypeptidase B, indicating that the plasmin effect was related to the exposure of carboxy terminal lysine residues on fibrin. These lysine termini, absent in undegraded fibrin, are known to be essential for the high affinity binding of plasminogen to fibrin. In contrast, clot lysis by t-PA was unaffected by plasmin pretreatment and little affected by carboxypeptidase B treatment of the fibrin substrate. Therefore, plasminogen bound to lysine termini on fibrin, although found to be essential for pro-UK, did not appear to serve as a substrate for t-PA. Selective activation of fibrin bound plasminogen has been attributed to the conformational change in Glu-plasminogen that occurs as a result of binding. The present findings suggest that this conformational change occurs when plasminogen is bound to a terminal lysine but not to an internal lysine. Plasminogen bound to the latter site on fibrin was activated by t-PA and therefore is involved in the ternary complex. This initiates lysis of the undegraded clot and exposes the plasminogen binding sites required by pro-UK. By their complementary activation of fibrin bound plasminogen, t-PA followed by pro-UK induces efficient and synergistic fibrinolysis, whereas each is relatively inefficient when used alone.

The Risk of Myocardial Infarction Associated with Antihypertensive Drug Treatment in Persons with Uncomplicated Essential Hypertension

We conducted a case‐control study based on computer‐recorded information accrued in the United Kingdom General Practice Research Database to assess and compare the relation between different antihypertensive drug therapies and myocardial infarction in patients with no known clinical or laboratory risk factors for myocardial infarction other than hypertension. Cases were treated hypertensive patients with no other known risk factors who developed a first acute myocardial infarction between January 1, 1993, and October 31, 1994. They were ascertained from a review of the clinical record together with a questionnaire filled out by the attending general practitioner. Controls were matched to each case for age, sex, general practice, and index date. Antihypertensive therapy was derived from the computerized patient record. The study consisted of 210 cases and 793 controls. Compared with users of β‐blockers alone, the adjusted relative risk (RR) estimates for all other treatment regimens were close to 1.0. A comparison of users of calcium channel blockers alone with users of β‐blockers alone yielded a RR estimate of 0.9 (95% CI 0.5, 1.7). We conclude that the risk of acute myocardial infarction in otherwise healthy, treated hypertensive patients is not materially associated with the particular drug they receive.

Comparative study of the efficacy and specificity of tissue plasminogen activator and pro-urokinase: demonstration of synergism and of different thresholds of non-selectivity

Clot lysis and non-specific plasminogen activation in human plasma by tissue tissue plasminogen activator (t-PA) and/or pro-urokinase (pro-UK) were studied. The fibrinolytic activity of pro-UK was expressed as latent units, i.e. measured after activation with plasmin on a fibrin plate against the reference standard. The t-PA unitage was assigned on a weight basis of a similar equivalence of 100,000 IU/mg. To simplify comparison, both activators were expressed in IU (1 IU = approximately 10 ng). At low concentration (1-50 IU/ml), t-PA induced more effective and more linear clot lysis, whereas pro-UK induced lysis was preceded by a lag phase. The two activators were equivalently effective at higher concentrations and saturated at the same lysis rate. Clots made from platelet rich plasma or whole blood were more responsive to lysis by pro-UK but not t-PA than corresponding platelet poor clots. At very low concentrations (2.5-5 IU/ml) of t-PA combined with moderate concentrations (25-50 IU/ml) of pro-UK, a synergistic effect on clot lysis, which was fibrin-specific, was observed. Plasminogen and fibrinogen and the appearance of plasmin-inhibitor complexes in plasma were measured after incubation with either activator with and without a clot present. Non-specific plasminogen activation occurred above a certain concentration of either activator but was found at lower concentrations of t-PA than pro-UK. In the absence of a clot, plasmin generation occurred with t-PA at about 30% of the concentration at which pro-UK induced a corresponding effect. It is concluded that there are important differences in the fibrinolytic and clot selective properties of t-PA and pro-UK, and that some of these properties may be complementary resulting in a fibrin specific, synergistic fibrinolytic effect.

Humoral factors in massive pulmonary embolism: An experimental study

Abstract Alterations in central venous pressure (CVP) were measured in response to embolization with standardized autologous venous thrombi. Platelets were counted before and after embolization and their accretion on the embolus was determined microscopically at autopsy. The rise in CVP could be blocked by heparin but only at sufficient doses to prevent the platelet accretion, or by aging the thrombus prior to embolization. conversely, the postembolic rise in CVP and mortality rates were augmented by an inhibitor of monoamine oxidase whose effect in turn was nullified by a serotonin antagonist. It is concluded that the release of biogenic amines from platelets, triggered by thrombin on the embolus, plays a significant role in the animals response to thromboemboli. Both in vivo and in vitro evidence is presented demonstrating that the dose requirements of heparin necessary to inhibit these events is greater than that generally used in treating pulmonary embolism. Certain clinical observations relating to these experimental findings are reviewed and the therapeutic implications discussed.

Bronchoconstriction in the Presence of Pulmonary Embolism

In a group of seven patients with pulmonary embolism, evidence of bronchoconstriction was found. Heparin appeared to relieve partially the observed airway obstruction. A possible common mechanism for the initiation of intravascular coagulation and the production of bronchoconstriction was considered. When other causes can be eliminated, the detection of bronchoconstriction may serve as a useful index for the early recognition of pulmonary emboli.

The blockage of the high-affinity lysine binding sites of plasminogen by EACA significantly inhibits prourokinase-induced plasminogen activation

Prourokinase-induced plasminogen activation is complex and involves three distinct reactions: (1) plasminogen activation by the intrinsic activity of prourokinase; (2) prourokinase activation by plasmin; (3) plasminogen activation by urokinase. To further understand some of the mechanisms involved, the effects of epsilon-aminocaproic acid (EACA), a lysine analogue, on these reactions were studied. At a low range of concentrations (10-50 microM), EACA significantly inhibited prourokinase-induced (Glu-/Lys-) plasminogen activation, prourokinase activation by Lys-plasmin, and (Glu-/Lys-) plasminogen activation by urokinase. However, no inhibition of plasminogen activation by Ala158-prourokinase (a plasmin-resistant mutant) occurred. Therefore, the overall inhibition of EACA on prourokinase-induced plasminogen activation was mainly due to inhibition of reactions 2 and 3, by blocking the high-affinity lysine binding interaction between plasmin and prourokinase, as well as between plasminogen and urokinase. These findings were consistent with kinetic studies which suggested that binding of kringle 1-4 of plasmin to the N-terminal region of prourokinase significantly promotes prourokinase activation, and that binding of kringle 1-4 of plasminogen to the C-terminal lysine158 of urokinase significantly promotes plasminogen activation. In conclusion, EACA was found to inhibit, rather than promote, prourokinase-induced plasminogen activation due to its blocking of the high-affinity lysine binding sites on plasmin(ogen).