Heparin-Induced Thrombocytopenia: Prevalence in a Large Cohort of Patients and Confirmed Role of PF4/Heparin Complex as the Main Antigen for Antibodies

Abstract

We performed a retrospective study on the prevalence of heparin-induced thrombocytopenia (HIT) in 233 patients receiving hog mucosa heparin therapy. Of these, 82 patients received s.c. calcium heparin, 130 patient received unfractionated (UF) i.v. heparin, and 21 patients received low molecular weight heparins (LMWH). An additional four patients, referred to our consultation and diagnosed by us as having clinically active type II HIT (HIT-II) were also studied. The mean platelet count of the 233 patients receiving heparin showed a significant decrease after 2 days of heparin treatment and a following significant increase 6 days later (basal: 257 &plusmn; 147 x 10 9 platelets/L; day 2: 239 &plusmn; 122, p < 0.0002; day 6: 286 &plusmn; 119, p < 0.004). Of the 212 patients receiving UF heparin, 13 (6%) fulfilled the criteria for HIT-II: seven of these had received i.v. heparin (mean daily dose 26,600 &plusmn; 4,082 IU & p l u s m n ; SD) and six had received s.c. heparin (mean daily dose 21,428:t 6,900 IU). Their mean basal platelet count was 226 & p l u s m n ; 100 SD &times; 10 9 platelets/L and the nadir during heparin treatment was 78 &plusmn; 39 x 10 9 platelets/L. Thrombotic complications occurred in four (30.7%) of the 13 patients with HIT-II. Since the immunological mechanism has been demonstrated for HIT-II and since platelet factor 4 (PF4) was identified as the co-factor for the binding of heparin-related antibodies, we set up our own enzyme-linked immunosorbent assay (ELISA) for testing antibodies against PF4/heparin complex bound through electrostatic bridges to the solid phase. The highest binding capacity of HIT-related IgG to the multimolecular complex was obtained at 20 &mu;g/ml for PF4 and 3 &mu;g/ml for heparin, corresponding to 250 ng of PF4 and 42 ng of heparin in each microtiter well. Such binding was inhibited in a dose-dependent manner by increasing amounts of heparin, protamine hydrochloride, and a monoclonal antibody anti-human PF4 clone 1OB2. We observed that HIT-related antibodies bound also to PF4/LMWH complexes but the optimal PF4/glycosaminoglycan ratio appeared more critical for LMWH (enoxaparin, fraxiparin, and pamaparin) than for UF heparin. Sera from eight patients with HIT-II were tested by PF4/heparin ELISA; six of these had IgG against the complex PF4/heparin and three also had IgM. The persistence of HIT-related antibodies was investigated in three patients: in one such antibodies were still detectable 3 years after the acute episode, while in the other two, they disappeared after 6 months and 1 year, respectively.

Heparin‐induced thrombocytopenia and thrombosis: a prospective analysis of the incidence in patients with heart and cerebrovascular diseases

“…The clinical significance of heparin-dependent antibodies is unknown, as is their incidence and the period during which they are detectable in plasma of patients without clinical signs of HITT. According to one report, the antibodies disappeared from the plasma after 2 months in two patients, whereas in another patient the antibody was still demonstrable 3 years after the occurrence of heparin-induced thrombocytopenia (Fabris et al, 1995). However, long-term existence of antibodies after previous heparin treatment cannot explain the presence of antibodies in our patient group, since all day 0 test samples were negative.…”

Risk factors for thrombosis in patients with immune mediated heparin‐induced thrombocytopenia

“…All patients with a platelet count drop concurrently underwent search for antiheparin/PF4 antibodies. All HIT patients met both the following diagnostic criteria: (1) thrombocytopenia, defined as a decrease in platelet count >30% 5 or more days after beginning heparin or in a shorter time in case of a previous exposure to heparin, and (2) a positive test for antiheparin/PF4 antibodies assayed by `inhouse ELISA' and by a commercially available polyanion‐PF4‐ELISA (GTI, WI, USA) [10, 11]. Optical Density (OD) was used as an index of antibody titre.…”