A role for von Willebrand factor proline residues 702-704 in ristocetin-mediated binding to platelet glycoprotein Ib

Mutant domains of von Willebrand factor (vWF) were constructed to determine the effects of altering net charge, and presumably conformation, within a peptide sequence (residues 694-708) previously shown to be involved in the platelet receptor glycoprotein (GP) Ib binding function of vWF. Non-conservative substitutions replaced a triplet of proline residues (proline702-704) with either a triplet of arginine (positively-charged) or aspartic acid (negatively-charged) residues. After establishing stable CHO cell transformants, we observed the secretion of covalently-linked dimeric molecules analogous to a domain with native sequence. Functional assays using immunopurified molecules revealed that the ristocetin-dependent binding to GP Ib was abolished with both charge mutants. However, in the absence of disulfide-bond dependent conformation both mutant molecules and the molecule with native sequence interacted with GP Ib. The results demonstrate that vWF proline702-704 are important for the ristocetin-mediated interaction between vWF and GP Ib, but are not essential residues of the GP Ib binding site within vWF.     

Ins405AsnPro mutation in the von Willebrand factor propeptide in recessive type 2A (IIC) von Willebrand's disease

The molecular defects of the von Willebrand factor (vWF) have been studied in the patient in whom the von Willebrand disease phenotype IIC was originally described. A six nucleotide insert, AATCCC, was found in exon 11 of the vWF gene, predicting the insertion of the amino acids asparagine and proline between phenylalanine 404 and threonine 405 of the vWF propeptide. The mutation was present in one allele. Analysis of amplification products derived from platelet vWF mRNA showed the other allele to be silent. The patient is thus a compound heterozygote for a null allele and the IIC allele, in accord with the recessive mode of inheritance of the IIC phenotype. Family studies indicated the IIC mutation to have occurred de novo, possibly as a result of a duplication event. In vitro mutagenesis and expression in COS-7 cells confirmed the detrimental effect of the mutation on vWF multimer assembly. Taken together with those of earlier studies the present findings suggest that the IIC phenotype may well be exclusively caused by mutations which result in changes of the amino acid sequence in certain regions of the vWF propeptide. Although in the recently revised classification of von Willebrand's disease variants, the IIC type is included in the 2A category, obviously it constitutes a very distinct subtype.     

Shear-dependent rolling on von Willebrand factor of mammalian cells expressing the platelet glycoprotein Ib-IX-V complex

Mural thrombi form on exposed arterial subendothelium by a two-step process of platelet adhesion and aggregation. At high shear stresses such as are found in stenotic arteries, both steps are mediated by von Willebrand factor (vWF). Platelets initially adhere on vWF affixed to the subendothelial matrix through the glycoprotein (GP) Ib-IX-V complex. To examine the role of the GP Ib-IX-V complex under dynamic conditions, we modeled initial platelet adhesion at shear stresses ranging from 2 to 40 dyn/cm2 using vWF-coated glass slides, mammalian cells expressing full or partial GP Ib-IX-V complexes, and a parallel plate flow chamber with phase contrast video microscopy and digital image processing. Mammalian cells expressing the full complex tethered and rolled on the vWF substrate, whereas control cells did not. The rolling was completely inhibited by the monoclonal GP Ib antibody, AK2, or the vWF antibody, 5D2, both shown previously to block vWF-dependent platelet aggregation. Other GP Ib antibodies, WM23 and SZ2, did not significantly change the number or mean velocity of rolling cells. At low levels of GP Ib surface expression, cells expressing the full complex rolled slower than cells expressing the complex without GP V, indicating that GP V strengthens the interactions with the vWF surface under these conditions. Preshearing vWF for 5 minutes at 40 dyn/cm2 immediately before introducing cells into the chamber did not significantly change the number or the mean velocity of rolling cells. Inhibiting sulfation of the tyrosine residues within the GP Ib subunit reduced the number but did not change the mean velocity of the rolling cells. Our results indicate that, under the conditions of these experiments, bonds between vWF and GP Ib constantly form and break under fluid shear stress. Additionally, our results suggest that GP Ib-IX-V complexes behave like selectin receptors in their ability to mediate smooth rolling while cells maintain continuous surface contact. Such a mechanism, in vivo, would allow platelets to slow down and eventually arrest on the blood vessel wall. The system described provides a valuable approach for investigating the structure-function relationship of individual receptors and ligands in the process of platelet adhesion and thrombosis.     

Polymorphisms of platelet membrane glycoprotein Ib associated with arterial thrombotic disease

Platelet membrane glycoprotein Ibalpha (GPIbalpha) is a major receptor for von Willebrand factor and thrombin, which plays a key role in the initial development of thrombi. Two polymorphisms (HPA-2 and VNTR) that affect phenotype have been described in GPIbalpha. The relevance of these polymorphisms to thrombotic disease was investigated by genotypic identification in three case-control studies: 104 case patients with acute cerebrovascular disease (CVD), 101 case patients with acute coronary heart disease (CHD), 95 patients with deep venous thrombosis (DVT), and one control age-, sex-, and race-matched for each case patient. Results show that the C/B genotype of the VNTR and the HPA-2b polymorphisms of GPIbalpha are strongly associated with increased risk of coronary heart disease and cerebral vascular disease but not with deep vein thrombosis. These two polymorphisms of GPIbalpha may represent newly identified risk factors for arterial thrombotic disease, but not for venous thrombosis.     

Conformational changes in the A1 domain of von Willebrand factor modulating the interaction with platelet glycoprotein Ibalpha

The interaction between von Willebrand factor (vWF) A1 domain and platelet glycoprotein Ib alpha occurs in the presence of high shear stress or when vWF becomes immobilized onto a surface but not appreciably in the normal circulation. To investigate the structural properties regulating A1 domain function, we have used recombinant fragments prepared either in cyclic form with oxidized Cys509-Cys695 disulfide bond or reduced and alkylated. Interaction with glycoprotein Ibalpha was assessed by testing inhibition of monoclonal antibody LJ-Ib1 binding to platelets and inhibition of shear-induced platelet aggregation mediated by native vWF. Fragments exposed to pH between 2.5 and 3.5 adopted the molten globule conformation with loosened tertiary structure intermediate between native and completely unordered state. Maximal receptor binding activity was observed when fragments kept at acidic pH, particularly after reduction of the Cys509-Cys695 disulfide bond, were subjected to quick refolding by rapid pH increase. In contrast, slow refolding by incremental pH change over several hours resulted in at least 20-fold lower activity. A specific single point mutation (I546V) resulted in enhanced receptor binding, whereas another mutation (S561G) caused markedly reduced binding. These results provide experimental evidence that conformational transitions can modulate function of the vWF A1 domain in solution.     

von Willebrand factor contained in factor VIII concentrates of different purities supports platelet adhesion in blood samples from a heterogeneous group of patients with von Willebrand disease

BACKGROUND AND OBJECTIVE: Plasma derived FVIII-VWF concentrates in which the VWF structure is reasonably maintained are recommended as substitutive therapy in VWD. Our aim was to assess platelet deposition and binding to subendothelial structures of VWF present in FVIII concentrates. DESIGN AND METHODS: Cryoprecipitate (CRY), intermediate-purity (IPC), or high-purity (HPC) FVIII concentrates were added in vitro to citrated blood samples from 11 patients affected by different subtypes of VWD, with the aim of normalizing VWF levels. Measurements of VWF:Ag, ristocetin cofactor (RiCof) activities, FVIII coagulant activity (FVIII:C), and platelet interaction with subendothelium under flow conditions (Baumgartner's perfusion method, computer-assisted morphometry, shear rate 1000 s-1, 10 min, 37 degrees C) were determined. Binding of VWF to the luminal surface of the perfused vessels was assessed by immunofluorescence microscopy. Paired t-test statistics were performed. RESULTS: Addition of FVIII-VWF preparations raised VWF:Ag from baseline (BSL) values of 0.3 (SD 0.2) to averages of 1.4 (SD 0.5, p < 0.001), 1.2 (SD 0.6, p < 0.001), and 0.4 (SD 0.3) IU mL-1 after CRY, IPC, and HPC, respectively. A positive labeling for VWF was observed by immunofluorescence in vessels perfused with blood containing any of the concentrates. Platelet adhesion of 13.2 (SD 7.6), 22.4 (SD 10.8), 24.8 (SD 7.8, p < 0.03), or 22.5 (SD 4.8)% was measured in BSL, CRY, IPC, or HPC tests, respectively. INTERPRETATION AND CONCLUSIONS: Our observations support the hypothesis above the mechanisms involved in the beneficial effects of commercial concentrates in von Willebrand disease: the VWF in these concentrates has functional capacity to bind to subendothelium and to support platelet adhesion.     

Type 2M von Willebrand disease: F606I and I662F mutations in the glycoprotein Ib binding domain selectively impair ristocetin- but not botrocetin-mediated binding of von Willebrand factor to platelets

von Willebrand disease (vWD) is a common, autosomally inherited, bleeding disorder caused by quantitative and/or qualitative deficiency of von Willebrand factor (vWF). We describe two families with a variant form of vWD where affected members of both families have borderline or low vWF antigen levels, normal vWF multimer patterns, disproportionately low ristocetin cofactor activity, and significant bleeding symptoms. Whereas ristocetin-induced binding of plasma vWF from affected members of both families to fixed platelets was reduced, botrocetin-induced platelet binding was normal. The sequencing of genomic DNA identified unique missense mutations in each family in the vWF exon 28. In Family A, a missense mutation at nucleotide 4105T --> A resulted in a Phe606Ile amino acid substitution (F606I) and in Family B, a missense mutation at nucleotide 4273A --> T resulted in an Ile662Phe amino acid substitution (I662F). Both mutations are within the large disulfide loop between Cys509 and Cys695 in the A1 domain that mediates vWF interaction with platelet glycoprotein Ib. Expression of recombinant vWF containing either F606I or I662F mutations resulted in mutant recombinant vWF with decreased ristocetin-induced platelet binding, but normal multimer structure, botrocetin-induced platelet binding, collagen binding, and binding to the conformation-sensitive monoclonal antibody, AvW-3. Both mutations are phenotypically distinct from the previously reported variant type 2MMilwaukee-1 because of the presence of normal botrocetin-induced platelet binding, collagen binding, and AvW-3 binding, as well as the greater frequency and intensity of clinical bleeding. When the reported type 2M mutations are mapped on the predicted three-dimensional structure of the A1 loop of vWF, the mutations cluster in one region that is distinct from the region in which the type 2B mutations cluster.     

Dominant type 1 von Willebrand disease caused by mutated cysteine residues in the D3 domain of von Willebrand factor

No defects have been reported in moderately severe type 1 von Willebrand disease (vWD) with a clear autosomal dominant inheritance pattern, and the mechanism underlying this form of vWD remains obscure. We have studied a type 1 vWD family with such a dominant phenotype. The entire coding sequence of the von Willebrand factor (vWF) gene was analyzed by direct sequencing of DNA fragments amplified by polymerase chain reaction. Only one candidate mutation T(3445)-->C in exon 26 was detected that predicts a replacement of cysteine (C) at position 386 of the mature vWF subunit by arginine (R). Both mutant and normal vWF alleles were expressed as shown by analysis of platelet mRNA. This substitution segregates with vWD in the family and was not found in 100 unrelated individuals. The recombinant mutant vWF(C386R) was characterized by expression in 293T cells. The secretion of vWF(C386R) was greatly impaired due to retention in the endoplasmic reticulum. In cotransfections of normal and mutant vWF constructs, the vWF(C386R) subunits caused a dose-dependent decrease in the secretion of vWF. The multimer pattern remained nearly normal and consistent with a dominant vWD type 1 phenotype. The importance of the cysteine residues in the D3 domain of vWF in the pathogenesis of dominant type 1 vWD was further shown by the detection of another cysteine mutation, Cys367-->Phe, in two additional unrelated patients with a similar dominant type 1 vWD phenotype. We conclude that the loss of cysteine pairing in the D3 domain, leaving one free cysteine, can induce a purely quantitative deficiency of vWF by dominantly suppressing the secretion of normal vWF.     

Identification of domains responsible for von Willebrand factor type VI collagen interaction mediating platelet adhesion under high flow

We have identified type VI collagen (Col VI) as a primary subendothelial extracellular matrix component responsible for von Willebrand factor (vWF)-dependent platelet adhesion and aggregation under high tensile strength. Intact tetrameric Col VI was the form of the collagen found to be capable of promoting vWF-mediated platelet adhesion/aggregation under this shear condition, whereas removal of the predominant portion of the terminal globules by pepsin treatment abrogated its activity. The inability of the pepsin-digested Col VI to support any platelet interaction at high flow was because of the failure of the A3(vWF) domain to bind to this form of collagen, suggesting a stringent requirement of a tridimensional conformation or of intactness of its macromolecular structure. In contrast, the A1(vWF) domain bound to both intact and pepsin-digested Col VI tetramers but, in accordance with the cooperating function of the two vWF domains, failed to support platelet adhesion/aggregation under high shear onto Col VI by itself. The putative A1(vWF) binding site resided within the A7(VI) module (residues 413-613) of the globular amino-terminal portion of the alpha3(VI) chain. Soluble recombinant A7(VI) polypeptide strongly perturbed the vWF-mediated platelet adhesion to Col VI under high shear rates, without affecting the binding of the vWF platelet receptor glycoprotein Ibalpha to its cognate ligand A1(vWF). The findings provide evidence for a concerted action of the A1(vWF) and A3(vWF) domains in inducing platelet arrest on Col VI. This is accomplished via an interaction of the A1(vWF) domain with a site contained in the alpha3 chain A7(VI) domain and via a conformation-dependent interaction of the A3(vWF) domain with the intact tetrameric collagen. The data further emphasize that Col VI microfilaments linking the subendothelial basement membrane to the interstitial collagenous network may play a pivotal role in the hemostatic process triggered upon damage of the blood vessel wall.     

Biochemical and functional characterization of recombinant von Willebrand factor produced on a large scale

Recombinant von Willebrand factor (r-vWF) was produced in serum-free medium on a large scale in recombinant Chinese hamster ovary cells and was purified from fermentation supernatant by a combination of anion exchange chromatography and heparin affinity chromatography. Heparin affinity chromatography yielded r-vWF polymers of different degrees of multimerization. r-vWF was analysed by qualitative and quantitative functional analysis. We could show that while binding of r-vWF to platelets did not depend on multimerization of the molecule, ristocetin-induced platelet aggregation, binding to collagen and binding to heparin correlated directly with the extent of multimerization. Binding of recombinant coagulation factor VIII (r-FVIII) to r-vWF was studied by real-time biospecific interaction analysis and surface plasmon technology. The data indicated that binding of r-FVIII did not depend on r-vWF multimerization. Real-time biospecific interaction analysis suggested a potential stoichiometry of 2 to 2.5 r-vWF subunits per r-FVIII molecule. Kinetic analysis of the r-vWF-r-FVIII interaction gave a binding rate constant of 3 x 10(6) M-1 s-1 and an association constant of 2.5 x 10(9) M-1. Reaction of r-vWF with carbohydrate-specific lectins demonstrated that r-vWF contained a high proportion of N-glycans composed of mannose, galactose, glucose, N-acetylglucosamine and terminal sialic acid. Carbohydrate moities were covalently bound to the protein structure and were quantitatively removed from r-vWF only after protein denaturation. The results demonstrated that r-vWF produced on large scale under serum-free culture conditions exhibited qualitative and quantitative functional properties comparable to human plasma-derived vWF.     

Electrospray ionisation mass spectrometry facilitates detection of fibrinogen (Bbeta 14 Arg --> Cys) mutation in a family with thrombosis

We report the first direct detection of a fibrinogen mutation by electrospray ionisation mass spectrometry. The propositus, from a family with a history of thrombosis, came to attention after a pulmonary embolism subsequent to a spontaneous abortion. Prolonged thrombin (41 s) and reptilase times (26 s) together with an impairment of fibrinopeptide B release suggested a mutation at the thrombin cleavage site of the Bbeta chain. Direct mass analysis of purified fibrin chains from a thrombin induced clot showed that 50% of the Bbeta chains remained uncleaved. The measured mass of the mono sialo isoform of this uncleaved chain was 54150 Da, compared to a value of 54198 Da for normal Bbeta chains. This decrease of 48 Da in the intact protein is indicative of either a Bbeta 14 Arg to Cys, or Arg to Leu substitution. Heterozygosity for the Bbeta 14 Arg --> Cys mutation was verified by PCR amplification and DNA sequence analysis.     

Missense mutation in a von Willebrand factor type A domain of the alpha 3(VI) collagen gene (COL6A3) in a family with Bethlem myopathy

OBJECTIVE: The purpose of this retrospective study was to examine the value of whole-body nuclear medicine imaging and to evaluate the typical scintigraphic pattern of sternocostoclavicular hyperostosis (SCCH) and/or pustulotic arthroosteitis (PAO). In this entity the correct diagnosis is frequently missed because of uncharacteristic changes in other imaging modalities. MATERIALS AND METHODS: Forty-nine patients (age range 15-65 years old, mean age 36 years) with sternocostoclavicular hyperostosis (SCCH) and/or pustulotic arthroosteitis (PAO) were examined with whole-body scintigraphy and conventional radiography. RESULTS: Forty-three of 49 patients with SCCH/PAO showed a characteristic "bullhead"-like high tracer uptake of the sternocostoclavicular region with the manubrium sterni representing the upper skull and the inflamed sternoclavicular joints corresponding to the horns (= bullhead sign). Scintigraphy revealed additional skeletal manifestations (spondylitis, sacroiliitis, osteitis) in 33 of 49 patients with SCCH and/or PAO. CONCLUSIONS: Bone scintigraphy is the imaging modality of choice for the diagnosis of skeletal involvement in PAO. Nuclear medicine reveals unexpected locations and shows the typical pattern of focal hot spots of the spine, sacroiliac joints and/or appendicular skeleton in the large majority of cases in combination with a bullhead-like tracer uptake of the sternocostoclavicular region. The bullhead sign is the typical and highly specific scintigraphic manifestation of SCCH and PAO in radionuclide bone scans and helps to avoid unnecessary biopsies.     

Effect of desmopressin on plasma factor VIII and von Willebrand factor concentrations in Greyhounds

OBJECTIVE: To determine the effect of 1-Deamino-8-D-arginine vasopressin on plasma concentrations of von Willebrand factor and factor VIII in Greyhound blood donors, and to compare the response of 1-Deamino-8-D-arginine vasopressin injection on plasma concentrations of von Willebrand factor between groups with different resting plasma concentrations of von Willebrand factor. ANIMALS: Fifteen Greyhound blood donors were used. Dogs were grouped into three categories depending on their von Willebrand factor concentrations. PROCEDURE: Desmopressin was administered subcutaneously at 1 microgram/kg [corrected] to all dogs. Plasma von Willebrand factor and factor VIII concentrations were measured before and 10, 20, 30, 45, 60, 90 and 120 min after desmopressin injection. RESULTS: The von Willebrand factor and factor VIII concentrations in all dogs increased significantly and remained higher than base-line throughout the 2 h period. CONCLUSION: Desmopressin is useful in increasing von Willebrand factor concentrations in Greyhound blood donors, including those with low resting concentrations.     

The physical exchange of factor VIII (FVIII) between von Willebrand factor and activated platelets and the effect of the FVIII B-domain on platelet binding

Normal hemostasis proceeds through the assembly of coagulant complexes on a lipid surface derived from activated platelets. The activation complex assembly is governed by multiple factors including the binding constants (Kd) of the coagulant factors for the lipid surface. The formation of the tenase complex requires delivery of factor VIII (FVIII) to the activated lipid surface by von Willebrand factor (vWF). Using electrophoretic quasi-elastic light scattering (ELS), we have examined the interaction of FVIII in the presence and absence of vWF with both resting and activated gel-filtered human platelets. Resting platelets do not bind FVIII. Platelets activated by thrombin, epinephrine, or SFLLRN, but not ADP or collagen, bind unactivated FVIII if vWF is not present. In the absence of vWF, unactivated FVIII binds to activated platelets with a Kd of 10.4 nM. B-domain deleted FVIII binds to activated platelets with a Kd of 5.1 nM. Thrombin -activated FVIII (FVIIIa) binds to activated platelets with a Kd of 1.7 nM. The activation of FVIII while bound to the platelet surface can be monitored as a function of time. In the presence of vWF, binding of unactivated FVIII to activated platelets was inhibited, but not the binding of FVIIIa. Displacement of bound unactivated FVIII from the platelet surface occurs when vWF is added to the FVIII-platelet complex. The binding of FVIII to activated platelets is affected by the B-domain, the state of FVIII activation, and the presence of soluble vWF and proceeds as a multistep process. FVIII binding by activated platelets is not affected by platelet gpIIb/IIIa or by platelet vWF.     

Porcine platelet von Willebrand antigen II (vW AgII): inhibitory effect on collagen-induced aggregation and comparative distribution with human platelets

Von Willebrand factor (vWF) is synthesized by human endothelial cells and megakaryocytes as a large precursor, the pre-provWF, which is finally cleaved into the propolypeptide of vWF (pp-vWF) and the mature vWF. We have purified in parallel the pp-vWF and the GP IIb-IIIa from porcine platelets. The N-terminus comparative analysis of porcine pp-vWF with respect to other species revealed more than a 75% and 65% homology with the bovine and human pp-vWF, respectively. Purified pp-vWF inhibited collagen-induced platelet aggregation in porcine platelet rich plasma (PRP) and specifically binds to collagen. Polyclonal antibody pabBp19 against the purified protein was prepared and characterized by ELISA, Western-blot and immunocytochemistry. The distribution of pp-vWF in soluble and membrane fractions from pig platelets has been performed by immunolabeling detection. pabBp19 did not blot human platelet lysates but human pp-vWF was detectable using the antibody Frieda 013091. Cell distribution and quantification studies of human and porcine platelet pp-vWF showed that the protein exists in the soluble and membrane fractions and its pattern is similar in both species.     

Relative importance of the glycoprotein Ib-binding domain and the RGD sequence of von Willebrand factor for its interaction with endothelial cells

Endothelial cell adhesion to von Willebrand Factor is mainly mediated through an interaction between the alpha vbeta3 integrin and the RGD sequence of von Willebrand factor (vWF). To define the potential involvement of glycoprotein Ib alpha (GPIb alpha) as an endothelial vWF receptor, we compared cell adhesion to three recombinant vWF, the wild-type (WT-rvWF) and two mutants, RGGS-rvWF (D1746G), defective for binding to platelet alphaIIb beta3, and deltaA1-rvWF with a deletion between amino-acids 478 and 716, which does not bind to platelet GPIb alpha. Adhesion of human umbilical vein endothelial cells to purified vWF recombinants was measured by automatized cell counting using an image analyzer. Whereas cell adhesion to delta A1-rvWF was unchanged compared with WT-rvWF, reaching a plateau of 40% total cells at a concentration of 2.5 microg/mL rvWF, adhesion to RGGS-rvWF was only 10% of total cells. Cell stimulation by tumor necrosis factor-alpha (TNF alpha), reported to upregulate the expression of the putative endothelial GPIb alpha, did not modify adhesion to these rvWF. Monoclonal antibodies to vWF or GPIb alpha, blocking vWF interaction with platelet GPIb alpha, were unable to inhibit endothelial cell adhesion to rvWF. In contrast, antibody 9 to vWF, blocking the alpha vbeta3-dependent endothelial cell adhesion to plasma vWF, inhibited adhesion to WT-rvWF as efficiently as to deltaA1-rvWF (50% inhibition at a concentration of 11 and 15 microg/mL, respectively). In agreement with the fact that endothelial cell adhesion to vWF appeared independent of the GPIb alpha-binding domain, we were unable to detect endothelial surface expression of GPIb alpha by flow cytometry or in cell lysates by immunoprecipitation followed by immunoblotting. Moreover, expression of GPIb alpha mRNA was undetectable in endothelial cells, even after stimulation by TNF alpha. These studies indicate that GPIb alpha is not expressed in human cultured endothelial cells and is not involved in adhesion to vWF-containing surfaces. Thus, in static conditions, cultured endothelial cells adhere to vWF through an alpha vbeta3-dependent, GPIb alpha-independent mechanism.     

Effect of anticoagulant therapy on the hypercoagulable state in patients carrying the factor V Arg506Gln mutation

The National Practitioner Data Bank (NPDB), created by the 1986 Health Care Quality Improvement Act, has been in operation since 1990. Hospitals and other credentialing bodies must query the NPDB when granting and renewing privileges. The NPDB receives about 25,000 reports of adverse actions against health practitioners each year. The NPDB was designed to be a flagging system providing information to licensing or credentialing authorities who would further examine practitioner records. Its purpose is to ensure that decision makers have information that might not otherwise be readily available, especially in the case of incompetent practitioners who move from hospital to hospital or state to state. Access to NPDB information is a concern for consumers and providers alike. Only 2% of matched reports to the NPDB made a difference in hospital privileging decisions. A limitation of NPDB information is that malpractice payments recorded in the NPDB do not necessarily constitute a comprehensive and definitive reflection of actual health care incompetence. All health care providers need to be aware of the NPDB, its mission, potential impact on their ability to be credentialed, and proposed additional uses of its information.