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Biomedical subjects

J G Kelton

Publications and source records attributed to J G Kelton.

At least 19 recordsLinked to original sources
PubMed1996

Splenectomy done during hematologic remission to prevent relapse in patients with thrombotic thrombocytopenic purpura.

OBJECTIVE: To assess whether splenectomy done during hematologic remission reduces the relapse rate in patients with relapsing thrombotic thrombocytopenic purpura. DESIGN: Consecutive case series. SETTING: Tertiary care teaching hospital. PATIENTS: 6 consecutive patients seen during a 10-year period who had had one or more relapses of thrombotic thrombocytopenic purpura. INTERVENTIONS: All patients had elective splenectomy while in hematologic remission and were followed after surgery for as long as 8.0 years. MEASUREMENTS: Attack rates (events per year) were calculated for each patient from time of presentation to time of splenectomy and from time of splenectomy to January 1996. RESULTS: A total of 26 episodes of thrombotic thrombocytopenic purpura occurred over 22.3 patient-years before splenectomy. After splenectomy, 3 acute episodes occurred over 22.7 patient-years. The attack rate (+/-1 SD) decreased from 2.3 +/- 2.0 events per year to 0.1 +/- 0.1 events per year. CONCLUSION: In patients who have had one or more relapses of thrombotic thrombocytopenic purpura, splenectomy done during hematologic remission reduces the frequency of acute relapse and the resulting need for medical therapy.

Humans↗
PubMed1996

An autosomal dominant, qualitative platelet disorder associated with multimerin deficiency, abnormalities in platelet factor V, thrombospondin, von Willebrand factor, and fibrinogen and an epinephrine aggregation defect.

Multimerin is a massive soluble, multimeric protein found in platelets and endothelial cells. Recent studies identified multimerin as a specific coagulation factor V binding protein, complexed with platelet, but not plasma, factor V. These findings led us to investigate individuals with inherited factor V deficiencies for possible multimerin abnormalities. Platelet proteins were evaluated using immunoassays, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblotting, immunoprecipitation, and direct binding studies. Patients with factor V Quebec, a disorder with abnormal platelet factor V, had a quantitative deficiency in multimerin (n = 11 tested; mean, 12.5%; range, 5% to 27% of the normal pool; normal range, 45% to 214%) with a normal multimer pattern. Quantitative and qualitative abnormalities were detected in their platelet factor V. An unrelated patient who was deficient in platelet and plasma factor V had normal platelet multimerin. The levels of platelet beta-thromboglobulin, von Willebrand factor, thrombospondin, and fibrinogen antigen were normal in the factor V Quebec patients. However, proteins with abnormal mobility were detected in their platelet lysate and releasate, and their platelet thrombospondin, von Willebrand factor, and fibrinogen showed evidence of proteolytic degradation. Platelet counts of the factor V Quebec patients ranged from mildly thrombocytopenic to low normal (mean, 159 x 10(9)/L; range, 104 to 198 x 10(9)/L). In addition, their platelets failed to aggregate in response to 6 to 10 micromol/L epinephrine despite normal numbers of platelet alpha 2-adrenergic receptors. These data indicate that patients with factor V Quebec have an inherited bleeding disorder distinct from other platelet disorders and associated with multiple abnormalities, including multimerin deficiency, abnormal platelet factor V, thrombospondin, von Willebrand factor, and fibrinogen, and an epinephrine aggregation defect.

Afibrinogenemia↗
PubMed1996

Isolation and characterization of cysteine proteinase in thrombotic thrombocytopenic purpura.

Thrombotic thrombocytopenic purpura (TTP) is an uncommon disorder characterized by thrombocytopenia, schistocytic haemolytic anaemia, fever, neurologic lesions, and renal failure. A platelet aggregating factor has been postulated to be responsible for this disorder, but its precise identity remains debated. Two different groups of investigators have provided evidence that the platelet aggregating factor is a cysteine proteinase. We have suggested that it was calpain, whereas others have suggested that it was calpain, whereas others have suggested that it was cathepsin L. To help resolve this issue, we have studied the platelet activating activity found in the acute serum samples from 10 different TTP patients as well as purified calpain and cathepsin L. The TTP activity was measured functionally (platelet serotonin release assay and casein lysis assay) and antigenically (immunodepletion using anti-calpain and anti-cathepsin L antibodies and antigenic analysis using SDS PAGE). The TTP serum paralleled the activity of the purified calpain and had optimal pH activity of 7.5. The purified cathepsin L activity had optimal activity at low pH (5.5) and activity was no longer measurable at pH 7.5. Similarly, a specific cathepsin L inhibitor (Z-Phe-Phe-CHN2) had no effect on the activity of the TTP samples nor on purified calpain, but it did abolish the activity activity of purified cathepsin L. The platelet activating of the TTP samples was detectable in the microparticle pellet following ultracentrifugation of TTP serum, and could be immunodepleted using antibodies to calpain but not to cathepsin L. These studies indicate that the microparticle-associated platelet activating factor in TTP corresponds to calpain.

Blood Platelets↗
PubMed1995

Characterization and localization of the Gova/b alloantigens to the glycosylphosphatidylinositol-anchored protein CDw109 on human platelets.

The Gova/b alloantigens are expressed on a 175-kD protein (GP175) on human platelets. Anti-Gov alloantibodies have been implicated in posttransfusion purpura and alloimmune neonatal thrombocytopenia. In this report we characterize the immunochemistry of the alloantigens and identify the platelet protein that expresses the Gov epitopes. Approximately 50% of GP175 containing the Gov epitope was released from platelets treated with phosphatidylinositol-specific phospholipase C, indicating that at least some of this protein exists as a glycosylphosphatidylinositol (GPI)-linked isoform. Radioimmunoprecipitation and immunodepletion studies indicated that the Gova/b alloantigens are expressed on the GPI-anchored CDw109 protein. The Gova/b epitopes were expressed on an extracellular, 120-kD soluble fragment (p120) of CDw109 produced by calcium-dependent protease cleavage. Anti-Gov immunoprecipitates of chymotryptic digests of p120 contained 70- and 52-kD fragments of CDw109. Deglycosylation of native CDw109 had no effect on recognition by Gov alloantisera; however, the epitopes were destroyed after exposure to sodium dodecyl sulfate. Gova/b alloantigens were expressed on platelets and PHA-activated T-cells, cultured human umbilical vein endothelial cells, and by many different tumor cell lines, consistent with the tissue distribution of CDw109.

Antibodies, Monoclonal↗
PubMed1995

Factor V is complexed with multimerin in resting platelet lysates and colocalizes with multimerin in platelet alpha-granules.

Factor V stored in platelets is an important source of factor Va for the prothrombinase complex. Investigations of potential platelet factor Va-binding proteins, using factor Va light chain affinity chromatography, identified a disulfide-linked multimeric protein with a reduced mobility of 155 kDa in the column eluate. Immunodepletion and immunoblotting indicated that this protein was multimerin. Multimerin specifically bound factors V and Va and the isolated factor Va light chain, but not the heavy chain of factor Va. Factor V stored in platelets, but not plasma factor V, was found to be complexed with multimerin. Multimerin immunodepletion of resting platelet lysates was associated with the removal of factor V and the loss of factor V coagulant activity. Immunoelectron microscopic studies colocalized factor V with multimerin in the alpha-granules of resting platelets. With thrombin-induced platelet activation, we observed dissociation of factor Va-multimerin complexes, multimerin-independent membrane binding of factor Va, and prothrombinase activity that was not inhibitable by multimerin antibodies. This study indicates that platelet factor V is stored as a complex with multimerin and suggests a possible role for multimerin as a carrier protein for factor V stored in platelets.

Animals↗
PubMed1995

The cDNA sequence of human endothelial cell multimerin. A unique protein with RGDS, coiled-coil, and epidermal growth factor-like domains and a carboxyl terminus similar to the globular domain of complement C1q and collagens type VIII and X.

Multimerin is a massive, soluble protein found in platelets and in the endothelium of blood vessels. Multimerin is composed of varying sized, disulfide-linked multimers, the smallest of which is a homotrimer. Multimerin is a factor V/Va-binding protein and may function as a carrier protein for platelet factor V. The cDNA for human multimerin was isolated from lambda gt11 endothelial cell libraries using antibodies, and the isolated cDNA clones were used to obtain the full sequence. The full-length multimerin cDNA was 4.2 kilobase pairs. Northern analyses identified a 4.7-kilobase transcript in cultured endothelial cells, a megakaryocytic cell line, platelets, and highly vascular tissues. The multimerin cDNA can encode a protein of 1228 amino acids with the probable signal peptide cleavage site between amino acids 19 and 20. The protein is predicted to be hydrophilic and to contain 23 N-glycosylation sites. The adhesive motif RGDS (Arg-Gly-Asp-Ser) and an epidermal growth factor-like domain were identified. Sequence searches indicated that multimerin is a unique protein. Analyses identified probable coiled-coil structures in the central portion of the multimerin sequence. Additionally, the carboxyl-terminal region of multimerin resembles the globular, non-collagen-like, carboxyl-terminal domains of several other trimeric proteins, including complement C1q and collagens type VIII and X.

Amino Acid Sequence↗
PubMed1995

Heparin-induced thrombocytopenia in patients treated with low-molecular-weight heparin or unfractionated heparin.

BACKGROUND: Heparin-induced thrombocytopenia, defined by the presence of heparin-dependent IgG antibodies, typically occurs five or more days after the start of heparin therapy and can be complicated by thrombotic events. The frequency of heparin-induced thrombocytopenia and of heparin-dependent IgG antibodies, as well as the relative risk of each in patients given low-molecular-weight heparin, is unknown. METHODS: We obtained daily platelet counts in 665 patients in a randomized, double-blind clinical trial comparing unfractionated heparin with low-molecular-weight heparin as prophylaxis after hip surgery. Heparin-induced thrombocytopenia was defined as a decrease in the platelet count below 150,000 per cubic millimeter that began five or more days after the start of heparin therapy, and a positive test for heparin-dependent IgG antibodies. We also tested a representative subgroup of 387 patients for heparin-dependent IgG antibodies regardless of their platelet counts. RESULTS: Heparin-induced thrombocytopenia occurred in 9 of 332 patients who received unfractionated heparin and in none of 333 patients who received low-molecular-weight heparin (2.7 percent vs. 0 percent; P = 0.0018). Eight of the 9 patients with heparin-induced thrombocytopenia also had one or more thrombotic events (venous in 7 and arterial in 1), as compared with 117 of 656 patients without heparin-induced thrombocytopenia (88.9 percent vs. 17.8 percent; odds ratio, 36.9; 95 percent confidence interval, 4.8 to 1638; P < 0.001). In the subgroup of 387 patients, the frequency of heparin-dependent IgG antibodies was higher among patients who received unfractionated heparin (7.8 percent, vs. 2.2 percent among patients who received low-molecular-weight heparin; P = 0.02). CONCLUSIONS: Heparin-induced thrombocytopenia, associated thrombotic events, and heparin-dependent IgG antibodies are more common in patients treated with unfractionated heparin than in those treated with low-molecular-weight heparin.

Confidence Intervals↗
PubMed1995

Bacterial infection-associated improvement of platelet counts in two patients with chronic and unresponsive idiopathic thrombocytopenic purpura with normal platelet survival studies.

Approximately 20% of adult patients with idiopathic thrombocytopenic purpura (ITP) do not respond to splenectomy and require alternative therapies to achieve a clinically safe platelet count. A small percentage of these patients have very refractory disease and are either unresponsive or poorly responsive to almost any therapy. In this report we describe two patients with chronic and unresponsive ITP with normal platelet survivals. Neither patient had responded to a large variety of treatments including corticosteroids, splenectomy, IVIgG, anti-D, chemotherapy, and ascorbic acid. However, both had a rapid, but short-lived, rise in their platelet count following a bacterial infection. One patient had a rise in platelet count for 6 months following the acute episode of bacteraemia. The second patient had a shorter response of 1 week. It is possible that these two patients represent a subset of patients with ITP who may benefit from cytokine therapy.

Aged↗
PubMed1995

The prenatal identification of fetal compatibility in neonatal alloimmune thrombocytopenia using amniotic fluid and variable number of tandem repeat (VNTR) analysis.

Most severe episodes of neonatal alloimmune thrombocytopenic purpura (NATP) are caused by antiplatelet alloantibodies against the HPA-1a (PlA1) antigen. However, half of subsequent fetuses produced from a HPA-1a/b father (genotypic frequency 28%) will result in a child who is not affected. Some investigators manage NATP by confirming the fetal platelet phenotype using percutaneous umbilical cord sampling, a procedure that carries a low but real risk of fetal morbidity and mortality. More recently, physicians determine the fetal platelet antigen genotype using DNA derived from amniotic fluid or chorionic villus samples. All therapy is withdrawn for a fetus who genotypes as HPA-1b/b. However, since the fetus is the same genotype as the mother, there can be uncertainty about the origin of the genetic material and thus the validity of the fetal genotype. The inappropriate withdrawal of therapy for a erroneously genotyped fetus could be fatal, and consequently many physicians advocate fetal HPA-1 phenotyping with confirmation using percutaneous umbilical blood sampling. In this report we describe the management of two pregnancies with previously affected infants due to anti-HPA-1a alloantibodies. Both husbands were HPA-1a/b. For the current pregnancies, amniotic fluid was collected at 20 or 29 weeks of gestation, and the platelet genotype indicated that the fetuses were HPA-1b/b. The fetal origin of the amniotic fluid derived DNA was confirmed by the forensic technique of DNA profiling using variable number of tandem repeat (VNTR) analysis. All therapy was withdrawn, percutaneous umbilical blood sampling was not performed, and both women vaginally delivered healthy non-thrombocytopenic infants. The application of platelet alloantigen genotyping using DNA from amniotic fluid cells identified the HPA-1b/b fetus, and VNTR analysis confirmed that the tissue was fetal derived, thus avoiding the necessity for percutaneous umbilical blood sampling. The use of this approach in patients at risk will avoid additional investigation and treatment in approximately one-seventh of all NATP pregnancies involving the HPA-1a antigen.

Amniotic Fluid↗
PubMed1995

A prospective study to determine the frequency and clinical significance of alloimmunization post-transfusion.

There is debate in the literature about the frequency and importance of delayed transfusion reactions. This uncertainty could reflect the endpoints used (clinical or serological) and the type of study (typically retrospective or case series). In this report we describe a prospective investigation to determine the frequency of alloimmunization post transfusion and whether the alloantibody production is a laboratory event or has clinical relevance. A total of 2490 patients were transfused 11,218 red cell concentrates. One or more blood samples were collected within 7 d post transfusion and screened for serological evidence of alloimmunization. If any antibody was detected the patient's post-transfusion sample was screened for biochemical evidence of haemolysis and the patient's chart reviewed for documentation of clinical signs of a transfusion reaction. Post transfusion alloimmunization occurred in 2.6% of the patients (95% CI 2.1-3.6%), who had no detectable alloantibody in pre-transfusion testing. For those 86 patients (3.5%) with alloantibodies detectable pretransfusion, 8.9% (95% CI 3.6-17.4%) developed additional aloantibodies. The most common alloantibodies detected were anti-Jka, anti-E and anti-K. Despite the high frequency of serological evidence of delayed transfusion reactions, only one patient (0.05%) had clinical evidence of a delayed haemolytic transfusion reaction (95% CI 0.0-0.27%). Serological evidence of a delayed transfusion reaction is common; however, these reactions rarely cause clinical symptoms.

Blood Group Incompatibility↗
PubMed1995

The use of anti-D to improve post-transfusion platelet response: a randomized trial.

Patients undergoing induction chemotherapy for acute leukaemia often become refractory to platelet transfusions. Increased clearance of transfused platelets due to alloimmune destruction has been identified as one of the primary mechanisms contributing to this refractory state. We performed a double-blind randomized trial to determine whether the administration of anti-D to Rh-positive individuals could prevent the refractory state and improve post-transfusion platelet response. Rh-positive patients with acute leukaemia undergoing induction chemotherapy and requiring platelet transfusions were allocated to weekly intravenous anti-D (20 micrograms/kg) or placebo. Platelets and red cell concentrates were administered according to standardized transfusion guidelines. Outcome measures included platelet transfusion utilization, red cell utilization, platelet recovery 18-24 h post-infusion, and the percentage of patients refractory to platelet transfusion. There were 43 patients studied: 21 received anti-D and 22 saline placebo. The mean number of platelet concentrates required per day of observation was 0.59 (SD 0.22) in the anti-D group and 0.61 (SD 0.22) in the placebo group, P = 0.86. No difference was detected between groups in terms of platelet recovery post-infusion, refractoriness to platelet transfusion or frequency of infection (P = 0.97). Red cell concentrate utilization was significantly increased in the anti-D group compared to the placebo group, 0.58 units per day versus 0.37 units per day respectively, P = 0.005. We conclude that the use of anti-D did not improve post-transfusion platelet response in Rh positive patients with acute leukaemia, but did result in an increased need for red cell transfusion.

Acute Disease↗
PubMed1995

Three examples of Rh haemolytic disease of the newborn with a negative direct antiglobulin test.

Typically the serological diagnosis of alloimmune haemolytic disease of the newborn (HDN) includes a positive direct antiglobulin test on the infant's red cells, and the presence of an IgG red cell alloantibody in both maternal and cord sera. HDN with a negative direct antiglobulin test has been reported with anti-A and anti-B, but not with other red-cell alloantibodies. In this report we describe four examples of HDN in infants whose red cells had a negative direct antiglobulin test. The first case was diagnosed retrospectively when the infant was admitted to hospital aged 3 weeks with severe anaemia and cardiac failure, and subsequently died. Maternal and infant sera were both shown to contain anti-C: however, the direct antiglobulin test on the infant's red cells was negative. Approximately 1 year later the mother of this infant gave birth to triplets: soon after birth one of the triplets required an exchange transfusion, one had hyperbilirubinaemia, and the third was unaffected. Anti-C and anti-e were detectable in the maternal serum at this time. The most probable Rh genotypes of the two affected infants were R1R2 (CDe/cDE), while the Rh genotype of the unaffected infant was R2R2 (cDE/cDE). Anti-c was implicated as causing HDN in a fourth infant (from a different family) who was a hydropic stillborn. The direct antiglobulin test on fetal blood was negative and other causes of non-immune hydrops were excluded. These four infants provide evidence that the direct antiglobulin test may be negative in some severely affected and even fatal cases of HDN.

Coombs Test↗
PubMed1995

Perinatal thrombocytopenia.

Maternal thrombocytopenia is common in normal pregnancies, but is a poor predictor of fetal thrombocytopenia even when the maternal thrombocytopenia is of pathologic etiology. The real risk to the fetus and the neonate is alloimmune thrombocytopenia, although identification of index cases is problematic and management of future pregnancies has not been defined.

Female↗
PubMed1995

The serological investigation of patients with autoimmune thrombocytopenia.

Techniques for measuring the plasma factor responsible for idiopathic thrombocytopenic purpura (ITP) have been sought for some 40 years, but unlike red cell serological techniques, platelet antibody testing has proved difficult and, at times, generated controversial results. The difficulty in measuring platelet antibodies probably reflects the intrinsic nature of the platelet membrane--to act as a sponge and nonspecifically bind plasma proteins. There have now been three different general types of techniques used to measure platelet autoantibodies: phase I assays measuring a platelet-dependent endpoint after incubating test serum and control platelets. These assays lack sufficient sensitivity and specificity to be useful. Except for special instances (the diagnosis of heparin-induced thrombocytopenia), they are no longer used. Phase II assays measure IgG on the surface of platelet or within the platelets following lysis. The immunoglobulin detected is termed platelet associated IgG. PAIgG assays are falling out of favour because they lack the sensitivity and specificity to be useful diagnostic tools. Most recently, the ability to isolate individual platelet glycoproteins and measure the binding of IgG to these glycoproteins has dramatically increased our understanding of ITP. These Phase III assays may prove useful diagnostic tools for the investigation and classification of immune thrombocytopenic disorders.

Antibodies, Monoclonal↗