Complications of haemophilia in babies (first two years of life): a report from the Centers for Disease Control and Prevention Universal Data Collection System.

AIM: To describe the prevalence and complications in babies ≤2 years with haemophilia. METHODS: We used a standardized collection tool to obtain consented data on eligible babies aged ≤2 years with haemophilia enrolled in the Centers for Disease Control and Prevention Universal Data Collection System surveillance project at US Hemophilia Treatment Centers (HTCs). RESULTS: Of 547 babies, 82% had haemophilia A, and 70% were diagnosed within one month of birth. Diagnosis was prompted by known maternal carrier status (40%), positive family history (23%), bleeding (35%) and unknown 2%; 81% bled during the first two years. The most common events were bleeding (circumcision, soft tissue, oral bleeding) and head injury. There were 46 episodes of intracranial haemorrhage (ICH) in 37 babies (7%): 18 spontaneous, 14 delivery related, 11 traumatic, 2 procedure related and 1 unknown cause. Of the 176 central venous access devices (CVADs) in 148 (27%) babies, there were 137 ports, 22 surgically inserted central catheters and 20 peripherally inserted central catheters. Ports had the lowest complication rates. Inhibitors occurred in 109 (20%) babies who experienced higher rates of ICH (14% vs. 5%; P = 0.002), CVAD placement (61% vs. 19%; P < 0.001) and CVAD complications (44% vs. 26%; P < 0.001). The most common replacement therapy was recombinant clotting factor concentrates. CONCLUSION: Bleeding events in haemophilic babies ≤2 years were common; no detectable difference in the rates of ICH by the mode of delivery was noted. Neonatal factor exposure did not affect the inhibitor rates. Minor head trauma, soft tissue and oropharyngeal bleeding were the leading indications for treatment.

Sites of initial bleeding episodes, mode of delivery and age of diagnosis in babies with haemophilia diagnosed before the age of 2 years: a report from The Centers for Disease Control and Prevention's (CDC) Universal Data Collection (UDC) project.

Lack of detailed natural history and outcomes data for neonates and toddlers with haemophilia hampers the provision of optimal management of the disorder. We report an analysis of prospective data collected from 580 neonates and toddlers aged 0-2 years with haemophilia enrolled in the Universal Data Collection (UDC) surveillance project of the Centers for Disease Control and Prevention (CDC). This study focuses on a cohort of babies with haemophilia whose diagnosis was established before the age of two. The mode of delivery, type and severity of haemophilia, onset and timing of haemorrhages, site(s) of bleeding, provision of prophylaxis with coagulation factor replacement therapy, and the role played by the federally funded Haemophilia Treatment Centers (HTC) in the management of these infants with haemophilia were evaluated. Seventy-five per cent of haemophilic infants were diagnosed early, in the first month of life, especially those with a family history or whose mothers were known carriers; infants of maternal carriers were more likely to be delivered by C-section. Involvement of an HTC prior to delivery resulted in avoidance of the use of assisted deliveries with vacuum and forceps. Bleeding from the circumcision site was the most common haemorrhagic complication, followed by intra- and extra-cranial haemorrhages and bleeding from heel stick blood sampling. Eight per cent of the infants were administered factor concentrate within 24 h of birth; more than half were treated to prevent bleeding. This study highlights the significant rate and the sites of initial bleeding unique to very young children with haemophilia and underscores the need for research to identify optimal evidence-based recommendations for their management.

Antibodies to human T-cell leukemia virus membrane antigens (HTLV-MA) in hemophiliacs.

Along with homosexual men, Haitians, and intravenous drug abusers, hemophiliacs are at high risk of contracting acquired immunodeficiency syndrome (AIDS). An earlier study revealed that 36 percent of a group of the AIDS patients had antibodies to cell membrane antigens associated with the human T-cell leukemia virus (HTLV-MA), whereas only 1.2 percent of matched asymptomatic homosexual controls had these antibodies. In the present experiments, serum samples from 172 asymptomatic hemophiliacs were examined for the presence of antibodies to HTLV-MA. Such antibodies were detected in 5 to 19 percent of the hemophiliacs examined from four geographical locations, but in only 1 percent or less of laboratory workers, normal blood donors, donors on hemodialysis, or donors with chronic active hepatitis.

Measurement of thrombopoiesis in rabbits using 75selenomethionine.

Incorporation of (75)selenomethionine ((75)SeM) has been used to study platelet production in rabbits. Radioactivity of platelets was low after the intravenous administration of (75)SeM and rose to a maximum approximately 3 days after administration. Platelet radioactivity was independent of concurrent plasma levels. The life span of rabbit platelets, as estimated with this technique, was 4-5 days. In vivo reutilization of (75)SeM previously incorporated into plasma proteins was not detected. In vitro incorporation of (75)SeM by platelets in platelet-rich plasma was not demonstrated. Acute hemorrhage 24 hr before administration of (75)SeM increased the incorporation of (75)SeM into platelets. Transfusion-induced thrombocytosis reduced the incorporation of (75)SeM to approximately 30% of that observed in control animals. Suppression of bone marrow function with nitrogen mustard resulted in decreased numbers of platelets in the circulation, and a decrease in incorporation of (75)SeM. Delayed appearance of (75)SeM was observed in circulating platelets during recovery from marrow suppression. Injection of 75-225 ml of plasma from thrombocytopenic donors into normal rabbits increased incorporation of (75)SeM into platelets while normal plasma did not have this effect. The rate of appearance of (75)SeM in circulating platelets appears to provide a sensitive and specific method for the study of production of platelets by megakaryocytes. The data suggest more rapid entry of platelets into the circulation, and a sustained increase in incorporation of (75)SeM into platelet protein after stimulation of platelet production. The results are consistent with the concept of a humoral agent (thrombopoietin) that acts on megakaryocytes to regulate platelet production.

Deficiency of protein C in congenital thrombotic disease.

A family with a history of recurring thrombosis was studied to determine if a plasma protein deficiency could account for the observed disease. Protein C levels in plasma were determined immunologically using the Laurell rocket technique. The propositus, his father, and his paternal uncle, who are severely affected, had 38-49% of normal levels of protein C antigen, whereas unaffected family members had normal levels. There was no familial deficiency of antithrombin III and plasminogen. Because activated protein C is a potent in vitro anticoagulant enzyme and an in vivo profibrinolytic agent, it is suggested that the recurrent thrombotic disease in this family is due to an inherited deficiency in protein C.

Thermal inactivation of the acquired immunodeficiency syndrome virus, human T lymphotropic virus-III/lymphadenopathy-associated virus, with special reference to antihemophilic factor.

The virus that causes the acquired immunodeficiency syndrome (AIDS), human T lymphotropic virus/lymphadenopathy-associated virus (HTLV-III/LAV), was incubated at temperatures from 37 degrees to 60 degrees C and virus titer (ID-50) was determined over time by a microculture infectivity assay. The rate of thermal decay was consistent with first-order kinetics, and these data were used to construct a linear Arrhenius plot (r = 0.99), which was used to determine inactivation time as a function of temperature. In the liquid state, thermal decay was little affected by matrix (culture media, serum, or liquid Factor VIII). In the lyophilized state, the time required to reduce virus titer 10-fold (1 log) at 60 degrees C was 32 min compared with 24 s in the liquid state. HTLV-III/LAV in liquid antihemophilic Factor VIII or IX was lyophilized and heated according to commercial manufacturers' specifications. Infectious virus was undetectable with these regimens. Heat treatment should reduce or stop transmission of HTLV-III/LAV by commercial antihemophilic Factor VIII or IX.

Studies of myeloperoxidase gene expression at the cellular level by in situ hybridization.

Recent studies have demonstrated myeloperoxidase (MPO) gene expression during granulocytic differentiation. Since these studies have been done exclusively by Northern and dot blot analysis and frequently with mixed populations of cells, quantitative changes in gene expression for particular populations of cells are difficult to assess. We therefore examined MPO expression at the cellular level in various normal and malignant hematopoietic cells by the in situ hybridization (ISH) technique. Using this approach, we demonstrated that inducing the promyelocytic HL-60 cell line to differentiate along either monocytic or granulocytic pathways decreases MPO mRNA expression. Similarly, when ISH was performed on normal bone marrow, relatively high levels of MPO mRNA were detected in myeloblasts, promyelocytes, and early eosinophilic precursors, whereas the expression was markedly decreased in more advanced stages of myeloid differentiation. These findings agree with the known decrease in MPO protein synthesis observed during granulocytic differentiation and suggest that regulation of MPO protein synthesis occurs at the level of MPO mRNA expression. We conclude by showing that ISH can detect MPO mRNA in myeloblasts of patients with acute leukemia and can be a potentially useful technique in the study of myeloid differentiation in acute leukemias.

Accelerated fibrinogen and platelet destruction after peritoneovenous shunting.

Peritoneovenous shunting with the LeVeen valve is generally recognized as an effective procedure for the treatment of intractable ascites and renal failure associated with severe liver disease. We recently observed a generalized hemorrhagic diathesis in patients receiving these valves. To investigate the mechanism of this hemorrhagic complication, we prospectively performed kinetic studies with 51Cr-labelled platelets and 125I-labelled fibrinogen in ten patients. When results of studies before and after valve insertion were compared, the following reductions were noted: fibrinogen concentration, 55%; the fibrinogen survival, 49%; the platelet count, 55% and the platelet survival, 35%. No endotoxin was detectable in ascitic fluid preoperatively, and there was no apparent relationship between ascitic fluid cell counts and changes in fibrinogen and platelet survival. Until the component or components of ascitic fluid responsible for accelerated consumption can be identified and steps are taken to modify the rates of platelet and fibrinogen consumption, it would seem prudent to select patients for surgery conservatively.

Characteristics of rat megakaryocyte colonies and their progenitors in agar culture.

The characteristics of megakaryocyte colonies that develop from megakaryocyte progenitors of rat bone marrow stimulated by rat spleen-conditioned medium (SCM) in agar culture were investigated. Colony frequency was optimal on day 7 and increased relative to both the number of cells plated and the concentration of SCM used. Plating efficiencies averaged 72 +/- 16 megakaryocyte colonies/10(5) cells with 0.1 ml SCM/culture. Colonies were categorized as small cell and big cell. Small-cell colonies had a greater proliferative potential, with a mean of 25 cells/colony. Big-cell colonies averaged 15 cells/colony. The ratio of big-cell to small-cell colonies was 0.69 +/- 0.29. Granulocyte-macrophage colonies, which were also stimulated by SCM, accounted for 70% +/- 15% of the total colonies in the cultures. Cytocidal experiments with tritiated thymidine reduced megakaryocyte colony formation by 45% and granulocyte-macrophage colony formation by 21%. The properties of rat, mouse, and human megakaryocyte progenitors as assayed in vitro are compared.

The effects of thrombopoietic activity of rabbit plasma fractions on megakaryocytopoiesis in agar cultures.

A plasma fraction that stimulates platelet production in vivo also stimulates megakaryocytopoiesis in vitro. The plasma activity is attributed to a humoral regulator, thrombopoietin. Addition of the thrombocytopenic plasma (TP) fraction to an agar system supporting megakaryocyte colonies increased the frequency of colony formation significantly over that stimulated by spleen cell-conditioned medium (SCM). TP had no effect on the size of the colonies and, in the absence of SCM, TP did not stimulate colony formation. In studies of single megakaryocytes, the numbers of small megakaryocytes, specifically those 5-10 microns in diameter, increased significantly after 3 days of incubation with TP alone. SCM supported not an increase in the numbers, but an increase in the proportion of larger (30- to 40-microns) megakaryocytes. A normal plasma fraction contained similar but consistently less activity than fractions containing TP. The findings indicated that TP stimulates differentiation of megakaryocyte precursors from unidentifiable to identifiable cells but does not alone support colony formation. Thus, TP appears to be a potentiator of megakaryocytopoiesis. However, the augmentation of colony frequency by TP further suggests that TP may also play a role in early colony development, either by enhancing progenitor responsiveness to a megakaryocyte colony-stimulating factor or by recruiting additional colony progenitors from a noncycling progenitor population. These studies establish a link between the stimulation of platelet production observed after TP administration in vivo and the effects of TP on early events in megakaryocytopoiesis.

Recombinant human granulocyte-macrophage colony-stimulating factor promotes megakaryocyte maturation in nonhuman primates.

Megakaryocytes are responsive to several nonlineage-specific cytokines in vitro. In this study, we examined the in vivo effects of recombinant human granulocyte-macrophage colony-stimulating factor (rh GM-CSF) on late stages of megakaryocytopoiesis in the rhesus monkey. Four rhesus monkeys were given 10 micrograms/kg body weight/day of rh GM-CSF s.c. in two divided doses daily for 8 days. Megakaryocyte maturation was evaluated serially by measuring nuclear ploidy and cytoplasmic size. GM-CSF-treated monkeys developed significant shifts in ploidy distribution from days 3 through 15 (p less than or equal to 0.001), with increased frequencies of 64N and 128N megakaryocytes. Mean megakaryocyte size increased 92.5% on day 9, paralleling the increase in DNA content, although megakaryocyte size within ploidy groups did not increase. Megakaryocyte number remained unchanged following rh GM-CSF treatment. The platelet count responses were variable, and mean platelet volume did not change. The present study demonstrates that therapeutic doses of rh GM-CSF stimulate megakaryocyte endomitosis and increase mean size. The data indicate that GM-CSF is effective in promoting the maturation stage of megakaryocyte development but does not result in a consistent thrombopoietic response.

Maintenance of normal platelet mass in anemic Belgrade rats and their response to iron.

The Belgrade (b/b) rat has severe hypochromic, microcytic anemia accompanied by mild thrombocytopenia and a 49% reduction in megakaryocytes (MKs). The platelet counts are decreased only 34%, but relative platelet size measured by two independent methods averages 50% greater than controls. Thus, the platelet mass of the b/b rat is within the normal range. The marrow MK progenitors (MK colony-forming units, CFU-MK) respond linearly to increased colony-stimulating activity in vitro, but they are reduced 68% and form smaller colonies than normal. Flow cytometric analysis of MK ploidy indicates that significantly more MKs are distributed into the low and high ploidy classes compared with the normal, and the mean ploidy is similar. The b/b rat maintains effective thrombocytopoiesis in spite of a severe reduction in MK progenitors, primarily by an increased rate of maturation of the endomitotic compartment. Iron treatment partially arrests the b/b anemia and is associated with a significant increase in CFU-MK, a normalized MK ploidy distribution, and a significant decrease in platelet size. The favorable response to iron therapy suggests that the megakaryocytopenia is secondary to the severe anemia and results from stem cell commitment to the erythroid lineage.

HIV and hemophilic children's growth.

The acquired immune deficiency syndrome (AIDS) often has profound effects on growth; however, the effects of human immunodeficiency virus (HIV) on asymptomatic children's growth are unknown. Before heat inactivation/HIV donor screening of factor concentrates, many hemophilic children became infected with HIV. We evaluated four hemophilic groups without AIDS, using age-standardized growth parameters: group 1, 41 HIV-seropositive children (median age of 13 years); group 2, 11 HIV-seronegative children (median age of 4 years); group 3, 20 children frequently receiving concentrates, evaluated before 1979 (median age of 9 years); and group 4, 11 children rarely receiving concentrates, evaluated before 1979 (median age of 6 years). Median height for age (HA), weight for age (WA), and weight for height (WH) of groups 1 and 2 exceeded the 50th percentile of referent norms. HA, WA, WH, and weight/height did not vary significantly by group, nor did these decline over periods of 11 to 70 months. However, for those less than 11 years of age in group 1, HA declined by 25 percentile points over at least a 3 year period. Also, group 1's T helper-to-suppressor cell ratios at 12 +/- 3 months following the latest growth evaluation were positively associated with both HA and WA at the last evaluation. Eight children were evaluated before 1979 and again after they seroconverted to HIV.(ABSTRACT TRUNCATED AT 250 WORDS)

The epidemiology of venous thromboembolism in Caucasians and African-Americans: the GATE Study.

The aim of this study was to assess, comprehensively, medical and genetic attributes of venous thromboembolism (VTE) in a multiracial American population. The Genetic Attributes and Thrombosis Epidemiology (GATE) study is an ongoing case-control study in Atlanta, Georgia, designed to examine racial differences in VTE etiology and pathogenesis. Between 1998 and 2001, 370 inpatients with confirmed VTE, and 250 control subjects were enrolled. Data collected included blood specimens for DNA and plasma analysis and a medical lifestyle history questionnaire. Comparing VTE cases, cancer, recent surgery, and immobilization were more common in caucasian cases, while hypertension, diabetes, and kidney disease were more prevalent in African-American cases. Family history of VTE was reported with equal frequency by cases of both races (28-29%). Race-adjusted odds ratios for the associations of factor V Leiden and prothrombin G20210A mutations were 3.1 (1.5, 6.7) and 1.9 (0.8, 4.4), respectively. Using a larger external comparison group, the odds ratio for the prothrombin mutation among Caucasians was a statistically significant 2.5 (1.4, 4.3). A case-only analysis revealed a near significant interaction between the two mutations among Caucasians. We found that clinical characteristics of VTE patients differed across race groups. Family history of VTE was common in white and black patients, yet known genetic risk factors for VTE are rare in African-American populations. Our findings underscore the need to determine gene polymorphisms associated with VTE in African-Americans.

The beta fibrinogen gene G-455-A polymorphism is a risk factor for Legg-Perthes disease.

Legg-Perthes disease is a pediatric hip disorder characterized by avascular necrosis of the femoral head. The etiology of Legg-Perthes disease may involve repeated interruptions of the blood supply to the proximal femur. Thus, the role of thrombosis in Legg-Perthes disease is of interest. The focus of this analysis is an evaluation of the relationship between Legg-Perthes disease and the beta fibrinogen gene G-455-A polymorphism in 55 cases of Legg-Perthes disease and 56 age, race, and gender-matched healthy controls. Parents of subjects completed a questionnaire about their child's lifestyle and medical history. Blood was obtained for plasma and DNA analysis. Study subjects were predominantly white (93%), male (77%) and under age 16 (70%). Cases were more likely to be exposed to passive smoke than were controls (odds ratio 5.6, 95% confidence interval 2.0-12.0). Assuming a dominant genetic model, individuals who possessed either the G/A or A/A genotype were over three times more likely to have Legg-Perthes disease compared to those without the polymorphism (odds ratio 3.4, 95% confidence interval 1.5-7.8). Separate analyzes by smoke exposure revealed that the excess risk of the G-455-A polymorphism occurred in those exposed (odds ratio 7.0) as opposed to those unexposed to passive smoke (odds ratio 1.9). Although this difference in the odds ratios is not statistically significant (P=0.2), it suggests a possible interactive effect of cigarette smoke and the b fibrinogen gene G-455-A polymorphism in the risk of developing Legg-Perthes disease.

Measurement of von Willebrand factor activity: relative effects of ABO blood type and race.

Tests based on three different principles are reported to measure the activity of von Willebrand factor (VWF): ristocetin cofactor (VWF:RCo), collagen binding (VWF:CB), and the so-called "activity ELISA" (VWF:MoAb). We measured these and other diagnostic parameters in a population of 123 randomly selected female study controls, age 18-45 years. Type O subjects had significantly lower levels than non-O subjects in each test. Race differences were seen in all tests except VWF:RCo, with Caucasians having significantly lower levels than African-Americans. ABO differences accounted for 19% of the total variance in VWF:Ag (P < 0.0001) and race for 7% (P < 0.0001), for a total of 26%. Both effects were mediated through VWF:Ag and were independent. VWF:Ag level was the primary determinant of VWF function, accounting for approximately 60% of the variance in VWF:RCo and VWF:CB and 54% of the variance in factor VIII. The ratio VWF:RCo/VWF:Ag differed significantly by race within blood group. The median ratios were 0.97 for type O Caucasians vs. 0.79 for type O African-Americans and 0.94 for non-O Caucasians vs. 0.76 for non-O African-Americans. The ratio VWF:CB/VWF:Ag did not vary. This suggests racial differences in the interaction of VWF with GP1b but not with subendothelium. Alternatively, VWF:RCo may be regulated to maintain a relatively constant plasma level in the presence of excessive VWF:Ag. This heterogeneity within the normal population is partially responsible for the difficulty in defining diagnostic limits for von Willebrand disease.

Platelet functional defects in women with unexplained menorrhagia.

Menorrhagia is a common clinical problem and is unexplained in more than 50% of women. Although studies suggest that von Willebrand's Disease (VWD) is found in a substantial number of women with unexplained menorrhagia, the prevalence of platelet defects in women with menorrhagia is unknown. To determine the prevalence of platelet and other hemostatic defects, we evaluated women ages 17-55 diagnosed with unexplained menorrhagia. Seventy-four women (52 white, 16 black, six other) were studied. Bleeding time was prolonged in 23 women (31.5%). Maximal percent platelet aggregation was decreased with one or more agonists in 35 (47.3%) women. The most commonly found platelet function defects were reduced aggregation responses to ristocetin in 22 women and to epinephrine in 16 women. Sixteen of 22 women with reduced ristocetin aggregation had von Willebrand ristocetin cofactor (VWF:RCo) and von Willebrand factor antigen (VWF:Ag) > 60%. Platelet ATP release was decreased with one or more agonists in 43 (58.1%) women. Of the black women studied, 11/16 (69%) had abnormal platelet aggregation studies compared with 20/52 white women (39%) (P = 0.06). Black women with menorrhagia had a higher prevalence of decreased platelet aggregation in response to ristocetin and epinephrine than did white women (P = 0.0075, P = 0.02). Ten women (13.5%) had VWF:RCo and/or VWF:Ag < 60%. Using race and blood group specific ranges, 5 (6.8%) women had decreased VWF:RCo, VWF:Ag and/or collagen binding (VWF:CB). Mild factor XI deficiency was found in two women and one woman with mild factor V deficiency and one hemophilia A carrier were identified. We conclude that the prevalence of platelet function defects and other inherited bleeding disorders is substantial in a multiracial US population of women with unexplained menorrhagia.

Human immunodeficiency virus infection in hemophilic children.

The following groups were compared: (1) children less than 18 years old who have hemophilia-associated acquired immunodeficiency syndrome (AIDS) with other children with AIDS and with adults who have hemophilia-associated AIDS and (2) asymptomatic HIV-infected hemophilic children with asymptomatic HIV-infected hemophilic adults. Children with hemophilia-associated AIDS were older than other children with AIDS (medians 13 and 1 years, respectively) and less frequently had lymphocytic interstitial pneumonitis (5% v 48%) but had similar incidences of Pneumocystis carinii pneumonia (51% v 53%) and similar case to fatality ratios (59% v 61%). Children with hemophilia-associated AIDS had P carinii pneumonia significantly less often than did adults with hemophilia-associated AIDS, but both had similar case to fatality ratios (adults 72% with P carinii pneumonia, 68% dead). Significantly more hemophilic children than adults with AIDS were nonwhite (30% v 14%) and resided in the tristate area of New York/New Jersey/Pennsylvania (43% v 25%). The immune effects of human immunodeficiency virus (HIV) to date on asymptomatic pediatric and adult hemophiliacs are similar but may be more severe in adults. It is concluded that, although some of the clinical manifestations of AIDS (eg, lymphocytic interstitial pneumonitis) occurring or not occurring in older children infected through blood factor products differ from those of other children with AIDS, disease outcome to date is equally poor. The reasons for the differences between hemophilic children and hemophilic adults with and without AIDS warrant further investigation.

Endothelial cell protein S synthesis is upregulated by the complex of IL-6 and soluble IL-6 receptor.

We have recently demonstrated that the proinflammatory cytokine, interleukin-6 (IL-6), could upregulate the production of protein S in the human hepatoma cell line, HepG-2, but not in endothelial cells. In this study, we have demonstrated that the combination of exogenous IL-6 and soluble IL-6 receptor (sIL-6R) could significantly upregulate protein S production in both primary human umbilical vein endothelial cells (HUVEC) and in the immortalized human microvascular endothelial cell line, HMEC-1. The IL-6/sIL-6R complex was also able to rapidly induce tyrosine phosphorylation of the IL-6 transducer, gp130. Neutralizing antibodies directed against either IL-6 or gp130 blocked protein S upregulation by the IL-6/sIL-6R complex. It was also observed that exogenous sIL-6R could also upregulate protein S by forming a complex with IL-6 constitutively produced by the endothelial cell. Two other cytokines which also utilize the gp130 receptor, oncostatin M (OSM) and leukemia inhibitory factor (LIF), were also able to upregulate endothelial cell protein S. This study demonstrates a mechanism that allows endothelial cells to respond to IL-6 and also illustrates the potential importance of circulating soluble receptors in the regulation of the anticoagulation pathway.