Efficacy and safety of Gammaplex(®) 5% in children and adolescents with primary immunodeficiency diseases.

This open-label multi-centre study evaluated Gammaplex(®) 5%, a human intravenous immunoglobulin (IVIG) 5% liquid, in 25 children and adolescent patients (aged 3-16 years) with primary immunodeficiency diseases (PIDs). Subjects received Gammaplex 5% (at doses of 300-800 mg/kg/infusion) for 12 months, with a 3-month follow-up. The primary efficacy end-point was the incidence of serious acute bacterial infections (SABIs) during the 12-month treatment period. Secondary objectives assessed safety and tolerability. Nineteen males and six females were treated using the same infusion schedule as their prior IVIG treatment (14 and 11 subjects on 21- and 28-day dosing schedules, respectively). Two SABIs of pneumonia were reported, resulting in an annual SABI event rate of 0·09 [upper one-sided 99% confidence interval (CI) = 0·36]. Twenty-one subjects (84%) experienced ≥ 1 infection during the study, with a median infective episode per subject/year of 3·08 (range = 0-10·4). Sixteen subjects (64%) missed ≥ 1 day of nursery or school because of infection or other illness. All trough immunoglobulin G levels exceeded 7·00 g/l after 15 weeks (mean = 9·69 g/l; range = 7·04-15·35 g/l). Product-related adverse events occurred in 14 subjects (56%); none were serious. Of 368 total infusions, 97 (26%) were associated temporally with an adverse event (≤ 72 h after infusion), regardless of causality. Laboratory test results and adverse-reaction data showed no evidence of product-related haemolysis or thromboembolic events. These data demonstrate that Gammaplex 5% is effective in preventing SABIs and well tolerated in children and adolescents with PID.

C1 esterase inhibitor concentrate in 1085 Hereditary Angioedema attacks--final results of the I.M.P.A.C.T.2 study.

BACKGROUND: The placebo-controlled study International Multicentre Prospective Angioedema C1-INH Trial 1 (I.M.P.A.C.T.1) demonstrated that 20 U/kg C1 esterase inhibitor (C1-INH) concentrate (Berinert®; CSL Behring, Marburg, Germany) is effective in treating acute abdominal and facial Hereditary Angioedema (HAE) attacks. METHODS: I.M.P.A.C.T.2 was an open-label extension study of I.M.P.A.C.T.1 to evaluate the safety and efficacy of long-term treatment with 20 U/kg C1-INH for successive HAE attacks at any body location. Efficacy outcomes included patient-reported time to onset of symptom relief (primary) and time to complete resolution of all symptoms (secondary), analysed on a per-patient and per-attack basis. Safety assessments included adverse events, vital signs, viral safety and anti-C1-INH antibodies. RESULTS: During a median study duration of 24 months, 1085 attacks were treated in 57 patients (10-53 years of age). In the per-patient analysis, the median time to onset of symptom relief was 0.46 h and was similar for all types of attacks (0.39-0.48 h); the median time to complete resolution of symptoms was 15.5 h (shortest for laryngeal attacks: 5.8 h; 12.8-26.6 h for abdominal, peripheral and facial attacks). Demographic factors, type of HAE, intensity of attacks, time to treatment, use of androgens and presence of anti-C1-INH antibodies had no clinically relevant effect on the efficacy outcomes. There were no treatment-related safety concerns. No inhibitory anti-C1-INH antibodies were detected in any patient. CONCLUSIONS: A single dose of 20 U/kg C1-INH concentrate is safe and provides reliable efficacy in the long-term treatment of successive HAE attacks at any body location.

Efficacy and safety of a new immunoglobulin G product, Gammaplex(®), in primary immunodeficiency diseases.

This open-label multi-centre study evaluated a new intravenous immunoglobulin, Gammaplex®, in the treatment of 50 patients with primary immunodeficiency and significant hypogammglobulinaemia. Patients treated previously with other intravenous immunoglobulins received Gammaplex® on their same infusion schedule for 1 year; 22 were on a 21-day and 28 on a 28-day regimen (300-800 mg/kg/infusion). There were no serious, acute bacterial infections, whereas six subjects (12·0%) had at least one such infection in the 6 months before enrollment. Forty subjects (80·0%) had at least one non-serious infection; the median number of infective episodes per subject per year was 3·07. Antibiotics were taken by 38 subjects therapeutically and prophylactically by 16 at some time. Fewer than half (46·0%) missed any time off work or school because of infection or other illness. Trough immunoglobulin (Ig)G levels were above 6·00 g/l in all subjects at all assessments after 15 weeks with two exceptions. Overall, 21·2% of infusions were associated with an adverse event up to 72 h after infusion. The frequency of adverse events increased with infusion rate. Headache was the most common product-related adverse event (7·5% of 703 infusions). In conclusion, Gammaplex® is effective in primary immunodeficiency and is well tolerated.

Cockroach allergy appears early in life in inner-city children with recurrent wheezing.

BACKGROUND: Cockroach allergy and exposure to high levels of this allergen are important in the increasing asthma-related health problems among young inner-city children. However, there are very little data regarding the prevalence of cockroach allergy in infants and young children with asthma. OBJECTIVE: This retrospective study was designed to test the hypothesis that cockroach allergy appears early in life in young children with recurrent wheezing. METHODS: We reviewed the medical records of all 196 children (ages 5 months to 16 years) evaluated between January 1995 and September 1997 at the Cook County Hospital Pediatric Allergy Clinic for recurrent wheezing. The patients were assigned into two age groups, less than 4 years old and 4 to 16 years old. The percentages of IgE skin tests positive for common aeroallergens were compared within and between the two age groups. All children in the younger age group were tested for cockroach and dust mites, cat, and dog when indicated by positive environmental history. All children in the older age group were tested for indoor and outdoor allergens. RESULTS: Sixty-three children were younger than 4 years of age, and of these, 15 (23.8%) had cockroach allergen sensitivity, compared with only eight patients (12.7%) who were skin test positive to dust mite allergen (P = .01). The youngest patient with a positive reaction to cockroach allergen was 6 months old. Patients with a single allergen skin reactivity were considered as monosensitized. Nine children younger than 4 years of age (14.3%) were monosensitized only to cockroach allergen in contrast to three children (4.8%) who were monosensitized to house-dust mites (P < .05). CONCLUSIONS: Our data suggest that cockroach allergen sensitivity starts early in life and may be the only sensitizing allergen in many young inner-city children.

Asthma care practices, perceptions, and beliefs of Chicago-area primary-care physicians. Chicago Asthma Surveillance Initiative Project Team.

INTRODUCTION: Although primary-care physicians were a principal target audience for the National Asthma Education and Prevention Program (NAEPP), there is little published information describing the postguideline asthma care practices of these physicians or their willingness to embrace the NAEPP guidelines. This study examines asthma care practices of Chicago-area primary-care physicians and assesses these practitioners' perceptions and beliefs about several aspects of the NAEPP guidelines. METHODS: In 1997, a self-administered survey was mailed to a randomly selected 10% sample of Chicago-area general pediatricians, internists, and family practitioners. RESULTS: Surveys were returned by 244 of the 405 eligible Chicago-area primary-care physicians (60.2%) in the sample. Of these, 66 (27.6%) were pediatricians, 83 (34.7%) were general internists, and 90 (37.7%) were family practitioners. Physicians reported that 54.6 +/- 2.7% (mean +/- SE) of patients with newly diagnosed asthma have spirometry performed as part of their initial evaluation. For patients with moderate persistent asthma, prescribing of inhaled corticosteroids varied by patient age, with 60.5% of physicians routinely prescribing them for patients < 5 years, compared with 95.7% of physicians prescribing them for patients > or = 5 years. Awareness of the NAEPP guide-lines among these physicians was high, with 88.5% reporting that they have heard of the guidelines, and 73.6% reporting having read them. Of patients with moderate or severe persistent asthma, physicians estimated that 47.7 +/- 2.7% were given written treatment plans. CONCLUSION: Several aspects of the NAEPP guidelines appear to have been incorporated into clinical practice by Chicago-area primary-care physicians, whereas other recommendations do not appear to have been readily adopted. This information suggests areas for interventions to improve primary care for asthma in the Chicago area.

Asthma care practices, perceptions, and beliefs of Chicago-area asthma specialists. Chicago Asthma Surveillance Initiative Project Team.

INTRODUCTION: Few studies have closely explored how well physicians who consider themselves specialists in asthma adhere to national guideline recommendations for the diagnosis and treatment of asthma. The purpose of this study is to characterize current knowledge, attitudes, beliefs, and self-reported treatment practices of the asthma specialists working in one large metropolitan area. METHODS: In 1997, a cross-sectional survey was mailed to asthma specialists (allergists or pulmonologists) engaged in direct patient care with a practice location in the Chicago area (Cook County or one of the five surrounding counties). An approximately 50% random sample of asthma specialists was surveyed. The survey included items on (1) asthma diagnosis; (2) clinical monitoring of asthma patients; (3) pharmacologic and nonpharmacologic asthma treatment; (4) opinions and beliefs about asthma treatment options and reasons for referrals; (5) involvement in continuing medical education; (6) experiences with managed care; (7) use of asthma practice guidelines; (8) demographic information about the respondents; and (9) characteristics of the practice settings. RESULTS: A total of 113 eligible surveys were returned (response rate, 72.0%). Ninety-nine percent of the respondents indicated they would prescribe inhaled corticosteroids for patients > or = 5 years old with moderate persistent asthma, and 85.5% would prescribe them for patients < 5 years old. The respondents reported that 71.2% of their patients with moderate or severe persistent asthma were routinely given written treatment plans. The use of these plans was reported more frequently by allergists than pulmonologists (77.6% vs 58.9%, p = 0.01). Nearly half of the respondents were involved in the development of hospital-based asthma programs; fewer (14.9%) were involved in developing asthma programs for managed care organizations. A majority (63.4%) of the physicians had given a formal professional education presentation on asthma in the past year. A majority of the respondents who care for patients under managed care contracts reported that these patients have encountered barriers to access in seeking specialty care. CONCLUSION: The results suggest that asthma specialists in the Chicago area are providing asthma care that is, in many ways, consistent with national guidelines. However, there are also important differences in care that are not consistent with the guideline recommendations. Perhaps even more notable are differences in reported asthma care between the two subspecialty groups of allergists and pulmonologists. The effect of these differences on the management of persons with asthma is not known. It is hoped that information from this community-based survey will serve to catalyze discussions among Chicago-area asthma specialists as to how they might envision improving care for persons with asthma in their community.

Prevalence and burden of illness for asthma and related symptoms among kindergartners in Chicago public schools.

BACKGROUND: Asthma mortality rates in poor communities of Chicago are among the highest in the country. Possible explanations include increased asthma prevalence, increased severity, and suboptimal health care. OBJECTIVE: To estimate the prevalence of asthma and asthma-related symptoms among inner-city kindergarten children, and to characterize their burden of illness, asthma-related health care access, and pharmacologic treatment. METHODS: Cross-sectional survey of parents of kindergartners was conducted in 11 randomly selected Chicago elementary schools. A self-administered 16-item questionnaire was given to parents of kindergartners. Parents who reported doctor-diagnosed asthma or at least one of several key asthma-related symptoms were then interviewed with a supplemental questionnaire examining asthma-related health care and medication use. RESULTS: Based on data from 638 children [mean age 5.7 (SD = 0.6) years], the prevalence of diagnosed asthma was 10.8%. Sixteen percent of the respondents reported that their child had wheezed in the past year. The prevalence of asthma-related symptoms unassociated with a diagnosis of asthma was 30.1%. The children with diagnosed asthma had evidence of a high burden of illness: over 40% were reported to have had sleep disturbance due to wheezing > or =1 to 2 nights/week and 86.6% reported acute care visits for respiratory symptoms in the past year. Self-reported access to medical care was high. Over 40% of the children with doctor diagnosed asthma were reported to have used a beta2-agonist in the preceding 2 weeks, and 12.2% used an inhaled anti-inflammatory. CONCLUSIONS: These data suggest that asthma prevalence in school-aged children in inner-city communities may be higher than US estimates. The burden of illness experienced by these children is substantial. Also, a large proportion of children were reported to have respiratory symptoms consistent with asthma, and no asthma diagnosis, suggesting possible undiagnosed asthma. While measures of health care access appear to indicate that the majority of children with asthma experience no identified barriers to health care, there is evidence to suggest undertreatment.

Respiratory syncytial virus stimulates neutrophil degranulation and chemokine release.

Neutrophil infiltration of the airways is a common finding in respiratory syncytial virus (RSV) bronchiolitis. Neutrophil-derived chemokines and neutrophil granule contents can cause further inflammation, hyperresponsiveness, and damage of the airways. In this study, peripheral blood neutrophils incubated with RSV (multiplicity of infection (MOI) = 10) induced IL-8, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, and myeloperoxidase (MPO) release. In contrast, LPS induced only chemokine but not MPO release. RSV-induced chemokine and MPO release was noncytotoxic as assessed by trypan blue exclusion. The mechanism of RSV-induced chemokine release was shown to be transcription dependent since cytokine mRNA synthesis was increased with RSV stimulation and the process was inhibited by actinomycin-D. In addition, the effect of dexamethasone (dex) on mediator release was also studied. Dex significantly inhibited chemokine release but did not inhibit MPO release. The mechanism of inhibition of the release of these chemokines is probably posttranscriptional since the mRNA synthesis was not inhibited by dex. We conclude that the release of chemokines (IL-8, MIP-1alpha, MIP-1beta) and granule enzymes (MPO) by RSV-stimulated neutrophils may contribute to the pulmonary pathology in RSV bronchiolitis. These in vitro findings showing that dex failed to consistently inhibit all the RSV-induced release of neutrophil inflammatory mediators may explain the variable efficacy of corticosteroids in the treatment of RSV bronchiolitis.

Stimulus-specific regulation of chemokine expression involves differential activation of the redox-responsive transcription factors AP-1 and NF-kappaB.

The promoters of the IL-8, MCP-1, and RANTES genes contain binding sites for the redox-responsive transcription factors AP-1 and NF-kappaB, which have been shown to be important for their expression. In this overview, we present evidence from our laboratories that the stimulus-specific regulation of these chemokines by the reactive oxidant H2O2, the proinflammatory cytokine TNF-alpha, and respiratory syncytial virus (RSV) is mediated in a cell type-specific manner involving different patterns of AP-1 and NF-kappaB binding activity. Our results demonstrate that H2O2 induction of IL-8 gene expression is linked with the selective binding of AP-1 to the IL-8 promoter, whereas TNF-alpha and RSV induction of IL-8 correlates with the activation of NF-kappaB binding. We propose that the differential activation and binding of inducible transcription factors to the promoter regions of chemokine genes provides a critical regulatory mechanism by which the CXC and CC chemokines can be selectively expressed in a cell type-specific and stimulus-specific manner. Such a regulatory mechanism of differential chemokine expression could critically influence the site-specific recruitment of distinct subsets of leukocytes to sites of inflammation and injury.

Myocardial infarction as the presenting manifestation of systemic lupus erythematosus with antiphosphatidylserine antibodies.

BACKGROUND: The presence of antiphospholipid antibodies has been associated with thrombotic events in systemic lupus erythematosus and other diseases. Antiphosphatidylserine antibodies have been shown more recently to have clinical implications being noted in thrombocytopenia, positive VDRL test, prolonged partial thromboplastin time, and recurrent fetal loss. OBJECTIVE: To report a patient with systemic lupus erythematosus who presented with a myocardial infarction resulting from a single clot in the distal left anterior descending coronary artery and to discuss the merits of evaluating such patients for the presence of antiphospholipid antibodies. METHODS: Case report and review of the literature. RESULTS: A 55-year-old woman with no risk factors for coronary artery disease presented with a myocardial infarction. A distal left anterior descending coronary artery clot was seen on angiography. Evaluation for the presence of a hypercoagulable state revealed evidence for systemic lupus erythematosus. Although lupus anti-coagulant and anticardiolipin antibody titers were negative, antiphosphatidylserine IgA antibodies were detected. CONCLUSION: Because antiphosphatidylserine antibodies of the IgA isotype have been shown to be associated with thrombosis and other manifestations of systemic lupus erythematosus, a complete evaluation for antiphospholipid antibodies should include testing for all three isotypes.

Analysis of signaling events associated with activation of neutrophil superoxide anion production by eosinophil granule major basic protein.

This study was undertaken to identify the signaling events involved in activation of neutrophil superoxide anion (O2-) production by eosinophil granule major basic protein (MBP). MBP did not produce an immediate increase in the cytosolic free calcium concentration ([Ca2+]i), characteristic of phospholipase C activation, but did cause a gradual increase in [Ca2+]i in cytochalasin B-treated cells. Preincubation with 0.01 to 3 micrograms/mL pertussis toxin did not inhibit MBP-stimulated O2- production, and MBP did not stimulate an increase in diradylglycerol levels. MBP did stimulate a low level of phospholipase D activity, as measured by a time-dependent increase in phosphatidic acid and, in the presence of 0.5% ethanol, phosphatidylethanol. Inhibition of MBP-stimulated O2- production by genistein and Western blot analysis using an antiphosphotyrosine antibody showed tyrosine kinase activation by MBP. Calmodulin antagonists (calmidazolium and W-7) caused up to 80% inhibition of MBP-stimulated O2- production. In agreement with the pharmacologic sensitivity, MBP did not stimulate any 51Cr release. These data indicate that tyrosine kinase and calmodulin-dependent steps are involved in the noncytotoxic stimulation of neutrophil O2- production by MBP.

Expression, purification, and characterization of the recombinant proform of eosinophil granule major basic protein.

The cDNA for the highly toxic eosinophil granule major basic protein (MBP) encodes a 25-kDa acidic precursor (proMBP) that is processed to form the 14-kDa mature MBP. To characterize the biochemical and biological properties of proMBP, and compare these to the known properties of MBP, we expressed recombinant proMBP in Chinese hamster ovary cells and purified the secreted form from supernatants. We developed a mAb specific for proMBP, J163-15E10, and by using a proMBP-specific RIA we found that recombinant proMBP was expressed quite efficiently at levels between 10 and 100 mg/l. By SDS-PAGE and immunoblotting analyses of bulk Chinese hamster ovary supernatants, recombinant proMBP was electrophoretically heterogeneous with an apparent molecular mass ranging from 3 x 10(4) to 1 x 10(5) daltons. Despite difficulties encountered because of the extreme molecular heterogeneity of the proform, two methods for purification of a predominant 33-kDa form of recombinant proMBP are presented. Glycosylation analysis of purified 33-kDa proMBP indicated that approximately 5 kDa is likely accounted for by the addition of one glycosaminoglycan group, three O-linked, and one N-linked complex type carbohydrate groups. Functional studies of purified recombinant proMBP were also conducted. Using amounts of proMBP determined to be optimal for MBP activity, it was shown that proMBP not only lacked the ability to inhibit protein synthesis in K562 cells, but it also lacked the ability to stimulate basophil histamine release or generate neutrophil superoxide anion release. Furthermore, proMBP inhibited in a dose-responsive manner the basophil histamine release and superoxide anion generation stimulated by MBP. The development of a mAb and RIA specific for proMBP will now make it possible to analyze biologic fluids for the presence of this protein, especially in pregnancy, when proMBP is increased.

Eosinophil major basic protein enhances the expression of neutrophil CR3 and p150,95.

BACKGROUND: We examined the effect of major basic protein (MBP) stimulation on the expression of the neutrophil B2-integrins: LFA-1 (CD11a/CD18), CR3 (CD11b/CD18), and p150,95 (CD11c/CD18). METHODS: Incubation of neutrophils with 0.75 to 3.0 mumol/L MBP for 30 minutes at 37 degrees C resulted in concentration-dependent increases in CR3 expression as detected by staining with the CD11b-specific monoclonal antibody, Leu-15. RESULTS: The expression of CR3 was significantly (p < 0.001) higher when neutrophils were stimulated with 1.5 mumol/L (53% +/- 9% increase) or 3.0 mumol/L (100% +/- 17% increase) MBP as compared with unstimulated neutrophils. The kinetics of MBP-stimulated CR3 expression were rapid and were similar to those of 100 nmol/L. N-formyl-methionyl-leucyl-phenylalanine-stimulated enhancement of CR3 expression. Incubation of neutrophils with reduced and alkylated MBP resulted in significantly lower (p < 0.05) increases in CR3 expression as compared with stimulation with native MBP. In addition, neither eosinophil cationic protein nor eosinophil-derived neurotoxin altered neutrophil CR3 expression. MBP stimulated minimal increases in LFA-1 expression. However, staining with the monoclonal antibody Leu-M5 (anti-CD11c) revealed that MBP stimulated significant increases in the expression of p150,95. CONCLUSIONS: These results indicate that MBP stimulates the increased expression of neutrophil adhesion molecules, which in turn may enhance the inflammatory role of neutrophils in the late-phase events of allergic diseases.

Repeated inhalation of nebulized albuterol did not induce arrhythmias in a patient with Wolff-Parkinson-White syndrome and asthma.

It has been assumed that in asthmatic patients with Wolff-Parkinson-White (WPW) syndrome, ablative therapy for the condition is necessary for the safe treatment of the asthma with beta 2-adrenergic drugs. The following case report illustrates that inhaled albuterol was safely administered to an asthmatic patient with electrocardiographic evidence of preexcitation, without the need of an ablative procedure. This case report is, to our knowledge, the first in the literature in which the cardiac rhythm of a patient with WPW syndrome was monitored during repeated inhalations of a beta 2-agonist.

Noncytotoxic activation of neutrophils by eosinophil granule major basic protein. Effect on superoxide anion generation and lysosomal enzyme release.

Eosinophil granule major basic protein (MBP) and neutrophils have each been implicated in the inflammatory late phase events of allergic disease. Based on this association and flow cytometric evidence presented in this report for MBP binding to neutrophils, we examined the ability of MBP to activate human neutrophils. Incubation of neutrophils with 0.5 to 3.0 microM MBP at room temperature produced a concentration-dependent chemiluminescence (CL) response that peaked after 50 to 70 min. Reduced-and-alkylated MBP, eosinophil cationic protein, and eosinophil-derived neurotoxin did not induce CL. MBP-induced CL was abrogated in the absence of Ca2+ and was absent in neutrophils isolated from two individuals with chronic granulomatous disease. MBP also stimulated release of superoxide anion (O2-) and lysozyme but not beta-glucuronidase or lactate dehydrogenase. Additionally, 1.5 microM MBP in combination with FMLP or platelet-activating factor stimulated a synergistic increase in O2- release from cytochalasin B-treated neutrophils. The degree of synergism with FMLP or platelet-activating factor was inversely related (p less than 0.005) to the level of MBP-induced O2- release. These results indicate that MBP activates neutrophils in a noncytolytic fashion and provide evidence that eosinophil-neutrophil collaboration may contribute to the pathogenesis observed in allergic late phase reactions.

Identification of an IgA inhibitor of neutrophil chemotaxis and its membrane target for the metabolic burst.

Affinity-purified IgA from the serum of an 8-year-old boy with a 5-year history of recurrent facial nodules, intermittent neutropenia and elevated immunoglobulin levels, inhibited the chemotaxis of polymorphonuclear neutrophils (PMN) from both patient and normal adults. Preincubation of normal PMN with IgA from the patient's serum (0.5 mg/ml) inhibited chemotaxis to C5a and to the chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine (FMLP) by 80%, while IgA or IgG from pooled human serum and IgG from the patient were without effect. Normal PMN chemotaxis was restored after IgA depletion of the patient's serum by affinity chromatography. The patient's IgA, but not IgA from pooled human serum, bound specifically to normal PMN by its antigen-binding sites and recognized a 62,000 MW membrane protein on normal neutrophils, which was distinct from the FMLP receptor, the C5a receptor, or the Fca receptor. Attachment of the patient's IgA to the 62,000 MW protein activated intracellular oxidative metabolism on a parity with phorbol myristate acetate (PMA) and resulted in a significant up-regulation of membrane receptors for FMLP. After the binding of patient (Pt) IgA, normal neutrophils were rendered significantly less responsive to subsequent stimulation with phorbol esters. These results characterize a novel mechanism of chemotactic inhibition by serum IgA and also identify a neutrophil membrane protein that is linked to intracellular oxidative metabolism.

Heparan sulfates from Swiss mouse 3T3 and SV3T3 cells: O-sulfate difference.

A difference in the extent of sulfation between the heparan sulfate isolated from Swiss 3T3 mouse cells and that from Swiss 3T3 cells transformed by the DNA virus SV40 has been reported previously. This variance is manifested by different chromatographic and electrophoretic properties. Heparan sulfates from the two cell types were treated with nitrous acid under conditions that gave selective deaminative cleavage of glucosaminyl residues with sulfated amino groups in order to define the nature of the difference in sulfation further. The O-sulfate containing fragments from the heparan sulfates were compared by gel filtration and ion-exchange chromatography. The results showed that the 3T3 heparan sulfate contains 8% more O-sulfate than does the SV3T3 heparan sulfate. Analysis of uronic acids revealed that both types of heparan sulfates contain 45% L-iduronic acid and 55% D-glucuronic acid. These and other observations indicate that the primary difference in sulfation between the 3T3 and SV3T3 heparan sulfates lies in the extent of O-sulfation.