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Biomedical subjects

R H Buckley

Publications and source records attributed to R H Buckley.

At least 37 records · Page 2Linked to original sources
PubMed1997

Mutation analysis of IL2RG in human X-linked severe combined immunodeficiency.

Severe combined immunodeficiency (SCID) is a syndrome of profoundly impaired cellular and humoral immunity. In humans, SCID is most commonly caused by mutations in the X-linked gene IL2RG, which encodes the common gamma chain, gamma c, of the leukocyte receptors for interleukin-2 and multiple other cytokines. To investigate the frequency and variety of IL2RG mutations that cause SCID, we analyzed DNA, RNA, and B-cell lines from a total of 103 unrelated SCID-affected males and their relatives using a combination of molecular and immunologic techniques. Sixty-two different mutations spanning all eight IL2RG exons were found in 87 cases, making possible correlations between mutation type and functional consequences. Although skewed maternal X chromosome inactivation, single-strand conformation polymorphism, mRNA expression, and cell surface staining with anti-gamma c antibodies were all helpful in establishing IL2RG defects as the cause of SCID, only dideoxy fingerprinting and DNA sequence determination each detected 100% of the IL2RG mutations in our series. Abnormal gamma c chains may be expressed in the lymphocytes of as many as two thirds of patients with X-linked SCID. Specific mutation diagnosis thus remains technically challenging, but it is important for genetic counseling and perhaps for helping to select appropriate subjects for retroviral gene therapy trials, This is a US government work. There are no restrictions on its use.

DNA Fingerprinting↗
PubMed1997

Successful formation of a chimeric human thymus allograft following transplantation of cultured postnatal human thymus.

Transplantation of cultured postnatal human thymus was performed in a patient with complete DiGeorge syndrome. Biopsy of the graft 3 mo after implantation revealed normal CD1+ thymocytes in thymic cortical epithelial regions and CD1- thymocytes in thymic medullary epithelial regions, respectively. HLA analysis of graft thymocyte and thymic microenvironment components demonstrated that developing thymocytes and thymic macrophages were recipient derived, while thymic epithelial components were of donor origin. The patient, who initially had no T cells and had profoundly defective T cell function, developed normal T cell responses to mitogens and Ags, tolerance to donor in a mixed lymphocyte reaction, and normal Ab titers after tetanus toxoid and pneumovax immunization. Thus, transplantation of cultured postnatal human thymic tissue in humans can form functional chimeric thymic tissue, and may provide a strategy to reconstitute the peripheral T cell pool in select congenital and acquired immune deficiency syndromes.

Chimera↗
PubMed1997

Human severe combined immunodeficiency: genetic, phenotypic, and functional diversity in one hundred eight infants.

OBJECTIVE: To determine the relative frequencies of the different genetic forms of severe combined immunodeficiency (SCID) and whether there are distinctive characteristics of the particular genotypes. STUDY DESIGN: The demographic, genetic, and immunologic features of 108 infants with SCID who were treated consecutively at Duke University Medical Center were analyzed. RESULTS: Eighty-nine subjects were boys and 19 were girls; there were 84 white infants, 16 black infants, and 8 Hispanic infants. Forty-nine had X-linked SCID with mutations of common cytokine receptor gamma chain (gamma c), 16 had adenosine deaminase (ADA) deficiency, 8 had Janus kinase 3 (Jak3) deficiency, 21 had unknown autosomal recessive mutations, 1 had reticular dysgenesis, 1 had cartilage hair hypoplasia, and 12 (all boys) had SCID of undetermined type. Deficiency of ADA caused the most profound lymphopenia; gamma c or Jak3 deficiency resulted in the most B cells and fewest natural killer (NK) cells; NK cells and function were highest in autosomal recessive and unknown types of SCID. CONCLUSIONS: Different SCID genotypes are associated with distinctive lymphocyte characteristics. The presence of NK function in ADA-deficient, autosomal recessive, and unknown type SCIDs, and low NK function in a majority of gamma c and Jak3 SCIDs indicates that some molecular lesions affect T, B, and NK cells (gamma c and Jak3), others primarily T cells (ADA deficiency), and others just T and B cells.

Adenosine Deaminase↗
PubMed1997

Gastroesophageal reflux and severe combined immunodeficiency.

BACKGROUND: Gastrointestinal and respiratory symptoms and failure to thrive not associated with infections or medications were noted in patients with severe combined immunodeficiency. OBJECTIVE: The aim of our study was to determine the frequency of gastroesophageal reflux in patients with severe combined immunodeficiency. METHODS: We studied the case histories of 73 pediatric patients who had been treated at Duke University Medical Center for severe combined immunodeficiency between 1982 and 1995. Charts were reviewed for documentation of gastroesophageal reflux on the basis of clinical course and results of barium swallow, esophageal pH probe monitoring, or endoscopy. To compare the incidence of gastroesophageal reflux in patients with severe combined immunodeficiency to known high-risk populations, we additionally tabulated the underlying diagnoses in an age-matched group of patients who underwent Nissen fundoplication from 1990 to 1995. RESULTS: We found clinically significant gastroesophageal reflux in 15 of the 73 patients (20.5%), much higher than has been reported in the normal population (0.1% to 0.3%, p < 0.001). Of patients treated between 1990 and the present, 10 of 36 (27.7%) had significant gastroesophageal reflux compared with five of 37 patients (13.5%) in the previous years. Thus with greater recognition and improved methods for diagnosis, the observed incidence of gastroesophageal reflux has increased greatly. The clinical presentations were not different from those of patients with other well-documented underlying diagnoses. Seven of the 15 patients (46.6%) did not respond to medical treatment with antacids, H2-blockers, and prokinetic agents and underwent surgical treatment. Indications for surgery included persistent esophagitis, vomiting, pneumonia, and growth failure. CONCLUSIONS: The reason for the high incidence of gastroesophageal reflux in patients with severe T-cell disorders remains unclear. Considering the frequency of this association, early recognition and treatment is important to enable adequate nutrition and prevent damage to the esophagus and lungs.

Adolescent↗
PubMed1997

Specificity and function of "natural" antibodies in immunodeficient subjects: clues to B cell lineage and development.

The origin of natural antibodies has long been a subject of controversy. Polyreactive natural antibodies recognize multiple ligands and are thought to arise from B1 B cells. Natural antibodies against carbohydrate antigens such as Gal alpha 1-3Gal or against blood groups A and B are thought to be "elicited" by gut bacteria, but their origin is uncertain. To explore the origin of naturally occurring anticarbohydrate antibodies, the specificity and function of the xenoreactive antibodies and isohemagglutinins were investigated in immunodeficient subjects. Subjects with defects in T cell-dependent antibody synthesis had normal levels of xenoreactive natural antibodies, most of which, like xenoreactive antibodies from normal individuals, were specific for Gal alpha 1-3Gal. On the other hand, some subjects with hyper-IgM syndrome who were able to synthesize abundant quantities of xenoreactive antibodies and polyreactive antibodies were devoid of anti-Gal alpha 1-3Gal antibodies. These results suggest that the lineages of B cells giving rise to anti-Gal alpha 1-3Gal antibodies and isohemagglutinins are distinct from B1 B cells or at least exist at a more "advanced" stage of development than those B1 B cells that give rise to polyreactive antibodies. The findings also suggest that B cells which synthesize anti-Gal alpha 1-3Gal antibodies and isohemagglutinins may be distinct from B2 B cells or exist at a more "primitive" stage of development than B2 B cells that synthesize elicited antibodies in normal individuals.

Adolescent↗
PubMed1997

Multicenter crossover comparison of the safety and efficacy of Intraglobin-F with Gamimune-N, Sandoglobulin, and Gammagard in patients with primary immunodeficiency diseases.

The safety and clinical efficacy of a liquid, beta-propiolactone-stabilized intravenous gamma-globulin, Intraglobin-F, was evaluated in a multicenter, double-blind study comparing Intraglobin-F to Gamimune-N, Sandoglobulin, or Gammagard. beta-Propiolactone stabilizes the IgG molecule to decrease aggregate formation and is a potent virucidal agent that reduces the risk of viral transmission by intravenous gamma-globulin (IVIG) preparations. Twenty-seven patients with primary immunodeficiency diseases were enrolled at three centers. Each patient received 6 months of therapy with either Intraglobin-F or the IVIG preparation that they had received during the preceding 3 months, then crossed over to the other preparation. Twenty-three patients completed the study. One patient withdrew because of an adverse event, generalized urticaria. A second patient withdrew because of fatigue and perceived decreased efficacy. Adverse reactions were comparable and occurred in 8.7% of the infusions of Intraglobin-F and 6% of the infusions with Sandoglobulin. None were severe or life-threatening. There was no discernible difference in efficacy between any of the products. The number of days when patients noted symptoms in their diaries was similar for Intraglobin-F and the comparison preparations, 4158 vs 4143. Similarly, there were no differences in the number of physician visits (33 vs 22), days missed from work or school (405 vs 404), days with fever (41 vs 47), or days of prophylactic antibiotics (675 vs 642). There was an increase in the number of days when antibiotics were given therapeutically (578 vs 451); most of the difference was attributable to one patient. There also was a difference in the number of days of hospitalization (21 vs 0), but 19 of the days were accounted for by two patients. When the patients were asked to score their feeling of well-being on a scale of 1 to 5, with 1 being entirely well, the mean score for the patients on Intraglobin-F was 1.86 (range, 1.0 to 3.0), compared to 1.85 (range, 1.0 to 3.2) for patients while on the comparison preparations. Trough IgG levels were slightly lower during the period when patients were treated with Intraglobin-F compared to the other products. There were no abnormalities in blood chemistries or hematologic parameters. Thus, Intraglobin-F is comparable to three of the marketed IVIG preparations in efficacy and safety, as well as patient acceptability, and offers the additional benefit of an extra virucidal step to reduce further the risk of transmitting viral infections.

Adolescent↗
PubMed1997

Normalization of the peripheral blood T cell receptor V beta repertoire after cultured postnatal human thymic transplantation in DiGeorge syndrome.

Complete DiGeorge syndrome is an immunodeficiency disease characterized by thymic aplasia and the absence of functioning peripheral T cells. A patient with this syndrome was transplanted with cultured postnatal human thymic tissue. Within 5 weeks of transplantation, flow cytometry, T cell receptor V beta sequence analysis, and cell function studies showed the presence of oligoclonal populations of nonfunctional clonally expanded peripheral T cells that were derived from pretransplantation T cells present in the skin. However, at 3 months posttransplantation, a biopsy of the transplanted thymus showed normal intrathymic T cell maturation of host T cells with normal TCR V beta expression on thymocytes. By 9 months postransplantation, peripheral T cell function was restored and the TCR V beta repertoire became polyclonal, coincident with the appearance of normal T cell function. These data suggest that the transplanted thymus was responsible for the establishment of a new T cell repertoire via thymopoiesis in the chimeric thymic graft.

Base Sequence↗
PubMed1996

Mutations in the mu heavy-chain gene in patients with agammaglobulinemia.

BACKGROUND: Most patients with congenital hypogammaglobulinemia and absent B cells are males with X-linked agammaglobulinemia, which is caused by mutations in the gene for Bruton's tyrosine kinase (Btk); however, there are females with a similar disorder who do not have mutations in this gene. We studied two families with autosomal recessive defects in B-cell development and patients with presumed X-linked agammaglobulinemia who did not have mutations in Btk. METHODS: A series of candidate genes that encode proteins involved in B-cell signal-transduction pathways were analyzed by linkage studies and mutation screening. RESULTS: Four different mutations were identified in the mu heavy-chain gene on chromosome 14. In one family, there was a homozygous 75-to-100-kb deletion that included D-region genes, J-region genes, and the mu constant-region gene. In a second family, there was a homozygous base-pair substitution in the alternative splice site of the mu heavy-chain gene. This mutation would inhibit production of the membrane form of the mu chain and produce an amino acid substitution in the secreted form. In addition, a patient previously thought to have X-linked agammaglobulinemia was found to have an amino acid substitution on one chromosome at an invariant cysteine that is required for the intrachain disulfide bond and, on the other chromosome, a large deletion that included the immunoglobulin locus. CONCLUSIONS: Defects in the mu heavy-chain gene are a cause of agammaglobulinemia in humans. This implies that an intact membrane-bound mu chain is essential for B-cell development.

Agammaglobulinemia↗
PubMed1996

Correction of purine nucleoside phosphorylase deficiency by transplantation of allogeneic bone marrow from a sibling.

Deficiency of the purine salvage pathway enzyme purine nucleoside phosphorylase causes a combined immunodeficiency and neurologic abnormalities and is usually fatal in childhood. We report the first successful transplantation of bone marrow from a sibling with identical class II human leukocyte antigens in this condition, demonstrating correction of both lymphocyte metabolic and functional abnormalities.

Bone Marrow Transplantation↗
PubMed1996

Practice parameters for the diagnosis and management of immunodeficiency. The Clinical and Laboratory Immunology Committee of the American Academy of Allergy, Asthma, and Immunology (CLIC-AAAAI)

In this brief review, only the most useful immunologic tests available for defining host defects that lead to susceptibility to infection have been emphasized. It should be pointed out that those evaluations and tests ordered by the physician will rule out the vast majority of the currently recognized defects. Finally, it is important that any patients identified as abnormal by these screening tests be characterized as fully as possible in centers specializing in these diseases before therapy is initiated, since what may appear to be a simple diagnosis on the surface may be an indicator of more complex underlying problems.

Antibody Formation↗
PubMed1996

Gastrointestinal pathology in patients with common variable immunodeficiency and X-linked agammaglobulinemia.

Review of the medical records of 43 patients with common variable immunodeficiency (CVID) and 23 patients with X-linked agammaglobulinemia (XLAG) revealed a high incidence of chronic gastrointestinal complaints, most commonly diarrhea. Thirty-eight biopsies, four small-bowel resection specimens, and one autopsy from 10 patients with CVID and one patient with XLAG showed a wide range of abnormalities. A pattern resembling acute graft-versus-host disease, with apoptotic bodies and lymphocytes in crypts, was seen in the stomach (four patients), small bowel (three patients), and colon (three patients). Small-bowel specimens from three CVID patients with malabsorption showed mild to severe villous atrophy. Three CVID patients had Giardia in biopsies. Two cases of small bowel lymphoma associated with nodular lymphoid hyperplasia were identified in CVID patients. One patient's small bowel contained foamy histiocytes in the lamina propria, resembling Whipple's disease or chronic granulomatous disease, with numerous apoptotic bodies in crypts. Ultrastructurally, the histiocytes contained cellular debris. The patient with XLAG had recurrent fissuring necrosis of small bowel resembling Crohn's disease; a patient with CVID had colitis with features similar to ulcerative colitis. Poorly formed granulomas were seen in the stomach (one CVID patient) and the colon (two CVID patients). Lymphocyte populations were dominated by T cells; B cells were scarce except in lymphoid follicles in CVID patients with nodular lymphoid hyperplasia. Patients with CVID and XLAG manifest a spectrum of abnormalities in the gastrointestinal tract, with patterns superficially resembling graft-versus-host disease, inflammatory bowel disease, and Whipple's disease, but often lacking some of the diagnostic features of the diseases. Many of the CVID patients with chronic gastrointestinal complaints (62%) also had evidence of autoimmune phenomena, suggesting that in some patients the inflammatory process in the gastrointestinal tract has an autoimmune component.

Adolescent↗
PubMed1995

Mutation of Jak3 in a patient with SCID: essential role of Jak3 in lymphoid development.

Males with X-linked severe combined immunodeficiency (XSCID) have defects in the common cytokine receptor gamma chain (gamma c) gene that encodes a shared, essential component of the receptors of interleukin-2 (IL-2), IL-4, IL-7, IL-9, and IL-15. The Janus family tyrosine kinase Jak3 is the only signaling molecule known to be associated with gamma c, so it was hypothesized that defects in Jak3 might cause an XSCID-like phenotype. A girl with immunological features indistinguishable from those of XSCID was therefore selected for analysis. An Epstein-Barr virus (EBV)-transformed cell line derived from her lymphocytes had normal gamma c expression but lacked Jak3 protein and had greatly diminished Jak3 messenger RNA. Sequencing revealed a different mutation on each allele: a single nucleotide insertion resulting in a frame shift and premature termination in the Jak3 JH4 domain and a nonsense mutation in the Jak3 JH2 domain. The lack of Jak3 expression correlated with impaired B cell signaling, as demonstrated by the inability of IL-4 to activate Stat6 in the EBV-transformed cell line from the patient. These observations indicate that the functions of gamma c are dependent on Jak3 and that Jak3 is essential for lymphoid development and signaling.

Amino Acid Sequence↗
PubMed1995

Two mutational hotspots in the interleukin-2 receptor gamma chain gene causing human X-linked severe combined immunodeficiency.

Human severe combined immunodeficiency (SCID), a syndrome of profoundly impaired cellular and humoral immunity, is most commonly caused by mutations in the X-linked gene for interleukin-2 (IL-2) receptor gamma chain (IL2RG). For mutational analysis of IL2RG in males with SCID, SSCP screening was followed by DNA sequencing. Of 40 IL2RG mutations found in unrelated SCID patients, 6 were point mutations at the CpG dinucleotide at cDNA 690-691, encoding amino acid R226. This residue lies in the extracellular domain of the protein in a region not previously recognized to be significantly conserved in the cytokine receptor gene family, 11 amino acids upstream from the highly conserved WSXWS motif. Three additional instances of mutation at another CpG dinucleotide at cDNA 879 produced a premature termination signal in the intracellular domain of IL2RG, resulting in loss of the SH2-homologous intracellular domain known to be essential for signaling from the IL-2 receptor complex. Mutations at these two hotspots constitute > 20% of the X-linked SCID mutations found by our group and a similar proportion of all reported IL2RG mutations.

Amino Acid Sequence↗
PubMed1994

Growth-dependent regulation of cellular ceramides in human T-cells.

The role of ceramide, a putative lipid second messenger in the regulation of cell growth, was investigated in T-lymphocytes. An inverse relationship between the cellular concentrations of ceramide and the proliferative capacity of human T-lymphocytes was observed for cells treated with either interleukin-2 or phorbol ester plus ionomycin. The same relationship between cellular ceramide concentrations and DNA synthesis also was observed for cells derived from a cultured T-cell line, the Jurkat T-cells. Alternative approaches for modulating the cellular ceramide concentrations were employed to determine the relationship between sphingolipids and cell growth. Treatment of normal T-lymphocyte cultures with exogenous cell-permeable ceramide analogues or sphingosine stereoisomers decreased DNA synthesis. A similar effect was seen with stearylamine. Cells treated with D,L-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol, an inhibitor of UDP-glucosyl:ceramide transferase, accumulated cellular ceramide concentrations and had decreased DNA synthesis. These results define a correlation between the concentration of cellular ceramides and the capacity of T-lymphocytes to proliferate. However, the addition of bacterial sphingomyelinase to the T-cell medium caused an increase in ceramide concentrations (presumably at the plasma membrane), which did not affect cell growth. These results support the hypothesis that functionally distinct pools of ceramide may reside within the T-cell.

Cell Division↗
PubMed1994

Breakthroughs in the understanding and therapy of primary immunodeficiency.

In the 40 years since Ogden Bruton discovered agammaglobulinemia, more than 50 additional immunodeficiency syndromes have been described. Until recently, there was little insight into the fundamental problems underlying a majority of these conditions. Recently, however, the molecular bases of three X-linked immunodeficiency disorders have been reported. These include X-linked immunodeficiency with hyper IgM, X-linked agammaglobulinemia, and X-linked severe combined immunodeficiency. These remarkable accomplishments have been made possible through a combination of new knowledge of molecular signaling mechanisms between and within cells of the immune system and greatly improved approaches to disease loci mapping within the human genome. Improvements in the therapy of immunodeficiency diseases have been impressive, and the development of generally safe and effective intravenous immunoglobulin preparations and T cell depletion techniques that permit the use of non-HLA-identical bone marrow donors have been the most important advances over the past 14 years. The identification and cloning of the genes for several of the primary immunodeficiency diseases have obvious implications for potential future somatic cell gene therapy for these patients. The rapidity of these advances suggests that soon there will be many more to come.

B-Lymphocytes↗
PubMed1994

Hyper IgM syndrome associated with defective CD40-mediated B cell activation.

Recent studies show that most patients with X-linked hyper IgM syndrome have defects in the gene for CD40 ligand. We evaluated 17 unrelated males suspected of having X-linked hyper IgM syndrome. Activated T cells from 13 of the 17 patients failed to bind a soluble CD40 construct. In these patients, the sequence of CD40 ligand demonstrated mutations. By contrast, T cells from the remaining four patients exhibited normal binding to the CD40 construct. Sequencing of the cDNA for CD40 ligand from these patients did not show mutations. The possibility that hyper IgM syndrome in these four patients was due to abnormalities in the B cell response to CD40-mediated signals was examined. Peripheral blood lymphocytes were stimulated with anti-CD40 alone, IL4 alone or anti-CD40 plus IL4. In comparison with B cells from controls or patients with hyper IgM syndrome and mutant CD40 ligand, B cells from the patients with hyper IgM syndrome and normal CD40 ligand were defective in their ability to secrete IgE (P < 0.02) or express activation markers, CD25 and CD23 (P < 0.02) in response to stimulation with anti-CD40. The failure of these B cells to respond to CD40-mediated activation could not be attributed to a generalized deficiency in B cell activation because IL4 induced normal up-regulation of CD23 and CD25 expression. These findings indicate that hyper IgM syndrome may result from defects in expression of CD40 ligand by activated T cells or defects in CD40-mediated signal transduction in B cells.

Antigens, CD↗
PubMed1993

Haploidentical bone marrow stem cell transplantation in human severe combined immunodeficiency.

From May 1992 to March 1993, 50 infants with severe combined immunodeficiency (SCID) were given bone marrow transplants at Duke University Medical Center. None received chemotherapy for conditioning or for graft-versus-host disease (GVHD) prophylaxis. Forty-one received haploidentical parental marrow depleted of T cells by soybean lectin and sheep red blood cell resetting, and nine received HLA-identical marrow. Forty (80%) survived from 1 week to almost 11 years posttransplantation, including nine of nine (100%) HLA-identical marrow recipients and 31 of 41 haploidentical recipients. T-cell function was present within 2 weeks after transplantation of unfractionated HLA-identical marrow, but not until 3 to 4 months after T-cell-depleted haploidentical marrow stem cells. All 37 patients who are more than 4 months posttransplantation have good T-cell function, and all but one have 100% donor T cells. B-cell function developed slowly or not at all in some recipients of haploidentical marrow. Fourteen (four HLA-identical and 10 haploidentical recipients) have some donor B cells; 19 patients are receiving intravenous immune globulin (IVIG) therapy.

B-Lymphocytes↗