Steroids and immunizations.
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Adrenal Cortex Hormones↗
Biomedical subjects
H D Ochs
Publications and source records attributed to H D Ochs.
The role of CD40 and its ligand in the regulation of the immune response.
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Amino Acid Sequence↗
A point mutation in gp91-phox of cytochrome b558 of the human NADPH oxidase leading to defective translocation of the cytosolic proteins p47-phox and p67-phox.
The superoxide-forming NADPH oxidase of human phagocytes is composed of membrane-bound and cytosolic proteins which, upon cell activation, assemble on the plasma membrane to form the active enzyme. Patients suffering from chronic granulomatous disease (CGD) are defective in one of the following components: p47-phox and p67-phox, residing in the cytosol of resting phagocytes, and gp91-phox and p22-phox, constituting the membrane-bound cytochrome b558. In an X-linked CGD patient we identified a novel missense mutation predicting an Asp-->Gly substitution at residue 500 of gp91-phox, associated with normal amounts of nonfunctional cytochrome b558 in the patient's neutrophils. In PMA-stimulated neutrophils and in a cell-free translocation assay with neutrophil membranes and cytosol, the association of the cytosolic proteins p47-phox and p67-phox with the membrane fraction of the patient was strongly disturbed. Furthermore, a synthetic peptide mimicking domain 491-504 of gp91-phox inhibited NADPH oxidase activity in the cell-free assay (IC50 about 10 microM), and the translocation of p47-phox and p67-phox in the cell-free translocation assay. We conclude that residue 500 of gp91-phox resides in a region critical for stable binding of p47-phox and p67-phox.
Amino Acid Sequence↗
The random inactivation of the X chromosome carrying the defective gene responsible for X-linked hyper IgM syndrome (X-HIM) in female carriers of HIGM1.
The molecular origin of X-linked hyper IgM syndrome has recently been identified as a defect in the ligand of CD40, gp39, a protein expressed on the surface of activated T cells. The availability of detailed pedigrees for three families with affected males allowed assessment of the random or nonrandom nature of the inactivation of the defective X chromosome as well as a determination of the origin of the mutation. X chromosome inactivation was studied because of the relevance to the ability to detect carriers of HIGM1 and the potential for phenotypic effect in the carriers. Using immunostaining, PCR, and DNA sequencing, we found that the defective gene for gp39 is not selectively inactivated. Even in the presence of extremely skewed inactivation, normal levels of serum Ig were found. In carriers in which the defective gene is predominantly expressed, staining alone revealed the carrier status reliably while cloning and sequencing of the cDNA was necessary when the normal gene was predominantly expressed. Unlike some other X-linked defects where extreme Lyonization may lead to disease, a small population of cells expressing the wild-type gp39 is sufficient to maintain normal humoral immunity and prevent the clinical symptoms of X-HIM.
Adolescent↗
The molecular basis of X-linked agammaglobulinemia, hyper-IgM syndrome, and severe combined immunodeficiency in humans.
The molecular basis for X-linked agammaglobulinemia, hyper-IgM syndrome, and severe combined immunodeficiency was recently identified. In X-linked agammaglobulinemia the molecular defect was found to reside in the gene encoding a novel cytoplasmic tyrosine kinase (bpk, atk, or btk) expressed by B and myeloid cells. This kinase belongs to a new subfamily of tyrosine kinases that contains SH1, SH2, and SH3 domains. A defect in the murine homologue of this kinase has been shown to be responsible for X-linked immunodeficiency in mice. Currently, the role of btk in B- and myeloid cell signaling is unknown. The molecular defect in X-linked hyper-IgM syndrome has been shown to reside in the gene encoding the T-cell activation protein gp39 (CD40L, TRAP). This protein binds to its counter receptor, CD40, on B cells and has been shown to participate in T-cell-dependent B-cell help leading to B-cell proliferation and isotype switching. X-linked severe combined immunodeficiency patients were found to have defects in the gene encoding the gamma-chain of the interleukin-2 receptor. This chain of the interleukin-2 receptor is constitutively expressed by T cells and is involved in the formation of high and intermediate affinity interleukin-2 receptor complexes. These two interleukin-2 receptor complexes are responsible for mediating interleukin-2-dependent signals.
Agammaglobulinemia↗
Specific antibody production to a recall or a neoantigen by SCID mice reconstituted with human peripheral blood lymphocytes.
To explore the extent of immune reconstitution of SCID mice by human peripheral blood lymphocytes (hu-PBL-SCID mice), we studied the production of immunoglobulin isotypes and specific antibody (Ab) by the engrafted human cells. Human IgG was detectable in 94% of hu-PBL-SCID mice. IgE synthesis by hu-PBL-SCID mice correlated with the IgE levels observed in human donors. All SCID mice receiving PBL obtained from human donors previously immunized with the T-cell-dependent Ag, bacteriophage phi x 174 (phage), produced phage neutralizing antibody. Quantity and quality (Ig isotypes) of phage-specific Ab produced by hu-PBL-SCID mice correlated with that observed in the donor serum. Human B cells alone failed to engraft, and T cells were required for the production of Ig and anti-phage Ab. Phage-specific Ab production occurred without direct Ag exposure of the hu-PBL-SCID mice, suggesting that the specific Ab production was induced directly by polyclonal activation of the engrafted human cells. Intravenous phage injections given 4 wk after cell transfer failed to further increase the anti-phage Ab titer. Phage neutralizing Ab production could not be boosted if spleen cells obtained from hu-PBL-SCID mice were cultured in the presence of Ag. However, hu-PBL-SCID mice produced increased amounts of anti-phage Ab, providing they were injected with phage at the time of cell transfer. Injection of phage at the time of cell transfer, but not 4 wk later, to mice receiving PBL from nonimmunized donors induced production of minute amounts of anti-phage Ab. We conclude that human peripheral blood lymphocytes transferred into SCID mice become maximally stimulated presumably by xenogeneic murine Ag, resulting in polyclonal expansion of the graft and spontaneous production of Ab to Ag the human donor was previously exposed to, and in loss of responses to subsequent Ag exposure. Ab production to neoantigen, however, can be induced and that to recall Ag can be modified if PBL are exposed to Ag at the time of cell transfer.
Adult↗
B cell activation via CD40 is required for specific antibody production by antigen-stimulated human B cells.
Costimulatory signals provided by T cells are required for B cells to produce specific antibody (Ab) to T-dependent antigen (Ag) bacteriophage phi x 174. In this study, we demonstrate that if cultured in the presence of anti-CD40, interleukin 10 (IL-10), and Ag, purified B cells can produce antiphage Ab in quantities comparable to those synthesized by B cells cocultured with Ag and T cells. Isotypes produced by B cells in this culture system correspond to those observed in sera of B cell donors. Culture of immunoglobulin (Ig)D- and IgD+ B cells reveals that Ag-induced production of antiphage Ab is restricted to IgD- subset of B cells. In the absence of Ag, anti-CD40/IL-10-stimulated B cells produce only minute amounts of antiphage Ab, indicating that Ag stimulation is indispensable and provides a signal that is synergistic with anti-CD40 and IL-10. Addition of a soluble form of the CD40 ligand (sgp39) to the culture system has a similar effect on specific Ab synthesis as anti-CD40; addition of the soluble construct, CD40 Ig, known to inhibit gp39/CD40 interaction, suppresses in vitro antiphage Ab production by Ag exposed peripheral blood mononuclear cells. Finally, in vivo requirement of gp39/CD40 interaction for specific Ab production was demonstrated by the finding that activated T cells from patients with x-linked hyper IgM syndrome express functionally defective gp39 and respond with depressed Ab titers and fail to switch from IgM to IgG after multiple phage immunizations. These observations illustrate that in vitro and possibly in vivo Ag-specific Ab synthesis requires the presence of Ag and IL-10, and activation signals via CD40.
Antibodies, Viral↗
Strain-dependent leakiness of mice with severe combined immune deficiency.
Mice with immunodeficiency provide an excellent in vivo model for cell transfer experiments. In this study, we compare the extent of immune deficiency of the original CB17 severe combined immune-deficient (SCID) mice with that of two other strains of immune-deficient mice, the recently developed C3H SCID mice and the beige/nude/X-linked immune-deficient (BNX) mice. Detectable levels of serum lg (higher than 0.4 microgram/ml) were found in 79% of CB17 SCID mice studied (n = 24) and in all BNX mice (n = 12); some leaky CB17 SCID mice had normal levels of Ig. In contrast, only 15% of C3H SCID mice (n = 61) had detectable serum lg; the highest Ig level in this strain was 9.6 micrograms/ml. Age had no effect on serum Ig concentrations of C3H SCID mice; in contrast, all old (> 1-year-old) CB17 SCID mice studied had detectable levels of serum Ig. Transfer of syngeneic, normal, neonatal thymocytes increased serum Ig of SCID mouse origin to near-normal levels in all CB17 SCID mice but had no effect on serum lg concentrations in C3H SCID mice. Treatment with anti-asialo-GM-1 antiserum to abrogate NK cell activity increased serum Ig levels in 37% of CB17 SCID mice but had no effect on Ig production in C3H SCID mice. Flow cytometric analysis failed to identify mature T or B cells in C3H SCID mice; in contrast, some leaky CB17 SCID mice had detectable numbers of T and B cells in the peritoneal cavity. After immunization with bacteriophage phi X 174, neither C3H nor CB17 SCID mice, including leaky mice, produced specific antibody to phage. In contrast, BNX mice produced small but significant amounts of anti-phage antibody. These results indicate that, of the three strains of immune-deficient mice, C3H SCID mice have the most severe immune defect. We predict that C3H SCID mice will be best suited for cell transfer experiments.
Animals↗
Costimulation via vascular cell adhesion molecule-1 induces in T cells increased responsiveness to the CD28 counter-receptor B7.
Optimal stimulation of CD4+ T cells in an immune response requires not only signals transduced via the CD3/TCR complex but also costimulatory signals delivered as a consequence of interactions between T-cell surface-associated costimulatory R and their counter-R on APC. CD28 plays a crucial role as a dominant costimulatory R during the induction of CD4+ T-cell proliferation by interacting with counter-R B7 on APC to sustain IL-2 production. The absence of CD28-mediated costimulation has been postulated to result in T-cell anergy or unresponsiveness. The costimulatory effects of CD28 can be generated with its natural counter-R B7 or mAb directed at CD28. Using soluble C gamma 1 chimeras of B7, ICAM-1, and VCAM-1, we have recently shown that B7 costimulates TCR-dependent proliferation of Ag-primed CD4+ T cells more efficiently than that of resting nonactivated CD4+ T cells. In contrast, proliferation of resting CD4+ T cells can be efficiently costimulated by either ICAM-1 or VCAM-1 via interactions with their R CD11a/CD18 (LFA-1/beta 2 integrin) and CD29/CD49d (VLA-4/beta 1 integrin), respectively. TCR-directed preactivation of resting CD4+ T cells with ICAM-1 can induce increased responsiveness to B7 costimulation. In this study, we show that prior TCR-directed activation of resting CD4+ T cells with VCAM-1 induced increased responsiveness to B7 costimulation. VCAM-1 also synergized with B7 to bring about supraoptimal proliferation of CD4+ T cells. In addition, costimulation of resting T cells with VCAM-1 significantly increased not only surface expression of CD28 but also CD28-mediated mobilization of intracellular free [Ca2+]i. Similar activation of T cells with fibronectin also resulted in increased B7 responsiveness, suggesting the involvement of VLA-4 molecule. VCAM-1 costimulation induced hyperresponsiveness to B7 costimulation in both CD18+ (normal) and CD18- (leukocyte adhesion deficient) T cells. Thus, VCAM-1 may play an important costimulatory role during the activation of resting T cells and, by augmenting responsiveness to B7, facilitate optimal development of immunological memory in addition to various regulatory and effector functions.
Antibodies, Monoclonal↗
Advances in X-linked immunodeficiency diseases.
The molecular basis for three well-defined X-linked diseases has recently been identified. In X-linked agammaglobulinemia, the gene encoding a novel cytoplasmic tyrosine kinase (btk) expressed by B cells is defective. This B-cell-specific kinase belongs to a new subfamily of tyrosine kinases. The molecular defect in X-linked hyper IgM affects the gene encoding the CD40 ligand (CD40L, gp39) on T cells. This protein binds to its natural receptor, CD40, expressed constitutively by B cells. The ligand-receptor interaction initiates B-cell proliferation and isotype switching. The molecular defect in X-linked severe combined immunodeficiency disease has been assigned to the gene encoding the gamma chain of the interleukin-2 receptor (IL-2R gamma), which is constitutively expressed by T cells and is involved in the formation of high and intermediate affinity IL-2R complexes. IL-2R-gamma is responsible for the failure of X-linked severe combined immunodeficiency disease T and B lymphocytes to respond to IL-2-dependent signals.
Adult↗
In vivo behavior of neutrophils from two patients with distinct inherited leukocyte adhesion deficiency syndromes.
The selectins and the beta 2-integrins (CD11/CD18) mediate distinct adhesive interactions between neutrophils and endothelial cells. Selectins are believed to initiate binding by mediating neutrophil rolling, whereas beta 2-integrins are required for subsequent activation-induced firm sticking and emigration. In vitro evidence suggests that two endothelial cell selectins, P- and E-selectin, can mediate rolling by binding to the carbohydrate ligand sialyl-Lewisx (sLex) on neutrophil surface glycoconjugates. To test the relative contribution of selectins and beta 2-integrins in vivo we used intravital microscopy to study the behavior of neutrophils from two patients with distinct inherited leukocyte adhesion deficiency syndromes. Neutrophils from a patient suffering from CD18 deficiency showed normal rolling behavior but were incapable of sticking or emigrating upon chemotactic stimulation. Neutrophils from a second patient with a newly described adhesion deficiency had normal CD18 but did not express sLex. These neutrophils rolled poorly and also failed to stick in venules under shear force. Under static conditions, however, chemoattractant-induced sticking and emigration could be observed. This demonstrates that both selectin-carbohydrate-mediated initiation of adhesion and subsequent activation-induced beta 2-integrin engagement are essential for the normal function of human neutrophils in vivo.
Animals↗
Activated B cells from patients with common variable immunodeficiency proliferate and synthesize immunoglobulin.
Most patients with common variable immunodeficiency (CVI) have normal numbers of circulating B cells but low concentrations of serum Ig. To determine if the hypogammaglobulinemia is caused by an intrinsic B cell defect, we studied B cell function of 22 CVI patients. Cultured B cells from all CVI patients underwent normal proliferation and synthesized normal quantities of IgE in the presence of anti-CD40 and IL-4. If cultured with anti-CD40 and IL-10, four patterns of Ig isotype synthesis were observed. Six CVI patients produced normal amounts of IgM, IgG, and IgA. Four patients produced normal quantities of IgM and IgG. Of the remaining 12 patients who failed to synthesize IgG and IgA, 8 produced normal and 4 synthesized decreased amounts of IgM. Analysis of the IgG subclasses produced by 10 patients with IgG-secreting B cells revealed that IgG4 was the most affected subclass, followed by IgG2; synthesis of IgG3 and IgG1 remained normal. Similarly, in the six IgA producing patients, IgA2 was more often affected than IgA1. The hierarchy of Ig isotype and subclass synthesis corresponds to Ig heavy chain constant region gene location on chromosome 14. Thus, circulating B cells of CVI patients are committed to synthesize one or more Ig isotypes or subclasses, and under proper conditions can proliferate, mature into Ig-secreting cells, and undergo class switch to IgE.
Adolescent↗
The role of adhesion molecules in the regulation of antibody responses.
We used the T-cell-dependent antigen, bacteriophage (phage) phi X174, to study antibody synthesis in patients, guinea pigs, and dogs with complement component deficiencies (C2, C4, C3, C7); in patients with adhesion molecule deficiencies (CD11/CD18 or sialylated Lewisx); and in patients with the hyper IgM (HIM) syndrome (absence of functional gp39 expression by activated T cells). Patients and guinea pigs deficient in early complement components, patients deficient in CD11/CD18, and patients lacking functional gp39 on activated T cells responded to repeated phage immunizations with depressed antibody titers, lack of or inadequate amplification, and failure to switch from IgM to IgG, suggesting that defective T-cell-B-cell interaction is the cause of the antibody deficiency observed in these patients.
Animals↗
Regulation of antibody responses: the role of complement and adhesion molecules.
To analyze the importance of cell surface-associated molecules in modulating the immune response by facilitating T/B cell interaction, we used the T cell-dependent antigen, bacteriophage phi X174. Taking advantage of "experiments of nature", we studied specific antibody synthesis in patients with deficiencies of complement components or of the adhesion molecule CD11/CD18 (leukocyte adhesion defect, LAD) and guinea pigs and dogs with early complement component deficiency. Following intravenous injection of bacteriophage phi X174 into normal subjects or animals, a primary response consisting of IgM, a secondary response consisting of IgM and IgG, and a tertiary, predominantly IgG response can be distinguished. Patients and guinea pigs deficient of early complement component and LAD patients responded to repeated phage immunization with depressed antibody titers, lack of or inadequate amplification, and failure to switch from IgM to IgG, suggesting a defect in generating antigen-specific memory cells. Several mechanisms have to be considered: (i) The complement portion of the antigen-antibody complement complex facilitates the accumulation and trapping of antigen in lymphoid organs, thus improving the response to Ag at low concentrations. (ii) Immune complexes preferentially bind to antigen-specific B cells, cells expressing Fc receptors, or CR2 and CR3, the receptors for C3bi. (iii) The weak binding established between the MHC-II/Ag complex and the TCR complex is strengthened through the binding of several adhesion molecule pairs. (iv) Receptor-ligand binding initiates activation signals. The concept of binding/signaling via interacting molecules is further supported by the observation that mAb 60.3, recognizing the beta chain of CD11/CD18, blocks in vitro synthesis of antibody to bacteriophage by primed PBMC.
Animals↗
Antibody responses to bacteriophage phi X174 in patients with adenosine deaminase deficiency.
Adenosine deaminase (ADA) deficiency and its biochemical consequences cause severe combined immunodeficiency (SCID). Treatment strategies, designed to correct the biochemical abnormalities, include transplantation of matched bone marrow or haploidentical bone marrow stem cells, repeated partial exchange transfusions with frozen irradiated human red blood cells (RBC), or weekly injection of polyethylene glycol-modified bovine ADA (PEG-ADA). To evaluate the effect of these therapeutic options, we studied in vitro T-cell function and in vivo antibody responses to the T-cell-dependent neoantigen, bacteriophage phi X174, in 10 children with ADA-deficient SCID. In untreated patients, T-cell function was severely depressed, and only minute amounts of antibacteriophage antibody were produced. Transplantation of bone marrow from a matched sibling (one patient) or a phenotypically matched parent (one patient) resulted in a stable graft, normal T-cell function, and substantial but subnormal antibody titers to bacteriophage, with reduced memory and impaired switch from IgM to IgG. Patients receiving T-cell-depleted haploidentical bone marrow stem cells had markedly depressed antibody responses for as long as 3 years posttransplantation, despite rapidly improving T-cell function that became normal in two of four patients. Two methods of enzyme replacement were explored. During treatment with human RBC transfusions, antibody responses to bacteriophage were as severely depressed as in untreated ADA-deficient patients. Treatment with weekly injections of PEG-ADA resulted in normalization of T-cell numbers in all four patients, normal or near-normal T-cell function in two, and mildly but variably improved T-cell function in the other two patients. Quantitatively and qualitatively normal antibody responses to bacteriophage were observed in three of four patients. Assessment of antibody responses to immunization with bacteriophage phi X174 is a useful method to monitor humoral immune function in treated ADA-deficient patients and can be used to estimate when intravenous immunoglobulin (IVIG) prophylaxis may be safely discontinued.
Adenosine Deaminase↗
Intravenous immunoglobulin therapy for active, extensive, and medically refractory idiopathic ulcerative or Crohn's colitis.
To determine whether intravenous immunoglobulin produces demonstrable clinical improvement in patients with refractory idiopathic inflammatory bowel disease, a pilot, open-label, nonrandomized, safety and therapeutic efficacy study was carried out at a tertiary care referral medical center. Twelve consecutive patients with refractory idiopathic colitis (nine ulcerative colitis, three Crohn's colitis) who were reluctant to receive immunosuppressive therapy or have surgical intervention were referred by physicians not participating as investigators in this study. Eleven patients were symptomatic for at least 6 months, with endoscopically moderate or severe mucosal inflammation despite medical therapy, including systemic corticosteroids in all cases, and one patient was dependent on oral prednisone to remain in clinical remission. Ten patients had extensive colitis, six of whom had pancolitis and four of whom had colitis extending to the hepatic flexure or transverse colon. Nine patients required hospitalization for treatment of colitis. Intravenous immunoglobulin was administered in one or two induction phases (2 g/kg over 2 or 5 days), followed by a maintenance phase (200-500 mg/kg every 2 wk for 12 or 24 wk). Tapering of systemic corticosteroid therapy was attempted, whereas other medications for idiopathic colitis were continued. Treatment response was assessed clinically and by colonoscopy with multiple biopsies whenever possible. Immunoglobulin therapy was well-tolerated and did not produce any biochemical abnormalities. In six patients who completed the treatment protocol, mean reductions +/- SE were achieved in subjective symptoms as quantified by a colitis activity score, 13.3 +/- 1.2 to 4.7 +/- 0.9 (p less than 0.001), and daily mg dose of prednisone, 41.7 +/- 8.0 to 1.9 +/- 1.2 (p less than 0.001). For all 12 patients, statistically significant reductions were achieved in the colitis activity score and daily prednisone dose. Of five patients who completed the treatment protocol and improved clinically, four underwent post-treatment colonoscopic and biopsy evaluations and had unequivocal reductions in the intensity of colonic mucosal inflammation. Three patients who had objective improvement with intravenous immunoglobulin experienced relapses of colitis after discontinuation of this therapy. Six patients did not complete the treatment protocol, two of whom required surgical intervention and four of whom withdrew to undergo colectomy electively. Intravenous immunoglobulin may be beneficial in subsets of patients with idiopathic colitis. The results of our pilot study justify the undertaking of a prospective, randomized controlled trial to determine the efficacy of intravenous immunoglobulin in carefully defined subsets of patients with idiopathic inflammatory bowel disease.
Adolescent↗
Immunoglobulin variable heavy chain cDNA sequence from a patient with X-linked agammaglobulinemia.
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Agammaglobulinemia↗
Maternal-fetal transmission of SIV in macaques: disseminated adenovirus infection in an offspring with congenital SIV infection.
To develop a nonhuman primate model for maternal-fetal transmission of HIV infection, we have inoculated pregnant Macaca nemestrina with uncloned SIVMne. Three animals inoculated during the third trimester delivered healthy infants. One of the three infants, a male born 31 days after the mother was inoculated with SIV, became virus-positive but failed to produce SIV-specific antibody and died with overt simian immunodeficiency and disseminated adenovirus (SV20) infection at age six and one-half months. SIV and adenovirus antigen could be demonstrated by immunohistochemical methods in multiple organ systems.
Adenoviridae Infections↗