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Biomedical subjects

J Inman

Publications and source records attributed to J Inman.

At least 19 recordsLinked to original sources
PubMed2001

IL-12 induction by a TH1-inducing adjuvant in vivo: dendritic cell subsets and regulation by IL-10.

IL-12 induction is critical for immune responses against many viruses and intracellular bacterial pathogens. Recent studies suggest that IL-12-secreting dendritic cells (DC) are potent Th1-inducing APC. However, controversy exists concerning the function of DC subsets. Murine studies have suggested that CD8(+) DC preferentially induce Th1 responses, whereas CD8(-) DC induce Th2 development; in this model, different DC subsets prime different responses. Alternatively, the propensity of DC subsets to prime a Th1 response could depend upon the type of initial stimulus. We used a prototypic Th1-inducing adjuvant, heat-killed Brucella abortus (HKBA) to assess stimulation of DC subsets, relationship between Ag burden and IL-12 production, and down-regulation of DC subset IL-12 production by IL-10. In this study, we show that DC were sole producers of IL-12, although most HKBA uptake was by splenic macrophages and granulocytes. More CD8(-) than CD8(+) DC produced IL-12 after HKBA challenge, whereas only CD8(+) DC produced IL-12 after injection of another Th1-promoting microbial substance, soluble Toxoplasma gondii Ags. Studies in IL-10-deficient mice revealed that IL-10 down-regulates frequency and duration of IL-12 production by both DC subsets. In the absence of IL-10, IL-12 expression is enabled in CD11c(low) cells, but not in macrophages or granulocytes. These findings support the concept of DC as the major IL-12 producers in spleens, but challenge the notion that CD8(+) and CD8(-) DC are destined to selectively induce Th1 or Th2 responses, respectively. Thus, the nature of the stimulating substance is important in determining which DC subsets are activated to produce IL-12.

Adjuvants, Immunologic↗
PubMed2001

The Medicago Genome Initiative: a model legume database.

The Medicago Genome Initiative (MGI) is a database of EST sequences of the model legume MEDICAGO: truncatula. The database is available to the public and has resulted from a collaborative research effort between the Samuel Roberts Noble Foundation and the National Center for Genome Resources to investigate the genome of M.truncatula. MGI is part of the greater integrated MEDICAGO: functional genomics program at the Noble Foundation (http://www.noble.org ), which is taking a global approach in studying the genetic and biochemical events associated with the growth, development and environmental interactions of this model legume. Our approach will include: large-scale EST sequencing, gene expression profiling, the generation of M.truncatula activation-tagged and promoter trap insertion mutants, high-throughput metabolic profiling, and proteome studies. These multidisciplinary information pools will be interfaced with one another to provide scientists with an integrated, holistic set of tools to address fundamental questions pertaining to legume biology. The public interface to the MGI database can be accessed at http://www.ncgr.org/research/mgi.

Computational Biology↗
PubMed2001

Ontogeny of Th1 memory responses against a Brucella abortus conjugate.

Protective immune responses to intracellular pathogens such as Brucella abortus are characteristically Th1-like. Recently we demonstrated that heat-killed B. abortus (HKBa), a strong Th1 stimulus, conjugated to ovalbumin (HKBA-OVA), but not B. abortus alone, can alter the antigen-specific cytokine profile from Th2- to Th1-like. In this report we study the ability of a single injection of B. abortus to switch a Th2 to a Th1 response in immature mice. One-day- and 1-week-old mice were given a single injection of B. abortus in the absence or presence of OVA, and at maturity mice were challenged with an allergenic preparation, OVA with alum (OVA-A). B. abortus given without OVA did not diminish the subsequent Th2 response in either age group. In contrast, mice receiving a single injection of B. abortus-OVA at the age of 1 week, but not those injected at the age of 1 day, had reversal of the ratio of OVA-specific Th1 to Th2 cells and decreased immunoglobulin E levels after allergen challenge as adults. Within 6 h both 1-day- and 1-week-old mice expressed interleukin-12 p40 mRNA following either B. abortus or B. abortus-OVA administration. However, only the 1-week-old mice exhibited increased expression of gamma interferon (IFN-gamma) mRNA. The absence of the early IFN-gamma response in 1-day-old mice may explain their inability to generate a Th1 memory response. These results suggest that at early stages of immune development, responses to intracellular bacteria may be Th2- rather than Th1-like. Furthermore, they suggest that the first encounter with antigen evokes either a Th1- or a Th2-like response which becomes imprinted, so that subsequent memory responses conform to the original Th bias. This has implications for protection against infectious agents and development of allergic responses.

Aging↗
PubMed2000

The genome sequence DataBase.

The Genome Sequence DataBase (GSDB) is a database of publicly available nucleotide sequences and their associated biological and bibliographic information. Several notable changes have occurred in the past year: GSDB stopped accepting data submissions from researchers; ownership of data submitted to GSDB was transferred to GenBank; sequence analysis capabilities were expanded to include Smith-Waterman and Frame Search; and Sequence Viewer became available to Mac users. The content of GSDB remains up-to-date because publicly available data is acquired from the International Nucleotide Sequence Database Collaboration databases (IC) on a nightly basis. This allows GSDB to continue providing researchers with the ability to analyze, query and retrieve nucleotide sequences in the database. GSDB and its related tools are freely accessible from the URL: http://www.ncgr.org

Databases, Factual↗
PubMed2000

The phytophthora genome initiative database: informatics and analysis for distributed pathogenomic research.

The Phytophthora Genome Initiative (PGI) is a distributed collaboration to study the genome and evolution of a particularly destructive group of plant pathogenic oomycete, with the goal of understanding the mechanisms of infection and resistance. NCGR provides informatics support for the collaboration as well as a centralized data repository. In the pilot phase of the project, several investigators prepared Phytophthora infestans and Phytophthora sojae EST and Phytophthora sojae BAC libraries and sent them to another laboratory for sequencing. Data from sequencing reactions were transferred to NCGR for analysis and curation. An analysis pipeline transforms raw data by performing simple analyses (i.e., vector removal and similarity searching) that are stored and can be retrieved by investigators using a web browser. Here we describe the database and access tools, provide an overview of the data therein and outline future plans. This resource has provided a unique opportunity for the distributed, collaborative study of a genus from which relatively little sequence data are available. Results may lead to insight into how better to control these pathogens. The homepage of PGI can be accessed at http:www.ncgr.org/pgi, with database access through the database access hyperlink.

Databases, Factual↗
PubMed2000

The role of phonology in a letter detection task.

In two experiments, we investigated whether onsets and rimes have a role in the processing of written English. In both experiments, participants detected letter targets (e.g., t) in nonwords like vult faster than in nonwords like vust. This finding is consistent with Selkirk's (1982) view that sonorants (e.g., the /l/ of vult) cohere with preceding short vowels and are part of the vowel nucleus. In contrast, the /t/ of vust is part of the syllable's coda st and so is harder to isolate. Experiment 2 demonstrated that the time required for one to detect single-member codas following vowel digraphs (e.g., the t in veet) was similar to the time to detect the same target letter following a postvocalic sonorant (e.g., the t in vult). No evidence was found for onsets. The results provide support for a phonological organization among letters of printed rimes.

Adult↗
PubMed1999

The Genome Sequence DataBase: towards an integrated functional genomics resource.

During 1998 the primary focus of the Genome Sequence DataBase (GSDB; http://www.ncgr.org/gsdb ) located at the National Center for Genome Resources (NCGR) has been to improve data quality, improve data collections, and provide new methods and tools to access and analyze data. Data quality has been improved by extensive curation of certain data fields necessary for maintaining data collections and for using certain tools. Data quality has also been increased by improvements to the suite of programs that import data from the International Nucleotide Sequence Database Collaboration (IC). The Sequence Tag Alignment and Consensus Knowledgebase (STACK), a database of human expressed gene sequences developed by the South African National Bioinformatics Institute (SANBI), became available within the last year, allowing public access to this valuable resource of expressed sequences. Data access was improved by the addition of the Sequence Viewer, a platform-independent graphical viewer for GSDB sequence data. This tool has also been integrated with other searching and data retrieval tools. A BLAST homology search service was also made available, allowing researchers to search all of the data, including the unique data, that are available from GSDB. These improvements are designed to make GSDB more accessible to users, extend the rich searching capability already present in GSDB, and to facilitate the transition to an integrated system containing many different types of biological data.

Animals↗
PubMed1998

HIV peptide conjugated to heat-killed bacteria promotes antiviral responses in immunodeficient mice.

Enhancement of immunity in the setting of HIV infection is difficult owing to loss of functional CD4+ T cells. The MHC class II-deficient mouse (II-/-) environment simulates that of the immunocompromised HIV-infected individual, since these mice have low CD4+ T cell numbers, defective CD4-dependent responses, and are susceptible to opportunistic infection. This strain was used to test whether heat-killed Brucella abortus (BA), covalently conjugated to the V3 peptide of HIV-1 (MN), could elicit anti-HIV responses. V3-BA, but not the T-dependent antigen V3-KLH, induced high levels of IL-12, IFN-gamma, and IL-10 mRNA in both wild-type (WT) and II-/- mice within 24 hr of injection. V3-BA-treated, but not V3-KLH-treated, II-/- mice developed serum IgG and IgA anti-V3 antibodies, with IgG2b and IgG3 as the predominant isotype. Viral neutralization studies, using a syncytium inhibition assay, demonstrated that the antibodies generated by V3-BA in II-/- mice were capable of neutralizing HIV. These experiments demonstrate that a heat-inactivated bacterium such as BA, when used as a carrier, can generate a cytokine environment that results in the production of neutralizing antiviral antibodies in an immunodeficient host. Such strategies could be important in the development of immunotherapies and vaccines for HIV-1 patients.

AIDS Vaccines↗
PubMed1997

Inhibition of primary and recall allergen-specific T helper cell type 2-mediated responses by a T helper cell type 1 stimulus.

Allergic responses are characterized by the production of Ag-specific IgE Abs that are dependent upon Th2-mediated T cell help. We determined whether heat-killed Brucella abortus (BA), an inducer of Th1 responses, could influence the allergic Th2-mediated IgE response to OVA adsorbed to alum (O/A). BA plus O/A, but not O/A alone, induced high levels of mRNA for IFN-gamma and IL-12 promptly after injection. Furthermore, initial treatment with BA plus O/A rendered both BALB/c and C57Bl/6 mice incapable of mounting high IgE responses even after repeated challenges with allergen alone. Long term abrogation of anti-OVA IgE correlated with an increased frequency of IFN-gamma-secreting OVA-specific cells and a decreased frequency of IL-4-secreting OVA-specific cells. Initial treatment with anti-IL-12 prevented BA-induced early IFN-gamma production and secondary IgG2a responses, but did not abrogate IgE suppression. Additionally, secondary OVA-specific IgE responses were down-regulated by BA conjugated to OVA or by BA given with O/A. BA-induced down-regulation of secondary IgE responses was associated with increased frequency of Ag-specific IFN-gamma-secreting cells. These results suggest the possibility that even recall Th2-mediated immune responses can be attenuated if Ag is given with a carrier or adjuvant that induces potent Th1-promoting cytokines.

Allergens↗
PubMed1996

Brucella abortus conjugated with a peptide derived from the V3 loop of human immunodeficiency virus (HIV) type 1 induces HIV-specific cytotoxic T-cell responses in normal and in CD4+ cell-depleted BALB/c mice.

We have previously shown that immunization of mice with human immunodeficiency virus (HIV)-derived proteins or peptides conjugated to inactivated Brucella abortus induces the secretion of virus-neutralizing antibodies, predominantly of the immunoglobulin G2a (IgG2a) isotype. In addition, B. abortus activates human CD4+ and CD8+ cells to secrete gamma interferon. Since these are both characteristics of a Th1-type immune response, which is associated with the development of cell-mediated immunity, it was important to determine if B. abortus conjugates would also act as a carrier to induce a cytotoxic T-lymphocyte (CTL) response. To test this hypothesis, we conjugated an 18-amino-acid peptide from the V3 loop of the MN strain of HIV-1 gp120 that contains both B- and cytotoxic T-cell epitopes to B. abortus (B. abortus-MN 18-mer). A 10-amino-acid fragment of this peptide has been shown to be the minimal CTL determinant presented by murine H-2Dd. It was found that two in vivo immunizations with 10(8) organisms of B. abortus-MN 18-mer followed by in vitro stimulation with peptide induced a virus-specific CTL response. Conjugation to B. abortus was required for in vivo priming, since there was no induction of memory CTLs when B. abortus was only mixed with peptide. Targets pulsed with peptide as well as those infected with a vaccinia virus encoding HIV gp160 were killed, demonstrating recognition of naturally processed envelope. Also, major histocompatibility complex-incompatible L cells which were infected with vaccinia viruses that encoded H-2Dd, but not H-2Kd, and pulsed with peptide were lysed. This demonstrated the appropriate major histocompatibility complex class I restriction. Treatment of the mice with anti-L3T4 prior to immunization caused a severe depletion of CD4+ lymphocytes, yet it did not decrease the CTL priming. Thus, inactivated B. abortus can induce non-CD4+ cells to produce the cytokines required for CTL induction. We conclude that B. abortus stimulates a cellular as well as a humoral immune response, even in the relative absence of CD4+ helper cells. It may be a particularly useful vaccine carrier in HIV-1-infected individuals or others with impaired CD4+ T-cell function.

AIDS Vaccines↗
PubMed1995

Monophosphoryl lipid A behaves as a T-cell-independent type 1 carrier for hapten-specific antibody responses in mice.

It is known that the lipopolysaccharide (LPS) of gram-negative bacteria, in addition to being a potent adjuvant, is an effective carrier for covalently associated haptens. However, the toxic nature of most forms of LPS precludes their use as adjuvants or carriers for human vaccines. 4'-Monophosphoryl lipid A (MLA), a derivative of LPS with attenuated toxicity, is currently being tested in humans as an immunological adjuvant. In this study, MLA was tested for its ability to function as a carrier for a small hapten, the trinitrophenyl group (TNP). MLA was first modified by addition of 6-aminocaproic acid to the 6' position of the disaccharide backbone (Cap-MLA). TNP was then attached to Cap-MLA via the free amino group, yielding TNP-Cap-MLA. Immunization of normal mice with TNP-Cap-MLA resulted in high-titer anti-TNP responses of immunoglobulin M and all immunoglobulin G subclasses. Furthermore MLA, like other T-cell-independent type 1 (TI-1) carriers, induced responses in athymic and X-linked immunodeficient mice. In all cases, immunization with either MLA alone or TNP-Cap plus MLA failed to induce measurable anti-TNP antibodies of any isotype, indicating that covalent association of MLA and hapten was necessary for MLA's carrier activity to be manifested. These properties of MLA make it a potential candidate as a carrier for vaccine subunit components, such as small peptides, especially for situations in which T-cell help is impaired, as occurs following human immunodeficiency virus type 1 infection.

Animals↗
PubMed1995

Brucella abortus conjugated with a gp120 or V3 loop peptide derived from human immunodeficiency virus (HIV) type 1 induces neutralizing anti-HIV antibodies, and the V3-B. abortus conjugate is effective even after CD4+ T-cell depletion.

Human immunodeficiency virus type 1 (HIV-1) infection is associated with loss of function and numbers of CD4+ T-helper cells. In order to bypass the requirement for CD4+ cells in antibody responses, we have utilized heat-inactivated Brucella abortus as a carrier. In this study we coupled a 14-mer V3 loop peptide (V3), which is homologous to 9 of 11 amino acids from the V3 loop of HIV-1 MN, and gp120 from HIV-1 SF2 to B. abortus [gp120(SF2)-B. abortus]. Our results showed that specific antibody responses, dominated by immunoglobulin G2a in BALB/c mice, were induced by these conjugates. Sera from the immunized mice bound native gp120 expressed on the surfaces of cells infected with a recombinant vaccinia virus gp160 vector (VPE16). Sera from mice immunized with gp120(SF2)-B. abortus inhibited binding of soluble CD4 to gp120, whereas sera from mice immunized with V3-B. abortus were ineffective. Sera from mice immunized with either conjugate were capable of blocking syncytium formation between CD4+ CEM cells and H9 cells chronically infected with the homologous virus. Sera from mice immunized with gp120(SF2)-B. abortus were more potent than sera from mice immunized with V3-B. abortus in inhibiting syncytia from heterologous HIV-1 laboratory strains. Importantly, in primary and secondary responses, V3-B. abortus evoked anti-HIV MN antibodies in mice depleted of CD4+ cells, and sera from these mice were able to inhibit syncytia. These findings indicate that B. abortus can provide carrier function for peptides and proteins from HIV-1 and suggest that they could be used for immunization of individuals with compromised CD4+ T-cell function.

AIDS Vaccines↗
PubMed1993

Lipopolysaccharide from Brucella abortus behaves as a T-cell-independent type 1 carrier in murine antigen-specific antibody responses.

In order to determine the carrier nature of lipopolysaccharide from Brucella abortus (LPS-BA) in evoking humoral responses, normal and immunodeficient mice were immunized with trinitrophenyl (TNP)-conjugated LPS-BA (TNP-LPS-BA) and the responses were compared with those to known T-dependent and T-independent antigens. TNP-LPS-BA, like T-independent type 1 (TI-1) antigens such as TNP-BA and TNP-LPS from Escherichia coli (TNP-LPS-EC), generated anti-TNP responses in BALB/c, athymic BALB/c nu/nu, and CBA/N mice. In contrast, N-2,4-dinitrophenyl-beta-alanylglycylglycyl-substituted keyhole limpet hemocyanin, a typical T-dependent antigen, was not immunogenic in athymic mice, and TNP-Ficoll (T-independent type 2) was ineffective in eliciting humoral responses in CBA/N mice. These results indicate that LPS from B. abortus acts as a TI-1 carrier in generating antibody responses. In C3H/HeJ mice, TNP-LPS-BA generated higher-titer immunoglobulin G1 (IgG1), IgG2a, and IgG2b anti-TNP antibodies than TNP-LPS-EC. Compared with those from BALB/c mice, pure resting B cells isolated from C3H/HeJ mice exhibited a 30-fold lower proliferative response to LPS-EC, whereas the LPS-BA response was reduced to a lesser extent (5-fold). This suggests that the disparity observed in antibody titers was due to different abilities of LPS from B. abortus and E. coli to stimulate C3H/HeJ B cells. The ability of LPS from B. abortus to act as a carrier in generating humoral immune responses indicates that LPS-BA can be substituted for whole B. abortus organisms in vaccine development.

Animals↗
PubMed1992

Role of antigen-specific T cell help in the generation of in vivo antibody responses. I. Antigen-specific T cell help is required to generate a polyclonal IgG1 response in anti-IgD antibody-injected mice.

A system in which injection of mice with an antibody to mouse IgD that they recognize as foreign stimulates a large, T cell-dependent IgG response was used to study whether Ag-specific T cell help is required to stimulate polyclonal (non-Ag-specific) IgG production in vivo. Igha x Ighb allotype heterozygous mice were injected with a conjugate of a foreign Ag coupled to a mAb specific for one of the two IgD allotypes expressed in these mice. This conjugate cross-links mIgD on B cells that express the recognized allotype. These cells process the conjugate and present the foreign Ag to Ag-specific T lymphocytes, which become activated. Thus, B cells of the recognized allotype can be stimulated by cross-linking of their mIgD, Ag-specific T cell help, non-Ag-specific cytokines, and non-Ag-specific contact with activated T cells. In contrast, B cells that express the Igh allotype not recognized by the Ag-anti-IgD antibody conjugate (bystander B cells) can be stimulated in this system only by non-Ag-specific cytokines and non-Ag-specific contact with activated T cells. Although both recognized and bystander B cells in conjugate-injected mice demonstrated substantial increases in size and Ia expression, only the recognized B cells were induced to synthesize DNA and to make a substantial polyclonal Ig response. Bystander B cells still failed to secrete IgG when mice were injected with an anti-IgD-Ag conjugate specific for the other Igh allotype as well as a mAb that cross-linked IgD of the bystander B cell allotype. These observations demonstrate that although non-Ag-specific cytokine and contact-mediated T cell help are sufficient to induce B cells to increase in size and Ia expression in anti-IgD antibody-injected mice, Ag-specific T cell help is required to stimulate the generation of an IgG response in these mice.

Animals↗
PubMed1992

Failure of signaling through a chimeric class I-immunoglobulin molecule expressed on the surface of transfected B lymphoma cells and cells of transgenic mice.

To test the possibility that the crosslinkage of molecules expressing a transmembrane region derived from the membrane form of the mu immunoglobulin heavy chain would be sufficient for signal transduction in B cells, a chimeric gene (Kk-mu) consisting of extracellular exons of the class I gene H-2Kk and the transmembrane and cytosolic exons of the mu constant region gene was introduced into WEHI-231 B lymphoma cells and into mouse blastocysts. A protein consistent with the predicted product of the Kk-mu gene was expressed in a transfected cell clone (S18) and in transgenic mice. Crosslinkage of Kk-mu protein with soluble, Sepharose-bound, or dextran-conjugated anti-H-2Kk antibodies failed to induce the accumulation of inositol phosphates or to elevate intracellular calcium concentrations in either S18 cells or B lymphocytes from transgenic mice. Furthermore, crosslinkage of Kk-mu did not inhibit growth of S18 cells or stimulate DNA synthesis by transgenic B cells, in the presence or absence of interleukin-4. The failure of crosslinkage of Kk-mu to initiate detectable intracellular biochemical change or to effect cellular growth suggests that simple crosslinkage of molecules expressing the mu transmembrane region is not sufficient to transduce signals in B cells.

Animals↗
PubMed1991

Biochemical analysis of the immune B cell defect in xid mice.

Previous studies have shown that B cells from xid immune defective CBA/N mice that are unresponsive do not proliferate after stimulation with unconjugated anti-Ig. The experiments in this manuscript demonstrate that dextran-anti-Ig conjugates, which induce extensive and prolonged sIg cross-linking, are able to stimulate proliferation of xid B cells. The ability of these conjugates to stimulate proliferation of xid B cells is not related to their ability to stimulate higher levels of PIP2 breakdown. Thus, high concentrations of unconjugated anti-Ig antibody, which are nonmitogenic for xid B cells, stimulate higher levels of PIP2 breakdown and of calcium transients than lower concentrations of dextran-conjugated anti-Ig, which are mitogenic. Although unconjugated anti-Ig does not provide a fully competent signal to stimulate proliferation of xid B cells, it induces a sufficiently stimulatory signal to enable them to enter DNA synthesis in the presence of the protein kinase C activator, indolactam. This suggests that the extent or duration of activation of protein kinase C by anti-Ig may be limiting in xid B cells. To examine whether another recently described pathway of B cell activation is defective in these mice, we studied the induction of early anti-Ig-mediated tyrosine kinase activity in xid B cells. Both unconjugated and dextran-conjugated anti-Ig antibody stimulated comparable but not identical patterns of tyrosine phosphorylation. These data taken together with other findings that the combination of phorbol ester and calcium ionophore stimulates high levels of proliferation in xid B cells suggests that the immune defect of xid B cells may be distal to surface Ig-mediated activation of tyrosine kinase and of PIP2 breakdown but proximal to PKC activation. Alternatively, the xid immune defect may not result from abnormalities in the early signal transduction pathways, but rather from more distal and/or as yet undefined pathways leading to B cell activation.

Animals↗
PubMed1988

T cell activation by purified, soluble, class I MHC molecules. Requirement for polyvalency.

To examine the nature of the interaction of the TCR with the MHC class I Ag, we have studied the stimulation requirements of an H-2Dd-reactive T cell hybridoma, using a homogeneous, purified preparation of a molecularly engineered soluble counterpart of the class I Ag, H-2Dd/Q10b. We demonstrate that this monovalent, soluble MHC Ag is incapable of stimulating the release of IL-2 from this T cell hybridoma. However, the same preparation of the purified protein can elicit a dose-dependent response when made multivalent either by covalent coupling to soluble, high m.w. dextran or to agarose beads, or by adsorption to polystyrene tissue culture plates.

Animals↗