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D M Estes

Publications and source records attributed to D M Estes.

At least 37 records · Page 2Linked to original sources
PubMed1999

Bovine CD4(+) T-lymphocyte clones specific for rhoptry-associated protein 1 of Babesia bigemina stimulate enhanced immunoglobulin G1 (IgG1) and IgG2 synthesis.

Optimal protective immunity against babesial infection is postulated to require both complement-fixing and opsonizing antibodies in addition to gamma interferon (IFN-gamma)-mediated macrophage activation. The rhoptry-associated protein 1 (RAP-1) of Babesia bigemina induces partial protective immunity and is a candidate vaccine antigen. Previous studies demonstrated that cattle immunized with native protein that were subsequently protected against challenge had a strong IFN-gamma and weaker interleukin-4 (IL-4) response in immune lymph node lymphocytes that reflected the cytokine profile of the majority of CD4(+) T-cell clones obtained from peripheral blood. RAP-1-specific T helper (Th) cell clones that coexpress IFN-gamma and IL-4 are typical of numerous parasite-specific clones examined. However, the function of such cells as helper cells to enhance immunoglobulin secretion by bovine B cells has not been reported. In cattle, both immunoglobulin G1 (IgG1) and IgG2 can fix complement, but IgG2 is the superior opsonizing subclass. Therefore, studies were undertaken to ascertain the functional relevance of RAP-1-specific, CD4(+) Th0 cells as helper cells to enhance IgG1 and/or IgG2 production by autologous B lymphocytes. For comparison, Th0 clones specific for the metazoan parasite Fasciola hepatica that expressed relatively more IL-4 than the B. bigemina-specific Th cells were similarly assayed. B. bigemina RAP-1-specific clones could enhance production of both IgG1 and IgG2 by autologous B cells, whereas Th cell clones specific for F. hepatica enhanced predominantly IgG1 production. The capacity to enhance IgG2 production was associated with production of IFN-gamma by Th cells cocultured with B cells, antigen, and IL-2. The in vitro helper T-cell activity of these T-cell clones was representative of the in vivo serologic responses, which were composed of a mixed IgG1-IgG2 response in B. bigemina RAP-1 immune cattle and a biased IgG1 response in F. hepatica-immune cattle.

Animals↗
PubMed1998

CD40 ligand-dependent signaling of bovine B lymphocyte development and differentiation.

The role of IFN-gamma in B cell differentiation in cattle has not been completely elucidated. We have previously investigated the role of IFN- in the control of antibody production by bovine B cells using anti-bovine IgM antibody in the solid-phase as a source of costimulation. Using this mimic of a T1-2 antigen, we demonstrated that IFN-gamma can enhance the production of IgG2 but not IgG1 from sIgM+ cells. The positive effects of IFN-gamma were enhanced by co-addition to cultures of rboIL-2. Under these activation conditions, the frequency of cells expressing mRNA for the IgG2 heavy chain also increased at least two-fold. In these studies, we investigated the role of IFN-gamma in antibody expression under T-dependent (TD) activation conditions using mouse fibroblasts transfected with boCD40L as a surrogate T cell. Under TD conditions, IFN-gamma had less dramatic effects on the production of IgG2 with IgM predominating in the cultures. Interestingly, the production of IgA was modestly enhanced with little effect on the production of IgG1 above baseline levels obtained with medium alone. In comparison to results with T1-2 conditions of activation, IL-2 did not increase total amounts of antibody above two-fold. Our results suggest that TH1 cells in cattle may be limited in their ability to provide B cell help to levels obtainable in a TH2 cytokine microenvironment due to the effects of IFN-gamma on bovine B cells co-activated via CD40.

Animals↗
PubMed1998

Bovine type 1 and type 2 responses.

The Th1/Th2 paradigm has provided a useful framework for understanding the observed bias in immune responses that are often dominated by either cell-mediated or humoral responses, and for devising therapeutic strategies to stimulate T cell- or antibody-mediated immunity. However this paradigm is an oversimplification of a much more complex immunoregulatory network. Studies with bovine Th cell clones and immunoregulatory cytokines support this viewpoint. This paper highlights the progress that has been made in defining type 1 and type 2 responses in cattle. Evidence is presented for the presence of different subtypes of antigen-specific Th cell clones of cattle which constitute a spectrum of cell phenotypes, and for cytokine-mediated regulation of Th cell responses that differs from that observed in mice. The majority of over 60 parasite antigen-specific Th cell clones coexpress IL-4 and IFN-gamma, and polarized cytokine profiles were rarely observed. Furthermore, IL-2 and IL-10 expression was not restricted to IFN-gamma or IL-4-producing cells, respectively. This lack of coordinate regulation of "Th1" and "Th2" cytokines strengthens the emerging viewpoint that Th1 and Th2 responses, per se, do not typify the immune response to most pathogens. In addition, we provide evidence that major regulatory cytokines, IL-4, IL-10, and IL-12, do not selectively exert their negative (IL-4 and IL-10) or positive (IL-12) effects on Th1-like cells.

Animals↗
PubMed1998

Kinetics of expression and subset distribution of the TNF superfamily members CD40 ligand and Fas ligand on T lymphocytes in cattle.

CD40 and Fas are members of the tumor necrosis factor receptor (TNFR) superfamily. CD40 and Fas play key roles in T cell-B cell interactions. Cross linkage of these molecules induces cell activation and cell death, respectively. The interaction of CD40 with its ligand (CD40L), which is expressed on activated T cells, plays a pivotal role in the generation of the T-dependent (TD) immune response, and FasL-bearing T cells, which have been shown to be predominantly of either the TH0 or TH1 type, have the potential to induce the apoptotic death of Fas expressing B cells. We investigated bovine CD40L mRNA expression in established T cell clones by RT-PCR and Southern blotting. T cells analyzed included CD4+ TH0 and TH1 cell subpopulations, CD8+, and gamma/delta T cells stimulated with either specific antigen or Con A. All CD4+ clones but not all CD8+ or gamma/delta T cell receptor (TCR)-bearing clones expressed mRNA for CD40L. To determine the activation requirements for CD40L expression in cattle, we examined the kinetics and induction requirements for CD40L transcription in peripheral blood T cells using a phorbol ester and/or ionomycin, immobilized mouse anti-bovine CD3, or Con A. Our results demonstrate that CD40L mRNA appears relatively early after activation (1 h) and peaks at 2-4 h poststimulation. A rise in intracellular calcium concentration mediated by ionomycin treatment alone was sufficient to induce CD40L mRNA expression at relatively high levels. Ionomycin treatment in combination with other agonists (anti-CD3, PMA) did not enhance CD40L mRNA expression above levels obtained with ionomycin alone. The bovine Fas ligand gene was partially cloned and mRNA expression determined by RT-PCR in a panel of T cell clones. Our results demonstrate that TH0 and TH1 bovine T cell clones expressed Fas ligand transcripts although only one gamma/delta T cell clone did. This expression was upregulated within 3 h after mitogen stimulation and reduced by 24 h.

Animals↗
PubMed1998

Effects of type I/type II interferons and transforming growth factor-beta on B-cell differentiation and proliferation. Definition of costimulation and cytokine requirements for immunoglobulin synthesis and expression.

In this report, we sought to determine the role of selected type I interferons [interferon-alpha (IFN-alpha) and interferon-tau (IFN-tau)], IFN-gamma and transforming growth factor-beta (TGF-beta) in the regulation of bovine antibody responses. B cells were stimulated via CD40 in the presence or absence of B-cell receptor (BCR) cross-linking. IFN-alpha enhanced IgM, IgG2 and IgA responses but did not enhance IgG1 responses. BCR signalling alone was more effective at inducing IgG2 responses with IFN-alpha than dual cross-linking with CD40. Recombinant ovine IFN-tau was less effective at inducing IgG2 responses when compared with IFN-alpha, though IgA responses were similar in magnitude following BCR cross-linking. At higher concentrations, IFN-tau enhanced IgA responses greater than twofold over the levels observed with IFN-alpha. Previous studies have shown that addition of IFN-gamma to BCR or pokeweed mitogen-activated bovine B cells stimulates IgG2 production. However, following CD40 stimulation alone, IFN-gamma was relatively ineffective at stimulating high-rate synthesis of any non-IgM isotype. Dual cross-linking via CD40 and the BCR resulted in decreased synthesis of IgM with a concomitant increase in IgA and similar levels of IgG2 production to those obtained via the BCR alone. We also assessed the effects of endogenous and exogenous TGF-beta on immunoglobulin synthesis by bovine B cells. Exogenous TGF-beta stimulates both IgG2 and IgA production following CD40 and BCR cross-linking in the presence of IL-2. Blocking endogenous TGF-beta did not inhibit the up-regulation of IgG2 or IgA by interferons.

Animals↗
PubMed1998

DNA and a CpG oligonucleotide derived from Babesia bovis are mitogenic for bovine B cells.

DNAs from bacteria and variety of nonvertebrate organisms, including nematodes, mollusks, yeasts, and insects, cause polyclonal activation of murine B lymphocytes. Similar studies have not been reported for bovine B cells, and to date no studies have reported mitogenic properties of protozoal DNA for any species. However, we and others have observed that protozoal parasite antigens can induce the proliferation of lymphocytes from nonexposed donors. Extending these studies, we now show that the mitogenic property of protozoal antigen preparations is in part attributable to parasite DNA and that Babesia bovis DNA is directly mitogenic for bovine B cells. DNase treatment of B. bovis extracts abrogated B. bovis-induced proliferation of peripheral blood mononuclear cells from nonexposed cattle. Like DNAs from other organisms that were mitogenic for murine B cells, B. bovis DNA is largely nonmethylated and induced a dose-dependent proliferation of bovine B cells, which was reduced upon methylation. Furthermore, B. bovis and E. coli DNAs enhanced immunoglobulin secretion by cultured B cells, inducing moderate increases in immunoglobulin G1 and stronger increases in immunoglobulin G2. Because certain nonmethylated CpG motifs present in bacterial DNA are known to stimulate proliferation of murine and human B cells, an 11-kb fragment of B. bovis DNA was analyzed for CG dinucleotide content and for the presence of known immunostimulatory sequences (ISS) centered on a CG motif. The frequency of CG dinucleotides was approximately one-half of the expected frequency, and several CpG hexameric sequences with known activity for murine B cells were identified. An oligodeoxynucleotide containing one of these ISS (AACGTT), which is present within the rhoptry-associated protein-1 (rap-1) open reading frame, was shown to stimulate B-cell proliferation. These ISS may be involved in host immune modulation during protozoal infection and may be useful as vaccine adjuvants.

Animals↗
PubMed1998

Immunodominant T-cell antigens and epitopes of Babesia bovis and Babesia bigemina.

Despite convincing evidence that T cells are critical for both cellular and humoral immunity against haemoprotozoan parasites, the difficulty of performing meaningful experiments in cattle that would define the role of T cells in immunity to Babesia spp. has impeded research in this area. However, experiments performed ex vivo with immune T cells can reflect in-vivo events, and provide valuable insight into the nature of immunogenic proteins and the responding lymphocytes. The progress made towards identification of the immunogenic proteins and epitopes that stimulate anamnestic CD4+ type-1 (interferon-gamma-producing) T-cell responses in cattle immune to challenge with Babesia bovis or B. bigemina is the subject of the present review.

Animals↗
PubMed1997

Cloning, expression and biological function of the bovine CD40 homologue: role in B-lymphocyte growth and differentiation in cattle.

Members of the tumour necrosis factor receptor superfamily play a key role in B-lymphocyte survival, proliferation, differentiation and programmed cell death. A member of this superfamily, the CD40 molecule plays an important role in the differentiation of B lymphocytes into effector cells and in early activation through cognate interaction with T lymphocytes. In this report, we describe a cDNA and its protein product identified in cattle with approximately 70% sequence conservation at the nucleic acid level with the human CD40 gene. Transcripts for the boCD40 molecule were identified in resting and activated B lymphocytes, some but not all CD4+ and CD8+ T lymphocyte clones, and peripheral blood-derived T lymphocytes. Coculture of resting B cells with simian virus 40 (SV40)-transformed NIH3T3 (MOP8) cells stably transfected with the ligand for CD40 (bovine CD40 ligand, boCD40L), resulted in proliferation which was enhanced by addition of rbo interleukin-4 (IL-4). Cross-linkage of CD40 on bovine B lymphocytes upon coculture with CD40L-transfected cells resulted in the increased production of secretory IgM and, to a lesser extent, of IgG. Addition of rboIL-4 to these cultures increased levels of IgM and IgG secretion approximately twofold over those induced by CD40L alone. Our results indicate that many of the functions described for human and mouse CD40 are also conserved in the bovine but that differences in subset distribution of expression of the CD40 molecule in lymphoid cell types in cattle may impact on regulation of the early activation steps in the acquired immune response.

Amino Acid Sequence↗
PubMed1996

Differentiation of B cells in the bovine. Role of cytokines in immunoglobulin isotype expression.

Cytokines such as IFN-gamma, IL-4 and TGF-beta, are known to regulate Ig class switching by positively or negatively regulating germline CH transcription in the mouse and human. Here we show at the protein level that similar regulatory mechanisms are in place in the bovine using various modes of B cell activation and cytokine combinations. Using antigen-receptor cross-linkage or mitogen (pokeweed mitogen) costimulation, we demonstrate that recombinant bovine IL-4 upregulates the expression of IgM, IgG1 and IgE in vitro. Upregulation of IgG1 and IgE production by rboIL-4 is inhibited in a dose-dependent manner by the addition or rboIFN-gamma to the cultures. Using similar methodologies, we have also investigated the effects of TGB-beta on IgA production in the presence or absence of various cofactors including IL-10 and IL-2. We find that TGF-beta will not solely upregulate IgA production from noncommitted precursors (sIgM+) in coculture with mitogen or with formalin fixed Staphylococcus cowan Strain I (SAC), but does have positive effects on IgA production in the presence of both IL-2 and IL-10. These observations suggest that different modes of T-independent costimulation using classical B cell activators does not result in differential IgG subisotype or IgE production by bovine B cells in the presence of cytokines. Together these results suggest that individual cytokines regulate isotype expression in cattle.

Animals↗
PubMed1996

Antigen-specific humoral immune responses to a hapten-carrier conjugate in SCID mice engrafted with bovine fetal hematopoietic tissues.

We describe the conditions necessary to reconstitute a bovine immune system in SCID mice that supports the generation of antigen-specific serum antibody responses to a hapten-carrier (DNP-KLH) conjugate. Sublethally irradiated SCID mice were engrafted with fetal bovine liver, lymph node, and thymus. Treatment groups were established in which human recombinant interleukin 7, autologous bovine bone marrow, and engraftment site (omentum or abdominal wall) were evaluated for their ability to enhance the reconstitution of the bovine immune system in SCID mice. Engrafted SCID mice were inoculated at approximately 6 and 12 weeks after engraftment with DNP-KLH. Vaccinated mice were evaluated by enzyme-linked immunosorbent assay for serum antibody specific for DNP-KLH and total concentrations of bovine serum immunoglobulin biweekly through 20 weeks after engraftment. The SCID mice with fetal tissues engrafted onto the abdominal wall from two different donors produced microgram quantities of bovine immunoglobulins and had >70% reconstitution rate. Administration of human interleukin 7 and/or fetal bovine bone marrow did not significantly increase the frequency of reconstitution. Subclass analysis of serum from mice receiving the abdominal implants suggested that class-switching occurred to a predominant non-IgM isotype response after the second inoculation. Pooled thymic and splenic cell populations obtained 20 weeks after engraftment from abdominally engrafted groups yielded 38 to 90% bovine cells, as determined by polymerase chain reaction in situ hybridization with bovine satellite DNA-specific primers. These results indicate that an antigen-specific bovine immune response can be generated in heterochimeric SCID-bo mice.

Animals↗
PubMed1995

Expression and biological activities of bovine interleukin 4: effects of recombinant bovine interleukin 4 on T cell proliferation and B cell differentiation and proliferation in vitro.

Interleukin 4 (IL-4) is a pleotropic cytokine affecting a wide range of cell types in both the mouse and the human. These activities include regulation of the growth and differentiation of both T and B lymphocytes. The activities of IL-4 in nonprimate, nonmurine systems are not well established. Herein, we demonstrate in the bovine system that IL-4 upregulates production of IgM, IgG1, and IgE in the presence of a variety of costimulators including anti-IgM, Staphylococcus aureus cowan strain I, and pokeweed mitogen. IgE responses are potentiated by the addition of IL-2 to IL-4. Culture of bovine B lymphocytes with IL-4 in the absence of additional costimulators resulted in the increased surface expression of CD23 (low-affinity Fc epsilon RII), IgM, IL-2R, and MHC class II in a dose-dependent manner. IL-4 alone increased basal levels of proliferation of bulk peripheral blood mononuclear cells but in the presence of Con A inhibited proliferation. In contrast to the activities of IL-4 in the murine system, proliferation of TH1- and TH2-like clones was inhibited in a dose-dependent manner as assessed by antigen-or IL-2-driven in vitro proliferative responses. These observations are consistent with the role of IL-4 as a key player in regulation of both T and B cell responses.

Animals↗
PubMed1994

IFN-gamma stimulates IgG2 production from bovine B cells costimulated with anti-mu and mitogen.

The effect of interferon-gamma (IFN-gamma) antibody production by anti-mu- or pokeweed mitogen-stimulated bovine B cells was studied. IFN-gamma induced IgG2 secretion in isolated bulk B cell populations and in B cells sorted for IgM expression. IgM production was suppressed by the presence of IFN-gamma alone but this effect was antagonized by interleukin 2 (IL2). The effects of IFN-gamma on secreted levels of IgM, IgG1, and IgG2 correlated with the frequencies of cells expressing transcripts of the respective isotypes when stimulated with IFN-gamma-containing T cell supernatants. These results indicate that IFN-gamma plays a key role in IgG2 production in the bovine by directly affecting suitably stimulated B cells. The ability of IL2 to synergize with IFN-gamma to augment both the IgM and IgG2 responses implicates a TH1-like subset in regulation of this isotype.

Animals↗
PubMed1993

Characterization of an unusual cell type (CD4+ CD3-) expanded by helminth infection and related to the parasite stress response.

Mice infected with the parasite Mesocestoides corti develop hypergammaglobulinemia, hepatomegaly, and splenomegaly. The immune response to M. corti infection is directed, in part, at molecules secreted by the organism. Two of these molecules have been shown to be hsp70 and hsp60 homologues. In this study it was found that incubation of splenocytes from infected animals with M. corti-secreted molecules or the isolated M. corti hsp70 results in the expansion of an unusual cell type with the morphology of large granular lymphocytes. The cell lines express Thy-1, CD4 (low), and CD45RB but lack TCR alpha beta, TCR gamma delta, CD3, CD8, and slg. The lack of a TCR suggested NK cells, but no cytolytic activity could be detected. In addition, the cell lines constitutively produce IL-6 and can be induced to express IL-2, but not IL-4, IL-5, or IFN-gamma. Given the phenotype of these cells, it is possible that they represent T lineage precursors or some type of effector cells. Notably, CD3- CD4+ cells appear to be expanded in the spleens and livers of M. corti-infected animals, suggesting an important role in infection. Moreover, the selective growth of this cell type with M. corti hsp70 suggests that the outgrowth and in vivo expansion of these cells may be related to the stress response of the parasite.

Animals↗
PubMed1991

Biochemical and functional analysis of extracellular stress proteins of Mesocestoides corti.

Previous studies of the serum antibody response in mice to Mesocestoides corti infection indicated that molecules released by the parasite influenced the production of IgM and IgG1 to the exclusion of other isotypes. Two proteins isolated from M. corti culture supernatants were found to be homologous to the 70-kDa heat shock proteins (hsp70) and Escherichia coli GroEL families of stress proteins. The proliferative responses of splenic lymphocytes from infected mice were assessed to unfractionated M. corti supernatants as well as the 70- and 60-kDa stress protein homologs isolated from supernatants. Lymphocytes from infected mice respond to complete supernatant and both of the isolated p70 and p60 stress protein homologs. In addition, supernatant from M. corti cultures stimulates an in vitro antibody response restricted to IgM and IgG1; the same isotypes induced during infection. These results suggest that stress proteins play an integral part in the immune response to M. corti and the associated isotype restriction.

Adenosine Triphosphate↗
PubMed1991

In vivo effects of anticytokine antibodies on isotype restriction in Mesocestoides corti-infected BALB/c mice.

Chronic infection of mice with the cestode Mesocestoides corti results in an antibody response restricted to immunoglobulins M (IgM) and G1 (IgG1). To determine which of the known lymphokines are involved in the restricted isotype response, we treated M. corti-infected mice with a panel of anticytokine monoclonal antibodies against interleukin-4 (IL-4), IL-5, IL-6, and gamma interferon. The effects of anti-IL-4 were of particular importance, since IL-4 is known to enhance IgG1 production and an IgG1 response predominates in infected animals. Interestingly, injection of anti-IL-4 alone had no effect on IgG1 levels at day 7 postinfection and actually enhanced levels at day 10. Decreases in IgM levels were observed in anti-IL-4-treated mice. Administration of anti-IL-5 inhibited IgM production early in infection, but no effects on IgG1 levels were observed. Treatment of infected mice with anti-gamma interferon had no effect on any of the isotypes analyzed. Treatment of infected mice with anti-IL-6 antibody had the most dramatic effects, with inhibition of IgM and IgG1 by day 14 of infection. The transient expression of IgG3, which is sometimes observed very early in the infection process, was also inhibited by anti-IL-6, suggesting that the inhibition observed was not isotype specific. To determine whether cytokines were acting in concert to effect the high IgM and IgG1 levels in infected animals, anticytokine antibodies were also injected in combinations. However, the only combinations that inhibited IgG1 levels contained anti-IL-6, and the extent of inhibition was not greater than that of anti-IL-6 alone. Results are discussed in terms of the effects of cytokines on parasite-induced isotype expression and the potential for IL-4-independent mechanisms of IgG1 production.

Animals↗
PubMed1990

Production and use of murine monoclonal antibodies reactive with bovine IgM isotype and IgG subisotypes (IgG1, IgG2a and IgG2b) in assessing immunoglobulin levels in serum of cattle.

Utilizing in-vitro and in-vivo immunizations, murine monoclonal antibodies (Mab's) that are specific for bovine immunoglobulin (IG)--IgM isotype and IgG subisotypes (IgG1, IgG2a and IgG2b)--have been produced. These Mab's were used to estimate the quantities of these bovine Ig isotype and IgG subisotypes in the sera of 56 cows and 24 bulls. The cows were all approximately 18 months of age and the bulls ranged from 12 to 18 months in age. The cows and bull were all apparently healthy and unimmunized. The cows were not pregnant and were nulliparous. Some differences were observed between the serum levels of IgM (higher in cows) and IgG1 (higher in bulls). No ready explanation can be offered for these observed differences.

Animals↗