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D L Mueller

Publications and source records attributed to D L Mueller.

At least 37 records · Page 2Linked to original sources
PubMed1996

Blocked signal transduction to the ERK and JNK protein kinases in anergic CD4+ T cells.

T cells activated by antigen receptor stimulation in the absence of accessory cell-derived costimulatory signals lose the capacity to synthesize the growth factor interleukin-2 (IL-2), a state called clonal anergy. An analysis of CD3- and CD28-induced signal transduction revealed reduced ERK and JNK enzyme activities in murine anergic T cells. The amounts of ERK and JNK proteins were unchanged, and the kinases could be fully activated in the presence of phorbol 12-myristate 13-acetate. Dephosphorylation of the calcineurin substrate NFATp (preexisting nuclear factor of activated T cells) also remained inducible. These results suggest that a specific block in the activation of ERK and JNK contributes to defective IL-2 production in clonal anergy.

Animals↗
PubMed1996

Differential regulation of bcl-2 and bcl-x by CD3, CD28, and the IL-2 receptor in cloned CD4+ helper T cells. A model for the long-term survival of memory cells.

In this study, we examined the molecular signals that control apoptosis in cloned CD4+ helper T cells. Resting T cells were highly resistant to spontaneous death in the absence of exogenous stress, and they expressed low levels of bcl-x protein and no detectable bcl-2. Upon exposure to gamma radiation, resting cells rapidly underwent apoptotic death. Incubation with IL-2 prevented this cell death and led to a large increase in bcl-2 protein expression and only a modest up-regulation in bcl-x. The combination of anti-CD3 and anti-CD28 mAbs was also effective in protecting the cells against radiation-induced apoptosis; however, this protection was associated predominantly with bcl-x up-regulation, and only a small induction of bcl-2 protein was observed. Finally, cyclosporin A blocked both IL-2 secretion and bcl-2 induction in response to CD3 plus CD28 stimulation, suggesting a role for endogenous lymphokine production in the induction of bcl-2. These data support a model in which memory T cells remain resistant to apoptosis because intermittent contact with Ag-bearing APC and IL-2R occupancy result in the expression of the life-proteins bcl-2 and bcl-x.

Animals↗
PubMed1996

Frequent aberrant immunoglobulin gene rearrangements in pro-B cells revealed by a bcl-xL transgene.

During B lymphocyte development, pro-B cells that fail to rearrange an immunoglobulin heavy (IgH) chain allele productively are thought to undergo developmental arrest and death, but because these cells are short-lived in vivo they are not well characterized. Transgenic mice expressing the apoptosis regulatory gene bcl-xL in the B lineage developed large expansions of pro-B cells in bone marrow. V(D)J rearrangements in the expanded populations were nearly all nonproductive, and DJH rearrangements were enriched for joints in DH reading frame 2 and for aberrant joints with extensive DH or JH deletions. Thus, the death of pro-B cells with failed immunoglobulin rearrangements occurs by apoptosis, and bcl-xL can deliver a strong survival signal at the pro-B stage. This analysis also demonstrated that immunoglobulin gene rearrangement is less precise than previously appreciated.

Animals↗
PubMed1995

Bcl-xL rescues WEHI 231 B lymphocytes from oxidant-mediated death following diverse apoptotic stimuli.

Developing lymphocytes undergo extensive cell death during selection of the immune repertoire. We investigated the influence of bcl-xL, a member of the bcl-2 family of apoptosis regulatory genes, on apoptosis in a model system for negative selection in the B lymphoid lineage. Overexpression of bcl-xL in WEHI 231 immature mouse B cells blocked apoptosis triggered by cross-linking of surface IgM. bcl-xL-transfected cells were also resistant to apoptosis following incubation in low serum medium or exposure to gamma-irradiation, the sphingomyelin ceramide, or compounds that increase intracellular levels of oxidants. Remarkably, the addition of antioxidants (catalase, N-acetylcysteine, or pyruvate) alone rescued the native WEHI 231 cells from apoptosis while having only minor effects on the viability of cells overexpressing bcl-xL. Anti-IgM cross-linking, ceramide, and gamma-irradiation treatments elevated intracellular peroxide production, which was prevented by treatment with antioxidants. Cells overexpressing bcl-xL had a similar rise in intracellular oxidants as control cells, indicating that bcl-xL modifies the cell's response to oxidants while having no detectable influence on the endogenous production of oxidants following apoptotic stimuli. These data implicate bcl-xL as a potent death repressor in B lymphocytes and support the hypothesis that bcl-xL regulates survival decisions within susceptible cells by functioning downstream of oxidant production.

Animals↗
PubMed1995

Molecular mechanisms underlying functional T-cell unresponsiveness.

Evidence from tissue culture experiments and in vivo studies indicates that certain forms of antigen presentation render T cells unresponsive to subsequent antigenic stimulation. Great progress has been made recently in understanding the induction, maintenance, and in vivo relevance of this unresponsive state, called clonal anergy.

Animals↗
PubMed1994

Cloning and molecular characterization of mouse bcl-x in B and T lymphocytes.

We report the cloning and initial characterization of the mouse homologues of the bcl-2-related gene, bcl-x. We compare these to the two major isoforms of human bcl-x that were previously identified, bcl-xLong (bcl-xL) which has death repressor activity similar to bcl-2, and bcl-xShort (bcl-xS), an alternative mRNA splice product that deletes a highly conserved domain shared by bcl-2 family members and promotes cell death in transfection studies. In addition to murine (m) bcl-xL and mbcl-xS, we have cloned a novel cDNA isoform that we designate mbcl-x delta TM. This cDNA deletes, by means of alternative splicing, the carboxy terminal transmembrane domain of bcl-x and is predicted to be a soluble, rather than membrane-bound, protein. We found that mbcl-x mRNA is highly inducible in splenocytes stimulated with either anti-CD3 Abs or LPS/dextran sulfate, and that the major species of mbcl-x mRNA in a variety of cell lines and tissues was the xL isoform. Transfection of mbcl-xL or mbcl-xS into Hela cells resulted in targeting primarily to mitochondria, whereas mbcl-x delta TM localized diffusely throughout the cytosol. Overexpression of the novel delta TM isoform in an IL-3-dependent cell line delayed the onset of apoptosis induced by growth factor withdrawal. Thus, a naturally-occurring form of mbcl-x present in lymphocytes that does not localize to the mitochondrial membrane is functional in preventing apoptotic cell death. Moreover, these data suggest that bcl-x provides a life signal in activated lymphocytes.

Amino Acid Sequence↗
PubMed1994

Subset of CD4+ T cell clones expressing IL-3 receptor alpha-chains uses IL-3 as a cofactor in autocrine growth.

In this paper we demonstrate that IL-3 can act as a cofactor for the growth of some CD4+ T cells. This lymphokine synergized with IL-4 to induce both a unique set of protein tyrosine phosphorylations and the vigorous proliferation of the keyhole limpet hemocyanin-specific and I-Ab-restricted CD4+ Th0 cell clone, E6. In addition, neutralizing anti-IL-3 Abs specifically inhibited the growth of E6 T cells to Ag or anti-CD3 mAb stimulation. Finally, this T cell clone was shown to express both the IL-3R alpha-chain and an IL-3R beta-chain (AlC2A). An examination of other CD4+ T cell clones determined that one Th1 clone (A.E7), two Th0 clones (16B.2 and L9A.1), and one Th2 clone (D10.G4.1) were not influenced by the addition of rIL-3. However, proliferation of the Th2 clones CDC25 and CDC35 to CD3-stimulation was significantly enhanced by IL-3. The sensitivity of these latter two clones to IL-3 was also found to be associated with expression of IL-3R alpha-chains. Because E6 T cells are highly dependent on IL-4 for autocrine growth similar to Th2 cells, these results suggest that IL-3 may synergize with IL-4 to enhance the proliferation of a subset of IL-4-dependent CD4+ T cells, and the study indicates that IL-3R alpha-chain expression may be a specific marker of this CD4+ T cell subset.

Animals↗
PubMed1991

Clonal anergy blocks the response to IL-4, as well as the production of IL-2, in dual-producing T helper cell clones.

In this report we extend the in vitro clonal anergy model to examine the regulation of proliferation in T cells that secrete both IL-2 and IL-4. Newly cloned Ag-specific murine T cells are shown to depend on both IL-2 and IL-4 synthesis for maximal proliferation. Whereas IL-2 responsiveness is constitutive in these cells, IL-4 responsiveness develops only after Ag and APC stimulation. Remarkably, proliferation of these cells to Ag is sensitive to inhibition by clonal anergy, even though IL-4 synthesis remains inducible. Anergy in these cells is associated with an inability to respond to IL-4, in addition to the development of an IL-2 production defect. The results suggest that anergy induction may be capable of preventing the clonal expansion of autoreactive T cells producing both IL-2 and IL-4 in vivo.

Animals↗
PubMed1990

An intracellular calcium increase and protein kinase C activation fail to initiate T cell proliferation in the absence of a costimulatory signal.

Resting T lymphocytes proliferate in response to a combination of a calcium ionophore and a phorbol ester. This observation suggests that an increase in intracellular calcium free ion concentration [Ca2+]i and activation of protein kinase C (PKC) are sufficient signaling events for the initiation of T cell proliferation. In contrast, an accessory cell-generated costimulatory signal, acting independently of the rise in [Ca2+]i and PKC activation, is required for Ag-induced proliferation of type I T cell clones. We now report that this costimulatory signal is unexpectedly also being delivered via a cell-cell interaction during the response to ionomycin and phorbol ester. In the absence of this signal (at limiting cell numbers), T cells fail to divide. We also demonstrate that proliferation in response to immobilized anti-CD3 mAb requires the cell-cell interaction. These results suggest a model of T cell stimulation in which activation of a costimulatory signaling pathway is important in the regulation of the IL-2 gene, and only in the presence of this (third) signal can an increase in [Ca2+]i and PKC activity induce T cell proliferation. Such a model predicts that IL-2-dependent expansion of T cell clones in vivo in response to Ag receptor occupancy requires the delivery of an independent accessory cell-derived co-stimulatory signal.

Animals↗
PubMed1990

Inhibition of antigen-specific proliferation of type 1 murine T cell clones after stimulation with immobilized anti-CD3 monoclonal antibody.

The effect of stimulating normal type 1 murine T cell clones with anti-CD3 antibody was examined in vitro. In the absence of accessory cells, anti-CD3 antibody immobilized on plastic plates stimulated inositol phosphate production, suboptimal proliferation, IL-2 and IL-3 production, and maximal IFN-gamma production. Addition of accessory cells augmented lymphokine production and proliferation when the effects of "high-dose suppression" were relieved by removing the T cells from the antibody-coated plates. Exposure of type 1 T cell clones to immobilized anti-CD3 antibody alone rapidly induced long-lasting proliferative unresponsiveness (anergy) to Ag stimulation that could be prevented by accessory cells. This anergic state was characterized by a lymphokine production defect, not a failure of the T cells to respond to exogenous IL-2 or to express surface Ti/CD3 complexes. In addition, anergy could not be induced in the presence of cyclosporine A. These results suggest that under certain conditions anti-CD3 antibodies may have potent immunosuppressive effects independent of Ti/CD3 modulation. Furthermore, our results support a two-signal model of type 1 T cell activation in which Ti/CD3 occupancy alone (signal 1) induces anergy, whereas Ti/CD3 occupancy in conjunction with a costimulatory signal (signal 2) induces a proliferative response.

Animals↗
PubMed1990

Percutaneous transgastric drainage of the lesser sac in children.

In four children (5-14 years old), lesser sac fluid collections (3 pseudocysts and 1 abscess) were externally drained using a percutaneous transgastric approach. With ultrasound or CT guidance and fluoroscopic monitoring, a sheathed needle is inserted through the stomach into the lesser sac fluid collection, the catheter is exchanged, secured in place and connected to external drainage. All four fluid collections were complications of pancreatitis (3 acute post-traumatic, 1 post-surgical). All four fluid collections were successfully drained without need for surgical intervention. There were no major complications. Three minor complications occurred as a result of reinsertion of catheters: transient gastric venous bleeding, hematuria, and bleeding into the pseudocyst. Percutaneous transgastric drainage can be performed successfully in children, and provides an alternative to surgical intervention of lesser sac fluid collections. Drainage early in the course of pancreatic pseudocysts may lessen the morbidity, potential mortality, and prolonged hospitalization usually necessary for standard medical and surgical management of these patients.

Abscess↗
PubMed1989

An accessory cell-derived costimulatory signal acts independently of protein kinase C activation to allow T cell proliferation and prevent the induction of unresponsiveness.

Presentation of Ag to type I CD4+ T cell clones by chemically fixed APC results in the induction of a long-lasting state of proliferative unresponsiveness in the T cell. Ag-specific TCR interactions do occur during this stimulation, as Ag- and Ia molecule-dependent increases in intracellular calcium free ion concentration can be demonstrated, yet free inositol phosphate generation is low and neither IL-2 synthesis nor proliferation occur. The addition of normal allogeneic accessory cells during this stimulation can restore the T cell proliferative response, as well as prevent the induction of unresponsiveness, thus defining an accessory cell-dependent costimulatory activity necessary for proliferation. We have now examined the biochemical effects of this costimulatory activity on early T cell activation. Normal accessory cell costimulatory activity was found to be incapable of augmenting the generation of free inositol phosphate in response to either fixed APC plus Ag or Con A alone. Furthermore, protein kinase C-dependent CD3 gamma-chain phosphorylation occurred in response to either fixed APC plus Ag or Con A alone, and the addition of normal accessory cells had no effect on the level of this phosphorylation. Finally, minimal CD3 zeta-chain tyrosine phosphorylation occurred during the induction of unresponsiveness with Ag and fixed APC alone and this also was not affected by the costimulatory activity. Our results demonstrate that T cell Ag receptor-mediated increases in intracellular calcium free ion concentration and protein kinase C activation occur independently of an accessory cell-derived costimulatory signal. In the absence of this costimulatory signal, these two intracellular second messengers are insufficient to induce a maximal proliferative response and in fact lead to a state of unresponsiveness.

Animals↗
PubMed1988

Acute scrotal abnormalities in children: diagnosis by combined sonography and scintigraphy.

Both scintigraphy and real-time sonography have been used to assess acute symptoms involving the scrotum. However, because of its high sensitivity and ability to document physiologic abnormalities, scintigraphy has been the procedure of choice. Scintigraphy, however, lacks specificity; its value lies mainly in serving to distinguish torsion from nontorsion. The purpose of this study was to supplement scrotal scintigraphy with sonography to determine if the combination improves diagnosis and management compared with scintigraphy alone. Forty-three scrotal scintigrams and sonograms were performed on 40 consecutive patients with acute scrotal symptoms. The interpretation of the scintigram was altered by sonography in six (14%) of the combined scans, directly affecting clinical management. In three patients with acute hydroceles diagnosed by sonography, exploratory surgery was avoided despite scintigraphic findings suggesting testicular torsion. Scintigraphy was normal in two patients with spontaneous testicular detorsion, whereas sonography showed recent spermatic-cord torsion that required subsequent orchiopexy. In a patient with epididymitis and orchitis, sonography showed a complicating scrotal abscess, which was not apparent on scintigraphy and which required antibiotic treatment. The addition of sonography to the scintigraphic evaluations of children with acute scrotal abnormalities changed the diagnosis and clinical management in 14% of the patients studied.

Acute Disease↗