bluuebunny/crossref_metadata_2025 huggingface dataset:
B cell lymphoma-2 transfected P815 cells resist reactive nitrogen intermediate-mediated macrophage-dependent cytotoxicity.
Authors: Albina J E , Martin B A , Henry W L , Louis C A , Reichner J S
Published: December 2022

Activated murine peritoneal macrophage cytotoxicity against P815 tumor cells has been shown to be mediated by the reactive nitrogen intermediates (RNI) produced by macrophages from L-arginine through nitric oxide (NO) synthase. Previous results from this laboratory indicated that NO-dependent killing of P815 fulfilled the criteria for apoptotic death. Work by others, in turn, demonstrated that the product of the bcl-2 gene confers protection against various inducers of apoptosis, including reactive oxygen intermediates. Experiments were performed to determine whether Bcl-2 could equally protect sensitive cells from RNI-dependent apoptosis within the context of a relevant biologic system such as the delivery of such RNI by activated macrophages. Results demonstrated that transfection of P815 cells with the human bcl-2 gene confers immunity from RNI-dependent, macrophage-mediated cytotoxicity. In contrast with wild-type or mock-transfected P815 cells, which do not contain detectable Bcl-2, bcl-2-transfected cells showed minimal DNA fragmentation and cell membrane failure when cocultured with activated macrophages. Additional findings indicate that Bcl-2 affords the transfected cells almost complete resistance to the DNA-fragmenting effects of chemically generated NO or H202 and partial protection from their cytolytic effects. These findings are consistent with the hypothesis that tumor cells expressing bcl-2 may escape destruction by macrophage-dependent immune surveillance mechanisms.
IFN-alpha beta reconstitutes the deficiency in lipid A-activated AKR macrophages for nitric oxide synthase.
Authors: Jiang H , Rummage J A , Zhou A , Chen Z , Herriot M J , Stewart C A , Kolosov M , Leu R W
Published: December 2022

AKR mouse peritoneal macrophages (PM) were previously found to have a defect in their response to lipid A for nitric oxide (NO)-mediated tumor cytotoxicity, which was related to a lower level of C1q synthesis and reconstituted by exogenous IFN-gamma or C1q. We used AKR-PM as a model to further define the role of IFN-alpha beta in modulation of induction of macrophage nitric oxide synthase (NOS) in response to lipid A. Studies have revealed that AKR-PM produced a significantly lower level of IFN-alpha beta than responsive C3H-PM in response to lipid A. AKR-PM failed to increase NOS mRNA synthesis and NO generation when exposed to lipid A, although they had normal levels of TNF-alpha bioactivity and mRNA expression. This partial deficiency of AKR-PM to lipid A stimulation was reconstituted completely by exogenous IFN-alpha beta for both synthesis of NOS mRNA and release of NO. The failure of AKR-PM to produce NOS to lipid A stimulation appears to be related to reduced secretion of IFN-alpha beta and the resultant failure to express TNF-alpha type II receptor (TNF-RII) mRNA, which in turn decreases TNF-alpha binding to its receptor for autocrine induction of NOS. Insufficient synthesis and secretion of endogenous IFN-alpha beta may be the primary reason for AKR-PM refractoriness to induction of NOS in response to lipid A. furthermore, the close correlation between lack of IFN-alpha beta secretion and decreased TNF-RII mRNA synthesis may implicate a critical role for IFN-alpha beta in the upregulation of macrophage TNF-RII receptor expression for autocrine induction of NOS during lipid A stimulation.
Human monocytes are unable to bind to or phagocytize IgA and IgG immune complexes formed with influenza virus in vitro.
Authors: Scott C B , Ratcliffe D R , Cramer E B
Published: December 2022

An in vitro model system was used to investigate how human monocytes recognize influenza A/WSN(H1N1)-infected epithelial cells, and the role that anti-influenza IgA and IgG Abs play in this interaction. Pretreatment of the monocytes with neuraminidase or F(ab')2 fragments of a mAb against the Ca1 epitope on the hemagglutinin (HA) molecule inhibited monocyte adherence to the infected cells. This suggested that monocytes bound to the sialic acid binding site on the HA molecule and not to other viral or epithelial cell Ags. In the presence of neutralizing concentrations of intact Abs (human serum IgA, secretory IgA, IgG, or mouse anti-HA mAb), monocytes were unable to bind to influenza-infected epithelium or to phagocytize the IgA or IgG immune complexes formed with influenza virus, even though they could use these same Abs to attach to or phagocytize inert particles. Under conditions of Ag excess and low concentrations of Ab, monocytes bound primarily to the viral HA molecule, but showed some recognition of the viral immune complexes. This dual binding did not increase monocyte adherence to the infected epithelium above that observed with virus alone. These findings indicate that neutralizing concentrations of IgA or IgG Abs, the predominant Abs found in the upper and lower respiratory tract, respectively, do not augment and can prevent monocyte recognition of influenza virus. This suggests that in vivo monocytes must use a non-Fc-mediated mechanism to adhere to and phagocytize these immune complexes.
Heat shock protein 65 induces CD62e, CD106, and CD54 on cultured human endothelial cells and increases their adhesiveness for monocytes and granulocytes.
Authors: Verdegaal M E , Zegveld S T , van Furth R
Published: December 2022

Expression of members of the heat shock protein 60 (hsp60) family in tissues has been reported to coincide with leukocyte infiltration, but it is not known whether these proteins are directly involved in the extravasation of leukocytes. Extravasation of leukocytes requires their adhesion to endothelial cells (EC) via an interaction between adhesion molecules expressed on both cell types. The aim of the present study was to investigate the effect of recombinant mycobacterial hsp65 on the adhesive characteristics of EC for monocytes and granulocytes. Incubation of EC with hsp65 induces a concentration- and time-dependent increase in adhesiveness of these EC for monocytes and granulocytes. The effect was maximal after incubation of EC with hsp65 for 4 to 6 h. In addition, incubation of EC with hsp65 induced the expression of endothelial CD62E (E-selectin), CD106 (vascular cell adhesion molecule-1) and CD54 (intercellular adhesion molecule-1). The increased adhesion of granulocytes to hsp65-stimulated EC was inhibited completely by blocking Ab against CD62E. mAb against endothelial CD62E, CD106, or CD54 or against the monocyte adhesion molecules CD14 or CD49d (very late Ag-4) did not inhibit the increased adhesion of monocytes to hsp65-stimulated EC; however, mAb against the monocyte adhesion molecule CD18 (beta2-integrin) inhibited monocyte adhesion to hsp65-stimulated EC to the same extent as monocyte adhesion to nonstimulated EC. Hsp65 did not exert its effect in an autocrine or paracrine fashion via the endogenous production of IL-1, TNF-alpha, or other factors or via contaminating LPS. Together these results indicate that hsp65 can play an important role in the adhesion of monocytes and granulocytes to EC at sites of inflammation via modulation of the adhesive characteristics of EC and thus may facilitate extravasation of these phagocytes.
5-Lipoxygenase metabolism in alveolar macrophages from subjects infected with the human immunodeficiency virus.
Authors: Coffey M J , Phare S M , Kazanjian P H , Peters-Golden M
Published: December 2022

Pulmonary infection represents a major source of morbidity and mortality in AIDS. One important component of pulmonary host defense is the elaboration by resident alveolar macrophages (AM) of proinflammatory leukotrienes (LT) and other 5-lipoxygenase (5-LO) metabolites of arachidonic acid (AA). In this study, we compared the 5-LO metabolic capacity of AM isolated from normal controls with two groups of HIV-infected subjects: (1) patients with low CD4 counts undergoing diagnostic evaluation for pulmonary indications, and (2) volunteers without pulmonary complaints stratified into normal (> 500) and low (< 200) CD4 count groups. Compared with AM from control subjects, AM from HIV-infected subjects with normal and low CD4 counts demonstrated a marked reduction in LT synthesis. This reduced metabolic capacity could not be attributed to in vivo activation because there was no increase in lavage fluid LTB4 levels. However, there was a reduction (approximately twofold) in 5-LO protein expression in both the normal and the low CD4 subsets. 5-LO-activating protein (FLAP) expression was unchanged in cells from the normal CD4 HIV group, but was decreased threefold in the two groups with low CD4 counts. These observations indicate that there is a graded defect in the 5-LO metabolic capacity of AM from HIV-infected subjects, with decreased expression of only 5-LO in the normal CD4 group, and decreased expression of both 5-LO and FLAP in the low CD4 group. This defect would be expected to compound the immunosuppression seen in these subjects.
Defective monocyte costimulation for IFN-gamma production in familial disseminated Mycobacterium avium complex infection: abnormal IL-12 regulation.
Authors: Frucht D M , Holland S M
Published: December 2022

We have described previously a family in which several members have disseminated Mycobacterium avium complex infection. PBMC from affected members produced abnormally low amounts of IFN-gamma upon stimulation with PHA. Using PHA-stimulated allogeneic cocultures of highly purified monocytes and T cells from familial patients and normal subjects, we have now demonstrated that familial patient monocytes are defective in accessory cell function for IFN-gamma production. Familial patient monocytes did not inhibit IFN-gamma production by normal cells, nor did inhibition of PG synthesis restore normal IFN-gamma production by familial patient cells. Familial patient cells responded to the addition of exogenous IL-12 by increasing IFN-gamma production, while addition of exogenous anti-IL-12 had an insignificant effect on their IFN-gamma production. IL-12 was undetectable in PHA-stimulated cocultures of familial patient monocytes with familial or normal T cells. In addition, IL-12 production by adherent cells from patients and their unaffected mothers was abnormally low following stimulation with fixed Staphylococcus aureus Cowan I strain. However, normal amounts of IL-12 were detected when adherent familial patient cells were stimulated with S. aureus Cowan I strain and IFN-gamma, suggesting abnormal regulation of IL-12 production by familial monocytes. This is the first report of defective IL-12 production associated with increased susceptibility to an infectious disease, a finding that supports the critical role of this cytokine in host defense.
Analysis of central B cell tolerance in autoimmune-prone MRL/lpr mice bearing autoantibody transgenes.
Authors: Rubio C F , Kench J , Russell D M , Yawger R , Nemazee D
Published: December 2022

The effect of the autoimmune prone MRL/lpr (H-2k) genetic background on central B cell tolerance was studied in mice bearing 3-83 (anti-H-2Kk) Ig heavy and light chain transgenes. B cells bearing the dominant, transgene-encoded anti-H-2Kk specificity were tolerized appropriately on the MRL/lpr genetic background. Nevertheless, mice developed disease traits characteristic of the MRL/lpr strain, including lymphadenopathy and elevated levels of IgG dsDNA autoantibodies. Two transgenic lines were examined in this analysis: 3-83 mu delta, which expresses IgM and IgD forms of the 3-83 Ab, and Tol 1, which expresses only the IgM form of 3-83. The results obtained differed somewhat between the two transgenic lines. Crosses using 3-83 mu(delta) mice never demonstrated any defects in B cell self-tolerance to H-2Kk. Similarly, no Kk autoantibody production was seen in Tol 1 mice that were backcrossed onto the MRL/lpr genetic background and maintained in a specific pathogen-free facility. However, a subset of Tol 1/MRL/lpr mice that were housed in a conventional mouse facility demonstrated significant transgene-derived anti-Kk autoantibodies. Overall, these results suggest that there is no general defect in central B cell tolerance in MRL/lpr mice, despite their defect in the fas gene. These findings suggest similarities between the MRL/lpr T and B cell systems, because both fail to manifest clear central tolerance defects, but they nevertheless promote hyperplasia and autoimmunity in the peripheral immune system.
Maintenance of nuclear factor-kappa B/Rel and c-myc expression during CD40 ligand rescue of WEHI 231 early B cells from receptor-mediated apoptosis through modulation of I kappa B proteins.
Authors: Schauer S L , Wang Z , Sonenshein G E , Rothstein T L
Published: December 2022

Engagement of surface IgM (sIgM) on WEHI 231 murine B lymphoma cells leads to abortive activation and apoptosis, suggesting that this cell line may represent a model for tolerance. Loss of viability in these cells is preceded by an early increase in c-myc RNA levels followed by a decline below control values, implicating c-myc in the control of apoptosis. Costimulation with CD40 ligand (CD40L) has been shown to rescue WEHI 231 cells from anti-sIgM-induced apoptosis, and therefore, the effects of CD40L on c-myc RNA and protein levels were measured. Treatment of these cells with the combination of CD40L and anti-sIgM led to induction and maintenance of elevated levels of c-myc RNA and protein. Since transcriptional regulation of c-myc is mediated through two nuclear factor-kappa B (NF-kappa B) sites in WEHI 231, the effects of CD40L on DNA binding by this family of transcription factors were evaluated. CD40L induced and sustained the levels of NF-kappa B binding to both of these sites, paralleling the changes in c-myc RNA levels. Elevated levels of NF-kappa B were partially achieved through a sustained decrease in the steady state amount of the NF-kappa B/Rel-specific inhibitory protein, I kappa B alpha, and a transient decrease in I kappa B beta. These data lend support to the hypothesis that anti-sIgM-induced apoptosis is caused by a drop in c-myc expression and that an appropriate second signal, such as that provided by CD40L, is able to rescue these cells by inducing NF-kappa B and thereby maintaining c-myc RNA levels.
Cutting edge: terra firma: a retreat from "danger".
Authors: Langman R E , Cohn M
Published: December 2022

The danger model differs from the associative recognition model solely with respect to the origin of effector T helpers (eTh). More properly, the danger model should be juxtaposed to the Ag-independent model for the origin of eTh cells (i.e., the primer question). The introduction of danger in no way challenges the need to make a self-nonself discrimination, nor, for that matter, does it challenge the need to define self as those Ags encountered in the absence of eTh and that persist, whereas nonself are those Ags encountered in the presence of eTh and that are transient. The clarification of these points is our goal here.
Protection against Fas/APO-1- and tumor necrosis factor-mediated cell death by a novel protein, sentrin.
Authors: Okura T , Gong L , Kamitani T , Wada T , Okura I , Wei C F , Chang H M , Yeh E T
Published: December 2022

Fas/APO-1 and TNF receptor 1 share a common signaling motif in their cytoplasmic tail called the "death domain." Using the death domain as bait in the yeast two-hybrid system, several death domain-containing proteins that participate in cell death signaling have been identified. Here we report the isolation of a novel protein, sentrin, which interacts with Fas/APO-1 and TNF receptor 1 but not with FADD/MORT1 or CD40. Two-hybrid interaction assays reveal that sentrin associates only with the signal-competent forms of Fas/APO-1 or TNF receptor 1 death domains. Sentrin is a novel protein of 101 amino acids with homology to ubiquitin, Nedd8, and a Saccharomyces cerevisiae protein, Smt3. When overexpressed, sentrin provides protection against both anti-Fas/APO-1 and TNF-induced cell death.
CD80 (B7.1) and CD54 (intracellular adhesion molecule-1) induce target cell susceptibility to promiscuous cytotoxic T cell lysis.
Authors: Nishio M , Spielman J , Lee R K , Nelson D L , Podack E R
Published: December 2022

Overexpression of B7 and intracellular adhesion molecule-1 (ICAM-1) on syngeneic cells triggered MHC-unrestricted lysis by MHC-restricted CD8+ CTL without TCR/CD3 engagement. Both CD28/B7 and LFA-1/ICAM-1 interactions were required for MHC-nonrestricted cytotoxicity. Cytotoxicity was measured in 4-h and 16- to 18-h cytotoxicity assays. B7-ICAM-1 overexpression triggered perforin- and Fas ligand-mediated lysis, while TNF-alpha was not elicited in MHC-unrestricted cytotoxicity. These results suggest that target cells may elicit MHC-unrestricted lysis from CTL through up-regulation of membrane adherence and costimulatory receptors.
Reciprocal cytotoxic T lymphocyte cross-reactivity interactions between two major epitopes within HIV-1 gp160.
Authors: Shirai M , Kurokohchi K , Pendleton C D , Arichi T , Boyd L F , Takahashi H , Margulies D H , Berzofsky J A
Published: December 2022

We have observed and analyzed an unexpected cross-reactivity of CD8+ CTL between two nonhomologous peptides of the HIV-1 IIIB gp160 envelope protein, P18 (residues 315-329) and HP53 (834-848, also called TH4.1), in the context of four different class I MHC molecules, Dd, Dp, Dq (or Lq), and H-2u. In strains expressing Dd, the cross-reactivity between peptides was bidirectional, whereas in other strains (H-2u, H-2p, and H-2q), the cross-reactivity was unidirectional; that is, P18-specific CTLs showed no killing against targets pulsed with HP53, although HP53 stimulated CTL showed cross-reactive lysis against P18-pulsed target cells. Cross-reactivity was also shown in immunization in vivo and with target cells endogenously expressing viral protein in vitro using two different recombinant vaccinia viruses expressing only the N-terminal portion of gp160, containing P18 but not HP53. Peptide cross-contamination was excluded. Cold target inhibition and single cell cloning experiments indicated that the same CTL was responding to both peptides. Using substituted and truncated peptides, we explored amino acid residues critical for cross-reactive CTL recognition, identified fine specificity similarities among all cross-reactive CTL lines but not non-cross-reactive lines, and mapped cross-reactivity to a 10-residue core of P18 and to an eight-residue core of HP53. A comparison of these peptide sequences and recent data on residues of P18 interacting with H-2Dd provided us with clues to residues involved in the interaction of the CTL with the MHC-peptide complex.
Alleles of highly homologous rat T cell receptor beta-chain variable segments 8.2 and 8.4: strain-specific expression, reactivity to superantigens, and binding of the mAb R78.
Authors: Asmuss A , Hofmann K , Hochgrebe T , Giegerich G , Hünig T , Herrmann T
Published: December 2022

This study addresses the molecular basis of a Tcrb-V polymorphism in the reactivity to the superantigens staphylococcus enterotoxin B (SEB) and the mtv-7 sag (MIs1a) of T cells recognized by the mAb R78, which reacts with the T cell receptor beta-chain variable segment 8.2 (Tcrb-V8.2) of Lewis (LEW) rats. Tcrb-V8.2-like sequences were isolated from liver DNA of the responder strain LEW (I) and the nonresponder strain DA (a) and alleles of the Tcrb-V8.2 and the highly homologous Tcrb-V8.4 were identified. Their expression was analyzed by RNase protection studies and cDNA clones were characterized. A comparison of thymocytes, activated R78+ cells, Con A-stimulated and SEB-stimulated cells allows the following conclusions: the newly identified Lewis allele of Tcrb-V8.4 (Trcb-V8.4I) is nonfunctional due to a frame shift induced by deletion of one nucleotide. The R78 epitope is expressed by Tcrb-V8.2I and Tcrb-V8.4a but not by Tcrb-V8.2a. The implication of this finding for mapping of the R78 epitope and the study of V region usage in experimental autoimmune encephalitis are discussed. Finally, the expression of both Tcrb-V8.2 alleles but not of Tcrb-V8.4a in SEB-stimulated cells defines a polymorphism of the CDR2 and/or CDR4 as the molecular basis of the differential superantigen reactivity.
Identification of a promoter element that regulates tissue-specific expression of the human CD80 (B7.1) gene.
Authors: Fong T C , Wu Y , Kipps T J
Published: December 2022

We isolated the promoter region of the gene encoding human CD80 to examine for elements responsible for the regulated expression of this important costimulatory molecule. Using CAT reporter constructs containing a heterologous general enhancer, we demonstrate that the CD80 promoter is active in CD80-expressing Raji cells, but has no significant activity in Jurkat cells that are CD80 negative. Transcriptional activity in Raji increases as the promoter is truncated from nucleotide position -906 to -84. However, truncation of this promoter to -41 significantly decreases its activity. Within this region is one stretch of DNA that is protected in DNase I footprint analysis and that shows some sequence similarity to the NF-kappaB element. Site-specific mutation of the 5' purine-rich portion of this element (B7-RE, or B7 regulatory element) abrogates expression. Nuclear extracts prepared from Raji, or from leukemic cells induced to express CD80, form a distinct complex(es) with B7-RE in electromobility shift assays. Moreover, a consensus NF-kappaB oligonucleotide can compete with B7-RE for nuclear extract binding. However, no super-shifted bands are observed when extracts are preincubated with Abs to p50, p65, or other Rel proteins. Moreover, we find that recombinant p49 (RelB), p50, p65 (RelA), or p49/p65 heterodimers do not bind B7-RE in vitro. These data indicate that B7-RE may help govern expression of genes independent of a tissue-specific enhancer and that this element is bound by nuclear factor(s) other than those that commonly bind NF-kappaB.
Alloreactive CD8+ T cells can recognize unusual, rare, and unique processed peptide/MHC complexes.
Authors: Malarkannan S , Gonzalez F , Nguyen V , Adair G , Shastri N
Published: December 2022

The identity and abundance of self-peptide/MHC class I complexes that serve as ligands for alloreactive T cells remain largely unknown. Using the Kb-restricted, alloreactive T cells as a probe, the Ag precursor gene, adenosine phosphoribosyl transferase (APRT), was isolated by expression cloning. Its naturally processed product was identified as the SLVELTSL (SEL8) octapeptide. The SEL8 peptide shared five residues with the previously identified SVVEFSSL (JAL8) peptide that stimulated the same T cell, but lacked the critical phenylalanine/tyrosine residue at the primary p5 anchor position. Despite the absence of this key conserved anchor residue, SEL8 was bound tightly by Kb MHC and yet was expressed at less than 10 copies/cell. Mutations in the donor APRT gene in the APC caused a concomitant loss in the ability of APCs to stimulate T cells. The results confirm that the display of peptide/MHC complexes in cells exceeds the predictions based upon consensus motifs, and that CD8+ alloreactive and conventional Ag-specific T cells are indistinguishable in their ability to recognize unique and rare peptide/MHC class I complexes.
Characterization of a light chain product of the human JC lambda 7 gene complex.
Authors: Niewold T A , Murphy C L , Weiss D T , Solomon A
Published: December 2022

The human light chain JC lambda locus is comprised of seven distinct segments, designated JC lambda 1, JC lambda 2, JC lambda 3, JC lambda 4, JC lambda 5, JC lambda 6, and JC lambda 7. Whereas three of these seven represent pseudogenes (psi Clambda 4, psi C lambda 5, and psi C lambda 6), the JC lambda 1, JC lambda 2, and JC lambda 3 complexes are functional, as demonstrated by the finding of their protein products through sequence analyses of lambda-type Bence Jones proteins and light chains derived from monoclonal Igs. Although the JC lambda 7 segment also appears functional, as evidenced through analysis of lymphocyte-derived mRNA, heretofore no monoclonal JC lambda 7-containing lambda-chains have been identified. Serologically, two distinct isotypic markers, Mcg and Oz, are associated, respectively, with JC lambda 1 and JC lambda 3 proteins, in contrast to JC lambda2 components, which do not express these determinants and represent a third isotype. Although another serologic marker, Ke (Kern), considered a fourth isotype, has been assigned to the JC lambda 7 complex, this relationship has been questioned. We now report the primary structural features of a lambda-type Bence Jones protein that include the four distinctive residues encoded by the JC lambda 7 gene segment. This protein, obtained from a patient with multiple myeloma and designated MCP, represents the first example of such a molecule and provides definitive evidence that the JC lambda 7 gene complex is functional. Additionally, comparison of the C lambda sequences of Mcg-/Oz- Bence Jones proteins MCP and KERN supports the contention that the Ke-associated one-residue amino acid variation at position 152 reflects a C lambda A2 polymorphism and that yet another isotypic marker, provisionally designated Mcp, is encoded by the JC lambda 7 gene segment. Thus, we posit that there are four human JC lambda isotypes, Mcg, Ke-Oz-/Ke+Oz-, Ke-Oz+, and Mcp, that represent, respectively, products of the JC lambda 1, JC lambda 2, JC lambda 3, and JC lambda 7 gene complexes.
An endogenous sialoprotein and a bacterial B cell superantigen compete in their VH family-specific binding interactions with human Igs.
Authors: Silverman G J , Pirès R , Bouvet J P
Published: December 2022

Staphylococcal protein A (SpA), a bacterial membrane protein, and protein Fv (Fv binding protein (pFv)), a human sialoprotein involved in gut-associated immunity, have both recently been shown to have unconventional V(H) family-restricted binding interactions with Igs. To determine whether these Ig binding proteins interact with related structures, we performed a series of comparative binding studies. The results confirmed that both molecules are bound by most V(H)3 IgM, but pFv is also recognized by V(H)3 and V(H)6 Ig that do not interact with SpA. We discovered that pFv and SpA (or a single domain of SpA) can compete for binding to a V(H)3 Ab, which suggests that they can recognize the same (or adjacent) V(H) sites. For both SpA and pFv, binding is less frequent among IgG than IgM. However, V(H)3 IgG more commonly possess Fab-mediated binding activity for pFv than for SpA. Binding studies of denatured Ig suggested that both pFv and SpA interact with conformationally dependent V(H) sites, although in certain cases pFv binding is more permissive than SpA binding. Taken together, these results indicate that the superantigen properties of SpA, a microbial protein, and those of pFv, an endogenous sialoprotein, involve binding interactions with overlapping and at times functionally equivalent sites in the V(H) domain, indicating that self and foreign proteins can employ highly conserved strategies to create superantigens for the Ag receptors of B lymphocytes.
Genes for chemokines MuMig and Crg-2 are induced in protozoan and viral infections in response to IFN-gamma with patterns of tissue expression that suggest nonredundant roles in vivo.
Authors: Amichay D , Gazzinelli R T , Karupiah G , Moench T R , Sher A , Farber J M
Published: December 2022

MuMig and Crg-2 are IFN-inducible murine chemokines whose human homologues, HuMig and IP-10, respectively, share activity in vitro as T cell chemoattractants. We analyzed the expression of the genes Mumig, crg-2, and IFN-gamma during experimental infections with Plasmodium yoelii, Toxoplasma gondii, and vaccinia virus. Mumig, crg-2, and IFN-gamma were induced in multiple organs. During the acute phase of each infection as well as after i.p. injection of rIFN-gamma, levels of Mumig mRNA in the liver were as high or higher than levels in any of the other organs. In contrast, the organs showing the highest expression of crg-2 and IFN-gamma varied among the experimental models, with induction of these latter two genes colocalizing. Differences in relative levels of expression of Mumig and crg-2 in liver and spleen were not demonstrably due to expression of the genes in different cell types within these organs. We showed that both Mumig and crg-2 are induced in the liver in hepatocytes and in the spleen in CD11b+ cells. IFN-gamma was necessary for induction of Mumig during infections with T. gondii or vaccinia virus. In contrast, induction of crg-2 was not completely dependent on IFN-gamma. These data demonstrate that despite the overlap in activities within chemokine subsets, chemokine genes show differences in their patterns of expression and in their responses to inducers that suggest nonredundant roles in vivo. Moreover, the pattern of induction of crg-2 is consistent with Crg-2 acting primarily locally, while the pattern for Mumig induction suggests that MuMig may have a systemic role during infection.
Role of monocyte chemoattractant protein-1 (MCP-1) in Th1 (mycobacterial) and Th2 (schistosomal) antigen-induced granuloma formation: relationship to local inflammation, Th cell expression, and IL-12 production.
Authors: Chensue S W , Warmington K S , Ruth J H , Sanghi P S , Lincoln P , Kunkel S L
Published: December 2022

The present study examined the contribution of monocyte chemotactic protein-1 (MCP-1) to granulomatous inflammation mediated by Th1- and Th2-related cytokines. Types 1 and 2 lung granulomas (GR) were respectively induced in presensitized CBA mice by embolization of beads coupled to purified protein derivative of Mycobacteria tuberculosis or soluble Ags derived from Schistosoma mansoni eggs. MCP-1 was spontaneously produced by intact GR, isolated GR macrophages, and draining lymph node cultures, but levels were greater in the type 2 than in the type 1 response. In vivo depletion of IFN-gamma augmented type 2 inflammation and local MCP production; IL-4 depletion had the opposite effect. These treatments had no significant effect on the type 1 response. Treatment with anti-MCP-1, but not that with anti-MIP-1alpha, Abs caused a 30% decrease in type 2 GR area. Neither treatment affected the type 1 GR. Intrinsic MCP-1 was detected immunohistochemically within lymph nodes and appeared to support IL-4-/IL-5-producing lymph node cells. In addition, MCP-1 inhibited IL-12 production by inflammatory macrophages. The latter was demonstrated as a potentially direct effect of MCP-1 on macrophages. These findings show that MCP-1 contributes more to type 2 than to type 1 cytokine-mediated inflammation and suggest a broader role for chemokines in regulating Th cell expression.
Stimulating properties of 5-oxo-eicosanoids for human monocytes: synergism with monocyte chemotactic protein-1 and -3.
Authors: Sozzani S , Zhou D , Locati M , Bernasconi S , Luini W , Mantovani A , O'Flaherty J T
Published: December 2022

The newly described products of 5-hydroxyeicosanoid dehydrogenase, 5-oxo-6,8,11,14-eicosatetraenoic acid (ETE) and 5-oxo-15(OH)ETE, induced directional migration and actin polymerization of human monocytes in vitro. At peak concentrations, the two eicosanoids had a chemotactic activity of about 40% of that observed in the presence of an optimal concentration of FMLP and twice the activity elicited by the related eicosanoid 5-hydroxy-6,8,11,14-eicosatetraenoic acid (5-HETE). 15-Oxo-ETE showed a very low but detectable chemotactic activity. All of these chemotactic responses were blocked by Bordetella pertussis toxin, but were resistant to LY255283, a leukotriene B4 (LTB4) receptor antagonist. 5-Oxo-ETEs and 5-HETE induced homologous desensitization of chemotactic response, but did not cross-desensitize to other chemotactic agonists (e.g., monocyte chemotactic protein (MCP)-1 and LTB4). 5-Oxo-ETEs increased in a synergistic fashion the monocyte migration to MCP-1 and MCP-3. In the same range of concentrations, 5-oxo-ETE increased MCP-1-induced release of arachidonic acid from labeled monocytes. No synergistic interaction was observed when FMLP was used as chemoattractant. Thus, this study identifies monocytes as cells responsive to 5-oxo-ETEs and shows that monocyte activation by 5-oxo-ETEs occurs through an LTB4 receptor-independent mechanism that associates with pertussis toxin-sensitive G proteins. The synergistic interaction between 5-oxo-ETEs and C-C chemokines, two families of mediators both synthesized by phagocytic cells, may be relevant in vivo for the regulation of monocyte accumulation at sites of allergic and inflammatory reactions.
Regional immunosuppression in esophageal squamous cancer: evidence from functional studies with matched lymph nodes.
Authors: O'Sullivan G C , Corbett A R , Shanahan F , Collins J K
Published: December 2022

Although production of immunosuppressive factor(s) by esophageal squamous cancer has been demonstrated, systemic immunosuppression occurs late. Whether local immunosuppression by tumor-derived factors occurs in vivo as a potential mechanism of escape from immune surveillance is unknown. We found that lymphocytes from nodes draining distal esophageal squamous tumors in 23 consecutive patients had depressed proliferative and cytotoxic responsiveness relative to both lymphocytes from a reference node outside the field of drainage and matched PBL from the same patient. In a subset of patients in which more than one tumor-draining node was examined, a radial or zonal immunosuppression relative to the primary tumor was evident. The findings were unrelated to surgery or anatomic location because all but 2 of 26 control patients with esophagogastric adenocarcinoma had normal or enhanced lymphocyte responsiveness in the tumor-draining node. The absence of overt or even micrometastatic nodal disease, as determined by immunostaining for cytokeratin expression, coupled with the long-term survival of several of the patients, strongly suggests that the immunosuppressive effect is due to mechanisms other than metastases, and may be a premetastatic occurrence. We conclude that regional immunesuppression does exist in patients with esophageal squamous cancer when systemic immunity is still well preserved. The local immune suppression inhibits the generation of lymphokine-activated killer (LAK) cells and may be an impediment to potential immunotherapeutic strategies.
Deficient expression of p56(lck) in Th2 cells leads to partial TCR signaling and a dysregulation in lymphokine mRNA levels.
Authors: al-Ramadi B K , Nakamura T , Leitenberg D , Bothwell A L
Published: December 2022

Activation of T lymphocytes through their TCR is regulated by a delicate balance of phosphorylation and dephosphorylation of protein substrates by protein tyrosine kinases (PTKs) and phosphotyrosyl phosphatases, respectively. One of the earliest steps in the activation pathway is thought to involve the Src family PTKs, p56(lck) (Lck) and p59(fyn) (Fyn); however, the precise contribution of each PTK in TCR-mediated signaling remains incompletely understood. To study the role of Lck in mature T cells, antisense RNA was used to inhibit its expression in a nontransformed Th2 clone. In this report, we demonstrate that specific inhibition of Lck expression in Th2 cells, in the presence of normal levels of functional Fyn PTK, has profound consequences on multiple events following TCR stimulation, including an altered pattern of tyrosine-phosphorylated substrates, defective phosphorylation of TCR-zeta and ZAP-70, defective Ca2+ mobilization, and a approximately 90% reduction in proliferative responses to antigenic and mitogenic stimuli. In contrast, Lck-deficient cells expressed constitutively elevated levels of lymphokine mRNA, including IL-4, IL-5, and IL-10, and were capable of secreting IL-4 upon activation through the TCR. These results demonstrate a dissociation in functional responses in Lck-deficient Th2 cells and suggest a role for Lck in the induction of a state of T cell unresponsiveness.
Tissue immunogenicity: the role of MHC antigen and the lymphocyte costimulator B7-1.
Authors: Coulombe M , Yang H , Guerder S , Flavell R A , Lafferty K J , Gill R G
Published: December 2022

Pancreatic islet transplantation represents a potential treatment for insulin-dependent diabetes mellitus. One approach to circumvent the requirement for recipient immune suppression is to reduce or eliminate the immunogenicity of the donor graft prior to transplantation. In this study, we have examined the relative contributions of graft MHC Ag expression and donor-derived costimulatory (CoS) activity to the rejection of islet allografts. Depletion of donor hemopoietic APCs from islet tissue facilitated long-term allograft survival even when donor class I MHC Ag expression was greatly increased by IFN-gamma treatment prior to grafting. Conversely, islet allografts from transgenic mice expressing the CoS molecule B7-1 (CD80) on islet beta cells were acutely rejected even when hemopoietic APCs were eliminated. Thus, B7-1 is sufficient to confer the capacity of islet parenchymal cells to stimulate allorejection. Taken together, these results point towards donor-derived CoS activity as a primary target of intervention therapy to modulate tissue immunogenicity.
IL-2 and TNF receptors as targets of regulatory T-T interactions: isolation and characterization of cytokine receptor-reactive T cell lines in the Lewis rat.
Authors: Mor F , Reizis B , Cohen I R , Steinman L
Published: December 2022

T cells are considered to be of prime importance in immune regulation of both B and T cell functions. The targets of recognition in T-T cell interactions are not clear. Most recent experimental work has focused on the idiotypic regulatory interactions mediated by TCR peptides. There is experimental evidence that regulatory cells exist that do not recognize the TCR. This type of regulation is selectively induced by activated T cells. Therefore, we designed this study to examine the possible role of cytokine receptors as targets of immune regulation. We tested two peptides of IL-2R alpha-chain, 2 of IL-2R beta-chain, and one of TNFR (p60). All peptides were found to be immunogenic at inducing T cell proliferation and four induced Abs in Lewis rats. We generated T cell lines to these five peptides, and tested them both in vitro and in vivo. We found that the T cells exhibited a proliferative response when cultured with activated, irradiated stimulator cells that were augmented upon addition of the cytokine receptor peptide. The cytokine profile of the lines was characterized as well as the Vbeta gene composition. One of the lines significantly protected against active encephalomyelitis. These results point at cytokine receptors as possible targets of immune regulation and T-T cell interactions.
Antigen-dependent regulation of IgE antibody production by human antigen-specific B cells.
Authors: Oshiba A , Gelfand E W
Published: December 2022

Ag-specific B cells from the peripheral blood of immunized donors were separated following rosette formation with tetanus toxin (TT)-coupled immunobeads to study the regulation of T cell-dependent Ab production. Surface IgM+ (sIgM+) B cells rosetting with TT-coated beads produced anti-TT IgM, IgG, and IgE Abs in cultures stimulated with EBV or anti-CD40 Ab in the presence of IL-4; this was associated with increased numbers of B cells positive for sIgG and sIgE. TT-binding sIgM+ B cells also produced significant amounts of anti-TT IgM, IgG, and IgE Abs when they were cultured with autologous T cells in the presence of IL-4. Addition of TT to the IL-4-stimulated, T cell-dependent cultures resulted in higher levels of TT-specific IgM and IgG Abs than observed with T cells alone, but the inclusion of TT with T cells suppressed anti-TT IgE production. Addition of TT Ag to cultures with allogeneic T cells failed to enhance IgM and IgG responses, and the inhibition of IgE production was reduced compared with cultures containing autologous T cells in the presence of Ag. These results demonstrate that the presence of Ag contributes to the regulation of Ab production, resulting in enhanced IgM and IgG responses, but down-regulation of IgE Ab production.
Disregulated expression of CD43 (leukosialin, sialophorin) in the B cell lineage leads to immunodeficiency.
Authors: Ostberg J R , Dragone L L , Driskell T , Moynihan J A , Phipps R , Barth R K , Frelinger J G
Published: December 2022

Leukosialin (CD43 or sialophorin) is a cell surface sialoglycoprotein implicated in cell adhesion and proliferation whose tightly regulated expression in B lymphocytes is likely important for their normal development and/or function. To examine the physiologic role of mouse CD43 (mCD43) in vivo, we exploited transgenic (TG) mice whose developmental expression of mCD43 was extended during B cell differentiation so that mCD43 was now expressed on peripheral B cells. Despite having increased B cells, localization of lymphocytes in the TG spleens appeared normal by immunocytochemistry with anti-CD4, anti-CD8, and anti-B220 mAbs. However, the numbers of splenic germinal centers and the resting sera Ig levels were decreased in the TG mice compared with littermate controls. TG mice had decreased humoral responses to the T-dependent Ags keyhole limpet hemocyanin and OVA, as well as reduced Ag-specific B cell numbers. In contrast, in vitro LPS stimulation of purified TG or control B cells resulted in similar proliferation and IgM responses. Thus, the alteration of B cell mCD43 expression that resulted in profound immunodeficiency in vivo was not due to absolute defects in B cell development or Ab production. However, TG B cells had a decreased ability to homotypically aggregate and to present Ag to the T cell hybridoma B3Z. These data suggest that the immunodeficiency seen in vivo is due to the anti-adhesive forces of mCD43 preventing normal T-B cell interaction. This likely reflects a general property of mucins in regulating cell interactions.
Molecular cloning of guinea pig membrane cofactor protein: preferential expression in testis.
Authors: Hosokawa M , Nonaka M , Okada N , Nonaka M , Okada H
Published: December 2022

Human membrane cofactor protein (MCP) is a widely distributed cell-associated complement-regulatory protein, and recent findings suggest that MCP may be involved in sperm-egg interaction. We have isolated four cDNA clones and one reverse transcriptase-PCR product homologous to human MCP from guinea pig testis. These clones defined five isoform classes generated from a single copy gene by alternative splicing. Reverse transcriptase-PCR revealed that two classes for the clones termed GMP1 and GM2 were predominant. GMP1 consisted of four short consensus repeats (SCRs), regions corresponding to the human serine/threonine/proline-rich C (STP(C)) domain and a human region of unknown significance, a hydrophobic region presumed to be a transmembrane domain, and a cytoplasmic region. Identity with human MCP in the SCR region was 56% at the amino acid level and 71% at the nucleotide level. GM2 had the same structure as GMP1, except that it lacked the fourth SCR, which is presumed to be essential for C3b binding of human MCP. Northern blotting analysis of various tissues revealed a significant level of MCP transcripts in testis. Guinea pig MCP is likely to have only one STP domain that is homologous to human STP(C) and is similar in this respect to human spermatozoa MCP. Gene analysis revealed a single base deletion and a lack of consensus sequences for splicing in the guinea pig regions corresponding to human STP(A) and STP(B), respectively. These results suggest that guinea pig MCP plays a more restricted role in reproduction than does human MCP.
Molecular cloning of a novel protein antigen of Leishmania major that elicits a potent immune response in experimental murine leishmaniasis.
Authors: Webb J R , Kaufmann D , Campos-Neto A , Reed S G
Published: December 2022

BALB/c mice are highly susceptible to infection with the protozoan parasite Leishmania major. This susceptibility has been attributed, in part, to the expansion of parasite-specific CD4+ Th2 cells that antagonize Th1 responses and promote humoral immunity. In the present study, we have utilized sera from L. major-infected BALB/c mice to screen an L. major amastigote cDNA expression library. One of the clones detected encodes a novel Ag designated as L. major stress-inducible 1 (LmSTI1). LmSTI1 contains six copies of the tetratricopeptide consensus motif and is highly related to a family of stress-inducible proteins that is conserved from yeast to humans. Sera from L. major-infected BALB/c mice have LmSTI1-specific Ab titers in excess of 1:200,000, comprised predominantly of IgG1, IgG2A, and IgG2B isotypes. Recombinant LmSTI1 protein elicited strong proliferative responses from draining lymph node cells of L. major-infected BALB/c mice at both early (10 days) and late (28 days) stages of infection and elicited production of high levels of IFN-gamma and low levels of IL-4. In contrast, soluble leishmanial lysate elicited high levels of IL-4 and low IFN-gamma production. Thus, we have identified an Ag of Leishmania capable of eliciting a mixed cellular response that is skewed toward a Th1 phenotype in susceptible BALB/c mice with advanced infections. In addition, analyses of sera from human patients with cutaneous, visceral, and post-kala azar visceral leishmaniasis indicated that a majority of individuals from all three clinical groups mounted strong humoral responses against LmSTI1.
Functions of domain 1 and 4 of vascular cell adhesion molecule-1 in alpha4 integrin-dependent adhesion under static and flow conditions are differentially regulated.
Authors: Abe Y , Ballantyne C M , Smith C W
Published: December 2022

To assess the potential contributions of endothelial vascular cell adhesion molecule-1 (VCAM-1) to leukocyte adhesion under conditions of flow, we evaluated the effects of cytokine stimulation of HUVEC on primary capture, rolling, and stable adhesion of mono- and lymphoblastoid cells in a parallel-plate flow chamber. Stimulation of HUVEC with IL-1beta or IL-4 for 24 h induced surface expression of VCAM-1, consistent with published results. Using blocking mAbs to inhibit specific adhesion molecules, we found that a significant level of primary capture at wall shear stresses between 0.5 and 2.0 dynes/cm2 could be attributed to alpha4 integrin and VCAM-1. This primary adhesion under flow was most often characterized by abrupt stationary adhesion with little evidence of rolling. Studies with IL-1beta-stimulated HUVEC revealed that domain 1 of VCAM-1 was solely responsible for alpha4 integrin-dependent primary capture under flow, but both domains 1 and 4 were utilized in alpha4 integrin-dependent adhesion under static conditions. In contrast to results obtained with these cytokines alone, stimulation of HUVEC with a combination of IL-1beta and IL-4 markedly reduced alpha4/VCAM-1-dependent adhesion under flow, even though the surface levels of VCAM-1 were greater than with either cytokine alone, and alpha4/VCAM-1-dependent adhesion under static conditions was intact. These results demonstrate that distinct functional differences exist between the interactions of VCAM-1 and alpha4 integrin under static and flow conditions, and they reveal the potential for endothelial modulation of adhesion under flow without altering VCAM-1-dependent adhesion under static conditions.