bluuebunny/crossref_metadata_2025 huggingface dataset:
Selective transformation of host lymphocytes in vivo by retrovirus-producing macrophages.
Authors: Bosco M C , Musso T , Gusella G L , Giovarelli M , Forni M , Soleti A , Masuelli L , Modesti A , Forni G , Varesio L
Published: December 2022

Macrophage-mediated retroviral transformation of host cells was studied in vivo utilizing the cloned murine macrophage-line GG2EE, generated by in vitro infection of bone marrow cells from C3H/HeJ mice (H-2k) with the acute transforming retrovirus J2 bearing the v-myc and v-raf oncogenes. Because GG2EE macrophages produce the J2 retrovirus, the development of secondary, J2 virus-induced tumors after the injection of the cell line into several strains of mice was evaluated. GG2EE cells proliferated and gave rise to histiocytic tumors in syngeneic mice and in allogeneic athymic Swiss mice. The inoculum of GG2EE cells in allogeneic DBA/2 mice (H-2d) and, to a lesser extent, in BALB/c (H-2d) and BALB/k (H-2k) mice gave rise to a small, solid mass at the injection site. Although the initial tumor was slowly rejected, secondary lymphomas belonging to the B or T cell lineage developed, leading to mouse death. Extensive phenotypic, functional, and chromosomal analyses proved that lymphomas were derived from host T and B cell transformation. Southern and Northern blot studies showed that J2 virus was integrated and expressed in lymphoma cells, demonstrating that the virus was transmitted to the host lymphocytes and suggesting that it was causal in lymphoma development. The existence of close and protracted interactions between GG2EE macrophages and allogeneic host lymphocytes and the presence of viral particles in the area of macrophage-lymphocyte contact were demonstrated by histologic and ultrastructural analysis. Rejection of J2 virus-infected lymphocytes in allogeneic mice suggested that host lymphocyte transformation was dependent upon the macrophage cell type. These results demonstrate that macrophage-derived J2 retrovirus transforms host lymphocytes in vivo in allogeneic mice and that a condition of host alloreactivity is critical for such event.
Differential expression of IL-4 receptors in human T and B lymphocytes.
Authors: Mozo L , Rivas D , Zamorano J , Gutierrez C
Published: December 2022

IL-4R have been described in unstimulated human T and B lymphocytes. However, a precise comparative study on the expression and regulation of IL-4R in isolated human T and B cell populations has not yet been fully assessed. We examined the mRNA levels and the cell membrane expression of IL-4R in freshly isolated T and B lymphocytes as well as in in vivo- and in vitro-stimulated cells. IL-4R protein expression and transcript levels were higher in tonsillar unstimulated B cells than in T cells. Splenic and peripheral blood B lymphocytes also expressed higher surface IL-4R in their membranes than T cells did. Large B lymphocytes from tonsils (in vivo-activated cells) obtained by Percoll gradient centrifugation displayed higher IL-4R levels than resting cells. On activation in vitro of T lymphocytes with IL-2 or PHA, slight increments on the IL-4R mRNA and protein levels were achieved. However, maximal levels of IL-4R expression were obtained on T cell incubation with IL-4 at a concentration of 100 U/ml. Similarly, the same concentration of this lymphokine up-regulated the surface IL-4R molecules and the IL-4R mRNA levels in purified B lymphocytes. Cross-linkage of surface Ig by insolubilized anti-IgM potentiated the effect of IL-4 in up-regulating IL-4R expression in B cells, probably by inducing outgrowth of IL-4R positive subpopulations. The B cell mitogen, Staphylococcus Aureus Cowan I, although inducing cell proliferation, was ineffective in promoting new receptor synthesis. Cell proliferation was not required for IL-4-dependent IL-4R up-regulation on both T and B lymphocytes.
Selective up-regulation of the 75-kDa tumor necrosis factor (TNF) receptor and its mRNA by TNF and IL-1.
Authors: Winzen R , Wallach D , Kemper O , Resch K , Holtmann H
Published: December 2022

In cells of the fibroblastoid line SV-80, rapid down-modulation of TNF binding in response to TNF itself, or to IL-1, was followed by a gradual recovery of binding, which occurred even in the continuous presence of the cytokines. Untreated cells carried mainly the 55-kDa receptor species. In cells treated with TNF or IL-1, the 55-kDa TNF-R, although increasing after initial down-modulation, remained lower than before treatment. Expression of the 75-kDa TNF-R species upon treatment with either cytokine was markedly increased. Both TNF and IL-1 also induced a strong increase of the mRNA for the 75-kDa receptors, whereas the amount of mRNA for the 55-kDa receptors decreased. The effects of TNF on cell surface expression of the TNF-R could not be blocked with antibodies to IL-1, nor could the effects of IL-1 be blocked with antibodies to TNF, indicating that each cytokine affects the cell surface expression of the receptors independently of the other. Applying both cytokines together resulted in much stronger increase in expression of the 75-kDa TNF-R than applying each alone. Similar changes in cell surface expression and mRNA levels of the two TNF-R as observed in SV-80 cells were also found in TNF and IL-1-treated human foreskin fibroblasts. It is suggested that these sustained changes in the pattern of receptor expression contribute to the adjustment of the cellular response to TNF when formation of TNF and IL-1 takes place over a prolonged period.
Rat class III Fc gamma receptor isoforms differ in IgG subclass-binding specificity and fail to associate productively with rat CD3 zeta.
Authors: Farber D L , Giorda R , Nettleton M Y , Trucco M , Kochan J P , Sears D W
Published: December 2022

Several new rat class III Fc gamma R isoforms are described here, extending the genetic complexity of this receptor family and further distinguishing rat CD16 from mouse CD16, represented by only one receptor isoform, and human CD16, represented by only two isoforms. RNase protection assays reveal that three rat tumor cell lines--RBL-1 basophilic leukemia cells, RM-SV1 macrophages, and CRNK-16 NK cells--all coordinately express multiple and probably identical rtFc gamma RIII-related transcripts in similar relative proportions but at significantly different levels. These results indicate that no single isoform predominates in these cell types but that the overall level of rtFc gamma RIII-related transcripts is differentially regulated. Two of the rtFc gamma RIII isoforms found to have extensive amino acid sequence differences in their second extracellular (EC2) domains are shown to bind rat and mouse IgG subclasses differently. This result suggests that the receptor isoform diversity in this species may function as a mechanism for extending the IgG-binding capacity of rat leukocytes. Cloned cDNA for the rat CD3 zeta protein was also isolated in this study and its ability to augment surface expression of class III Fc gamma R was tested by rosetting of cDNA-transfected COS cells. Like the structurally homologous mouse CD3 zeta, rat CD3 zeta fails to promote surface expression of Fc gamma RIII, sharply contrasting the efficient receptor expression produced by human CD3 zeta. Variations in the transmembrane amino acid sequences correlate with the divergent capacities of these CD3 zeta molecules to augment receptor expression. The high levels of CD3 zeta message expressed in rat NK cells may indicate that other unidentified hetero-subunits are required for assembly of rat CD3 zeta into functional CD16 receptors.
Functional importance of the cyclic AMP response element-like decamer motif in the CD8 alpha promoter.
Authors: Gao M H , Kavathas P B
Published: December 2022

Expression of the CD8 gene is highly regulated during lymphocyte differentiation and in a tissue-specific manner. We characterized the human CD8 alpha promoter region to determine whether tissue specificity resides within the promoter and to define important regulatory elements. A set of six fragments 5' of the CD8 alpha gene were linked to a luciferase reporter gene. The luciferase activity was then measured in a transient transfection assay. We found that CD8 alpha promoter activity can be detected from a 146-bp fragment upstream of the translation start site, but not from a 133-bp fragment. The cyclic AMP response element (CRE)-like site within the 10 bp from -143 to -133 is critical for promoter activity. Mutation of the CRE/decamer in the context of a 429-bp fragment causes loss of activity. Tissue specificity does not reside in the 146-bp fragment because this fragment directs expression in both T and non-T cell lines. Fragments longer than 146 bp are generally expressed less well in the cell lines suggesting the potential existence of negative regulatory elements upstream of -146. Using a CRE/decamer-containing oligomer as a probe in an electrophoresis mobility shift assay, three retarded bands formed by proteins binding to the DNA were detected using nuclear extracts from two T cell lines. Two of the three bands contain proteins of the CRE-binding protein (CREB)/activating transcription factor (ATF) family. Because the CRE-binding protein/activating transcription factor proteins play a role in the expression of many other T cell-specific genes, our work strengthens the hypothesis that the CRE motif is important for regulating the expression of T cell-specific genes.
Alteration of Leishmania donovani infection levels by selective impairment of macrophage signal transduction.
Authors: Moore K J , Labrecque S , Matlashewski G
Published: December 2022

Leishmania donovani is an obligate intracellular protozoan which residues and multiples in macrophages. The molecular basis for this host-parasite interaction is poorly understood. Targeting a signal transduction pathway in the macrophage would allow this parasite to manipulate cellular gene expression, and this may aid in ensuring its survival. We demonstrate that in macrophages infected with L. donovani for 18 h, c-fos gene expression mediated through protein kinase A was unaffected under conditions where there was an impairment of protein kinase C (PKC)-mediated c-fos gene expression. This selective impairment of PKC-mediated c-fos gene expression was substantially augmented in macrophages put in contact with L. donovani promastigotes or amastigotes for only 1 h. Treatment of macrophages with L. donovani-conditioned media was not sufficient to significantly impair signal transduction. These data revealed that L. donovani selectively impaired the transmission of information from the cell surface to the nucleus and that this effect is induced very soon after macrophage-parasite contact. The biologic significance of this altered signal transduction in the macrophage with respect to infection with L. donovani was then examined by treating macrophages with various protein kinase inhibitors prior to infection with amastigotes. Macrophages that were treated with PKC inhibitors demonstrated an increase in the initial uptake of the parasite and carried heavier infection levels than did controls. In contrast, treatment of macrophages with an inhibitor of calmodulin-dependent protein kinase (CaM-PK) did not show significant differences in the initial uptake of parasite, but prolonged impairment of CaM-PK resulted in a decrease in the level of macrophage infection. Further experiments revealed that promastigote proliferation was severely impaired by the CaM-PK inhibitor but not any of the other inhibitors.
Mechanistic differences between migration inhibitory factor (MIF) and IFN-gamma for macrophage activation. MIF and IFN-gamma synergize with lipid A to mediate migration inhibition but only IFN-gamma induces production of TNF-alpha and nitric oxide.
Authors: Herriott M J , Jiang H , Stewart C A , Fast D J , Leu R W
Published: December 2022

Previously we found that murine macrophage migration inhibition (MMI) was mediated by IFN-gamma-priming and lipid A triggering. With the recent availability of human recombinant migration inhibitory factor (MIF), which is distinctly different from IFN-gamma and other cytokines, we have now attempted to explore possible mechanistic differences between IFN-gamma and MIF to mediate MMI. Neither MIF not IFN-gamma were active alone, but effectively primed murine inflammatory macrophages for subsequent triggering by lipid A to mediate MMI. A specific neutralizing antibody for rMIF abrogated MMI mediated only by MIF and not by IFN-gamma-primed macrophages. Distinct differences were also found between the mechanisms by which MIF and IFN-gamma synergized with lipid A for activation in that IFN-gamma-primed and lipid A triggered macrophages produced TNF and nitric oxide (NO), whereas MIF-primed cells did not. Macrophages primed with IFN-gamma and triggered by rTNF were inhibited in their migration, whereas MIF failed to synergize with rTNF for MMI. An inhibitor of NO production NG-monomethyl-L-arginine inhibited MMI mediated by higher activating concentrations of lipid A and by IFN-gamma-primed and lipid A triggered macrophages, but had no effect on MIF-primed cells in concert with lipid A for increased expression of both TNF-alpha mRNA and NO synthase mRNA. Taken together, our results indicate that both MIF and IFN-gamma prime macrophages to synergize with lipid A to mediate MMI but by different mechanisms. The activation process by IFN-gamma to mediate migration inhibition appears to resemble requirements for rendering macrophages tumor cytotoxic in the production of TNF for autocrine-mediated NO generation by primed macrophages. In contrast, MIF-mediated MMI was independent of requirements for either TNF or NO production.
Modulation of lipopolysaccharide-induced cytokine gene expression in mouse bone marrow-derived macrophages by muramyl dipeptide.
Authors: Le Contel C , Temime N , Charron D J , Parant M A
Published: December 2022

Previous studies have shown that an i.v. injection of muramyl dipeptide (MDP) before a LPS challenge strongly potentiated serum TNF and IL-6 release in mice. Therefore the direct action of MDP was examined on TNF-producing cells, namely in macrophages stimulated or not by LPS. The level of TNF-alpha, IL-1 alpha, and IL-6 mRNA was determined in bone marrow-derived macrophages (BMM). A marked TNF-alpha mRNA accumulation was found between 1 and 6 h after stimulation with MDP or LPS. LPS-induced IL-1 alpha mRNA transcript was delayed (3 h) than those after MDP induction (1 h). Conversely, kinetic induction of the IL-6 mRNA transcript was delayed in MDP-treated BMM as compared with LPS-stimulated cells. MDP pretreatment of BMM for 3 h not only enhanced the total level of LPS-induced TNF-alpha, IL-1 alpha, and IL-6 mRNA (respectively 2.9-, 1.6-, and 2.4-fold increase), but it also delayed the kinetics of IL-1 alpha and IL-6 species accumulation. The enhancement induced by MDP pretreatment at the level of cytokine mRNA accumulation was correlated with an increase in LPS-induced TNF and IL-6 biologic activity production in supernatant fluids. In addition, in BMM from C3H/Hej mice MDP pretreatment enhanced the weak effect of LPS on TNF mRNA transcript accumulation and was required to produce LPS-induced TNF bioactivity. Our results suggest that MDP and LPS could act through distinct pathway(s) to induce cytokine gene expression. Moreover, the priming effect displayed by MDP could result in modulation of the LPS-induced cytokine gene expression at the transcriptional and/or post-transcriptional level.
Calcium mobilization-associated and independent cytosolic acidification elicited in tandem with Na+/H+ exchanger activation in target cell-adherent human NK cells.
Authors: Edwards B S , Hoffman R R , Curry M S
Published: December 2022

Flow cytometry was used to identify mechanisms by which human NK cells regulate intracellular pH (pHi) and to investigate the relationship between NK cell pHi and cytolytic function. Temporally resolved determinations of pHi were simultaneously made in NK cells that formed conjugates with target cells (NKC) and unconjugated NK cells (NKU) on the basis of the red/orange fluorescence emission ratio of the pH indicator seminapthylrhodafluor-1. Two pHi regulatory mechanisms were identified in NK cells: 1) a HCO3-/Cl- antiport that promoted pHi changes in response to variation of extracellular HCO3- and Cl- but not Na+ ion concentrations, was sensitive to stilbene derivatives 4,4'-diisothiocyanatostilbene sulfonic acid and 4-acetamido-4'-isothyocyanotostilbene-2-2'-disulfonic acid, and exhibited similar activity in NKC and NKU, and 2) a Na+/H+ exchanger that promoted pHi changes in response to extracellular Na+ ion concentration changes and was sensitive to dimethylamiloride (DMA), permeable to Na+ and Li+ but not K+ or N-methyl-D-glucamine, and quiescent in NKU but activated in target cell-adherent NKC. When Na+/H+ exchange was blocked with 10 microM DMA, the pHi of NKC bound to NK-sensitive K562 target cells progressively decreased for 3 to 4 min, then stabilized at 0.1 to 0.15 pH units below the pHi of NKU. A significant temporal decline in NKC pHi also occurred in the nominal absence of extracellular Ca2+ (0.07 +/- 0.02 pH units) and when NKC formed conjugates with NK-resistant B lymphoblastoid target cells that failed to mobilize NK cell Ca2+ (0.07 +/- 0.01 pH units) (mean +/- SD). However, the magnitude of the NKC cytosolic acidification response was consistently reduced (42 +/- 4 and 44 +/- 6%, respectively) under these Ca2+ flux response-limiting conditions. Thus, adhesion to target cells triggered two pHi-related responses in NK cells: 1) a decline in pHi which exhibited both Ca2+ mobilization-dependent and -independent components, and 2) Na+/H+ exchanger activation by which acid production was neutralized. Manipulations of HCO3- and DMA that clamped NKC pHi at values ranging from 6.8 to 7.2 failed to significantly influence NK cell cytolytic function. By contrast, isosmotic replacement of extracellular Na+ with NMDG resulted in a 57 +/- 22% inhibition of NK cell-mediated cytolysis (n = 8, p < 0.01). NKC pHi declined to 6.8 to 6.9 under these conditions, but this was not the apparent basis of impaired cytolysis because sustained elevation of NKC pHi by addition of dimethylamine failed to significantly reverse inhibition.(ABSTRACT TRUNCATED AT 400 WORDS)
Large cytoplasmic inclusion body kappa-chain has unusual intrachain disulfide bonding.
Authors: Jäck H M , Sloan B , Grisham G , Reason D , Wabl M
Published: December 2022

The Ig kappa L chain synthesized by the mouse hybridoma line F10 forms large fibrils in the lumen of the endoplasmic reticulum. Despite the formation of these inclusion bodies, free kappa-chain is secreted. Both intracellular and secreted kappa-chains in this line migrate faster on SDS gels; thus, the F10 kappa-chain seems to be more compact than normal Ig L chain. In normal kappa-chain, four cysteine residues form intrachain disulfide bonds and the fifth connects the L chain to the H chain. Although the five cysteine residues of the aberrant kappa-chain are in the normal positions, they display an unusual gel pattern when the intrachain disulfide bonds are opened with 2-ME; that is, the intrachain disulfide bonding pattern of F10 kappa-chain seems to be unusual. It is suggested that the abnormal folding pattern favors fibril formation.
Nitric oxide-mediated apoptosis in murine peritoneal macrophages.
Authors: Albina J E , Cui S , Mateo R B , Reichner J S
Published: December 2022

Nitric oxide (NO) synthase, the enzyme responsible for the generation of the cytotoxic compound NO from L-arginine, is induced in macrophages during activation. Previous work demonstrated that the cytotoxicity of NO extends to the macrophages that produce it, because the activity of NO synthase in these cells correlates inversely with their life span in culture. Data presented here demonstrate that the NO-dependent death of murine peritoneal macrophages activated in vitro with IFN-gamma and LPS is mediated through apoptosis. Evidence in this direction was provided by microscopic examination of the cells, which revealed the presence of nuclear and cytoplasmic alterations characteristic of apoptosis, and by the specific pattern of internucleosomal DNA fragmentation detected by electrophoresis. That these alterations resulted from the production of NO was confirmed by the preventive effects of cell activation in L-arginine-restricted medium or in medium containing an inhibitor of NO synthase, NG-monomethy L-arginine, and more directly by the induction of apoptosis by exposure of the cells to authentic NO gas. Additional results demonstrated that glucose starvation, the inhibition of the tricarboxylic acid cycle with fluorocitrate or of glycolysis with iodoacetate, but not the suppression of the electron transport chain with potassium cyanide, also induced macrophage apoptosis. The potential role of metabolic inhibition as a mechanism for NO-mediated apoptosis, as well as the relationship of these findings with events occurring in wounds and other sites of macrophage infiltration are discussed.
Short time exposure to lipopolysaccharide is sufficient to activate human monocytes.
Authors: Gallay P , Jongeneel C V , Barras C , Burnier M , Baumgartner J D , Glauser M P , Heumann D
Published: December 2022

Very little is known about early events in LPS binding and about the duration of LPS exposure required to activate monocytes. In the present study, we have investigated the kinetics of LPS binding to human monocytes and the time of exposure required to trigger the synthesis of TNF-alpha. We directly followed the binding of FITC-labeled LPS to monocytes by flow cytometry or confocal laser microscopy. LPS was able to bind to the cell surface within 1 min of exposure, and was internalized within 5 min. Equilibrium was reached within 15 min at all but the lowest LPS concentration tested (10 ng/ml). Binding was dependent on the presence of LPS-binding protein, supplied either as a plasma component or in purified form, and inhibited by an anti-CD14 mAb (MY4). Polymyxin B, an inhibitor of LPS-mediated activation, essentially abrogated the LPS-binding protein- and CD14-dependent binding of LPS to monocytes. Using either the anti-CD14 mAb or polymyxin B to block further LPS binding, we found that 5 to 10 min of exposure was sufficient to trigger maximal TNF-alpha release. Similarly, monocytes washed after 5 to 15 min exposure to eliminate LPS also produced high levels of TNF-alpha transcripts without further presence of LPS in the medium. We conclude that a few minutes of exposure to physiologically relevant concentrations of LPS are sufficient to trigger both maximal binding and activation of monocytes.
Cytokine mediation of the suppressive effect of ferritin on colony-stimulating factor-1-dependent monocytopoiesis.
Authors: Kreisberg R , Detrick M S , Osmand A P , Moore R N
Published: December 2022

Peptide-specific IgG from a rabbit immunized with an alanine-lysine-proline-arginine ((ALA1)-tuftsin) containing 14-mer "ferritin" peptide neutralized rat liver ferritin inhibition of in vitro CSF-1-dependent monocytopoiesis. Antiferritin IgG similarly neutralized the inhibitory effect of ferritin but did not neutralize peptide inhibition of the in vitro myelopoietic response. No cross-reactivity between the respective antibodies and Ag was detected either by Western immunoblot or by competitive ELISA. Depletion of adherent cells before marrow cell culture significantly reduced the inhibitory effect of ferritin but did not influence peptide inhibition of CSF-1-stimulated colony formation. Adherent marrow cells and P388D1 cells treated with both CSF-1 and ferritin, but not either alone, produced inhibitory supernatant culture media that were neutralized by antipeptide but not antiferritin IgG. High resolution molecular sieve chromatography of the inhibitory adherent marrow cell and P388D1 supernatants resolved two peaks of 50 to 60 kDa and approximately 30 kDa in each. The inhibitory activity in all four peaks was neutralized by antipeptide but not antiferritin IgG. The ferritin/CSF inhibitors were not further characterized although identity with IL-6, IL-8, TNF-alpha, transforming growth factor-beta, and IFN-alpha/beta could be eliminated. The results indicate that ferritin inhibition of CSF-1-dependent monocytopoiesis is mediated by an endogenously produced inhibitor, or inhibitors, that shares antigenic similarity with the (ALA1)-tuftsin-containing 14-mer peptide and that adherent marrow cells, most likely monocytes or macrophages, produce the endogenous inhibitors in response to both CSF-1 and ferritin.
Impairment of function in aging neutrophils is associated with apoptosis.
Authors: Whyte M K , Meagher L C , MacDermot J , Haslett C
Published: December 2022

Neutrophil apoptosis or programmed cell death permits neutrophil recognition and ingestion by macrophages and represents a mechanism capable of promoting resolution of inflammation. The consequences of apoptosis for neutrophil function are the subject of these investigations. A direct relationship between apoptosis and loss of cytoskeletal functions, phagocytosis, degranulation, and respiratory burst was demonstrated by counterflow centrifugation of neutrophils (aged for 24 h in culture) into fractions with varying proportions of apoptosis. Apoptotic neutrophils displayed a loss of background functions: ability to spread and change shape and random migration. They also showed a reduced ability to respond to deliberate stimulation with the receptor-dependent stimulus, FMLP, by undergoing shape change, chemotaxis, degranulation, and respiratory burst, and showed an inability to phagocytose opsonized zymosan. Loss of FMLP binding to apoptotic neutrophils was demonstrated by analysis of FML[3H]P binding and by autoradiography. Superoxide anion production, but not shape change or degranulation response, to the receptor-independent stimulus, PMA, was preserved in apoptotic neutrophils, implying some retention of intracellular signaling pathways relevant to superoxide production. Apoptosis thus marks a loss of neutrophil functional responsiveness and defines the cell as effectively "isolated" from its external milieu. Neutrophil apoptosis may represent an important event in the control of inflammation, marking the neutrophil both for disposal and as a cell with profound loss of its capacity to generate and release histotoxic products on external stimulation.
Topical application of T-2 toxin inhibits the contact hypersensitivity response in BALB/c mice.
Authors: Blaylock B L , Kouchi Y , Comment C E , Pollock P L , Luster M I
Published: December 2022

T-2 toxin, a trichothecene mycotoxin, has previously been shown to alter immune functions and promote skin tumors. We demonstrate that topically applied T-2 toxin reduces the ear swelling response to oxazolone challenge in BALB/c mice. For this reduction in ear swelling to occur, toxin application must be at, or within, 1 h after challenge. Dose-response studies showed a 44% reduction in ear swelling with 30 ng of T-2 toxin as compared with a similar reduction with 300 ng of dexamethasone. T-2 toxin did not affect Ag transport from the challenge site to the draining lymph nodes as measured by FITC transport. However, T-2 toxin significantly reduced both MHC class II (Ia) expression and Ag presentation at the same concentrations. Because T-2 toxin, a known protein synthesis inhibitor, was found to inhibit protein synthesis in epidermal cell cultures as measured by [3H]-leucine incorporation, cycloheximide was also examined. Cycloheximide reduced both oxazolone-induced ear swelling and Ag presentation in a similar manner to T-2 toxin. One mechanism of action for T-2 toxin in reducing the contact hypersensitivity response is via inhibition of protein synthesis and effective Ag presentation by epidermal Langerhans cells. This may involve alterations in Ia Ag expression, although a role for class II in the induction phase of the contact hypersensitivity response has not been established definitively.
Correlation of oncostatin M secretion by human retrovirus-infected cells with potent growth stimulation of cultured spindle cells from AIDS-Kaposi's sarcoma.
Authors: Radka S F , Nakamura S , Sakurada S , Salahuddin S Z
Published: December 2022

Oncostatin M (OM), a 30-kDa glycoprotein, recently was identified as a major growth-promoting factor in the conditioned medium (CM) of the 38-0 cell line, a CD4,+ chronically human T lymphotropic virus type (HTLV)-II-infected, transformed T cell line. CM 38-0 induced the proliferation of spindle cells cultured in vitro from AIDS-associated Kaposi's sarcoma (AIDS-KS) cells. To determine how much of the AIDS-KS cell growth activity present in 38-0 CM was because of the presence of OM, we depleted OM by using specific mAb-affinity chromatography. OM purified from this CM stimulated AIDS-KS cell growth in a concentration-dependent fashion. The effluent, completely depleted of OM, failed to induce growth of AIDS-KS cells. To detect the constitutive release of OM by cells acutely or chronically infected with either HTLV-I, HTLV-II, or HIV-1, we utilized an enzyme-linked immunoassay. Whereas the chronically infected cells released significant levels of OM, the acutely infected cells released little or no OM. The presence of OM in HIV-1-infected T-cell CM correlated completely with AIDS-KS cell growth activity. Infrequently, low level AIDS-KS cell growth activity was seen in the absence of OM. This correlated with relatively high levels of IL-6 in the CM. In a CM-containing OM in the absence of detectable IL-6, a neutralizing antibody to OM completely abrogated KS cell growth activity. The presence of specific oncostatin M receptors on the KS cell lines was confirmed by cross-linking experiments. The results shown here suggest that T cells chronically infected with HIV-1 can secrete OM, which may play a role in the initiation or progression of AIDS-KS lesions, either alone, or in concert with IL-6.
Dissection of the function of HLA class II and costimulation in B cell-mediated and toxic shock syndrome toxin-1-induced T cell proliferation.
Authors: Dennig D , O'Reilly R J
Published: December 2022

In this study we have demonstrated that costimulation for TSST-1-induced T cell proliferation is provided by HLA class II negative accessory B cells. In addition, TSST-1 binding to these HLA class II negative accessory B cells could not be detected by immunocytofluorescence, or in a system in which T cell proliferation depended on TSST-1 pretreated HLA class II negative B cells. These findings suggest that accessory signals can be supplied by B cells that are not involved in the activation of T cells via Ag-HLA/TCR-CD3 complex interaction: TCR ligation by a TSST-1/HLA complex and costimulation need not be delivered by the same cell. In the presence of HLA class II negative B cells, T cell proliferation in response to TSST-1 could be achieved despite addition of a mAb that can block HLA class II binding sites for TSST-1. These findings suggest the possibility that TSST-1 may directly ligate TCR. In the presence of anti-BB1-treated HLA class II negative accessory cells, TSST-1-induced T cell proliferation was inhibited by over 50% indicating the involvement of the BB1/CD28 pathway. The leukemic pre-B cell lines NALM-6, and BLIN-1, both of which express surface HLA class II Ag, but not the costimulatory B cell activation Ag BB1, were deficient in their accessory function for T cell proliferation in response to the superantigen TSST-1 and the mitogen Con A.
Characterization of a novel cell-surface molecule expressed on subpopulations of activated T and B cells.
Authors: Laszlo G , Hathcock K S , Dickler H B , Hodes R J
Published: December 2022

A mAb, GL7, is described that reacts with a 35-kDa protein on subsets of activated mouse B cells as well as activated CD4+ and CD8+ peripheral T cells. In normal mice analyzed by flow cytometry, GL7 bound at low surface density to 0 to 9% of splenic B cells and 0 to 1% of splenic T cells. In contrast, GL7 bound at high density to a subpopulation comprising approximately 20% of TCR-bright thymocytes, and to B220+ cells in the bone marrow. The activation of B cells by various stimuli resulted in high levels of expression of the surface molecule identified by GL7 on up to 70% of B cells after 48 h; the remaining B cells expressed low or undetectable levels of this molecule, despite evidence of other activation-specific changes in cell-surface phenotype. The GL7-positive population of B cells induced by IL-5 stimulation exhibited high levels of both proliferative and IgM secretory activity, whereas the GL7-negative population showed little of either activity. Activation of splenic T cells with Con A for 48 h resulted in the expression of this determinant at high density on both CD4+ and CD8+ cells. GL7 thus appears to identify a previously uncharacterized cell-surface molecule expressed selectively on subpopulations of activated B and T cells as well as on discrete subpopulations of T and B lineage cells in vivo.
Variable IgH chain enhancer activity in Burkitt's lymphomas suggests an additional, direct mechanism of c-myc deregulation.
Authors: Jain V K , Judde J G , Max E E , Magrath I T
Published: December 2022

The deregulation of the c-myc gene in small non-cleaved cell lymphomas (SNCL) with 8;14 translocations is thought to be due to the juxtaposition of this gene with the IgH chain locus, but exactly how the Ig locus contributes to the deregulation is unclear. One widely considered hypothesis is that Ig gene enhancers, when moved near c-myc, might stimulate inappropriate transcription of this gene. To evaluate this hypothesis, we have tested the ability of the two known H chain enhancers, the JH-C mu intronic enhancer and the 3' alpha enhancer, to support transcription from c-myc promoters, by using transient transfection assays in a panel of 32 SNCL cell lines with 8;14 translocations. The activity of the JH-C mu intronic enhancer varied widely among the cell lines tested and correlated with the presence of nuclear factors binding to the E4/octamer regions of the enhancer. The 3' alpha enhancer was much less active than the intronic enhancer in all of our cell lines and seems unlikely to account for the c-myc deregulation (although the enhancer we tested was from the rat, because the human homologue is unidentified at present). A marked difference was also seen in the ability of individual cell lines to support transcription from the unenhanced c-myc constructs. Cell lines supporting the lowest enhancer activity tended to support the highest level of transcription from unenhanced c-myc promoters. The differences in transcription from c-myc promoters were confirmed by stably transfecting constructs containing c-myc promoters in SNCL cell lines. Our data strongly suggest that the importance of Ig enhancers for c-myc deregulation varies markedly in different cell lines and that, for many, trans-acting regulators of the c-myc promoters are as important or more important, than Ig enhancers in the deregulation of the c-myc gene.
Partitioning of responder CD8+ T cells in lymph node and lung of mice with Sendai virus pneumonia by LECAM-1 and CD45RB phenotype.
Authors: Hou S , Doherty P C
Published: December 2022

Patterns of LECAM-1 and CD45RB expression have been analysed for mediastinal lymph node (MLN) and bronchoalveolar lavage (BAL) populations from C57BL/6J mice with primary Sendai virus pneumonia. The findings indicate that virus-specific CD8+ CTL precursors differentiate in the regional lymph node and become effector CTL after localization to the virus-infected respiratory tract. Relatively few of the MLN CD8+ T cells were LECAM-1-, and all were CD45RB-hi throughout the acute and recovery phases of this disease process. The CD8+ CD45RB-hi and LECAM-1+ T cells characteristic of the MLN were more apparent in the BAL during the 1st wk after exposure to virus, with this shifting to a predominant CD8+ LECAM-1- CD45RB-lo phenotype from day 10 after infection. In contrast, the CD4+ set was generally CD45RB-lo LECAM-1- in the BAL, although CD4+ CD45RB-lo and LECAM-1- cells maintained at relatively high levels in the MLN. The virus-specific CD8+ effectors found in the BAL from day 8 after infection were uniformly LECAM-1-, but varied in the level of CD45RB expression. Evidence of CTL activity was minimal for the MLN, though virus-specific CTLp were present from day 5 after infection and reached maximum numbers within a further 2 days. The ratio of LECAM-1-:LECAM-1+ CTLp in the MLN ranged from 25:1 to > 200:1, with this pattern being maintained for memory spleen cells recovered 3 mo later. Lack of expression of LECAM-1 is thus characteristic of the great majority of Sendai-virus-specific CD8+ T cells, identified either as effector CTL or as precursors that can be stimulated in vitro under limiting dilution conditions. These lymphocyte populations cannot be discriminated on the basis of hi or lo CD45RB expression.
Overexpression of LFA-1 and ICAM-1 in Down syndrome thymus. Implications for abnormal thymocyte maturation.
Authors: Murphy M , Insoft R M , Pike-Nobile L , Derbin K S , Epstein L B
Published: December 2022

The interaction between lymphocyte function-associated antigen 1 (LFA-1) on thymocytes and intercellular adhesion molecule 1 (ICAM-1) on the thymic stroma plays an important role in thymocyte maturation. Consequently, we examined the constitutive expression of LFA-1 and ICAM-1 in postnatal human thymus from children with Down syndrome (trisomy 21, DS) and age-matched control children. We studied DS thymuses because this aneuploid condition is associated with abnormal thymic anatomy and patterns of thymocyte maturation and the affected individuals have a greatly increased incidence of infection. In addition, the beta-chain for LFA-1 is encoded on human chromosome 21, suggesting that trisomy 21 thymocytes may overexpress this adhesion molecule. Using immunofluorescence and flow cytometry, LFA-1 beta expression was evaluated in eleven pairs of DS and age-matched control thymocytes. The mean channel of fluorescence for LFA-1 beta expression was significantly higher in DS thymocytes than in the controls (p < or = 0.01). Six of the 11 pairs were examined for LFA-1 alpha chain expression. DS thymocytes also showed significantly higher levels of LFA-1 alpha expression (p < or = 0.05), which is consistent with findings that surface expression of the LFA-1 alpha chain is dependent on beta-chain expression. Using immunohistochemical analysis and quantitative video imaging, we examined the level of ICAM-1 expression on frozen sections from four pairs of DS and control thymuses, and found nearly twofold higher levels of ICAM-1 expression in the DS thymuses (p < or = 0.05). DS thymuses also showed a diffuse pattern of ICAM-1 expression with elevated staining in both cortex and medulla and poor demarcation of staining at the cortico-medullary junctions. Given our recent observation that DS thymuses overexpress mRNA for IFN-gamma and TNF, and the fact that both of these cytokines induce ICAM-1 expression on cultured human thymic epithelial cells, we propose that increased levels of IFN-gamma and TNF contribute to the enhanced expression of ICAM-1 in DS thymuses. Our findings support a role for cytokines in the regulation of adhesion molecule expression in the thymus and suggest that the increased expression and abnormal distribution of adhesion molecules in DS thymuses alters the interaction between developing thymocytes and the thymic stroma and results in the abnormal thymocyte maturation observed in DS.
The human Ig-beta cDNA sequence, a homologue of murine B29, is identical in B cell and plasma cell lines producing all the human Ig isotypes.
Authors: Hashimoto S , Gregersen P K , Chiorazzi N
Published: December 2022

The B cell Ag receptor complex consists of at least two disulfide-linked, heterodimeric structures: the clonally restricted membrane Ig (mIg) molecule and the nonpolymorphic Ig-alpha:Ig-beta protein dimer. The latter molecule is encoded by two separate genes, mb-1 and B29. The DNA sequences of murine and human mb-1 and murine B29 have been determined previously. This study describes the sequence of the full-length human cDNA homologue of the murine Ig-beta/B29 message. The human sequence codes for a protein that displays the typical subunit features of a transmembrane member of the Ig superfamily. The transmembrane and intracytoplasmic domains exhibit striking nucleotide and amino acid sequence similarity between the two species. These regions show almost complete conservation of areas presumed to be involved in noncovalent interactions with other members of the receptor complex and with intracellular kinases and cytoskeletal components. The only sequence dissimilarity seen in these presumed critical areas involves the Y-E-G-L-N motif, a potential target for tyrosine phosphorylation. In contrast, the extracellular portion is much more divergent. Inasmuch as similar patterns of species diversity have been reported for Ig-alpha, the Ig-alpha and Ig-beta molecules may have coevolved to maintain species-specific extracellular interactions between one another and with mIg. Similar to the Ig-alpha molecule, the Ig-beta sequence is identical in B lineage cells expressing all five Ig isotypes. However, in contrast to the Ig-alpha molecule, the Ig-beta sequence is expressed at apparently similar levels in terminally differentiated, mIg- plasma cells as well as in mIg+, mature B cells. These data suggest that Ig-beta has functions in addition to those associated with surface mIg expression.
A role for gamma delta + T cells during experimental infection of mice with Leishmania major.
Authors: Rosat J P , MacDonald H R , Louis J A
Published: December 2022

To assess the importance of gamma delta + T lymphocyte responses in experimental murine cutaneous leishmaniasis, the expression of gamma delta + TCR on the surface of lymphocytes in spleen and draining lymph nodes of mice infected with Leishmania major was examined. In both susceptible BALB/c and resistant CBA/J mice, an increase in gamma delta + T cells was observed after s.c. infection. Whereas in chronically infected BALB/c mice, these cells can represent up to 35% of the CD3+ cells in the spleen, in CBA/J mice the percentage of gamma delta + T cells returns to a lower level after resolution of lesions. Further experiments, in which the disease outcome was modified in susceptible BALB/c and in resistant CBA/J mice by chemotherapy or immunointervention, have confirmed that a correlation existed between the multiplication of the parasites and the expansion of gamma delta + T cells in the spleen. Interestingly, in BALB/c mice infected for 4 mo, these gamma delta + T cells represent 80% of the activated blast population and a large proportion of them express the alpha-chain of the CD8 molecule. In an attempt to assess a potential role for the gamma delta + T cells in the control of infection with L. major, the effect of injection of mAb against gamma delta TCR on the course of infection was studied. In both BALB/c and CBA/J mice, blocking of the gamma delta TCR resulted in the development of larger lesions that contained increased numbers of parasites. This treatment significantly delayed the healing of cutaneous lesions in otherwise resistant CBA/J mice. Taken together these results indicate that gamma delta + T cells are expanded during experimental infection of mice with L. major and could be involved in host defense against this parasite.
Reduced contact sensitivity reactions in mice treated with monoclonal antibodies to leukocyte function-associated molecule-1 and intercellular adhesion molecule-1.
Authors: Scheynius A , Camp R L , Puré E
Published: December 2022

We have investigated the effect of the administration of mAb against leukocyte function-associated molecule-1 (CD11a/CD18) and intercellular adhesion molecule-1 (CD54) on the delayed-type hypersensitivity reaction in 2,4-dinitro-1-fluorobenzene-sensitized CD2F1 mice. An i.p. injection of the mAb FD441.8 against CD11a at the time of ear challenge led to an almost complete inhibition of ear swelling compared with control animals. Administration of anti-CD54 mAb, 3E2 or YN1/1.7.4, before challenge, resulted in approximately 50% reduction of the delayed-type hypersensitivity response. The decrease in ear swelling reflected a profound inhibition of the edema and the cell infiltration in ears from animals treated with anti-CD11a, and a partial inhibition with anti-CD54 treatment. In addition, the threefold increase in the number of cells recovered from the draining lymph nodes 24 h after challenge in sensitized mice injected with normal IgG was ablated in mice treated with anti-CD11a and partially reduced in anti-CD54-treated animals. Immunohistochemistry and flow cytometry analysis demonstrated that the in vivo administered anti-CD11a mAb was associated with the surface of the majority of the cells in the lymph nodes 24 h after injection and challenge, whereas the anti-intercellular adhesion molecule-1 mAb reacted preferentially with the vascular endothelium. It is concluded that leukocyte function-associated molecule-1 and intercellular adhesion molecule-1 contribute to the generation of an optimal delayed-type hypersensitivity response.
Aberrant expression of the very early activation antigen on MRL/Mp-lpr/lpr lymphocytes.
Authors: Sobel E S , Yokoyama W M , Shevach E M , Eisenberg R A , Cohen P L
Published: December 2022

MRL/Mp-lpr/lpr (MRL/lpr) mice develop marked lymphadenopathy, characterized predominantly by Thy1+CD3+CD4-CD8- cells ("double negative T cells"; DNT). It is paradoxical that DNT proliferate poorly in vitro when stimulated through CD3 or by mitogens. The hamster mAb H1.2F3, raised against dendritic epidermal DNT, recognizes a very early activation (VEA) Ag, which is generally absent on resting cells but expressed soon after T cell activation with ConA or phorbol ester. Cross-linking of this disulfide-linked dimer in the presence of APC and phorbol ester induces proliferation of normal T cells. Therefore, we tested to see whether MRL/lpr DNT expressed this Ag and whether it might play a role in DNT expansion. Unmanipulated cells from enlarged MRL/lpr lymph nodes expressed VEA in an age-dependent manner, peaking at 3 to 4 mo of age. Only limited expression in a small subset of lymphocytes from the congenic MRL/Mp(-)+/+ strain was seen. VEA expression on freshly harvested MRL/lpr lymphocytes was seen mainly on DNT, yet double staining of the DNT for VEA Ag and three other markers known to be present on lpr DNT showed that the DNT were a heterogeneous population. In addition, some CD4+ T cells expressed VEA Ag. Despite their constitutive VEA Ag expression, MRL/lpr DNT showed no proliferative response to cross-linking with the H1.2F3 antibody. Furthermore, unlike normal T cells, they failed to respond to the antibody even when phorbol ester was added. The addition of supplementary cytokines did not correct this defect. These studies indicate that MRL/lpr DNT constitutively and aberrantly express VEA but do not respond when it is cross-linked. These abnormalities may result from the failure to express Fas, the recently reported apoptosis-inducing receptor defective in lpr mice.