bluuebunny/crossref_metadata_2025 huggingface dataset:
Separation and comparison of two monokines with lymphocyte-activating factor activity: IL-1 beta and hybridoma growth factor (HGF). Identification of leukocyte-derived HGF as IL-6.
Authors: Van Damme J , Van Beeumen J , Decock B , Van Snick J , De Ley M , Billiau A
Published: December 2022

Supernatants of mitogen-stimulated human leukocytes contain two biologically related cytokines, IL-1 and hybridoma growth factor (HGF). IL-1 beta is a potent inducer of HGF in fibroblasts but has little stimulating effect on monocytes that spontaneously produce HGF. Leukocyte-derived HGF and IL-1 were separated by the use of affinity chromatography on specific antibodies and discriminating assay systems for both cytokines. They had different Mr upon gel filtration and SDS-PAGE. In contrast to IL-1 beta, HGF showed heterogeneity on a cation-exchange column. IL-1 beta and HGF were purified to homogeneity by a sequence of four and five purification steps, respectively. Leukocyte-derived HGF was characterized by analysis of its NH2-terminal amino acid sequence. This revealed complete homology with fibroblast-derived HGF, 26-kDa protein, IFN-beta 2, and B cell stimulatory factor 2, molecules which have collectively been designated as IL-6. IL-1 beta exerted an antiviral and growth-promoting effect of fibroblasts, whereas HGF/IL-6 did not. Both IL-1 and IL-6 possessed lymphocyte-activating factor activity, which could be neutralized only by an anti-serum against the corresponding cytokine.
Certain lymphoid cells contain the membrane-associated component of the phagocyte-specific NADPH oxidase.
Authors: Pick E , Gadba R
Published: December 2022

Anionic amphiphiles such as long chain unsaturated fatty acids and SDS were shown to activate the superoxide (O2-) producing NADPH oxidase in a cell-free system derived from sonically disrupted phagocytes (macrophages and granulocytes). O2- production required the cooperation of a membrane associated component sedimenting at 48,000 X g (pi) and a cytosolic factor (sigma). The purpose of the present investigation was to find out whether components pi and sigma were also present in non-phagocytic cells that do not produce O2- when stimulated. It was found that the 48,000 X g pellets of guinea pig lymph node and thymus cell sonicates contained significant amounts of component pi, as shown by their ability to support SDS-elicited NADPH-dependent O2- production when supplemented with macrophage cytosol. Lymph node and thymus pi could be extracted from the membrane by 30 mM octyl glucoside, just as its macrophage-derived equivalent. Combining lymph node and thymus 48,000 X g pellet with autologous cytosol did not yield an active enzyme preparation. Also, cytosol from lymph node and thymus cells could not cooperate with macrophage 48,000 X g pellet, indicating that component sigma was lacking in lymphoid cells. Neither pi nor sigma could be detected in guinea pig kidney, the mouse myeloma cell line MOPC 315 and the canine cell line Cf2Th. The 48,000 X g pellet of all nonphagocytic cells examined contained a b-cytochrome that resembled, by its spectral characteristics, the cytochrome b559 thought to be characteristic of phagocytes. In macrophages, cytochrome b559 represented 80% of b-cytochrome content of the 48,000 X g pellet, whereas in non-phagocytic cells, the equivalent material represented only 50 to 60%. There was no correlation between the presence and quantity of the cytochrome b559-like chromophore in the 48,000 X g pellet of a particular cell type and its ability to cooperate with macrophage cytosol in SDS-elicited O2- production.
T cell receptor beta-chain genes in BW5147 and other AKR tumors. Deletion order of murine V beta gene segments and possible 5' regulatory regions.
Authors: Lee N E , Davis M M
Published: December 2022

The AKR thymoma BW5147 has rearranged both of its TCR beta-chain loci, using the same J beta region (J beta 2.5) in each, but with different V beta gene segments. Although the two rearrangements are expressed approximately equally in cytoplasmic RNA, the principle of allelic exclusion is maintained because only one rearrangement is in-frame and capable of encoding a functional protein. In hybridomas made with BW5147 as the fusion partner, this protein may combine with the alpha-chain protein derived from the normal cell to form new Ag/MHC specificities. An analysis of the sequences upstream from the BW5147 rearrangements and additional V regions suggests that two conserved sequences, 10 and nine nucleotides in length and located adjacent to each other 70 to 100 nucleotides 5' of the initiation codon, may be important in the expression of TCR beta-chain genes. Although B and T cells derive from common stem cells, no sequences are observed in T cells that are homologous to the octamer located 5' of all Ig genes. This implies that at least some of the sequences that regulate transcription are not shared in the two major types of lymphocytes. A survey of BW5147 and six other AKR thymomas using probes for 10 of the 18 known V region families indicates a distribution of V beta rearrangements in the tumors consistent with that found in thymocytes. Four of these tumors have apparent VDJ rearrangements on both chromosomes, with the deletion of other V beta gene segments. These data suggest that the primary mechanism of VDJ beta rearrangement is by looping out and excision of the intervening DNA and that most of the V regions are located 5' to the C region. These data were also used to develop a deletion order of the V beta gene segments in the TCR beta-chain locus.
I.29 lymphoma cells express a nonmutated VH gene before and after H chain switch.
Authors: Klein D , Nietupski J , Sirlin S , Stavnezer J
Published: December 2022

The I.29 B cell lymphoma consists of IgM+ and IgA+ cells which express the same germ-line VH gene. IgA+ cells of the I.29 lymphoma were derived from the IgM+ cells by a typical H chain switch recombination event. The IgM+ cells can be induced with LPS to undergo H chain switching in culture. It has been proposed that the somatic hypermutation process is activated during H chain switch, since V genes expressed in IgG+ and IgA+ cells have more frequently undergone mutation than those expressed in IgM+ cells. We have investigated this question by sequencing VH genes expressed before and after H chain switch in the I.29 lymphoma. We have also sequenced the germ-line VH gene corresponding to the gene expressed by I.29 cells to determine whether the VH gene expressed in the IgM+ cells had already undergone somatic mutation. Our results indicate that somatic mutation was not activated in the precursor cell for the I.29 lymphoma, nor during isotype switch in I.29 cells. It is possible that cells of the I.29 lymphoma, or their precursor, have not received the signal which induces somatic mutation, or that I.29 cells belong to a subset of B cells that cannot be induced to undergo any (or much) somatic mutation.
Idiotype vaccination leads to the emergence of a stable surface Ig-negative variant of the mouse lymphoma BCL1, with different growth characteristics.
Authors: George A J , Spellerberg M B , Stevenson F K
Published: December 2022

Vaccination of mice with tumor-derived idiotypic IgM from the B cell lymphoma, BCL1, induces an anti-idiotypic immune response which suppresses tumor development. One of the mechanisms by which tumor cells can escape attack is by failing to express significant levels of idiotypic immunoglobulin at the cell surface, and a stable variant of this phenotype has been isolated. The variant, termed SNAG 1, continues to synthesize idiotypic IgM, which can be detected in the cytoplasm, but it neither secretes nor expresses IgM on the cell surface (less than 10% of the levels of the original BCL tumor), even though the H and L chains show no gross structural changes. The SNAG 1 cells resemble the parent BCL cells in morphology, in expression of MHC class I and II Ag and in bearing FcR. A significant difference between the BCL lymphoma cells and the variant cells is that the latter fail to respond to LPS by either DNA synthesis or secretion of IgM, suggesting that surface Ig might be required for such a response. The variant has a slower rate of division than the parent tumor both in vitro and in vivo, and a rather different organ distribution. Study of such variants might allow analysis of the mechanisms involved in surface Ig expression and its possible role in tumor cell growth and migration.
T cell recognition of Mlsc,x determinants.
Authors: Abromson-Leeman S R , Laning J C , Dorf M E
Published: December 2022

Among a large number of cow insulin-specific T cell clones derived from both C57BL/10 and B10.A strains, several were found to react to non-MHC-linked gene products of a number of allogeneic strains. The stimulatory moiety for three of these clones correlates, in part, with expression of Mlsc, as defined by mouse strains C3H/HeJ and A/J. In addition, all three of these clones are stimulated by cells from strain PL/J, which has the poorly defined Mlsx allele. The data strongly suggest that Mlsx may, in fact, be Mlsc or is, at least, highly cross-reactive with Mlsc. Segregation analysis by using (B10.D2 X PL/J)F2 mice demonstrates that the Mlsx gene is genetically independent of the Mlsa linked Ly-9 marker on chromosome 1. Further studies with the use of these Mlsc,x-reactive clones reveal that they also recognize a gene product present in many mouse strains including DBA/2 which were previously phenotyped as Mlsa. However, testing of BxD recombinant inbred lines excludes Mlsa as being the stimulatory moiety. We therefore propose reclassification of the Mls phenotypes of several mouse strains based upon a two-locus model for Mls.
Clonal analysis of the BALB/c secondary B cell repertoire specific for a self-antigen, cytochrome c.
Authors: Jemmerson R , Blankenfeld R
Published: December 2022

mAb to rat cytochrome c (cyt c), totaling 556, were produced by individual clones of secondary B lymphocytes from nine groups of five BALB/c mice each in vitro using the splenic focus culture system. Inasmuch as rat and mouse cyt c are identical, these B cells can be considered specific for a self-antigen. The mAb were categorized into specificity groups based on their reactivities with a panel of seven cyts c that differ at two to six amino acid residues. The number of distinct specificities for the native protein was restricted to fewer than 20. Different groups of mice expressed the same specificities at comparable frequencies, including a single dominant one, and the total number of secondary cyt c-specific B cells was constant among groups of mice. This suggests that the acquisition of the secondary B cell specificity repertoire for this self-antigen is regulated. However, it is indeed possible that each specificity group may comprise a number of distinct mAb molecules that have arisen stochastically. Specificities expressed by as few as 1% of the total mAb were observed. Thus, it is likely that the identified specificities reflect the secondary B cell specificity repertoire for rat cyt c. The dominant specificity expressed by 50% of the mAb was characterized by elimination of antigen recognition as a result of replacement of aspartic acid by glutamic acid at position 62. Minor specificities expressed by 19% of the mAb were characterized by more subtle affects of an amino acid change at position 62 and/or an amino acid substitution from rat cyt c at position 60. Antibodies in other specificity groups reacted with epitopes in the region of residues 44 and 47. Whereas substitutions at positions 44, 47, 60, and 62 eliminated recognition by most of the mAb, changes at position 92 and at 103 also appeared to affect the binding of some mAb in the region around residues 60 and 62. The amino acid residues implicated in the recognition by murine mAb of murine cyt c have been shown previously to be involved in the epitopes of foreign mammalian cyt c. Therefore, self-tolerance cannot fully explain the restriction of the epitopes to these regions on foreign mammalian cyt c.
Antigen as a positive and negative regulator of proliferation in cytotoxic lymphocytes. A model for the differential regulation of proliferation and lytic activity.
Authors: Russell J H , Manning D E , McCulley D E , Meleedy-Rey P
Published: December 2022

The goal of these experiments has been to capitalize on the functionally distinct responses of heteroclitic CTL to cross-reactive Ag to explore the role of Ag in regulating CTL proliferation and lytic function. The experiments here have examined one of these clones in detail but several such clones appear similar. The principal findings were: 1) proliferation and lysis may be optimally stimulated by quantitatively different interactions, 2) Ag can have negative as well as positive effects on the proliferative potential of the responding cell, and 3) both positive and negative effects on lysis can be mimicked by agents which are similar to the second messengers produced by stimulating the TCR.
A T cell surface molecule different from CD2 is involved in spontaneous rosette formation with erythrocytes.
Authors: Bernard A , Aubrit F , Raynal B , Pham D , Boumsell L
Published: December 2022

Increasing evidence indicates that rosettes which spontaneously from between human T cells and E might be of physiologic relevance. We show here that another T cell-surface molecule than CD2 is involved in rosette formation. Four mAb have been obtained reacting with human T cells that block rosettes with E from many species, including autologous cells. They react with a molecule, we termed E2, which is actively synthetized by T and monocytic cells. Immunoprecipitation revealed a major 32-kDa band. Immunoblots revealed a major 32-kDa band and a minor 20-kDa band. This molecule was detected on all T cells tested--and present at high densities on corticothymocytes, but at low densities on medullary thymocytes. It was also found on monocytes but not on B cells. However B-CLL cells did carry this molecule. E2 molecules were also detected on nonhematologic cells. Together with the recent evidence that 3 molecules from the erythrocyte surface are also involved for rosettes, intricate molecular interactions would account for adhesion of T cells to autologous E and possibly autologous nucleated cells.
Clearance and organ localization of small DNA anti-DNA immune complexes in mice.
Authors: Emlen W , Burdick G
Published: December 2022

DNA anti-DNA immune complexes (IC) play a major role in the pathogenesis of SLE. We studied the clearance and organ localization of small DNA anti-DNA IC formed at different Ag/antibody ratios in normal mice. IC formed at Ag excess, containing areas of "exposed" DNA not covered by IgG, showed rapid Ag-mediated clearance from the circulation by the liver. DNAse digestion of these IC in vitro yielded small IC devoid of exposed DNA that were cleared more slowly from the circulation. IC formed at antibody excess were cleared by an Ag-independent mechanism at rates proportional to the number of IgG in the IC. None of the IC studied bound significantly to complement receptors on circulating cells in vivo or in vitro. For all IC, after initial rapid clearance, 10 to 20% of the injected material persisted in the circulation. Analysis of these IC showed that they were processed in vivo to yield complexes similar to those generated by in vitro DNAse digestion. We conclude that IC containing exposed DNA are removed rapidly from the circulation by Ag-mediated clearance. However, in vivo processing of IC occurs to yield smaller IC that are cleared slowly. We propose that these IC containing small DNA may persist in the circulation and accumulate in tissues, thereby playing an important role in the pathogenesis of tissue injury in SLE.
HLA-DR4 restricted lymphocyte proliferation to a Mycobacterium tuberculosis extract in rheumatoid arthritis and healthy subjects.
Authors: Palacios-Boix A A , Estrada-G I , Colston M J , Panayi G S
Published: December 2022

The ability of an antigenic extract of Mycobacterium tuberculosis to induce proliferation of PMBC from rheumatoid arthritis (RA) patients and normal controls was examined. The subjects were further classified as bearing or not bearing the HLA-DR4 phenotype, since this specificity is regarded as a genetic determinant commonly associated with RA. The mycobacterial extract induced significantly higher proliferative responses in lymphocytes from all HLA-DR4 positive as compared to HLA-DR4 negative subjects regardless of whether they had RA or not. This response was maximal at day 6 of incubation and could be abrogated by anti-DR/DQ mAb added to the culture. SDS-PAGE of this extract revealed three major protein bands located at Mr 14, 47, and 65 kDa. After fractionation, Western blotting, and resuspension of protein-laden nitrocellulose particles, only the 14- and 47-kDa proteins retained the original proliferative capacity of the mycobacterial extract. The band separating with a Mr of 47 kDa was found to be the most strongly associated with the HLA-DR4 restricted lymphocyte proliferation, and represents a newly identified M. tuberculosis Ag of relevance in T cell responses. These data provide insight into the pathogenic potential of certain bacterial Ag which could trigger or perpetuate inflammatory disorders when presented in the appropriate genetical background.
Recombinant murine IL-3 fails to stimulate T or B lymphopoiesis in vivo, but enhances immune responses to T cell-dependent antigens.
Authors: Kimoto M , Kindler V , Higaki M , Ody C , Izui S , Vassalli P
Published: December 2022

We have explored the in vivo effect of IL-3 on the lymphopoiesis and humoral responses of mice bearing osmotic minipumps loaded with murine rIL-3 for 1 to 4 wk. A marked splenomegaly due to the accumulation of hemopoietic precursors was seen, but no increase was found in the lymphoid organs in the total number of cells belonging to the T or B lymphocyte lineage, i.e., of L3T4+ or Lyt-2+, or of allospecific cytotoxic T lymphocyte precursor for the T lineage, or of sIg+ or B220+ cells, or of B colony-forming cells for the B lineage; total activity of natural killer and lymphokine-activated killer cells was decreased. In contrast to the splenomegaly, a marked diminution in the number of thymocytes was observed, suggesting that rIL-3 in large amounts does suppress the T lymphopoiesis, perhaps as the result of the selective stimulation of early progenitor cells toward the hemopoietic pathway. rIL-3 perfusion during immunization increased the IgM and IgG responses to a T cell-dependent antigen, human IgG, and prevented tolerance induction by the deaggregated human IgG, although in the same conditions it did not modify the response to a T cell-independent antigen. Our results suggest that in vivo IL-3 does not act directly on lymphocytes or their precursors, but may potentiate the humoral immune response to T cell-dependent antigens, presumably by acting on accessory cells.
Response of resident murine peritoneal macrophages to in vivo administration of granulocyte-macrophage colony-stimulating factor.
Authors: Morrissey P J , Bressler L , Charrier K , Alpert A
Published: December 2022

The effect of s.c. inoculation of purified recombinant derived granulocyte-macrophage (GM)-CSF on resident murine peritoneal macrophages was assessed in this study. From 18 to 24 h after s.c. administration of GM-CSF to normal mice, the resident peritoneal macrophages were harvested and the levels of membrane-bound IL-1, FcR, Mac-1 cell-surface Ag, and class II MHC expression were assessed. Peritoneal cells from GM-CSF-inoculated mice had significantly greater levels of membrane-bound IL-1 than did control mice. In addition when resident peritoneal macrophages from normal mice were purified by adherence and grown in the presence of GM-CSF, they produced greater levels of both membrane-bound and secreted IL-1. The peritoneal cells from GM-CSF-inoculated mice did not differ from controls in the expression of class II MHC-encoded Ag. This observation was confirmed by the finding that GM-CSF was unable to induce class II MHC expression on P388D1 cells, whereas a secondary mixed leukocyte culture supernatant was. Peritoneal cells from GM-CSF-inoculated mice also exhibited greater levels of expression of FcR and the Mac-1 cell-surface Ag. This resulted in an increase in their ability to phagocytose opsonized SRBC in vitro.
Expression of the tumor necrosis factor locus is not necessary for the cytolytic activity of T lymphocytes.
Authors: Jongeneel C V , Nedospasov S A , Plaetinck G , Naquet P , Cerottini J C
Published: December 2022

In order to test whether tumor necrosis factors alpha (TNF-alpha) or beta (TNF-beta, also known as lymphotoxin) are involved in the lysis of target cells by cytolytic T lymphocytes, we probed for the presence of the TNF mRNAs in several quiescent and activated CTL clones. No TNF mRNA could be found in constitutively cytolytic Lyt-2+ clones, and only two out of three clones tested accumulated TNF mRNA after stimulation with phorbol myristate acetate and ionomycin. Of two L3T4+ clones that can be induced to become cytolytic by a combination of antigen and IL-1, only one accumulated TNF-beta mRNA in the process. The PC60 rat X mouse T cell hybrid, which becomes cytolytic in response to a combination of IL-1 and IL-2, also failed to accumulate TNF mRNA after stimulation with these agents. Our results strongly suggest that TNF-alpha or -beta are not necessary agents of the cytolytic activity exhibited by antigen-specific T lymphocytes.
Production and properties of histamine-releasing factor of guinea pigs.
Authors: Alam R , Lewis D M , Olenchock S A
Published: December 2022

The production of histamine-releasing factor (HRF) by human mononuclear cells has previously been reported. In this paper we describe the production of HRF by guinea pig spleen cells, thymocytes, and PBMC. Guinea pig lymphoid cells were cultured either alone or in the presence of mitogens (PHA and Con A) or specific Ag(OVA and keyhole limpet hemocyanin) and the dialyzed cell-free supernatant was tested for histamine-releasing activity on guinea pig lung mast cells and blood basophils. Lung mast cells were isolated by enzymatic digestion and partially purified by countercurrent elutriation and discontinuous Percoll gradient centrifugation. Guinea pig spleen cells, thymocytes, and PBMC spontaneously produced significant amounts of HRF. The production was enhanced upon stimulation with PHA or specific Ag in animals immunized with Ag in CFA. Two distinct species of HRF were identified with m.w. of 50,000 to 70,000 and 5000 to 8000 by gel chromatography. HRF is a trypsin- and chymotrypsin-sensitive heat-stable protein. It does not bind to Con A-Sepharose and its production is not inhibited by tunicamycin. HRF-induced histamine release from lung mast cells is a temperature-dependent process and is complete in 10 min at 37 degrees C. Intradermal injection of HRF caused an immediate ear-swelling reaction in guinea pigs. The most severe ear-swelling reactions did not resolve within 1 h, but instead evolved over a period of 12 to 24 h.
Inhibitory effect of exogenous arachidonic acid on alveolar macrophage 5-lipoxygenase metabolism. Role of ATP depletion.
Authors: Peters-Golden M , Shelly C
Published: December 2022

Although a variety of agonists have the capacity to stimulate the release of endogenous arachidonic acid (AA) from macrophage lipids and its subsequent metabolism to both cyclooxygenase and 5-lipoxygenase eicosanoids, including leukotrienes (LT), previous work suggests that exogenously supplied arachidonate is metabolized preferentially to cyclooxygenase products and mono-hydroxyeicosatetranenoic acids rather than LT. We have compared the metabolism of endogenous and exogenous AA in cultured rat alveolar macrophages and have further examined the effect of exogenous fatty acids on eicosanoid synthesis stimulated by the calcium ionophore A23187 and the particulate agonist zymosan. As reported by others, exogenous AA was metabolized to large amounts of cyclooxygenase products, 12-hydroxyeicosatetraenoic acid and 15-hydroxyeicosatetraenoic acid, but far lesser amounts of all 5-lipoxygenase products. However, whereas exogenous AA augmented the synthesis of cyclooxygenase metabolites in both A23187- and zymosan-stimulated cells, it dose-dependently inhibited the synthesis of LT. Inasmuch as unsaturated fatty acids, including AA, uncouple oxidative phosphorylation, and because 5-lipoxygenase activity depends on ATP, we examined the possibility that ATP depletion was responsible for the inhibition of LT synthesis by exogenous AA. Arachidonate depleted cellular ATP in a dose-dependent fashion similar to its inhibition of LT synthesis. In addition, the other unsaturated fatty acids, linolenic and eicosatrienoic, but not the saturated fatty acids, palmitic or stearic, both depleted ATP and inhibited A23187-induced LT synthesis. These data suggest that the relative lack of synthesis of LT from exogenous AA is related to the ability of this unsaturated fatty acid to function as an inhibitor, as well as a substrate, of 5-lipoxygenase. Our results further suggest that this inhibition of 5-lipoxygenase may be mediated by ATP depletion.
Activation of HLA-restricted EBV-specific cytotoxic T cells does not require CD4+ (helper) T cells or exogenous cytokines.
Authors: Fishwild D M , Benike C J , Engleman E G
Published: December 2022

MHC-restricted, viral Ag-specific "memory" CTL are thought to play a decisive role in the defense against pathogenic viruses. However, the requirements for activating such CTL remain controversial. In particular, the role of CD4+ helper cells and their soluble products (e.g., IL-2) are uncertain. To approach these questions as they relate to EBV-specific CTL, highly purified CD8+ T cells from healthy EBV-seropositive individuals were cultured with autologous irradiated EBV-transformed B lymphoblastoid cell lines (LCL), in the presence or absence of autologous CD4+ cells or 1 to 10 U/ml purified rIL-2. The results indicate that the induction of CTL requires neither Th cells nor exogenous IL-2. The CTL generated from isolated CD8+ cells were HLA class I restricted as demonstrated by their ability to lyse targets sharing at least one HLA-A or -B Ag with the stimulating autologous LCL. Furthermore, a mAb (W6/32) to a common determinant on HLA class I Ag blocked both the generation and effector phases of killing, whereas an HLA class II directed mAb had no effect. Addition of an IL-2R-specific antibody (anti-Tac) to the culture medium blocked induction of CTL, suggesting that endogenously produced IL-2 plays an obligatory role in this system. Paraformaldehyde fixation of LCL abrogated their ability to function as stimulator cells; however, addition of 2 U/ml exogenous IL-2 to fixed LCL cultured with CD8+ cells allowed for the induction of highly specific CTL. These results indicate that EBV-specific memory CTL can be activated in the absence of CD4+ helper cells or their soluble products, but nonetheless require Ag and IL-2.
Identification of antigens and antibodies specific for Pneumocystis carinii.
Authors: Kovacs J A , Halpern J L , Swan J C , Moss J , Parrillo J E , Masur H
Published: December 2022

To increase understanding of Pneumocystis carinii and its interaction with its hosts, Ag specific for rodent and human P. carinii were identified by the immunoblot method after PAGE of P. carinii organism extracts. The m.w. of the major Ag of rat P. carinii were 45,000, 110,000, and a broad band of 49,000 to 64,000, and of human P. carinii were 22,000, 24,000, and a broad band of 35,000 to 45,000 daltons. Human and rat pneumocystis were not antigenically identical. Specific antibodies against rat P. carinii Ag were found in 18 of 79 rats by the immunoblot method. Specific antibodies against human P. carinii Ag were found in 32 of 33 adult human sera, but in only 1 of 8 sera from infants and children. Specific antibodies were found in sera of 13 of the 14 adults with no history of P. carinii pneumonia, and all 19 patients with recently diagnosed P. carinii pneumonia, including 9 patients with P. carinii pneumonia associated with AIDS. The results of this study support previous suggestions that a large proportion of adults have been exposed to P. carinii and provide a basis for the further investigations of host-P. carinii interactions.
Target cell directed NK inactivation. Concomitant loss of NK and antibody-dependent cellular cytotoxicity activities.
Authors: Abrams S I , Brahmi Z
Published: December 2022

We investigated the inactivation of human NK cells, a population of large granular lymphocytes (LGL), with K562, an NK-sensitive target cell (TC) and KLCL, an NK-resistant TC, but which can be lysed by NK cells via antibody (Ab)-dependent cellular cytotoxicity. NK-enriched effector cells (ECc) were first treated with either K562 or Ab-coated KLCL (Ab-KLCL). After incubation, ECc were separated from their TC then examined for residual NK and ADCC activities, phenotypic changes, and changes in LGL morphology. K562-treated ECc and Ab-KLCL-treated ECc, when retested against the inactivating TC, respectively, lost greater than 90% of their lytic activities. However, K562-treated ECc lost 60 to 70% of their activity against Ab-KLCL, whereas Ab-KLCL-treated ECc lost less than 10% of their activity against K562. In contrast to what we observed with K562-treated ECc, we detected significant reductions in plasma membrane expression of Leu-11a and Leu-11b on Ab-KLCL-treated ECc. Although the proportion of OKM1+ cells remained unchanged after the inactivation process, the density of OKM1 on both K562-treated ECc and Ab-KLCL-treated ECc increased significantly. Morphologic analysis revealed no apparent differences in the percentages of LGL before and after treatment with K562 or Ab-KLCL. Finally, IL-2 restored lytic potential to both K562-treated ECc and Ab-KLCL-treated ECc and, in addition, IL-2-induced enhancement of Ab-KLCL-treated ECc was accompanied by a partial reexpression of Leu-11a. These data support the hypothesis that NK-cell-mediated cytotoxicity and antibody-dependent cellular cytotoxicity may result from a common lytic mechanism, although the initiation steps and regulation of the pathway are distinct.
Use of colony assays and anti-T cell immunotoxins to elucidate the immunobiologic features of leukemic progenitor cells in T-lineage acute lymphoblastic leukemia.
Authors: Uckun F M , Myers D E , Ledbetter J A , Swaim S E , Gajl-Peczalska K J , Vallera D A
Published: December 2022

The specific binding of radioiodinated rIL-2 to fresh marrow blasts from T-lineage acute lymphoblastic leukemia (ALL) patients was initially investigated. The estimated number of radioiodinated rIL-2 molecules bound per blast ranged from undetectable to 1948. In colony assays, 72% of 32 cases analyzed showed a significant proliferative response to rIL-2, which depended on PHA-stimulated lymphocyte conditioned medium activation. Colony stimulation indices correlated with the number of radioiodinated rIL-2 molecules bound per blast but not with expression of CD25/Tac Ag on fresh marrow blasts or primary colony blasts. These findings provide evidence that in T-lineage ALL functional IL-2R proteins are expressed on leukemic progenitor blasts which may be distinct from Tac Ag. We used the mAb 35.1, T101, and G3.7 to test for expression of CD2, CD5, and CD7 on fresh marrow blasts from 126 T-lineage ALL patients. CD2, CD5, and CD7 were expressed in 84%, 93%, and 99% of cases, respectively. Furthermore, colony blasts that represent the early progeny of leukemic progenitor blasts were also CD2+CD5+CD7+. Ricin conjugates of 35.1, T101, and G3.7 mAb were used as Ag-specific cytotoxic probes to test for expression of CD2, CD5, and CD7 at the level of T-lineage leukemic progenitor blasts. Each immunotoxin was able to selectively eliminate greater than 99% of leukemic progenitor blasts, providing unique and direct evidence that these cells co-express CD2, CD5, and CD7. Neither mixtures of anti-CD5 and anti-CD7 nor anti-CD2, anti-CD5, and anti-CD7 immunotoxins were more effective against blast progenitor cells than the individual immunotoxins alone, confirming that CD2, CD5, and CD7 are not expressed on non-overlapping progenitor cell subpopulations.
Differential in vitro activation of CD8-CD4+ and CD4-CD8+ T lymphocytes by combinations of anti-CD2 and anti-CD3 antibodies.
Authors: Yang S Y , Rhee S , Welte K , Dupont B
Published: December 2022

Purified peripheral blood T lymphocytes and the CD8-CD4+ and CD4-CD8+ T cell subsets, exhaustively depleted of APC have been studied for their capacity to respond to mAb directed against the CD3-Ti Ag-specific TCR complex and against the CD2 SRBCR. It is demonstrated that high affinity IL-2R can be readily induced by either anti-CD3 and/or anti-CD2 stimulation. However, IL-2 production can be observed only in the CD4+CD8- T cell subset. These results clearly contrast data obtained with purified CD4-CD8+ T cells, which are able to proliferate when the CD3-Ti complex is activated in the presence of APC. The data presented in the present study demonstrate that a simplified model for T cell activation and clonal expansion of the two major T cell subsets involve only the CD3-Ti complex and the CD2 Ag. Under conditions where the activation signals for the T cells are restricted only to the activation of CD3-Ti and CD2, the CD4+CD8- T cells respond with IL-2 production and expression of high affinity IL-2R, whereas the CD4-CD8+ T cell subset depends on exogenous IL-2 provided by the CD4+CD8- cells. These data do not, however, exclude an involvement of other cell-surface signals for regulation and control of T cell activation and T cell effector functions.
2H1--a novel antigen involved in T lymphocyte triggering.
Authors: Morimoto C , Rudd C E , Letvin N L , Hagan M , Schlossman S F
Published: December 2022

A mAb anti-2H1, was produced against PHA-activated T cells from a lower primate, Aotus trivirgatus. Anti-2H1 reacted with 90% of peripheral T cells but was found to react with only 10% of thymocytes and some, but not all, leukemic T cell lines. 2H1 expression was dramatically increased when thymic cells were activated by Con A plus PMA. In contrast, anti-2H1 did not react with B cells, macrophages, null cells, or hematopoietic stem cell lines. More importantly, anti-2H1 antibody can induce T cell activation and proliferation in synergy with PMA or anti-T11(3). SDS-PAGE analysis of polypeptides immunoprecipitated with anti-2H1 showed two major polypeptides of 140 and 105 kDa. Thus, the 2H1 Ag can be distinguished from T3, T11, and 9.3 Ag. These results indicate that the 2H1 Ag appears to be involved in the activation of T lymphocytes.
Loss of CD45R and gain of UCHL1 reactivity is a feature of primed T cells.
Authors: Akbar A N , Terry L , Timms A , Beverley P C , Janossy G
Published: December 2022

A group of mAb recognizing the 200- and/or 220-kDa determinants (CD45R) of the leukocyte common Ag such as 2H4, WR16, MD4.3, and SN130 cross-block each other showing that they recognize a closely related epitope. The antibody UCHL1 reacts with a 180-kDa determinant of the leukocyte common Ag and exhibits a reciprocal T subset distribution pattern to the CD45R group. Peripheral blood T cells were 40% positive for UCHL1 and 58 to 65% positive for the CD45R antibodies; less than 1% of cells stained for both. On activation of CD45R+,UCHL1- T cells by PHA, up to 40% of cells became positive for both CD45R and UCHL1 by day 3. By day 7, CD45R+,UCHL1- cells fell from 90 to less than 21% whereas UCHL1+,CD45R- cells rose from 2 to 93%. Conversely, PHA-stimulated UCHL1+,CD45R- cells remained UCHL1+,CD45R- during the 7 days in culture showing that phenotypic change was unidirectional from CD45R+ to UCHL1+. In primary allogeneic mixed lymphocyte reactions, activated CD45R+ T cells also showed a change to UCHL1+. When these cells were rechallenged by the original alloantigen, the UCHL1+ cells showed 7- to 20-fold greater proliferation than the CD45R+ cells on day 3 after rechallenge. The recovery of virtually all alloantigen induced secondary proliferative response in the UCHL1+,CD45R- T cell population suggests that UCHL1 identifies a primed population of T cells which may include memory cells.
A monoclonal antibody detects macrophage maturation antigen which appears independently of class II antigen expression. Reactivity of monoclonal EBM11 with bovine macrophages.
Authors: Bielefeldt-Ohmann H , Sabara M , Lawman M J , Griebel P , Babiuk L A
Published: December 2022

A mAb EBM11, raised against human macrophages [M phi] was found to detect a bovine M phi diameter-subpopulation. The Ag was strictly intracellular and was expressed in M phi only at a certain state of maturation. Its expression was regulated independently of the activation state of the cells, as revealed by treating M phi in vitro with bovine rIFN-alpha I1, IFN-gamma, or TNF-alpha, all potent M luminal diameter activators. Such treatment had no apparent effect on Ag expression. The Ag was present in 1 to 5% of peripheral blood leukocytes, i.e., up to 20% of circulating blood monocytes, in both normal noninfected cattle and in cattle persistently infected with bovine viral diarrhoea virus. Blood monocytes of the latter group were activated in vivo, but apparently did not reach a more mature state than found in noninfected animals. After an acute infection with bovine herpes virus type 1, the frequency and total number of EBM11+ cells decreased dramatically in inverse relationship to an equally significant increase in frequency and total number of circulating monocytes. In cryostat sections of normal tissues, the EBM11 mAb reacted with sinus M phi and M phi in germinal centers of lymphoid tissues, with alveolar M phi and liver Kupffer cells. In skin it reacted with few scattered M phi in the dermis, but not with epidermal Langerhans cells. This latter feature distinguishes the bovine system from the human. In virus-induced inflammatory processes in skin and keratinized epithelia EBM11+ cells constituted a subpopulation of the infiltrating M phi. The data obtained suggest that EBM11 mAb could be useful both for the elucidation of differentiation/maturation pathways of cells of the monocyte-macrophage lineage as well as for studies of M phi-virus interactions in virus infections. In either aspect cattle could provide a useful comparative model.
Immunoblot analysis of IgG subclasses of multiple lupus autoantibodies.
Authors: Bonfa E , Llovet R , Elkon K
Published: December 2022

Sera from 35 patients with systemic lupus erythematosus were analyzed for the subclass distribution of IgG autoantibodies to saline-soluble intracellular proteins. To assess the response to all Ag, an immunoblot technique was used, and strips were sequentially probed with patient sera, monoclonal anti-subclass sera, and a labeled anti-mouse reagent. The relative proportions of each subclass reactive with a specific Ag was determined semi-quantitatively by densitometric scanning. Overall, all of the IgG subclasses were involved in the autoantibody response, although the frequency of detection was highest for IgG1 and lowest for IgG4. When the subclass responses to different Ag were compared, IgG1 was the major subclass reactive with the Ro, La, and U1 ribonucleoprotein Ag, whereas IgG1 and IgG2 were almost equally represented in the responses to the Sm BB' and D Ag as well as to the ribosomal P proteins. Individual patient sera frequently showed discordance between the dominant subclass reactive against apparently unrelated proteins and even against proteins within the same antigenic particle (e.g., the Sm BB' and D proteins). These observations indicate that there are two major patterns of subclass response to the common lupus autoantigens but that considerable variation between patients and even within the same patient (to different Ag) occurs.
Transforming growth factor-beta 1 (TGF-beta 1) and recombinant human tumor necrosis factor-alpha reciprocally regulate the generation of lymphokine-activated killer cell activity. Comparison between natural porcine platelet-derived TGF-beta 1 and TGF-beta 2, and recombinant human TGF-beta 1.
Authors: Espevik T , Figari I S , Ranges G E , Palladino M A
Published: December 2022

We have investigated the ability of porcine-platelet-derived transforming growth factor-beta 1 (TGF-beta 1) to inhibit the generation of lymphokine-activated killer (LAK) cells by human rIL-2. The results demonstrate that TGF-beta 1, in a dose-related manner, significantly inhibits rIL-2-induced LAK cell activity against Daudi and COLO target cells and, to a lesser degree, against K-562 cells. Maximal inhibition was obtained by the addition of TGF-beta 1 at the time of culture initiation and, to a lesser degree, on day 1. Only minimal inhibition was obtained when TGF-beta 1 addition was delayed until day 2 of culture or when added directly into the LAK cell assay. Additional studies demonstrated that porcine platelet-derived TGF-beta 2 and human rTGF-beta 1 inhibited LAK cell generation similar to that obtained with TGF-beta 1. The inhibition of LAK cell activity by TGF-beta 1 was reversed by the addition of human rTNF-alpha at the initiation of culture. In addition, rTNF-alpha synergized with suboptimal levels of rIL-2 in the generation of LAK activity. After stimulation with rIL-2, LAK cells produced significant levels of IFN-gamma, TNF-alpha, and TNF-beta. TGF-beta 1 inhibited the production of these cytokines in a dose-related manner. The results extend the previous known activities for human rTNF-alpha and TGF-beta 1 and further demonstrate the reciprocal relationship between these two molecules in the regulation of certain immune functions.
Inability of murine peritoneal macrophages to convert linoleic acid into arachidonic acid. Evidence of chain elongation.
Authors: Chapkin R S , Somers S D , Erickson K L
Published: December 2022

Various murine macrophage populations synthesize and secrete large amounts of arachidonic acid (20:4n-6) derived eicosanoids (cyclo-oxygenase and lipoxygenase products). These metabolites are known to possess a wide variety of functions with regard to the initiation and regulation of inflammation and tumorigenesis. Because the dietary intake of 20:4n-6 is usually low, tissues are largely dependent upon dietary linoleic acid (18:2n-6) as an initial unsaturated precursor for the biosynthesis of 20:4n-6. The purpose of these experiments was to determine whether resident or responsive murine macrophages possess desaturase and elongase activities capable of in vitro conversion of 18:2n-6 into 20:4n-6. Peritoneal exudate macrophages were purified by adherence and incubated in serum-free medium containing fatty acid-free BSA with [1-14C] 18:2n-6. Approximately 90 to 98% of the [14C]18:2n-6 at 4 and 16 h was recovered in phosphatidylcholine and phosphatidylethanolamine. The metabolism of [14C]18:2n-6 was determined after transesterification and separation of the 14C-fatty acid methyl esters by argentation TLC, reverse phase HPLC, and electron impact gas chromatography/mass spectrometry. Resident and responsive macrophages lacked the capacity to transform [14C]18:2n-6 into 20:4n-6. In addition, prelabeled macrophages incubated with soluble, calcium ionophore A23187 or phorbol myristate, or particulate, zymosan, membrane perturbing agents also lacked delta 6 desaturase activity. All macrophages tested were capable of elongating [14C]18:2n-6 into [14C]20:2n-6. These observations suggest that 20:4n-6, present in macrophage phospholipids, is biosynthesized elsewhere and transported to the macrophage for esterification into the phospholipids. In addition, these findings demonstrate that elongase activity is present in both the resident and responsive peritoneal macrophage.
Human monocytes generate basophil histamine-releasing activities.
Authors: Schulman E S , McGettigan M C , Post T J , Vigderman R J , Shapiro S S
Published: December 2022

Human peripheral blood monocytes generated activities during 24-h culture that were capable of triggering histamine release from 17 of 18 human basophil donors. Monocytes and their in vitro transformed macrophages continued to elaborate these basophil histamine-releasing activities for at least 3 wk in culture. In the 18 basophil donors tested, maximum histamine release induced by monocyte supernatants was 33.8 +/- 5.9% (mean +/- SEM) of total basophil histamine content; optimum anti-IgE-induced release was 38.8 +/- 6.2%. Basophil histamine release in response to monocyte activities was optimal at 37 degrees C and at calcium concentrations of 2 to 5 mM. Release was greater than 90% complete 1 min after challenge and was inhibited by anti-allergic drugs. The mechanism of release appeared to be independent of IgE binding. Gel filtration of supernatants derived from both day 1 (monocyte stage) and day 14 (macrophage stage) cultures demonstrated activity peaks with approximate m.w. of 12,000 and 30,000. In contrast to the marked responsiveness of basophils, only 2 of 10 human lung mast cell preparations responded; release in those preparations was low: 3% and 13% histamine release, respectively. Thus, monocytes produce potent histamine-releasing activities with differential actions on basophils and mast cells.
Binding of staphylococcal enterotoxin A to accessory cells is a requirement for its ability to activate human T cells.
Authors: Carlsson R , Fischer H , Sjögren H O
Published: December 2022

Staphylococcal enterotoxin A (SEA) activates human T cells at extremely low concentrations corresponding to 1 to 5 molecules/T cell. SEA, in contrast to other polyclonal activators, is absolutely dependent on accessory cells to interact with and activate T cells. Only cells that can bind SEA can act as accessory cells. Monocytes, B cells, and B lymphomas have these properties. Cells of the EBV-transformed B lymphoma Raji are particularly efficient as accessory cells. T cells do not bind soluble SEA. Probably the SEA molecule has to be attached to the accessory cell to become mitogenic. It is presently not known whether the T cell recognizes a molecular complex between the SEA molecule and the putative receptor moiety on the accessory cell or the SEA molecule itself modified by its binding to the accessory cell.
Thy-1- and Ly-6-mediated lymphokine production and growth inhibition of a T cell hybridoma require co-expression of the T cell antigen receptor complex.
Authors: Sussman J J , Saito T , Shevach E M , Germain R N , Ashwell J D
Published: December 2022

Repetitive subcloning of an Ag-specific T cell hybridoma yielded a variant that lacked functional mRNA for the Ag receptor (Ti, T cell Ag receptor alpha/beta heterodimer) beta-chains and failed to express CD3/Ti on the cell surface. Transfection with the original Ti alpha- and beta-chain genes restored CD3/Ti expression to normal levels. Whereas the parental T cell hybridoma produced IL-2 when stimulated with mAb against CD3, Thy-1, and Ly-6, the CD3/Ti negative cell failed to do so. Reconstitution of CD3/Ti expression restored normal IL-2 production in response to these mAb. A separate response to activation, the inhibition of transformed growth, was also dependent upon co-expression of CD3/Ti. These data demonstrate that cell surface expression of CD3/Ti is required for IL-2 production and growth inhibition initiated by two distinct activating molecules, and suggest that CD3/Ti may be a final common pathway for many transmembrane activation signals.