bluuebunny/crossref_metadata_2025 huggingface dataset:
Pharmacokinetics and pharmacodynamics of three different prednisolone prodrugs: effect on circulating lymphocyte subsets and function.
Authors: Frey B M , Walker C , Frey F J , de Weck A L
Published: December 2022

To be pharmacologically active, i.v. prednisolone disodium phosphate (prednisolone phosphate), prednisolone sodium tetrahydrophthalate (prednisolone phthalate), and oral prednisone have to be converted in vivo into prednisolone. The purpose of the present study was to establish whether these agents are functionally equivalent when given in equimolar doses. The peak concentrations of total and unbound prednisolone assessed by HPLC and equilibrium dialysis in five normal volunteers were five to 12 times higher after i.v. prednisolone phosphate than after i.v. prednisolone phthalate or oral prednisone (p less than 0.001). The area under the plasma concentration vs time curves (AUC) of total and unbound prednisolone was higher after i.v. prednisolone phosphate than after oral prednisone or i.v. prednisolone phthalate (p less than 0.001). The immunosuppressive activity of the plasma samples was determined as the percentage inhibition of the mixed lymphocyte reaction, and the area under the inhibition vs time curve of the MLR (AUIC), a measure of immunosuppressive capacity over time, was calculated. After i.v. prednisolone phthalate or oral prednisone, the mean AUC of unbound prednisolone were 45 and 61% of that obtained after i.v. prednisolone phosphate, and the corresponding AUIC were 76 and 68%, respectively, indicating lower systemic availability of prednisolone and less of an immunosuppressive effect in plasma after i.v. prednisolone phthalate or oral prednisone than after i.v. prednisolone phosphate. These differences in biologic activity of the plasma were corroborated by determining ex vivo the cell cycle progression within the G1 phase, and the interleukin 2 (IL 2) release after the activation of peripheral lymphocytes obtained at various time points after administration of the three steroids. After steroid doses corresponding to 1 mg/kg of prednisolone, a transient inhibition of 80% of IL 2 production was observed with all three glucocorticoids given, whereas the maximal inhibition of the IL 2 production was 60, 30, and 22% after a low dose (0.1 mg/kg) of prednisolone phosphate, prednisone, or prednisolone phthalate, respectively. After the low dose, only minimal changes in the percentages of the OKT4+ and OKT8+ cells were detectable, whereas after the high dose of all three steroids the ratio of OKT4+ to the OKT8+ cells dropped, the decline being more pronounced after prednisolone phosphate than after prednisolone phthalate or prednisone.(ABSTRACT TRUNCATED AT 400 WORDS)
Complete heavy and light chain variable region sequence of anti-arsonate monoclonal antibodies from BALB/c and A/J mice sharing the 36-60 idiotype are highly homologous.
Authors: Juszczak E , Near R I , Gefter M L , Margolies M N
Published: December 2022

Structural and serologic studies on murine A/J monoclonal anti-arsonate antibodies resulted in the identification of a second idiotype family (Id36-60) in addition to the predominant idiotype family (IdCR). Id36-60, unlike IdCR, is a dominant idiotype in the BALB/c strain but is a "minor" idiotype in the A/J strain. The complete heavy and light chain variable region (VH and VL) amino acid sequences of a representative Id36-60 hybridoma protein from both the A/J and BALB/c strains have been determined. There are only four amino acid sequence differences between the VH of antibody 36-60 (A/J) and antibody 1210.7 (BALB/c). Two of these differences arise from single nucleotide changes in which the A/J and BALB/c Id36-60 VH germline gene sequences differ. The two other differences are the result of somatic mutation in hybridoma protein 36-60. In addition, Id36-60 heavy chains employ the same D and JH3 segments in both strains. The entire Vk2 VL of 36-60 and 1210.7 differ by only two amino acids, suggesting that like the heavy chains, they are derived from highly homologous VL genes. The same Jk segment is used in both antibodies. A comparison of the amino acid sequence data from Id36-60-bearing hybridomas suggests that a heavy chain amino acid difference accounts for the diminished arsonate binding by the 1210.7 hybridoma protein. Because the 1210.7 heavy chain is the unmutated product of the BALB/c VH gene, somatic mutation in VH may be required to enhance Ars affinity in this system.
Prostaglandin-sensitive adenylate cyclase of a murine macrophage-like cell line (P388D1): II. Isolation and partial characterization of PGE2-binding proteins.
Authors: Fernandez-Botran R , Suzuki T
Published: December 2022

Properties of prostaglandin (PG) E2 binding sites of a murine macrophage cell line (P388D1) were investigated. The specific binding of [3H]-PGE2 to intact P388D1 cells at 4 degrees C in the presence of cytochalasin D (10 micrograms/ml) approached saturation at concentration greater than 7.5 X 10(-9) M, and could be displaced most effectively by unlabeled PGE2 and less effectively by unlabeled PGI2. The Scatchard analysis of the binding data clearly indicated the heterogeneity with respect to the PGE2 binding affinity and showed the presence of about 3.9 fmol/10(8) cells of the high affinity sites (KD = 1.1 X 10(-9) M) and about 24 fmol/10(8) cells of the low affinity sites (KD = 2 X 10(-8) M). PGE2-binding proteins were isolated from the detergent lysate of the radiolabeled P388D1 cells by affinity chromatography on Sepharose 4B coupled to PGE2. The affinity-isolated materials were further purified by successive use of Sephadex G-100 gel filtration and isoelectric focusing in the presence of dithiothreitol (1 mM) and Triton X-100 (0.5%). The final step yielded about 0.25% of the original radioactivity, which sharply focused as a single peak at pH near 6.5. The electrofocused PGE2-binding proteins migrated as a single band with a m.w. of 95,000 during SDS-PAGE. The electrofocused PGE2-binding proteins bound specifically to [3H]-PGE2 but showed again the heterogeneity with respect to their affinity.
The modulation of expression of polypeptide surface antigens on developing schistosomula of Schistosoma mansoni.
Authors: Simpson A J , Payares G , Walker T , Knight M , Smithers S R
Published: December 2022

Five low m.w. polypeptide antigens are expressed on the surface of freshly transformed schistosomula of Schistosoma mansoni, and were reproducibly identified by surface labeling with 125I by using IODOGEN and immunoprecipitating with immune mouse sera. These molecules have approximate m.w. of 38,000, 32,000, 20,000, 17,000, and 15,000. They correspond to antigens recognized previously by lactoperoxidase-catalyzed iodination. Analysis of the surface of developing schistosomulum demonstrated that the 38,000 and 17,000 dalton antigens were lost from the parasite surface during 48 hr of in vitro culture. This process was not dependent on the presence of host serum. The two antigens were not lost due to shedding into the culture medium but were apparently sequestered to a site where they were no longer available for surface labeling. The 32,000, 20,000, and 15,000 dalton antigens, however, remained exposed on the schistosomulum surface for up to 2 days of in vitro culture. The expression of two new antigens was also induced by culture in vitro: a doublet of approximately 45,000 daltons and an antigen of approximately 11,000 daltons. The expression of the former was dependent on the presence of serum. These results demonstrate that the development of the schistosomula surface is a complex process, with events both dependent and independent of the presence of serum. In addition, the expression of polypeptide antigens is not coordinated, and antigens are lost, retained, or appear on the schistosomulum surface during the early stages of maturation.
Syngeneic responses by murine thymocytes: a role for non-MHC and non-MLS genes.
Authors: Wu S , Thomas D W
Published: December 2022

A subpopulation of thymocytes from adult mice that is nonadherent to macrophage monolayers showed dramatically increased syngeneic mixed leukocyte responses (SMLR). Cloned cells were derived by limiting dilution from these SMLR-primed BALB/c thymocytes and were maintained and subcloned by repeated stimulation with syngeneic BALB/c spleen cells without the addition of exogenous interleukins. The cloned thymocytes were tested for their reactivities against H-2- and Mls-identical BALB/c and B10.D2 spleen cells (H-2d, Mlsb). We found that BALB/c and B10.D2 stimulator cells differed significantly in their capacity to restimulate the cloned BALB/c thymocytes. In addition, polyclonal syngeneic mixed leukocyte cultures (SMLC) of BALB/c thymocytes also showed differential restimulation by BALB/c and B10.D2 stimulators. Taken together, our data indicate a role for the product(s) specified by non-MHC and non-Mls gene(s) in the autorecognition by thymocytes.
Immunoregulatory pathways in adult responder mice. II. Regulation of delayed-type hypersensitivity responses by GAT-specific suppressor factors present in GAT-tolerant adult responder mice.
Authors: Jenkins M K , Waltenbaugh C , Miller S D
Published: December 2022

We studied the effects of T cell extracts from adult responder BALB/c mice tolerized with poly(Glu60Ala30Tyr10) (GAT)-coupled syngeneic spleen cells (GAT-SP) on delayed-type hypersensitivity (DTH), T cell-proliferative (Tprlf), and plaque-forming cell (PFC) responses. Adult responder mice injected i.v. with GAT-SP develop Lyt-1-2+ suppressor T cells (Ts), which suppress the induction of GAT-specific DTH and PFC, but not Tprlf responses. Sonicates from these Ts contain an afferent-acting, soluble factor(s) (GAT-TsFdh) that specifically suppresses the same responses as the intact Ts (i.e., DTH and PFC, but not Tprlf). Immunosorbent chromatography studies were employed to determine the molecular nature of the suppressive material active on both cellular and humoral responses. In both assay systems, GAT-TsFdh was found to bear determinants encoded by the I subregion of the H-2 complex and a receptor(s) for GAT. BALB/c-derived GAT-TsFdh suppressed the induction of GAT DTH in syngeneic BALB/c and H-2-compatible B10.D2, but not in allogeneic C57BL/6 or CBA/Cum, suggesting a possible H-2 restriction in the suppression. It was also shown that one target of functional regulation by GAT-TsFdh is the T helper cell for DTH responses (DTH-Th). The results suggest that similar Ts and TsF regulate humoral and cell-mediated responses, perhaps by affecting a target common to both pathways (e.g., the T helper cell). The resistance of Tprlf responses to suppression by GAT-TsFdh indicates that the effector DTH-Th target is not a major component of the proliferative response. These data are discussed with respect to GAT-specific TsF-regulating PFC responses, which have been identified in nonresponders and in responders tolerized as neonates with GAT.
Defective IFN-gamma production in the human neonate. I. Dysregulation rather than intrinsic abnormality.
Authors: Wakasugi N , Virelizier J L
Published: December 2022

Cord blood leukocytes (CBL) stimulated with PHA, Con A, or with the monoclonal antibody OKT3 proliferate normally but produce very low titers of IFN-gamma. This defect was not observed with maternal leukocytes collected at the time of delivery, indicating that the defective production of IFN-gamma in CBL is not a mere consequence of a hormonal change associated with labor. CBL produced large amounts of IFN-gamma (comparable to those observed in adult control and in mothers) after stimulation with staphylococcal enterotoxin A (SEA). Furthermore, gamma-irradiation with as little as 500 or 1000 rad, or incubation at 37 degrees C for 24 hr, reversed the defect in PHA-induced IFN-gamma secretion. This finding indicates that the defective secretion of IFN-gamma of CBL is not intrinsic, but rather is the consequence of a subtle dysregulation. We could not find evidence for a defective accessory function with cord blood monocytes, because the addition of adherent cells from adult donors did not reverse the defect. In co-cultures of adult leukocytes and CBL, PHA-induced IFN-gamma secretion was comparable to that of adult cultures tested alone. Nonirradiated CBL were not able to suppress IFN-gamma secretion by irradiated autologous leukocytes. Together, our results suggest that the defective PHA-induced IFN-gamma secretion of CBL is the result of an original type of dysregulation and is associated with an excessive sensitivity to suppressive signals rather than excessive suppressor function.
Human immune responses to synthetic peptides from the Epstein-Barr nuclear antigen.
Authors: Rhodes G , Carson D A , Valbracht J , Houghten R , Vaughan J H
Published: December 2022

Humans infected with Epstein-Barr virus (EBV), the causative agent of infectious mononucleosis, develop antibodies against a nuclear antigen (EBNA) that is present in virally transformed B lymphocytes. The EBNA protein contains a unique glycine-alanine repeating sequence. We have synthesized peptides corresponding to various regions of the EBNA molecule within and near this sequence. Rabbit antibodies against the peptides within the sequence reacted directly with the EBNA protein, as detected by Western blotting. The sera of individuals with antibodies against Epstein-Barr virus contained abundant antibodies also reactive with one or several of the synthetic peptides within the sequence. Moreover, human antibodies against these simple peptides were induced specifically early in the course of infectious mononucleosis. When compared with normal controls, antibody levels to the glycine-alanine peptides were significantly higher in patients with rheumatoid arthritis and progressive systemic sclerosis, but not in patients with two other autoimmune diseases. These results document that i) antibodies against the peptides detect the EBNA protein, ii) humans infected with EBV produce high titers of antibodies reactive with these synthetic antigens, and iii) antibody titers against the peptides are abnormally elevated in certain autoimmune diseases.
Functional distinctions between the LFA-1, LFA-2, and LFA-3 membrane proteins on human CTL are revealed with trypsin-pretreated target cells.
Authors: Gromkowski S H , Krensky A M , Martz E , Burakoff S J
Published: December 2022

We asked whether we could distinguish the roles of the human lymphocyte membrane proteins LFA-1, LFA-2, and LFA-3 in the function of CTL-mediated killing. Little is known about the functions of these molecularly distinct proteins beyond the facts that i) binding of a monoclonal antibody (MAb) to any one of them is sufficient to inhibit killing, ii) that in each case inhibition involves prevention of CTL-target cell conjugate formation, and iii) that MAb to LFA-1 and LFA-2 inhibit best when bound to the CTL, whereas anti-LFA-3 inhibits only when bound to the target cell. This latter is despite the fact that (in our test system) LFA-1 and LFA-3 are expressed both on the CTL and on the target. When the target cells were pretreated with trypsin, the sensitivity of CTL-mediated killing was affected in a different way for each site. Inhibition of anti-LFA-1 was increased by approximately 20-fold. Inhibition by anti-LFA-2 was unaffected. Inhibition by anti-LFA-3 was abolished. Trypsin did not remove the specific antigens recognized by the various CTL, HLA-A,B,C or HLA-DR. Nor did it remove LFA-1 from the target cell. It did, however, selectively remove LFA-3 from the target cell. These results indicate, for the first time, that LFA-1 and LFA-2 have functionally distinct roles. They suggest that an unidentified trypsin-sensitive target cell molecule, operationally designated the "trypsin-sensitive counter blocker" (TSCB), plays an important role in the function of LFA-1, possibly by providing a target cell binding site for LFA-1 on the CTL. The hypothesis that this TSCB is identical to LFA-3 (and the related possibility that LFA-1 and LFA-3 are mutual ligands) is not favored by our data, but is not excluded. Finally, the data indicate that the mechanisms by which MAb inhibit killing differ at the LFA-1 and LFA-3 sites. They are consistent with LFA-1 providing adhesion strengthening by binding to another site (the TSCB?) and with LFA-3 delivering an inhibitory signal when provoked with MAb.
OKT3 monoclonal antibody induces production of colony-stimulating factor(s) for granulocytes and macrophages in cultures of human T lymphocytes and adherent cells.
Authors: Platzer E , Rubin B Y , Lu L , Welte K , Broxmeyer H E , Moore M A
Published: December 2022

OKT3 monoclonal antibody (mab) recognizes a membrane antigen associated with the T cell antigen recognition receptor, and is known to be mitogenic and to induce lymphokine production. Our studies demonstrate the ability of OKT3 mab to induce from cultures of human T lymphocytes supplemented with adherent cells the production of colony-stimulating factor(s) for granulocytes and macrophages (GM-CSF) and interferon-gamma (IFN-gamma), an inhibitor of clonal growth of hematopoietic progenitor cells. As has been shown for the mitogenic and IFN-gamma-inducing activity of OKT3 mab, the induction of GM-CSF release in cultures of T cells is strictly dependent on the presence of adherent cells. However, the concentrations of OKT3 mab required for optimal GM-CSF production (50 ng/ml) were found to be 80-fold higher than those sufficient for maximal IFN-gamma production, proliferation, and interleukin 2 production. IFN-gamma activity induced by OKT3 mab partially inhibited colony and cluster formation from progenitor cells of granulocytes and macrophages in vitro. Therefore, neutralization of the IFN-gamma by monoclonal anti-human-IFN-gamma antibody before assay of conditioned medium in bone marrow cultures significantly enhanced the detection of GM-CSF. Kinetic studies demonstrated maximal cumulative GM-CSF production in response to optimal OKT3 mab concentrations on days 4 through 6 in cultures of T cells supplemented with 15% adherent cells. Highly enriched OKT4+ and OKT8+ T cell subsets co-cultured with adherent cells in the presence of OKT3 mab both produced GM-CSF and IFN-gamma and showed similar dose-response curves to OKT3 mab. The requirement for the presence of adherent cells could not be overcome by the addition of purified interleukin 1 or macrophage supernatants. Studies using irreversible inhibitors of DNA (mitomycin C) or protein biosynthesis (emetine-HCl) revealed the necessity of intact DNA synthesis and translation in mononuclear cells to produce GM-CSF in response to OKT3 mab. Loss of GM-CSF production was observed when either adherent cells or T lymphocytes were treated with emetine before co-culture with untreated cells of the other population in the presence of OKT3 mab. In contrast, mitomycin C reduced GM-CSF production significantly when T cells, but not adherent cells, were pretreated. These results suggest that T lymphocytes and adherent cells closely cooperate in the production of GM-CSF induced by OKT3 mab.
Cytofluorometric isolation of I937, an Ia antigen-bearing variant of the Ia-negative human monocytic cell line U937.
Authors: Gitter B D , Finn O J , Metzgar R S
Published: December 2022

Cells of the human monocyte cell line U937 are generally considered devoid of any Ia antigens on their surface. In analyzing U937 cells with a large panel of monoclonal anti-human Ia antibodies by flow cytometry, we detected a small number of cells that appeared to react with antibodies to HLA-DR and HLA-DS/DC molecules. These Ia-positive cells were isolated and were cloned, resulting in a human monocyte cell line that expresses high levels of Ia antigens. We analyzed these antigens by one- and two-dimensional polyacrylamide gel electrophoresis, after radiolabeling and immunoprecipitation. Three distinct Ia molecules, alpha 1 beta 1, alpha 1 beta 3 (HLA-DR-like), and alpha 2 beta 2 (HLA-DC/DS-like) are synthesized by I937 cells, alpha 1 beta 3 molecule being the predominant species. The Ia antigen-bearing human monocyte cell line is expected to be useful for studying events involved in antigen presentation.
Reconstitution of nude mouse T cell function in vivo: IL 2-independent effect of human T cells.
Authors: Dosch H M , White D , Grant C
Published: December 2022

The i.p. injection of 1 to 5 X 10(6) heavily irradiated human T lymphocytes resulted in the lasting reconstitution of T cell functions in young mice bearing the nu/nu mutation. IgM and IgG responses to immunization with sheep red cells or ovalbumin, splenic lectin responses, and the expression of easily detectable Thy-1 determinants on up to 20% of spleen cells could be documented for several months after the injection of human cells. Only a narrow cell dose range was effective. Injection of larger cell numbers not only failed to induce immune reconstitution but also resulted in the development of resistance to subsequent treatments. Only mature T cells, but not thymocytes, could induce nude mouse T cell development. Lymphoblasts from one patient with acute T cell leukemia consistently immune-reconstituted nude recipients. These cells were completely unable to produce IL 2 in vitro. In contrast, the IL 2-producing T cell line Jurkat was ineffective, indicating that the abilities to produce IL 2 and to induce nude mouse T cell development are independent. In an extension of earlier models of the nude mouse immune defect, two distinct T precursor cell pools are proposed as the major components of an extrathymic differentiation pathway. As an adequate trigger of differentiation, interaction with thymus-processed T cells guides the development of precursors in the first cell pool towards populating the second IL 2-responsive pool of T precursor cells.
Role of interleukin 1 in experimental pulmonary granuloma in mice.
Authors: Kobayashi K , Allred C , Cohen S , Yoshida T
Published: December 2022

Pulmonary granulomas were induced in immunized BALB/c mice by the intratracheal injection of antigen-coated and plain agarose beads. Prominent lesions developed within 24 hr, reached peak intensity within 3 days, and gradually declined in size thereafter. The hypersensitivity granulomas induced in sensitized mice by antigen-coated beads were much larger than the lesions induced by plain beads. Minimal inflammation was produced in unsensitized mice injected with antigen-coated or plain beads. Aqueous extracts prepared from pulmonary granuloma lesions induced in sensitized mice by antigen-coated beads contained high levels of interleukin 1 (IL 1) and migration inhibition factor (MIF) activities. The kinetics of appearance of these mediators were similar. Lower but detectable activity of both mediators was detected in extracts prepared from sensitized mice injected with plain beads. Neither interleukin 2 (IL 2) nor IL 2 neutralizing activities were detected in the extracts. The presence of IL 1 and MIF in extracts prepared from early and peak pulmonary granulomatous lesions suggests that these soluble factors are produced by cells within the lesions, and that they are involved in mediating the expression and/or maintenance of the granulomas.
Strain dependence of muramyl dipeptide-induced LAF(IL 1) release by murine-adherent peritoneal cells.
Authors: Abehsira-Amar O , Damais C , Parant M , Chedid L
Published: December 2022

The capacity of MDP to stimulate LAF(IL 1) production by adherent peritoneal cells (APC) from different strains of mice was examined. It was observed that MDP could stimulate thioglycollate-induced APC from DBA/2, C57B1/6, and CBA/CA mice, but not from C3H mice. Resident APC from DBA/2 or C3H/HeJ mice were also responders and nonresponders, respectively, to MDP for LAF(IL 1) production, the positive effect of MDP being more marked on DBA/2 cells in the presence of indomethacin. Because a LPS high-responder C3H substrain (C3HeB/Fe) did not respond to MDP, it was concluded that the strain dependence with respect to the induction of LAF(IL 1) seen with both MDP and LPS was not linked. Moreover, the unresponsiveness of C3H mice to MDP was not linked to their MHC haplotype, because a second H-2k strain (CBA/CA) was a good responder to MDP stimulation.
A "lymphokine-like" soluble product that induces proliferation and maturation of B cells appears in the serum-free supernatant of a T cell hybridoma as a consequence of mycoplasmal contamination.
Authors: Proust J J , Buchholz M A , Nordin A A
Published: December 2022

The serum-free supernatant of a cloned murine T cell hybridoma supports the proliferation and maturation to Ig secretion of purified B cells (mu+ cells) from BALB/c nu/nu mice, but has no effect on the proliferation of nylon wool-selected BALB/c nu/+ splenic T cells. Although the supernatant activates B cells without co-stimulation, it synergizes with anti-mu for the proliferative response. The induction of B cell proliferation and maturation to Ig secretion is directly related to contamination of the hybridoma by Mycoplasma hyorhinis. Hybridoma cells freed of mycoplasma by detergent treatment fail to produce active supernatant, and reinfection of the treated cells reconstitutes the activity. Furthermore, deliberate infection of a mycoplasma-free unrelated T cell hybridoma, as well as the monocytic cell line P388D1, results in the production of supernatants with B cell proliferating activity. Mycoplasma organisms isolated from the supernatant induce B cell proliferation without subsequent maturation to Ig secretion. Gel filtration chromatography of the supernatant from mycoplasma-contaminated hybridoma cells yields two peaks of activity. The first peak, found at the exclusion limit of the gel, results in B cell proliferation without maturation and may be attributed to mycoplasma organisms. The second peak (average m.w. 90,000) results in B cell proliferation as well as differentiation to Ig secretion. A "lymphokine-like" soluble product released by Mycoplasma hyorhinis is most likely responsible for this B cell activation, because fractionation of the supernatant from deliberately contaminated P388D1 cells gives essentially the same results, and gel filtration of mycoplasma-free supernatants does not generate any active fractions. The possibility should be considered that mycoplasma-derived soluble products may be among the many factors controlling in vitro B cell growth and maturation.
Differential glycosylation requirements for the cell surface expression of class I molecules.
Authors: Landolfi N F , Rich R R , Cook R G
Published: December 2022

The importance of asparagine-linked glycosylation in the cell surface expression of several class I molecules was examined. C57BL/6 (B6) T cell blasts were treated with tunicamycin (TM), an antibiotic that inhibits N-linked glycosylation. The levels of various class I molecules on these cells were examined by flow cytometry and were compared to the levels of the same molecules on untreated cells. A 12-hr TM treatment did not significantly alter the levels of H-2Kb, Db, or Qa-2; however, such treatment decreased the surface expression of the Qa-1b allelic product to undetectable levels. A time-course study indicated that a decrease in the level of Qa-1.2 expression was apparent after only 4 hr of TM treatment. An examination of T cell blasts prepared from mouse strains possessing the Qa-1a, Qa-1c, and Qa-1d alleles indicated that all allelic products of this locus demonstrated a marked decrease in cell surface expression on TM treatment, whereas other class I molecules (H-2Ks, TL) exhibited slight or no decrease. Two-dimensional polyacrylamide gel electrophoresis analysis of immunoprecipitates from detergent lysates of surface-iodinated TM-treated B6 blasts revealed the presence of the unglycosylated form of the H-2Kb molecule on the cell surface. No such form of the Qa-1.2 molecule could be detected by similar analysis. To establish that the above observations were not simply a result of the inability of the Qa-1-specific alloantisera to react with the unglycosylated Qa-1 molecule, lysates of surface-iodinated B6 blasts were digested with endoglycosidase F, which cleaves N-linked carbohydrate moieties. Immunoprecipitation analysis indicated that the antisera could react with the unglycosylated form of the Qa-1 molecule. These results indicate that N-linked glycosylation has differential importance in the cell surface expression of class I molecules.
Resistance to phagocytosis by group A streptococci: failure of deposited complement opsonins to interact with cellular receptors.
Authors: Weis J J , Law S K , Levine R P , Cleary P P
Published: December 2022

The previous finding that phagocytosis-resistant M+ group A streptococci bear quantities of C3 which are sufficient for phagocytosis of their M- derivatives was investigated at two levels. It was first established that the C3 associated with M+ streptococci was not able to promote adherence to cells bearing the complement receptors CR1 and CR3 under conditions in which M- streptococci readily attached. The molecular form of C3 bound to M+ and M- streptococci was then defined by adding 125I-C3 to serum used for opsonization. C3 eluted from the bacteria by chaotropic and hydrolytic agents was analyzed by SDS-PAGE, and revealed that both cell types bound the opsonic forms of C3, C3b, and iC3b. Furthermore, approximately 80% of the C3b and iC3b associated with both cell types was covalently bound to a surface component, although most of the C3 bound to M+ streptococci was detergent-extractable, whereas greater than 50% of that bound to M- streptococci was not. These findings demonstrate that the M+ surface is interfering with the receptor binding of deposited C3b and iC3b, and that this contributes to resistance to phagocytosis by these organisms.
The murine lymphocyte receptor for IgE. III. Use of chemical cross-linking reagents to further characterize the B lymphocyte Fc epsilon receptor.
Authors: Lee W T , Conrad D H
Published: December 2022

Cross-linking reagents were used to further characterize the murine B cell receptor for the Fc portion of IgE (Fc epsilon R) and compare this receptor to the well-characterized high-affinity Fc epsilon R on rat basophilic leukemia (RBL) cells. The disulfide cleavable and noncleavable reagents 3,3'-dithiobis(sulfosuccinimidyl) propionate (DTSSP) and bis(sulfosuccinimidyl) suberate (BS3) were used. With these reagents, efficient cross-linking of the alpha component of the high-affinity RBL Fc epsilon R to the membrane-buried beta and gamma components occurred only if the membrane was solubilized before the cross-linking reaction. In studies with purified murine B cells, IgE could be cross-linked to the Fc epsilon R on intact cells with either DTSSP or BS3. Under the same conditions, up to 10% of the B cell surface immunoglobulin (sIg) (both IgM and IgD) was also found to cross-link to a portion of the IgE/Fc epsilon R complex, suggesting that on the intact murine B cell the Fc epsilon R is frequently in close association with sIg. The B cell Fc epsilon R was also examined for the presence of receptor-associated proteins. Under conditions where the high-affinity RBL Fc epsilon R was substantially cross-linked to the alpha, beta, gamma complex, no evidence was seen for similar cross-linking of the B cell Fc epsilon R. Cross-linking experiments on affinity-purified Fc epsilon R preparations also gave no evidence for receptor-associated proteins with the B cell Fc epsilon R, although evidence for receptor-receptor association was seen. Thus, these data further support the concept that there may be little relationship between the high-affinity mast cell/basophil Fc epsilon R and the low-affinity lymphocyte Fc epsilon R.
Arachidonic acid metabolism by murine peritoneal macrophages infected with Leishmania donovani: in vitro evidence for parasite-induced alterations in cyclooxygenase and lipoxygenase pathways.
Authors: Reiner N E , Malemud C J
Published: December 2022

Leishmania donovani is an obligate intracellular protozoan that resides within mononuclear phagocytes of infected mammals. Affected human and rodent hosts commonly show abnormalities of T cell function, which may be related to altered macrophage physiology resulting from intracellular parasitism. To examine this possibility, we studied the metabolism of endogenous arachidonyl-phospholipids and [3H]-arachidonyl-phospholipids by murine peritoneal exudate macrophages infected with amastigotes of L. donovani. Our results indicated that infected cells synthesized increased amounts of both cyclooxygenase and lipoxygenase metabolites of arachidonic acid. Increased synthesis of immunoreactive prostaglandin (PG)E2 was evident as early as 1 to 4 hr after infection, was correlated with the fraction of cells infected, and was inhibited by sodium meclofenamate (0.2 and 20 microM) but not nordihydroguaiaretic acid (3 microM). As determined by thin-layer chromatography, infected cells also produced markedly increased amounts of prostaglandin F2 alpha (also inhibited by sodium meclofenamate) with insignificant increases in thromboxane B2 and the stable metabolite of prostacyclin, 6-oxo-PGF1 alpha. In contrast, stimulation of cells with opsonized zymosan resulted in significantly increased synthesis of all four eicosanoids. L. donovani infection was also found to induce marked increases in synthesis of lipoxygenase metabolites of arachidonic acid by infected cells. This was evidenced by increased amounts of [3H]-labeled material in cell extracts that co-migrated with authentic standards of 5 and 12/15-hydroxy-eicosate-traenoic acids in thin-layer chromatograms. Increased synthesis of these products was largely inhibited by both NDGA (3 microM) and sodium meclofenamate (20 and 0.2 microM). Additional evidence for augmentation of 5-lipoxygenase by Leishmania was provided by the demonstration of increased leukotriene-C4 in conditioned medium from infected cells. These results indicate that macrophages infected with L. donovani produce increased amounts of arachidonic acid metabolites with the potential for influencing cellular immune function and the inflammatory response to infection.
Nerve growth factor: a chemotactic factor for polymorphonuclear leukocytes in vivo.
Authors: Boyle M D , Lawman M J , Gee A P , Young M
Published: December 2022

Previous studies have shown that mouse submandibular gland nerve growth factor (NGF) stimulates chemotactic migration of human polymorphonuclear leukocytes in vitro. The results of the present study demonstrate that subdermal injection of NGF in mice also stimulates rapid and marked chemotactic recruitment of leukocytes. This property of NGF is manifest in the nanomolar range of concentrations, it requires the known serine class protease activity of the growth factor, and it does not require participation of the fifth component of complement. Another, as yet unrecognized, C5-independent pathway must be involved. Chemotactic stimulation of cells involved in the early inflammatory response to injury may help to explain earlier observations that NGF can accelerate the rate of contraction of experimentally induced wounds in mice.
H-2K-restricted granuloma formation by Ly-2+ T cells in antibacterial protection to facultative intracellular bacteria.
Authors: Näher H , Sperling U , Hahn H
Published: December 2022

Cellular, genetic, and antigenic requirements for granuloma formation in murine listeriosis were determined by using adoptive transfer of granuloma formation. Granuloma formation was restricted by a class I MHC antigen (H-2K) and critically depended on a Ly-2+ (Ly-1+2+) T cell. Expression of granuloma formation required living bacteria; heat-killed bacteria was not sufficient. H-2K-restricted transfer of granuloma formation was associated with a high degree of protection. Markedly less protection, presumably due to macrophage activation by T cells, was found under conditions of H-2 I-A homology. It is concluded that two T cell populations are involved in protection against L. monocytogenes: protection associated with granuloma formation depends on Ly-2+ (Ly-1+2+) T cells, is restricted by H-2K, and requires products of living bacteria to be expressed, whereas protection based on macrophage activation depends on H-2 I-A-restricted T helper cells.
Natural killer cells vs cytotoxic T cells in the peripheral blood of virus-infected mice.
Authors: Stitz L , Althage A , Hengartner H , Zinkernagel R
Published: December 2022

The cell-mediated immune response of mice against various enveloped RNA and DNA viruses expressed by immune lymphocytes from the spleen and the peripheral blood (PBL) were compared. PBL from mice of various strains infected with vaccinia virus, vesicular stomatitis virus (VSV), or lymphocytic choriomeningitis virus (LCMV) were tested on histocompatible or incompatible target cells infected with the homologous virus. PBL from immune mice showed clear H-2 restriction, but additionally, they expressed high natural killing (NK) activity on YAC-1 cells. The high NK-cytolytic activity of PBL on YAC-1 differed significantly from that expressed by splenic lymphocytes. In both lymphocyte populations lysis was detected as early as 1 day after infection; NK activity decreased in the spleen after day 4 post infection, whereas that of PBL persisted at high levels for up to 10 days after infection. Treatment of mice with anti-asialo GM1 in vivo abrogated NK activity in PBL effector cells tested in vitro. These results may explain some of the difficulties to observe MHC-restricted cytotoxic T cells in PBL from humans or primates during primary infections with virus.
Transcriptional regulation of HLA class II and invariant chain genes.
Authors: Levine F , Erlich H A , Mach B , Pious D
Published: December 2022

Class II (Ia) antigens are coded for by a family of genes located in the human MHC (HLA). These genes are regulated in a complex manner, being constitutively expressed, inducibly expressed, or not expressed, depending on the cell type examined. 6.1.6 is a variant of a normal B lymphoblastoid line that has lost expression of all class II molecules and has previously been shown to have a defect in the regulation of class II genes. In this report, we have examined those genes by Southern and Northern blotting and have found that 6.1.6 is severely deficient in mRNA for all class II genes examined, although the genes are structurally intact. P30, a partial revertant of 6.1.6, re-expresses mRNA for a subset of class II genes. mRNA for the class II-associated invariant chain is substantially reduced but not absent in 6.1.6.
Dissociation of inductive from differentiative signals transmitted by C8-substituted guanine ribonucleosides to B cells from SJL mice.
Authors: Goodman M G , Weigle W O
Published: December 2022

A hitherto unknown defect in the immune responsiveness of B lymphocytes from SJL mice has enabled us to distinguish two qualitatively distinct classes of signal delivered to B cells by C8-substituted guanine ribonucleosides. This defect renders B cells from SJL mice unresponsive to the inductive (early acting) signal of 8-mercaptoguanosine (8MGuo) that culminates in mitogenesis and nonspecific secretion of immunoglobulin. Unresponsiveness is not attributable to a shift in either the dose-response or kinetic profiles, nor can the presence of suppressor cells be demonstrated. In striking contrast, however, SJL B cells exhibit normal responsiveness to the differentiative (T cell-like, or late acting) signal provided by the substituted nucleoside. This signal enables SJL B cells, depleted of T cells, to respond to T cell-dependent antigens, and synergizes with T cell-derived lymphokines. These data suggest 1) that nonspecific secretion of immunoglobulin is dependent on both inductive and differentiative signals, 2) that antigen alone can supply an effective inductive signal for antigen-specific responses, and 3) that the SJL mouse will provide a useful model for selective study of inductive vs differentiative events.
A monoclonal antibody-based immunoradiometric assay for detection of circulating antigen in Bancroftian filariasis.
Authors: Forsyth K P , Spark R , Kazura J , Brown G V , Peters P , Heywood P , Dissanayake S , Mitchell G F
Published: December 2022

A monoclonal antibody designated Gib 13 has been used in an immunoradiometric assay (IRMA) to detect circulating antigen in the sera of Wuchereria bancrofti-infected subjects from an endemic area of Papua New Guinea. A clear association between the presence of patent infection and the Gib 13 target epitope in serum was established because 93% of microfilaremic individuals were antigen-positive. Moreover, there was a significant correlation between levels of serum antigen and blood microfilarial counts. Detection of circulating antigen in amicrofilaremic subjects with acute symptoms of lymphatic filariasis, and 53% of asymptomatic amicrofilaremic subjects, but not in nonendemic controls, suggests that the Gib 13 IRMA will also be of value in the diagnosis of occult filariasis. However, as in all IRMA based on detection of potentially immunogenic molecules in man, antibodies can be expected to be the major contributor to reduced sensitivity of the assay.
Monoclonal antibodies to parasite antigens found in the serum of Dirofilaria immitis-infected dogs.
Authors: Weil G J , Malane M S , Powers K G , Blair L S
Published: December 2022

We recently reported that parasite antigens are detectable in the serum of Dirofilaria immitis-infected dogs by counterimmunoelectrophoresis (CIE). Hybridoma cell lines that produce monoclonal antibodies specific for these antigens were obtained by immunizing mice with a partially purified antigen preparation, fusing spleen cells with SP-2 myeloma cells, and screening cell culture supernatants for antibody by ELISA and CIE inhibition. Antibodies specific for two epitopes shared by the two major circulating parasite antigens were identified. Immunoperoxidase studies showed that the epitopes recognized by the monoclonals were widely distributed in D. immitis, but the female uterus and eggs were particularly strongly labeled. A monoclonal antibody-based ELISA was developed to measure parasite antigens in dog sera. Parasite antigens were detected in 45 of 46 sera from infected dogs but were absent in sera from uninfected dogs and sera from dogs infected with Dipetalonema reconditum. Serum antigen content was significantly correlated with the number of female worms recovered from infected dogs (r = 0.82, p less than 0.001). Antigenemia was first detected 6 mo after infection, and antigen levels remained fairly stable between 9 and 21 mo after infection. Parasite antigen detection with this monoclonal antibody-based ELISA appears to be superior to previously described diagnostic methods for canine dirofilariasis in terms of sensitivity, specificity, and relation to infection intensity.
Immunoprecipitation analysis of radiolabelled protein antigens biosynthesized in vitro by S. mansoni. I. Identification of antigens uniquely recognized by protective antibodies.
Authors: Barker R H , Srivastava B S , Suri P , Goldberg M , Knopf P M
Published: December 2022

Protein antigens from 4-wk worms were metabolically radiolabelled with [3H]leucine or [35S]methionine. Three freeze-thaw cycles released a large proportion (50% to 60%) of the TCA-precipitable radioactivity from the worms. Immune serum from twice-infected Fischer rats (F-2x), which was shown to confer resistance in a passive immunization assay, and immune serum from twice-infected Wistar Furth rats (W-2x), which does not confer resistance, were used for analyzing antigens in this worm fraction. Antibodies in these antisera differed in their titers to the freeze-thaw released antigens (W-2x greater than F-2x) and in their relative affinities for these antigens (F-2x greater than W-2x). Gradient slab gel electrophoresis of immunoprecipitates of radiolabelled antigens under denaturing conditions revealed many components, which could be categorized into two main types: unique antigens, recognized only by F-2x antibodies, and nonunique antigens, recognized by both F-2x and W-2x antibodies. The potential relevance of these antigens in resistance was further examined by antibody absorption experiments in which 4-wk worms were used as an immunoabsorbent to remove 90% to 95% of the immunoprecipitating activity and 65% to 70% (p less than 0.005) of the capacity to confer resistance in a passive immunization assay. It was concluded that loss of both anti-schistosome activities was specific since antigen released by worms during absorption could account for only 16% of the reduction in antigen-binding capacity and the titer of antibodies directed against beta-galactosidase did not significantly change during absorption. Antigens recognized uniquely by F-2x antibodies are therefore candidates for immunization studies examining induction of resistance against Schistosoma mansoni.
Hemagglutination assay of polypeptide coded by the pre-S region of hepatitis B virus DNA with monoclonal antibody: correlation of pre-S polypeptide with the receptor for polymerized human serum albumin in serums containing hepatitis B antigens.
Authors: Okamoto H , Imai M , Usuda S , Tanaka E , Tachibana K , Mishiro S , Machida A , Nakamura T , Miyakawa Y , Mayumi M
Published: December 2022

The receptor for polymerized human as well as chimpanzee serum albumins has been identified on the 55-amino acid polypeptide coded by the pre-S region of hepatitis B virus DNA. Monoclonal antibodies were raised against a synthetic polypeptide of 19 amino acid residues representing a hydrophilic region of the pre-S amino acid sequence deduced from hepatitis B virus DNA. Sheep erythrocytes fixed with glutaraldehyde were coated with monoclonal antibody against the synthetic polypeptide to develop a hemagglutination assay for pre-S polypeptide. The pre-S polypeptide was detected in the serum containing hepatitis B surface antigen particles along with hepatitis B e antigen, with titers in parallel with those of the receptor for polymerized human serum albumin.
Protection and recovery in influenza virus-infected mice immunosuppressed with anti-IgM.
Authors: Kris R M , Asofsky R , Evans C B , Small P A
Published: December 2022

BALB/c mice, immunosuppressed from birth with goat anti-mouse IgM, were able to recover from influenza virus infection in the absence of detectable serum and nasal antibody. Recovery was delayed a few days when compared with control animals. Antibody-deficient mice, that had recovered from an initial influenza virus infection, i.e., convalescent mice, were subsequently rechallenged with homologous influenza virus in order to study the importance of nasal and serum antibody in prevention of infection. Convalescent mice were susceptible to reinfection when nasal and serum antibody were not detectable. The mice were resistant to reinfection when serum and/or nasal antibody was detectable by radioimmunoassay. Normal mice that were passively immunized with high titer mouse anti-influenza virus serum were susceptible to challenge with homologous influenza virus. The serum antibody levels in these mice were higher than most of those found in the immune convalescent mice suppressed with anti-IgM, thereby suggesting that the serum antibody, found in convalescent suppressed mice, is not protective. We conclude that 1) mice can recover from influenza virus infection in the absence of detectable levels of nasal and serum antibody, thus indirectly confirming the role of cell-mediated immunity in recovery; 2) serum IgM, IgG2A, IgG2B, IgG3, and probably IgG1 antibody levels are not responsible for protection against influenza virus infection of the upper respiratory tract; and 3) nasal IgA antibody correlates best with protection against reinfection of the upper respiratory tract, but some other locally protective agent cannot be excluded.