Biomedical subjects

J Van Snick

Publications and source records attributed to J Van Snick.

At least 109 records · Page 6Linked to original sources

PubMed1987

Identification of the human 26-kD protein, interferon beta 2 (IFN-beta 2), as a B cell hybridoma/plasmacytoma growth factor induced by interleukin 1 and tumor necrosis factor.

A factor that promotes the growth of certain B cell hybridomas and of plasmacytomas is shown to be produced by normal human fibroblasts and by a line of human osteosarcoma cells (MG-63) after treatment with IL-1 or TNF. The hybridoma-plasmacytoma growth factor (HPGF) is identified with a 26 kD protein whose mRNA was previously shown to be induced in the same cells by the same inducers. First, poly(A)-rich RNA extracted from IL-1-treated cells could be enriched in HPGF-mRNA content by hybridization to 26 kD cDNA. Second, MG-63-derived HPGF purified to electrophoretic homogeneity was subjected to amino acid sequence analysis, whereby the NH2-terminal sequence was found to match the nucleotide sequence of a 26 kD cDNA clone.

Amino Acid Sequence↗

PubMed1987

IgG2a restriction of murine antibodies elicited by viral infections.

The isotypic distribution of IgG antibodies was determined in the serum of mice after infection with a panel of RNA and DNA viruses representative of 11 different genera. The antiviral response induced by all these viruses showed a striking preponderance of the IgG2a subclass whatever the strain of mice tested or the time elapsed after infection. Together with the predominance of IgG1 in antiprotein and of IgG3 in anticarbohydrate response, this IgG2a restriction of antiviral antibodies strongly suggests the existence of highly specific mechanisms for the regulation of individual subclasses in the mouse.

Animals↗

PubMed1987

Interleukin 1 and poly(rI).poly(rC) induce production of a hybridoma growth factor by human fibroblasts.

Cultures of normal diploid fibroblasts and of a human osteosarcoma cell line (MG-63) are shown to be able to produce a factor which promotes the growth of B cell hybridomas (hybridoma growth factor, HGF). The induction is stimulated by treatment of the cells with interleukin 1 (IL 1) (alpha or beta) or polyriboinosinic-polyribocytidylic acid [poly(rI).poly(rC)]. Combined treatment with cycloheximide and actinomycin D also stimulates production and enhances production induced by IL 1 or poly(rI).poly(rC). Extremely small doses of IL 1 (0.1 units/ml) are active as inducer of HGF. Also, under optimal conditions the yield of HGF can attain as much as 10(4) units/ml. Tumor necrosis factor (TNF-alpha), which otherwise shares various properties with IL 1, is a weak inducer of HGF. Although there is a superficial resemblance between induction of HGF and that of interferon-beta, the two activities are serologically distinct and conditions for their induction are quite different. In fact, conditions for induction of HGF are indistinguishable from those described for the induction of the mRNA of the so-called 26-kDa protein (also known as interferon-beta 2). Finally, the HGF derived from IL 1- or poly(rI).poly(rC)-treated fibroblasts is serologically not distinguishable from that produced by mitogen-stimulated peripheral blood leukocytes.

Animals↗

PubMed1987

Interleukin-HP1-related hybridoma and plasmacytoma growth factors induced by lipopolysaccharide in vivo.

Serum of lipopolysaccharide (LPS)-treated mice was found to support the growth of interleukin-HP1 (HP1)-dependent hybridoma and plasmacytoma cell lines. This growth-promoting activity, which was undetectable in normal serum, rose more than 1000-fold within 2 h after i.v. injection of the toxin and disappeared in less than 1 day. It could be traced to a single component, which behaved like HP1 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reverse phase high performance liquid chromatography, and was completely inhibited by a rabbit anti-HP1 antiserum. The LPS-induced factor was apparently not of T cell origin, as indicated by the strong activity found in the serum of LPS-treated nude mice. In vitro, LPS also considerably enhanced the production by macrophages of a factor with similar characteristics.

Animals↗

PubMed1987

B cell growth modulating and differentiating activity of recombinant human 26-kd protein (BSF-2, HuIFN-beta 2, HPGF).

The human "26-kd protein' is a secreted glycoprotein expressed, for example, in (blood) leukocytes, in epithelial cells treated with various inducers, but most strongly in interleukin-1 (IL-1)-treated fibroblasts. After finding it has antiviral and 2-5A synthetase-inducing activity, one group of authors called this protein IFN-beta 2. However, recently the full-length 26-kd cDNA sequence was shown to be identical with that of a B-cell-differentiating lymphokine called BSF-2, and another report suggested that the 26-kd protein could support the growth of some transformed murine B cell lines. To define its biological activities, we expressed the recombinant 26-kd protein by translating in Xenopus laevis oocytes a pure, synthetic chimeric mRNA containing the 26-kd protein coding region surrounded by Xenopus laevis beta-globin untranslated regions. A similar construction, but containing the HuIFN-beta cDNA coding region, was used to produce HuIFN-beta by the same procedure. Both recombinant glycoproteins were secreted, glycosylated, and their amounts were measured by [35S]methionine incorporation by the oocyte. Here we show that the recombinant 26-kd protein exhibits a high growth factor activity when assayed on an IL-HP1-dependent murine B cell hybridoma (sp. act. approximately 2 X 10(8) U/mg) as well as a potent differentiating activity on human CESS cells (sp. act. approximately 5 X 10(7) U/mg). While rHuIFN-beta was inactive in the latter two assays, it had the expected antiviral activity of 1-5 X 10(8) U/mg. The parallel recombinant 26-kd protein preparations had no detectable antiviral activity (i.e. a maximal specific activity of 1-3 X 10(2) U/mg, if any). The 26-kd protein is thus clearly an interleukin, and considering the confusing nomenclature now in use, this factor may better be renamed "interleukin 6'.

Animals↗

PubMed1986

Reaction of phosphofructokinase 2/fructose 2,6-bisphosphatase with monoclonal antibodies. A proof of the bifunctionality of the enzyme.

Monoclonal antibodies were derived from mice immunized against homogeneous chicken liver phosphofructokinase 2/fructose 2,6-bisphosphatase. Of 112 clones, 30 were found to secrete antibodies that specifically reacted with the antigen in enzyme-linked immunoabsorbant assay (ELISA) while 17, which were ELISA-negative, produced antibodies that affected the enzymic activity of the antigen. Four clones were subcloned and used for an extensive investigation of the reaction of the corresponding antibodies with the supposedly bifunctional enzyme. A definite proof of the bifunctionality of the enzyme was obtained from the two following observations. First, the two activities were similarly retained by the four antibodies that had been coupled to Sepharose. Second, one of the antibodies inhibited both activities with the same efficiency. Furthermore, the antigen-antibody reaction led to the formation of aggregates with an apparent molecular mass of several megadaltons, showing that the two subunits of the antigen reacted with the same antibody and were therefore identical. The four monoclonal antibodies affected the activity of phosphofructokinase 2. This effect was seen as an up to 17-fold activation as well as an up to 85% inhibition. Only one of the four antibodies (antibody 10) had inhibitory effects on fructose 2,6-bisphosphatase, an effect which was in part explained by a decrease in the rate of formation of the intermediary phosphoenzyme. All the effects described above were obtained on both the chicken liver and the pigeon muscle enzymes but with lower doses of antibody in the case of the former enzyme. Antibody 10 was also shown to react with mouse liver phosphofructokinase 2/fructose 2,6-bisphosphatase, and with phosphofructokinase 2 from chicken brain, heart and testis and from frog skeletal muscle and liver. None of the four antibodies cross-reacted with phosphofructokinase 2 from Saccharomyces cerevisiae or from spinach leaves.

Adsorption↗

PubMed1986

Variable region sequences of murine IgM anti-IgG monoclonal autoantibodies (rheumatoid factors). A structural explanation for the high frequency of IgM anti-IgG B cells.

The nucleotide sequences of heavy and light chains from 10 monoclonal IgM anti-IgG1 (RF) antibodies were determined and reported here as translated amino acid sequences. Only three families of VK light chains were used in these antibodies: VK1 (two examples), VK8 (three examples), and VK19 (four examples). This represents a significant nonrandom selection of light chains. In contrast, all other variable region gene segments (i.e., VH, DH, JH, and JK) were used in a pattern consistent with random selection from the available pool of germline genes. In two cases, the same anti-IgG1 specificity was generated by a combination of very homologous light chains with unrelated heavy chains. We infer from this that the light chain is the segment used by these antibodies to bind IgG1. The nature of these sequences provides an explanation for the curious observation that as many as 15% of splenic B cells in normal mice may be expressing IgM anti-IgG; if, as our data suggest, certain light chains in combination with many different heavy chains can be used in assembling the anti-IgG specificity, then, because of combinatorial association in which the heavy chain is not relevant for specificity, the fraction of IgM-producing B cells expressing these light chains should approximate the fraction of B cells making IgM anti-IgG. We calculate, based on data presented in several other studies, that 5-17% of B cells express one of the VK types observed in monoclonal RF. This agrees well with estimates for the number of B cells making IgM anti-IgG. In addition, our findings could rule out other explanations of the high percentage of B cells making RF, such as constant stimulation by antigen or presence of numerous antigenic epitopes since it was shown that IgM anti-IgG1 antibodies are not somatically mutated and that they are structurally homogeneous. We aligned the VK sequences of the RF in hopes of finding some primary sequence homology between the represented VK families which might point to residues involved in the binding interaction. Although we found no such homology in the hypervariable regions, we did find significant and unexpected homology in the FR2 and FR3 of these light chains. We noted that these regions are exposed in the Ig structure and postulate that they may be involved in a unique type of binding interaction between two Ig family domains, i.e., VK binding to a constant region domain of IgG.

Amino Acid Sequence↗

PubMed1986

Rheumatoid factor production in 129/Sv mice: involvement of an intestinal infectious agent.

High titers of IgG2a-specific rheumatoid factors (RF) are frequently observed in colonies of 129/Sv mice. The involvement of a transmissible agent in this phenomenon was shown by the following findings: (i) cesarean-derived and isolator-reared offsprings of RF-positive dams were free of RF, ii) a single intragastric inoculation with intestinal fluid from RF-positive donors elicited chronic RF production in RF-negative recipients, and iii) intestinal fluid collected from these primary recipients induced a comparable RF response in a second set of animals. The nature of this RF-inducing agent, however, remained elusive. Although its ability to pass through filters that efficiently retained bacteria could be unequivocally established, systematic serological analysis failed to detect any significant correlation between RF production and antibody responses to common mouse viruses or Mycoplasma. Moreover, all attempts to identify the RF-inducing agent by electron microscopy or to grow it in nude or newborn mice, as well as in cell cultures, remained unsuccessful.

Animals↗

PubMed1986

Transfer of immune complexes from lymphocytes to follicular dendritic cells.

Antigens in the form of immune complexes are retained on the membranes of follicular dendritic cells (FDC) for long periods of time. To examine how immune complexes reach germinal centers, where FDC are located, we injected mice with anti-2,4-dinitrophenyl (DNP) antibodies complexed to DNP-myoglobin-coated gold particles. The distribution of the particles in spleens or draining lymph nodes was then determined with the electron microscope. The vast majority of the particles were cell bound. Shortly after injection they were phagocytized by macrophages or fixed on lymphocytes. The latter were found even in the corona of lymph follicles but not in germinal centers. Already 30 min after injection, FDC in contact with the corona were faintly positive but were negative in the center. FDC precursor cells were occasionally observed but in too small a number to account for the transport of immune complexes to the germinal centers. Twenty-four hours after injection colloidal gold particles were found in phagolysosomes of macrophages or on cytoplasmic extensions of FDC in all parts of the germinal centers. Experiments performed on isolated FDC showed that they are not only able to take up free immune complexes but are also able to adsorb immune complexes from pulsed lymphocytes. These results strengthen the idea that lymphoid cells binding immune complexes by their Fc receptors may transport these complexes inside germinal centers.

Animals↗

PubMed1986

Monoclonal antibodies against isotypic and isoallotypic determinants of human IgA1 and IgA2: fine specificities and binding properties.

We have analysed and compared the fine specificity and behavior in various immunoassays of 10 mouse monoclonal antibodies, from three independent laboratories, directed against IgA1, IgA2 or non-IgA2m(2). The following observations were made. (1) Although all of the monoclonal antibodies were specific for a particular IgA subclass or isoallotype in a radioimmunoassay, three of them were not specific when tested in indirect immunofluorescence on plasma cells derived from pokeweed-activated peripheral blood lymphocytes. In this highly sensitive system, contrary to direct immunofluorescence previously performed using formalin-fixed lymphoid tissue, the anti-IgA1 69.114 reacted with some of the IgA2 plasma cells, the anti-IgA2 DLDB7 reacted with some of the IgA1 plasma cells and the anti-IgA2 16.512 dimly reacted with all IgM plasma cells. (2) Among the eight anti-IgA subclass antibodies, seven were directed against the CH2 domain of IgA whereas the anti-IgA1 1-155-1 recognised an epitope destroyed by Streptococcus sanguis IgA1 protease and localised in the hinge region of IgA1. The two anti-isoallotype antibodies were directed against epitope(s) probably localised in the 65 C-terminal amino acid residues of the alpha-CH3 domain. All of the 10 antibodies were able to react with endogeneously produced surface IgA on B-cells. (3) Using monoclonal anti-IgA subclass antibodies in radioimmunoassay may be hazardous in the absence of knowledge of their affinity constants and of careful control experiments: some of the antibodies were not sensitive in radioimmunoassays designed to measure the serum titer of specific IgA1 and IgA2 antibodies. Moreover, major differences were observed between the different monoclonal reagents with respect to the influence of the size of IgA on a solid-phase sandwich radioimmunoassay. While three of the anti-IgA1 underestimated dimeric IgA relative to monomeric IgA, the fourth anti-IgA1 and all the anti-IgA2 overestimated dimeric IgA relative to monomeric IgA, by a factor sometimes close to 7.

Animals↗

PubMed1986

Purification and NH2-terminal amino acid sequence of a T-cell-derived lymphokine with growth factor activity for B-cell hybridomas.

A T-cell-derived lymphokine was identified by its ability to support the growth of a subset of B-cell hybridomas. Hybrids that failed to survive in the absence of this molecule represented a major proportion of rat-mouse hybridomas but were very rare among mouse-mouse B-cell hybrids. Stable factor-dependent B-cell hybridomas were used to monitor the purification of the growth factor from the supernatant of a clonotypically stimulated mouse helper T-cell clone. Sequential fractionation using gel filtration, anion-exchange chromatography, and reversed-phase HPLC resolved the factor from other B-cell growth factors and yielded a single-chain protein characterized by a major charge (pI = 5-7) and molecular mass (22- to 29-kDa) heterogeneity, probably due to variations in glycosylation. The NH2-terminal amino acid sequence of this protein, which is active on B-cell hybridomas in the 0.1 pM range, showed no significant homology with that of known lymphokines. Because the purified factor also supported the growth and survival in vitro of murine plasmacytomas (to be published elsewhere), it was provisionally designated interleukin-HP1 (where H stands for hybridoma and P stands for plasmacytoma).

Amino Acid Sequence↗

PubMed1986

The murine antibody response to lactate dehydrogenase-elevating virus.

Mice infected with lactate dehydrogenase-elevating virus (LDV) were found to produce high titres of IgG anti-LDV antibodies that remained elevated for more than 1 year. This response, which was T-dependent, showed a striking preponderance of IgG2a with, from one strain to another, variable proportions of IgG2b and IgG3 but always very little IgG1. The binding of these antibodies to viral protein blots showed a major reaction with VP3, the heterogeneous glycosylated material of the viral envelope. A minor reaction was also noted with VP1, the nucleocapsid protein, but no antibodies were detected against VP2, the non-glycosylated envelope protein of LDV. A similar preponderance of anti-VP3 antibodies was also observed in a large set of anti-LDV hybridomas. Analysis of VP3 with monoclonal antibodies suggested that, despite its heterogeneity, this material has a common polypeptide moiety that apparently carries two major epitopes.

Animals↗

PubMed1986

Retention of immune complexes by murine lymph node or spleen follicular dendritic cells. Role of antibody isotype.

Using monoclonal anti-trinitrophenyl (TNP) antibodies complexed to TNP-myoglobin-coated gold particles, we analysed at the ultrastructural level the retention by follicular dendritic cells (FDC) of immune complexes containing various antibody isotypes. Gold-labelled immune complexes were injected subcutaneously or intravenously into naive mice and, after 24 h, germinal centres of draining lymph nodes or spleen were examined by electron microscopy. FDC generally retained complexes containing IgG2a and IgG2b better than those formed with IgG1 or IgG3. IgM was rarely retained. FDC isolated from lymph nodes or spleens were incubated in vitro with gold-labelled complexes in a serum-free medium. IgG2a and IgG2b complexes were also retained in vitro in large quantities by FDC; IgG1 and IgG3 complexes were retained in smaller quantities or in highly variable quantities compared with IgG2; IgM complexes were rarely seen on FDC. There was no difference between FDC isolated from lymph nodes or from spleen with respect to the Ig isotypes required for Fc-mediated retention of immune complexes.

Animals↗

PubMed1985

A simple panning method for the selection of cell surface antigen transfectants.

Mouse Ltk- (H-2k) cells that became reactive with anti-H-2d monoclonal antibodies (McAb) after transfection with DNA from H-2d-positive cells were isolated by an indirect panning method. The cells were transfected with DNA from a plasmid containing the selective thymidine kinase (tk) gene in addition to total cell DNA, so that a first selection could be carried out in hypoxanthine/aminopterin/thymidine (HAT) medium. The HAT-selected tk+ transfectants were incubated with anti-H-2d McAb, washed and transferred to dishes coated with purified anti-immunoglobulin (anti-Ig). This 2-step method of panning has the advantage that only the anti-Ig reagent requires purification. Transfected cells clearly reactive with either anti-Ld/Rd or anti-Kd McAb were isolated after only 1 or 2 cycles of panning.

Animals↗

PubMed1985

Rheumatoid factor (RF) production during anamnestic immune responses in the mouse. III. Activation of RF precursor cells is induced by their interaction with immune complexes and carrier-specific helper T cells.

IgG1 immune complexes were identified as the humoral stimuli responsible for the synthesis of IgG1-specific IgM rheumatoid factor (RF), which occurs in the mouse during the early stages of secondary immune responses to protein antigens. The specificity of this phenomenon was illustrated by the fact that complexes made with IgG1 F(ab')2 fragments or with antibodies of a different isotype failed to induce significant anti-IgG1 RF synthesis. The importance of immune complexes in the induction of RF was further underscored by the substantial increase in the titers of isotype-specific RF observed in the serum of mice immunized with IgG1- or IgG2a-complexed antigen rather than with antigen alone. The RF-inducing capacity of the complexes varied with the antigen/antibody ratio: it was maximal in antibody excess or at equivalence, but dramatically reduced in large antigen excess. The importance of T cell priming in RF precursor cell activation by immune complexes was demonstrated by the failure of T cell-deprived spleen cells to reconstitute the capability of irradiated mice to produce RF, and by the optimal RF responses observed after reconstitution of irradiated recipients with primed T cells and naive B cells. The involvement of T cells in this process could not be explained by the release of nonspecific B cell activators, because antigenic stimulation of primed T cells failed to enhance the activation of RF precursor cells by immune complexes of unrelated antigen.

Animals↗

PubMed1985

Isotypically restricted activation of B lymphocytes by lactic dehydrogenase virus.

Lactic dehydrogenase elevating virus (LDV) was found to selectively stimulate IgG2a synthesis in infected mice. Within one week after infection, the production of IgG2a increased nearly 50-fold whereas that of IgM, IgA, IgG1 and IgG3 remained virtually unchanged. IgG2b synthesis was also enhanced but to a lesser extent. Several observations suggested that this stimulation of IgG2 production resulted from a polyclonal B cell activation: (a) the isoelectric focusing patterns of IgG2a before and after LDV infection were exactly the same, (b) the frequency of clones with anti-LDV activity in hybridoma collections derived from infected mice was extremely low (less than 4/1000) and (c) the proliferative response elicited by LDV in unsensitized animals was comparable with that induced by lipopolysaccharide. The effect of LDV on immunoglobulin synthesis was drastically reduced in nude mice but was not affected by the X-linked B lymphocyte defect of animals carrying the xid mutation.

Animals↗

PubMed1985

In vivo suppression of T-dependent antibody responses by treatment with a monoclonal anti-L3T4 antibody.

Minute amounts of the anti-L3T4 antibody designated GK1.5 were found to deeply suppress in vivo antibody responses to T-dependent antigens. Primary responses to sheep erythrocytes were completely inhibited even when GK1.5 was administered up to 6 days before or 4 days after the antigen. Secondary responses to potent immunogens like sheep erythrocytes or keyhole limpet hemocyanin were also completely abolished by a single injection of GK1.5 just before the boost. This treatment had no effect on T-independent reactions such as the polyclonal activation of B lymphocytes with lipopolysaccharide or the anti-2,4,6-trinitrophenyl (TNP) response elicited by injection of TNP-Ficoll.

Animals↗

PubMed1985

Enhancement of IgG anti-carrier responses by IgG2 anti-hapten antibodies in mice.

Isologous IgG2a and IgG2b anti-hapten antibodies injected along with trinitrophenylated or fluoresceinated keyhole limpet hemocyanin (KLH) were found to considerably stimulate primary IgG responses to the carrier protein in mice. No such stimulation was observed with IgG1, IgM and IgA anti-hapten antibodies. Depending on the antigen antibody combination, the amplification obtained after a single injection of IgG2-complexed haptenated KLH ranged from 20- to 1000-fold. By comparison, secondary responses were only marginally enhanced (approximately 3-fold) when IgG2-complexed rather than free antigen was used to boost irradiated recipients previously reconstituted with primed spleen cells. The IgG2-mediated enhancement was effective over a wide range of antigen/antibody ratios, did not require the use of high affinity antibodies and occurred whatever the route of injection. The stimulation was specific for the complexed antigen and developed to the same extent whether complexes were formed in vitro or in vivo. A comparable stimulation of the anti-carrier response was obtained with several other haptenated proteins but with formaldehyde-treated diphtheria and tetanus toxoids inhibition rather than stimulation was observed.

Animals↗