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J C Roder

Publications and source records attributed to J C Roder.

At least 55 records · Page 3Linked to original sources
PubMed1994

Systemic administration of ciliary neurotrophic factor induces cachexia in rodents.

Ciliary neurotrophic factor (CNTF) has previously been shown to promote the survival of several classes of neurons and glial. We report here that in addition to its effects on the nervous system, CNTF can induce potent effects in extra-neural tissues. Implantation of C6 glioma cells engineered to secrete CNTF either subcutaneously or into the peritoneal cavity of adult mice, or systemic injections of purified rat or human recombinant CNTF, resulted in a rapid syndrome of weight loss resulting in death over a period of 7-10 d. This weight loss could not be explained by a reduction in food intake and involved losses of both fat and skeletal muscle. CNTF also induced the synthesis of acute phase proteins such as haptoglobin. Implantation of C6 lines expressing a nonsecreted form of CNTF, or the parental C6 line itself, did not result in wasting effects. Analysis of this CNTF-induced wasting indicates similarities with the previously described cachectins, tumor necrosis factor, interleukin 6, and leukemia inhibitory factor, but does not involve the induction of these cytokines.

Animals↗
PubMed1993

Derivation of completely cell culture-derived mice from early-passage embryonic stem cells.

Several newly generated mouse embryonic stem (ES) cell lines were tested for their ability to produce completely ES cell-derived mice at early passage numbers by ES cell <==> tetraploid embryo aggregation. One line, designated R1, produced live offspring which were completely ES cell-derived as judged by isoenzyme analysis and coat color. These cell culture-derived animals were normal, viable, and fertile. However, prolonged in vitro culture negatively affected this initial totipotency of R1, and after passage 14, ES cell-derived newborns died at birth. However, one of the five subclones (R1-S3) derived from single cells at passage 12 retained the original totipotency and gave rise to viable, completely ES cell-derived animals. The total in vitro culture time of the sublines at the time of testing was equivalent to passage 24 of the original line. Fully potent early passage R1 cells and the R1-S3 subclone should be very useful not only for ES cell-based genetic manipulations but also in defining optimal in vitro culture conditions for retaining the initial totipotency of ES cells.

Animals↗
PubMed1993

Effect of vector topology on homologous recombination at the CHO aprt locus.

The Chinese hamster ovary aprt gene was used as a model for studying the effect of vector topology on gene targeting frequency. A single recombination vector containing 2.7 kb of isogenic DNA homologous to the aprt gene was digested with eight separate restriction enzymes to generate a variety of both replacement- and insertion-type recombination substrates. The frequency of homologous recombination, normalized by cotransfection with a linearized neo' marker, was assayed by the correction of a mutant hemizygous aprt allele and was not found to reflect vector topology. Southern analysis of representative recombination products suggests that the gene targeting events occurred predominantly by double crossover/gene conversion.

Adenine Phosphoribosyltransferase↗
PubMed1992

Intra-rectal injection of tumour cells: a novel animal model of rectal cancer.

The purpose of this study was to develop an animal model of rectal cancer. Three murine-derived cell lines, B16 melanoma, CT26 and MCA38 colon carcinoma, as well as the human colon cancer cell line LS174T were injected into the submucosa of the mouse rectum. Subcutaneous CT26 anbd B16 tumours and intra-caecal CT26 tumours served as controls for tumourigenicity of the cell lines. B16 melanoma produced a locally aggressive rectal tumour as well as skin and para-aortic lymph node metastases. CT26 produced local tumour when injected intra-rectally and colon tumours and liver metastases when injected into the caecum. MCA38 and LS174T intra-rectal injections resulted in large rectal carcinomas without metastases. We believe that growth of a colon cancer cell line in the rectum approximates the human disease more closely than other models of colorectal cancer. We would expect that the model could similarly be utilized to assess the effects of novel adjuvant treatments for rectal cancer as well as in the study of the tumour biology of rectal cancer.

Adenocarcinoma↗
PubMed1992

Production and characterization of a novel monoclonal antibody inhibitory for murine natural killer cell activity.

Natural killer (NK) cells are considered to play an important role in tumor surveillance. The killing of tumor target cells by NK cells is the result of a complex series of sequential binding, signal processing and lytic events. However, the mechanism which NK cells use to recognize tumor targets is poorly understood. To further study the cell-surface molecules involved in tumor recognition, we immunized rats against cloned murine T cells with NK activity (DBA/2.1) and generated rat-mouse hybridomas which were screened for the ability to block lytic activity of DBA/2.1 effector cells. Culture supernatants from one IgM-producing hybridoma, designated S1C4, were found to consistently inhibit DBA/2.1-mediated lysis of YAC-1 target cells. Endogenous splenic NK activity was also diminished in the presence of S1C4 monoclonal antibody (mAb) while alloantigen-specific cytotoxic T lymphocyte (CTL) activity was not affected. S1C4 mAb appears to react with effector cell-surface structures involved in the recognition/adhesion phase of NK activity since pretreatment of effector cells with mAb S1C4 inhibits their ability to bind to YAC-1 target cells. ELISA studies revealed that the S1C4 antigen is expressed by a range of lymphoid cell lines, as well as by DBA/2.1 cells and fresh splenic NK cells. S1C4 mAb were shown to react with 22, 24, 30, and 46 kiloDalton (kDa) DBA/2.1 cell membrane components on immunoblots performed under reducing conditions. These structures do not correspond to any known recognition/adhesion molecules, suggesting that mAb S1C4 defines novel cell membrane components involved in NK cell function.

Animals↗
PubMed1991

Comparative biodistribution and antibody-dependent cellular cytotoxicity of native and heavy chain chimeric antibody.

We have recently chimerized the heavy chain of the pan-carcinoma monoclonal antibody (mAb) B72.3. Studies were undertaken to compare the IgG1 chimeric antibody, B72.3-1-3 with native murine B72.3 (nB72.3). Using fluorescence-activated cell sorting analysis, B72.3-1-3 demonstrated specific binding to fresh LS174T tumor cells. Biodistribution of 131I B72.3-1-3 was similar to 131I nB72.3 in nude mice bearing LS174T xenografts. Peak radiolocalization indices were noted on day 6 for B72.3-1-3 and day 8 for nB72.3. Both antibodies were capable of imaging LS174T tumors by radioimmunoscintigraphy. Antibody-dependent cellular cytotoxicity of LS174T by human peripheral blood lymphocytes was tested in 8h 51Cr release assays. With either no antibody or nB72.3, lymphocytes were not capable of killing LS174T cells. However, B72.3-1-3 at a concentration of 5 and 50 micrograms/ml mediated significant lysis of tumor cells by human lymphocytes. These results suggest that chimeric antibodies retain their binding properties to tumor cells and display biodistribution patterns similar to their unmodified counterparts. Such modifications may reduce the deleterious human antimouse antibody response to murine mAbs as well as augment antibody-dependent cellular cytotoxicity of tumor cells by human effectors.

Animals↗
PubMed1990

Comparison of cellular immunotherapies and anti-CD3 in the treatment of MCA-38-LD experimental hepatic metastases in C57BL/6 mice.

An experimental model of hepatic metastases in C57BL/6 mice was used to compare the antitumor effects of lymphokine-activated killer (LAK) cells, anti-CD3-activated T-cells (ATC), and anti-CD3 alone. Liver metastases were produced by in vivo passage of MCA-38-LD adenocarcinoma via the ileocolic vein. LAK cells and ATC were generated by 3-day in vitro incubation of spleen cells in interleukin 2 and anti-CD3, respectively. Percentage of tumor volume in livers was determined with a morphometric technique. With less than therapeutic LAK cell doses (0.5-1.0 x 10(7) cells), no effect was seen in mean (+SE, -SE) percentage of tumor volume of control [23.3 (29.3, 18.5)] compared to LAK cell-treated [21.6 (29.3, 15.9)] animals. The same number of ATC significantly reduced the mean percentage of tumor volume [2.7 (4.7, 1.4)] (P less than 0.005). High dose interleukin 2 also significantly decreased tumor volume. More strikingly, a single dose of anti-CD3 alone had a beneficial effect on mean percentage of tumor volume when given i.p. [1.0 (1.9, 0.4)] or i.v. [1.2 (1.7, 0.7)] (P less than 0.0003). A total of 33% of anti-CD3-treated mice had no detectable liver metastases. In 51Cr release assays, the cytotoxicity of ATC was shown to be partially mediated by nylon wool-adherent accessory cells. The effectiveness of anti-CD3 in this immunotherapy model suggests that a similar approach may be taken to immunotherapy of human malignancies, without the requirements for in vitro-generated killer cells or exogenously administered interleukin 2.

Adenocarcinoma↗
PubMed1990

Class I (H-2Kb) gene transfection reduces susceptibility of YAC-1 lymphoma targets to natural killer cells.

A "hybrid gene" (MTKb) comprised of the human metallothionein IIA promoter ligated to the genomic sequence of the major histocompatibility complex class I (H-2Kb) gene was subcloned into the expression vector pSV2neo and transfected into the natural killer (NK) cell-sensitive YAC-1 lymphoma. The Kb gene product was readily detectable on the cell surface of G418-resistant transfectants using both Kb-specific monoclonal antibodies and H-2b-specific cytolytic T cells. Unlike control pSV2neo transfectants, MTKb-pSV2neo transfectants were relatively resistant to lysis by NK cells from H-2a, H-2b, H-2k or H-2 (a x b)F1 haplotype mice. These data strongly suggest that the effects of MHC expression on susceptibility to NK cells can be mediated by a single and well-defined class I molecule, Kb.

Animals↗
PubMed1989

A functional and phenotypic comparison of murine natural killer (NK) cells and lymphokine-activated killer (LAK) cells.

Lymphokine-activated killer (LAK) cells have been defined as interleukin 2 (IL-2)-activated cytolytic effector cells exhibiting non-MHC restricted killing against a wide range of NK-sensitive and NK-resistant tumor cells. There has been considerable debate as to whether LAK cells are derived from NK cells or from a unique precursor population. In the present study, we compare LAK cells derived from T-cell-depleted nylon-wool-non-adherent spleen cells with endogenous NK cells and NK cells activated with the interferon inducer polyl.C., in terms of their phenotype and functional characteristics. The predominant splenic LAK precursor in the mouse was found to be a nylon-wool-non-adherent, thy1-, MICG-, J11d.2-, asialo GM1+ cell. This phenotype is shared by endogenous NK cells. A significant number of activated NK cells express macromolecular insoluble cold globulin (MICG) in addition to asialo GM1. Neither endogenous nor activated NK cells express a heat-stable antigen found on bone-marrow cells, immature T cells, and most B cells and defined by monoclonal antibody (MAb)J11d.2. However, J11d.2 is expressed on some LAK precursor and effector cells. The asialo GM1 marker is common to all LAK effector cells, while many are also thy1+, and/or MICG+. LAK effector cells are therefore a heterogeneous population sharing some phenotypic characteristics in common with NK cells. In addition, there is a positive correlation between LAK and NK activity in "high NK" and "low NK" mouse strains, suggesting that NK cells and LAK cells share a common lineage. Monoclonal antibodies to the alpha and beta chains of LFA-I inhibit LAK and activated NK function, while endogenous NK and CTL killing is affected only by anti-alpha chain antibodies. LAK cells, like MHC-restricted and non-restricted CTL clones, express mRNA transcripts of the C11 serine protease gene. We conclude that LAK cells share several features in common with cells of the NK lineage and may therefore represent NK cells in a unique state of activation. LAK cells appear to employ cytolytic machinery common to other lytic cell types.

Animals↗
PubMed1989

Synthesis of soluble myelin-associated glycoprotein in insect and mammalian cells.

The myelin-associated glycoprotein (MAG) has an extracellular domain containing five sequences which are homologous to the immunoglobulin-fold motif. Adhesive interactions mediated by the MAG extracellular domain are involved in the development of the myelin sheath. The MAG cDNA has been modified to introduce a stop codon immediately before the transmembrane domain. Expression of the modified cDNA in insect cells and murine NIH-3T3 cells resulted in secretion of the soluble MAG extracellular domain. Treatment of soluble MAG with glycopeptidase F and endoglycosidase H showed significant differences in glycosylation for the insect and mammalian cell-expression systems. The soluble form of MAG has been purified from insect-cell supernatants by adsorption to a lentil-lectin support. The soluble MAG will provide a powerful new approach for studies of MAG-adhesive interactions during brain development.

Amino Acid Sequence↗
PubMed1989

Amelioration of experimental lung metastasis in mice by therapy with anti-CD3 monoclonal antibodies.

Binding of CD3-specific antibodies to the TcR-CD3 complex results in T cell activation without the need for occupation of the T cell receptor (TcR) by its ligand. Murine T cells activated in this manner will kill a broad range of tumor targets but not normal lymphoblasts. We report here that non-specific cytolytic activity can be induced in vivo by a single i.p. injection of nonlytic 145-2C11 anti-CD3 monoclonal antibody. At least three populations of effector cells are activated in these mice. These are non-MHC (major histocompatibility complex) restricted cytotoxic T lymphocytes, activated natural killer cells, and lymphokine-activated killer cells. Anti-CD3 treatment is effective in significantly reducing the number of lung tumor nodules which form in mice inoculated with oncogenic ras-transfected syngeneic 10T1/2 fibroblasts. Anti-CD3-activated killer cells may, therefore, find a future role in cancer immunotherapy.

Animals↗
PubMed1989

Murine anti-CD3 monoclonal antibody induces potent cytolytic activity in both T and NK cell populations.

Antibodies specific for the CD3 complex have the capacity to both stimulate and inhibit a variety of T cell functions. We show here that a monoclonal antibody to the epsilon chain of CD3 can induce efficient non-MHC-restricted cytolytic activity in murine lymphocytes with peak activity occurring after 48 hr of incubation. In a panel of targets, the anti-CD3-activated effectors lysed tumor cells but not normal lymphoblasts. Cytolysis was not dependent on the presence of the antibody in the cytolytic assay. Moderate to high cytolytic activity was elicited from lymph nodes, spleen, and thymus by anti-CD3 treatment in vitro, whereas only low activity was apparent in bone marrow. The precursors of anti-CD3-activated cells consisted largely of mature T cells, although a smaller component of immature T cells was also involved. Thus, separation of thymocytes based on adhesion to peanut agglutinin revealed that both positive (immature) and negative (mature) fractions could be activated, while cytotoxic pretreatment of spleen cells with an antibody (J11d) to immature T cells before anti-CD3 activation significantly decreased the resulting cytotoxicity. The majority of precursors in spleen were Thy 1+ and CD8+ and/or AGM1+. Antibody depletion studies showed that the effector cells have both a T and a NK component consisting of Thy 1+, CD5+, CD8+, CD4-, and AGM1- cells and Thy 1-, CD5-, CD8-, CD4-, and AGM1+ cells, respectively. The production of significant amounts of IL-2 and TNF in culture following anti-CD3 treatment, along with the synergistic effect of exogenously added IL-2, suggests that one or both of the effector cell types could be induced by lymphokines. The intraperitoneal administration of the anti-CD3 antibody induces cytolytic activity in vivo. Therefore, the direct activation of cytolysis by anti-CD3 antibody and the additional effects, both direct and synergistic, of lymphokines produced by the activated lymphocytes could conceivably provide a potent anti-tumor therapy.

Animals↗
PubMed1989

The role of T cell receptors in non-MHC-restricted cytotoxicity.

The relationship between natural killer cells (NK) and cells of the T lineage has been obscured by the existence of poorly characterized clones of presumed NK origin. We have analyzed nine of these cloned cell lines displaying varying levels of cytotoxic activity against NKS YAC-1 target cells for rearrangement and expression of the genes encoding the alpha, beta, and gamma chains of the T cell receptor for antigen. Rearrangements at both the TcR beta and gamma loci were detected in all clones often at both alleles. Rearrangement patterns at the TcR beta locus were identical in several clones, despite different degrees of cytotoxicity. T cell receptor alpha, beta, and gamma genes were expressed as full length transcripts in all clones regardless of their levels of cytotoxic activity. To explore the involvement of cell surface molecules in the cytolytic events, studies were undertaken to determine whether cytotoxic activity could be inhibited by antibodies against CD3, LFA-I, and H-2KdDd. In two selected clones, both alpha and beta chains of the LFA-I molecule were expressed but only monoclonal antibodies against the alpha chain significantly blocked cytotoxicity. Cytotoxicity was also inhibited by monoclonal antibodies against epitopes of H-2KdDd and CD3, the extent of inhibition correlating with the level of surface expression on both clones. These data suggest that conventional alpha/beta heterodimers may be necessary but not sufficient for target cell recognition by these clones. Since T cell receptor rearrangement and expression occur normally in the T cell lineage but not the NK lineage, these results also indicate that a subpopulation of cells with non-MHC-restricted killer activity lies on the T cell differentiation pathway and is selected by in vitro growth with IL-2. The limited rearrangement pattern observed can be explained if only a small subpopulation of T cells is capable of non-MHC-restricted killing, and if certain rearrangements favor self-MHC recognition which is known to block cytolysis in the NK system.

Animals↗
PubMed1989

Decreased p21 levels in anti-sense ras transfectants augments NK sensitivity.

We have shown that the oncogenic EJ-ras gene, under the control of a metallothionein-I (Mt) promoter, can be induced to cause an increased susceptibility of transfected 10T1/2 fibroblasts to cytolysis mediated by natural killer (NK) cells. This effect may be specific to the ras gene family, since other oncogenes that we have tested here (src) and elsewhere (myc) do not show this effect. We have now examined the effect of modulating the level of p21 in both a positive or negative manner. The level of p21 ras was decreased by two independent mechanisms. First Zn2+ was removed from Mt-EJ-ras transformed cells. In the second approach we transfected 10T1/2 cells with a Mt-anti-sense c-H-ras construct which reduced p21 expression, slowed the growth rate and altered the morphology of 10T1/2 cells when induced with Zn. Surprisingly, the decrease in p21 ras levels by both approaches caused a marked increase in NK susceptibility (NKS) which was equivalent to that observed when the p21 ras levels were increased either by inducing EJ-ras or removing Zn2+ from Mt-anti-sense c-H-ras containing cells. The kinetics of induction of NK sensitivity due to decreasing normal p21 ras levels was identical to that observed for increasing mutated p21 ras levels. Peak enhancement of NKS was observed 24 hr after ras perturbation. These results suggest that either a positive or negative change in the steady-state level of p21 ras is sufficient to induce NK sensitivity, and NK sensitivity is not inextricably linked to cellular transformation by the ras gene.

Animals↗
PubMed1989

Recombinant myelin-associated glycoprotein confers neural adhesion and neurite outgrowth function.

Myelin-associated glycoprotein (MAG) cDNA clones for the small (p67) and large (p72) forms were expressed in heterologous cells. Purified recombinant MAG protein was incorporated into fluorescent liposomes, and both forms were shown to bind predominantly to neurites in DRG or spinal cord cultures. This adhesion was completely blocked by Fab fragments of monoclonal anti-MAG antibody. Liposomes prepared with the control protein glycophorin or no protein failed to bind neurites. Small cerebellar neurons, which are not myelinated in vivo, failed to bind MAG liposomes. In a second test of function, p67 MAG-transfected fibroblasts were markedly enhanced in their ability to promote DRG neurite extension over a 2 day culture period compared with control fibroblasts not expressing MAG. Neurite extension was blocked by anti-MAG antibodies. These results show that both forms of MAG can facilitate the interactions between glial cells and neurites that ultimately lead to myelin formation.

Axons↗
PubMed1989

Modulated adhesion: a proposal for the role of myelin-associated glycoprotein in myelin wrapping.

Myelin-associated glycoprotein (MAG) is a 100-kDa integral membrane glycoprotein expressed by oligodendrocytes and Schwann cells in the central and peripheral nervous systems, respectively. It is found first in loosely wrapped myelin and then periaxonally after compaction. Clinical findings, structural analysis, and cell assays indicate a role for MAG in adhesion. We propose that the phosphorylation state of MAG modulates its adhesion and that a minimum spatial requirement for the separation of the kinase and phosphatase activities postulated by this model may explain the correlation between axon size and myelination state.

Cell Adhesion↗