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Biomedical subjects

C Sorg

Publications and source records attributed to C Sorg.

At least 217 records · Page 12Linked to original sources
PubMed1983

Monoclonal antibodies against human malignant melanoma.

Monoclonal antibodies were raised against cultured melanoma cells and selected for their reactivity with melanoma-associated antigens phenotypically expressed on various melanoma lines and their sublines. Of 34 monoclonal antibodies tested which were all reacting with melanoma lines only 19 antibodies were found to bind to melanoma tissues. Of the 19 antibodies 10 displayed broad cross-reactivity with other cells and structures whereas 9 had a restricted reactivity. Of these antibodies 5 types could be defined which reacted with nevocellular antigens (I and II) with neural, endothelial and epidermal cells. The antibodies revealed a great heterogeneity of primary and metastatic melanoma. So far no clear association of the antigens with either primary or various forms of metastatic melanoma could be established.

Animals↗
PubMed1982

Purification of human blood monocytes by hypotonic density gradient centrifugation in Percoll.

Leukocyte cell concentrates, obtained by continuous flow leukapheresis from single donors, were separated on a continuous hypotonic (260 mosM) Percoll gradient. On average, 86% of monocytes were recovered in a sharp band at a purity of up to 91% (average 76%). By this procedure 1-2 X 10(9) monocytes may be obtained from an individual donor. Hypotonic gradient purification, as compared with isotonic (295 mosM) conditions, proved superior with regard to capacity, speed of performance, yield and monocyte purity.

Cell Separation↗
PubMed1982

Heterogeneity of macrophages in response to lymphokines and other signals.

Macrophages which are intimately involved in acute and chronic inflammatory reactions are functionally heterogeneous not only with regard to the expression of constitutive functions but also in their response to lymphokine signals. The biological basis of this heterogeneity is poorly understood. Whether we are dealing with true subpopulations or with intermediately stable phenotypes has not been resolved. To study these questions we adopted a bone marrow liquid culture system in which bone marrow cells--in the presence of a colony-stimulating factor--proliferate and differentiate into macrophages. This culture system was taken here as a model to study the expression of various functions by macrophages in the course of maturation. Several tests were performed daily and in parallel from the same batch of cells. It was found that certain functions were expressed early and were also characteristic for mature macrophages such as Fc receptors, phagocytosis of latex beads and unspecific esterase. Other functions appeared and disappeared in an ordered sequence, such as the response to macrophage migration inhibitory factor and chemotactic factor as well as the production of interferon and plasminogen activator. The time course of functional expression was strongly dependent on proliferation of precursor cells as well as on proliferation of differentiated macrophages. It is concluded that the transient phenotypic expression of functions during differentiation is the basis for the functional heterogeneity of macrophages.

Animals↗
PubMed1982

Use of a multiwell assembly for chemotaxis and evaluation by enzyme-linked immunosorbent assay (ELISA).

A multiwell chamber assembly for chemotaxis tests was designed, which integrates the established microtiter system. A microtiter plate is covered with a plastic plate containing up to 96 holes of the diameter of the microtiter wells. Between the plates, a Nucleopore filter sheet (5 micron) and a silicon rubber gasket is placed. As a model system, human monocytes and lymphocyte-derived chemotactic factors were used. As it was observed that monocytes migrate through the membrane and settle on the bottom of the microtiter wells, an ELISA was adapted for quantitation of cells. After washing and incubation with a xenoantiserum against human monocytes, the bound antibody was quantitated using protein-A-conjugated alkaline phosphatase and p-nitrophenyl phosphate as detection system. The plates were read in a multichannel photometer. Cell numbers were determined directly from a calibration curve established before with varying numbers of monocytes. Current experience allows the following conclusions: The chemotaxis test in microtiter plates is simpler, faster and uses less material than conventional Boyden chambers. Evaluation by ELISA is much faster and more accurate than by microscopy.

Animals↗
PubMed1982

Modulation of macrophage functions by lymphokines.

Macrophage migration inhibitory factors (MIFs) of mouse and guinea pig have been thoroughly characterized with regard to molecular weight and isoelectric points. Several molecular weight species have been identified. In a comparative study with purified MIFs it was found that these molecules were distinct from a series of other lymphokines, particularly so from macrophage activating activities. Investigations on the molecular weight heterogeneity of MIF have led us to a transglutaminase-like activity which was found to be expressed in certain subsets of macrophages. The question whether low molecular weight factors are polymerized by this enzyme to oligomers is further investigated. Studies on the induction by lymphokines of interferon and plasminogen activator revealed a great heterogeneity of responding macrophages. In studies on the biological basis of the functional heterogeneity of macrophages, the question was investigated whether the heterogeneity was due to different macrophage subpopulations or to intermediate relatively stable phenotypes on their way to maturity and senescence. To approach this question, the bone marrow liquid culture system was used as a developing system. Our data are summarized in a unifying model which takes into account the different constitutive and inducible functions during the cell cycle. Accordingly, lymphokines may act either as differentiation signals, as mitogens or activating signals.

Animals↗
PubMed1981

Transglutaminase as a marker for subsets of murine macrophages.

Transglutaminase was detected either at the single cell level by fluorescent staining with dansylcadaverine or in cell homogenates by incorporation of [14C] putrescine into alpha-casein. In the mouse it was found that erythrocytes, granulocytes, thymocytes or lymphocytes with or without concanavalin A stimulation were negative in the fluorescence test. Normal peritoneal washout macrophages and peritoneal exudate cells stained positive to varying degrees (induced with mineral oil 64%, with thioglycollate 50%, with proteose peptone 22%, normal washout 1%). Macrophages from bone marrow liquid cultures were 20% positive at day 3 and 100% at day 17. Promonocytes and monocytes were negative. Positively stained cells also phagocytosed opsonized sheep erythrocytes. The degree of staining varied considerably in the macrophage-like cell lines IC21 (100%), J774.2 (75%), P388-D1 (50%). This result and those from autoradiography studies indicate that expression of transglutaminase is not associated with the S-phase of the cell cycle. The fluorescence test correlates quantitatively with the [14C] putrescine incorporation test. The enzyme is Ca2+-dependent and appears neither to be on the outer cell surface nor being released into the culture medium. Circumstantial evidence indicates that it is also not compartmentalized in cytoplasmic vesicles. While the induction and modulation of enzyme expression is still under study, it is concluded that transglutaminase is a new marker for macrophages of a certain differentiation or activation state.

Animals↗
PubMed1981

Tumor production in the nude mouse, fibrinolytic activity and cross-reactivity with antimelanoma sera of various human tumor cell lines.

It has been reported previously that antisera produced in non human primates against many cultured human melanoma lines recognize tumor-associated antigens on the melanoma cells. The reactivity correlated with other parameters of malignancy, such as rapidity of tumor growth in nude mice and release of plasminogen activator. In this study we have investigated whether these antisera also react with non melanoma cancer lines and searched for a correlation between this reaction and the other parameters of malignancy. There was cross-reactivity of the ten sera with 4-43% of the 46 non melanoma lines compared to 62-100% of the 16 melanoma lines, indicating a preferential reactivity with melanoma lines. For the non-melanoma lines no correlation was found between the release of plasminogen activator, the degree of tumorigenicity in the nude mouse or the reactivity of surface antigens as detected with antimelanoma sera.

Animals↗
PubMed1981

Influence of various detachment procedures on the functional state of cultured murine macrophages.

Macrophages were obtained from seven-day-old bone marrow liquid cultures to which a colony-stimulating factor from L-cell-conditioned medium had been added. It was found that the yield of macrophages from the bone marrow liquid culture was dependent on the type and brand of culture dishes. Highest yields were obtained in teflon membrane bag cultures. The cumulative yield after serial passage of macrophages was 700-fold of the cell input after two months. After mechanical detachment from plastic culture dishes, the survival rate of cells was related to the brand of the plastic dish. Viability also varied greatly after pretreatment of cells with scandicain (28%), proteinase K (78%), or pronase (99%). After mechanical detachment of macrophages or pretreatment with scandicain, adherence to plastic, latex phagocytosis and chemotaxis was not or only slightly impaired, whereas the same functions after protease treatment were greatly reduced or even abolished and were only recovered after another culture period. Scandicain, proteinase K, and pronase treatment was mitogenic to macrophages in contrast to mechanical detachment. Pronase treatment also induced release of plasminogen activator activity. The results demonstrate that macrophages respond extremely sensitive to environmental conditions and to various insults by changing their functional state.

Animals↗
PubMed1980

Sequential expression of functions during macrophage differentiation in murine bone marrow liquid cultures.

In the presence of a colony-stimulating factor, murine bone marrow cells proliferate and differentiate into macrophages. This culture system was taken as a model to study the expression of various functions by macrophages in the course of maturation. Several tests were performed daily and in parallel from the same batch of cells. It was found that certain functions were expressed early and were also characteristic for mature macrophages such as Fc receptors, phagocytosis of latex beads and unspecific esterase activity. Other functions appeared and disappeared in an ordered sequence, such as the response to macrophage migration inhibitory factor and chemotactic factor as well as the production of interferon and of plasminogen activator. The time course of functional expression was strongly dependent on proliferation of precursor cells as well as proliferation of differentiated macrophages. It is suggested that the phenotypic expression of functions during differentiation is the basis for the functional heterogeneity of macrophages.

Animals↗
PubMed1980

Use of an enzyme-linked immunosorbent assay (ELISA) for screening of hybridoma antibodies against cell surface antigens.

A micro-ELISA for screening of antibodies from hybridoma cultures against surface antigens of human melanoma is described. The technique employs alkaline phosphatase-conjugated protein A and target cells attached to poly-L-lysine-coated microtiter plates. The micro-ELISA is equally sensitive as the radioimmunoassay. Mild glutaraldehyde treatment of cells did not lead to an appreciable loss of antigen activity. The fixed cells can be stored at 4 degrees C for at least 6 weeks. It is concluded that the ELISA is superior to the radioimmunoassay in the following aspects: (1) exclusion of radioactive hazards, (2) speed of performance, and (3) lower costs.

Animals↗
PubMed1980

Interferon production by Corynebacterium parvum and BCG-activated murine spleen macrophages.

Macrophages were identified to be a major source of interferon produced in murine spleen cell cultures after intravenous injection of Corynebacterium parvum (C. parvum), strain CN 6134 or Bacille Calmette Guérin (BCG). Another strain of C. parvum, CN 5888, which lacks RES stimulating activity and adjuvant activity in vivo, was not effective when injected intravenously. Protein synthesis was required for interferon activity to be produced and protein synthesis was also required for the antiviral state to be expressed. The antiviral activity was relatively stable to pH 2 and neutralized by an antiserum against virus-induced fibroblast interferon, thus exhibiting some properties of type I interferon. In vitro only CN 6134, the biologically active strain, could induce small amounts of interferon in spleen macrophage cultures. Macrophages from CN 6134 or BCG-infected athymic nu/nu mice produced similar interferon titers as their controls. It is concluded that infection with certain immunomodulators can activate splenic macrophages via a predominantly T-cell independent mechanism. Interferon in turn may operate locally as a mediator of immunoregulation.

Animals↗
PubMed1980

Chemical characterization of macrophage cytotoxicity factor, macrophage migration inhibitory factor, T-helper cell-replacing factor and colony-stimulating factor from culture supernatants of concanavalin A-stimulated murine spleen cells.

Supernatants from Concanavalin A-stimulated murine spleen cells were subjected to hydrophobic interaction chromatography on phenyl-Sepharose. Macrophage cytotoxicity factor (MCF), macrophage migration inhibitory factor (MIF), T-helper cell-replacing factor (TRF) and colony-stimulating factor (CSF) were bound at high ionic strength and were released stepwise at low ionic strength. CSF thus could be separated from MCF, MIF and TRF and the bulk of other proteins. Chromatograhy of pools containing MCF, MIF and TRF on Sephadex did not lead to a separation of the three activities which were all found in a molecular weight range of 25.000-55.000. Isoelectric focusing of these pools in pH range from 4 to 9 gave two peaks for MCF in a single sharp peak at pH 5.3. The results demonstrate that the four biological activities can be distinguished on a chemical basis and are accessible for purification and chemical characterization.

Animals↗
PubMed1980

Detection of plasminogen activator in macrophage culture supernatants by a photometric assay.

Plasminogen activator is usually detected indirectly through the lysis of radioiodinated fibrin. Here, it was investigated whether the fibrinolysis test may be substituted by a photometric assay using a synthetic chromogenic substrate (H-D-Val-Leu-Lys-p-nitroanilide; S-2251). Supernatants from cultured murine macrophages served as source of plasminogen activator. It is shown that under standard conditions the photometric assay is about twice as fast and 2--4-fold more sensitive than the fibrinolysis assay. Furthermore, reproducibility of the photometric test was found within 1.5--3.5% standard deviation compared to the fibrinolysis test which was found between 10--20% standard deviation.

Animals↗