Maintenance of granuloma macrophages in serum-free medium

Fauve, Robert M.; Jusforgues, Hélène; Hévin, Brigitte

doi:10.1016/0022-1759(83)90442-8

Cited by 58 publications

(30 citation statements)

References 7 publications

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“…Mouse bone marrow-derived M (BMM) were cultured in RPMI 1640 medium supplemented with 15% L-cell-conditioned medium as described previously (51). Mouse resident peritoneal M (RPM), Biogel-elicited peritoneal M (BioM), thioglycollate-elicited peritoneal M (ThioM), and thioglycollate-elicited peritoneal neutrophils (ThioPMN) were obtained using standard protocols described previously (52,53). For the cytokine treatment of RPM, cells were cultured for 1 h for adherence, washed, and then treated with or without IL-4 (20 ng/ml), IL-10 (10 ng/ml), TNF-␣ (10 ng/ml), and inter- 2 Lin et al, manuscript in preparation.…”

Section: Methodsmentioning

confidence: 99%

EMR4, a Novel Epidermal Growth Factor (EGF)-TM7 Molecule Up-regulated in Activated Mouse Macrophages, Binds to a Putative Cellular Ligand on B Lymphoma Cell Line A20

Stacey¹,

Chang²,

Sanos³

et al. 2002

Journal of Biological Chemistry

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A novel member of the EGF-TM7 family, mEMR4, was identified and characterized. The full-length mouse EMR4 cDNA encodes a predicted 689-amino acid protein containing two epidermal growth factor (EGF)-like modules, a mucin-like spacer domain, and a seventransmembrane domain with a cytoplasmic tail. Genetic mapping established that mEMR4 is localized in the distal region of mouse chromosome 17 in close proximity to another EGF-TM7 gene, F4/80 (Emr1). Similar to F4/80, mEMR4 is predominantly expressed on resident macrophages. However, a much lower expression level was also detected in thioglycollateelicited peritoneal neutrophils and bone marrowderived dendritic cells. The expression of mEMR4 is up-regulated following macrophage activation in Biogel and thioglycollate-elicited peritoneal macrophages. Similarly, mEMR4 is over-expressed in TNF-␣-treated resident peritoneal macrophages, whereas interleukin-4 and -10 dramatically reduce the expression. mEMR4 was found to undergo proteolytic processing within the extracellular stalk region resulting in two protein subunits associated noncovalently as a heterodimer. The proteolytic cleavage site was identified by N-terminal amino acid sequencing and located at the conserved GPCR (G protein-coupled receptor) proteolytic site in the extracellular region. Using multivalent biotinylated mEMR4-mFc fusion proteins as a probe, a putative cell surface ligand was identified on a B lymphoma cell line, A20, in a cell-binding assay. The mEMR4-ligand interaction is Ca 2؉ -independent and is mediated predominantly by the second EGF-like module. mEMR4 is the first EGF-TM7 receptor known to mediate the cellular interaction between myeloid cells and B cells.

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Section: Methodsmentioning

confidence: 99%

EMR4, a Novel Epidermal Growth Factor (EGF)-TM7 Molecule Up-regulated in Activated Mouse Macrophages, Binds to a Putative Cellular Ligand on B Lymphoma Cell Line A20

Stacey¹,

Chang²,

Sanos³

et al. 2002

Journal of Biological Chemistry

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“…A quantitative method for measuring the intensity of AIR based on the determination of leukocyte and protein concentration in the local exudate was described by Fauve et al [21,221. The local AIR is induced by subcutaneous injection of a suspension of polyacrylamide microbeads, which is an entirely insoluble, chemically inert, nonantigenic substrate, acting as a neutral foreign material.…”

Section: Introductionmentioning

confidence: 99%

Genetics of nonspecific immunity: I. Bidirectional selective breeding of lines of mice endowed with maximal or minimal inflammatory responsiveness

et al. 1992

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The genetic regulation of acute inflammatory reaction (AIR) was studied by the method of bidirectional selective breeding, used to produce a line of mice giving the maximal and a line of mice giving the minimal inflammatory reaction (AIR max and AIR min, respectively). The AIR was triggered by subcutaneous injection of a neutral substrate (suspension of polyacrylamide microbeads), and measured by the leukocyte and serum protein accumulation in the exudate. The two parameters are positively correlated and present a normal frequency distribution. The highly genetically heterogeneous foundation population was produced by the equipoised intercrossing of eight inbred strains of mice, and selective breeding carried out by assortative matings of extreme phenotypes. The response to selection in 11 consecutive generations was highly asymmetrical: a marked AIR increase in the AIR max and no change in the AIR min line occurred. The mean value of realized heritability in the AIR max line was 0.26 and 0.18 for cell and protein concentrations, respectively. The response to selection must have resulted from the interaction of seven to nine independent gene loci endowed with additive effects. The lack of response to selection of the AIR min line is discussed. The large inter-line difference opens new possibilities for studying the biochemistry and molecular genetics of inflammation, and also for investigating the beneficial or detrimental effect of inflammatory responses.

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“…3LL cells and macrophages were then added at a ratio of 1:10 and incubated for 24 hr. Cells were then pulsed-labeled with 0.04 ,uCi of 125I-labeled deoxyuridine (specific activity, 5 Ci/mg;…”

Section: Methodsmentioning

confidence: 99%

Immunostimulatory human urinary protein.

Fontan

Saklani-Jusforgues

Fauve

1992

Proc. Natl. Acad. Sci. U.S.A.

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We have previously extracted a protein from inflammatory mouse granuloma that fully protects normal mice against lethal Listeria monocytogenes infection. We now report that polyclonal antibodies directed against this protein react with a human urinary fraction that also provides full protection of normal mice against L. monocytogenes. Murine monoclonal antibodies completely inhibit the protective activity of the human urinary fraction. We have purified to apparent homogeneity a human glycoprotein of 43 kDa (HGP.43), pI = injection of C57BL/6 mice with Bio-Gel P-100 (Bio-Rad), the gel was withdrawn and the inflammatory cells were recovered by filtration through a 400-mesh screen. The cells were centrifuged (250 x g for 10 min), washed twice with ice-cold Dulbecco's modified Eagle's medium (DMEM), incubated (1 x 106 cells per ml, 0.5 ml per culture tube) for 24 hr, and washed to remove polymorphonuclear cells. Lewis tumor cells (3) were isolated from murine lung metastases and cultured in plastic Petri dishes in DMEM containing 10% (vol/vol) fetal calf serum at 3TC in an atmosphere of 7.5% C02/92.5% air.Antibodies. Polyclonal rabbit anti-mouse granuloma protein antibodies were prepared as described by Fontan et al. (1). Polyclonal antibodies against the human urinary protein were raised in rabbits by s.c. injection of 10 ,ug in Freund's complete adjuvant and twice-monthly booster injections. Immunoglobulins were purified by precipitation in (NH4)2SO4 at 33% saturation and DEAE-gel chromatography. Antibodies against human al-AGP were purified by means ofimmunoaffinity chromatography using pure a1-AGP (Sigma) coupled to CNBr-activated Sepharose. Monoclonal antibodies against the human urinary protein were prepared as follows. Female BALB/c mice were immunized by s.c. injection of 50 jug of a semipurified active human urinary fraction. Spleen cells were fused with the SP2/0 BALB/c myeloma cell line. Wells with hybrid growth were screened for reactivity with the urinary fraction by using an ELISA. Positive wells were subcloned twice by limiting dilution. One of the subclones (K52.5G2) was chosen for the neutralization test. This clone produces an IgG1 that was purified by (NH4)2SO4 precipitation and DEAE-gel chromatography. The supernatants and antibody-free controls were tested in mice challenged with a lethal inoculum ofL. monocytogenes.All the materials, solutions, and tissue culture media were free of endotoxin.Statistics. Results are expressed as the mean ± SEM. The significance of the differences between experimental groups was analyzed by Student's t test. A P value of <0.05 was considered significant. RESULTS Purification and Characterization of the Urinary Protein. After Amicon ultrafiltration concentration of the urines, the material was chromatographed on a DEAE-Sephacel anionexchange column and eluted with 0.2 and 1 M NaCl. All the immunostimulating activity was eluted with 0.2 M NaCI (Fig. 1). The active fractions were pooled, dialyzed, and loaded onto a Cibacron blue column to remove contaminat...

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Maintenance of granuloma macrophages in serum-free medium

Cited by 58 publications

References 7 publications

EMR4, a Novel Epidermal Growth Factor (EGF)-TM7 Molecule Up-regulated in Activated Mouse Macrophages, Binds to a Putative Cellular Ligand on B Lymphoma Cell Line A20

EMR4, a Novel Epidermal Growth Factor (EGF)-TM7 Molecule Up-regulated in Activated Mouse Macrophages, Binds to a Putative Cellular Ligand on B Lymphoma Cell Line A20

Genetics of nonspecific immunity: I. Bidirectional selective breeding of lines of mice endowed with maximal or minimal inflammatory responsiveness

Immunostimulatory human urinary protein.

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