Limits...
Selective suppression of interleukin-12 induction after macrophage receptor ligation.

Sutterwala FS, Noel GJ, Clynes R, Mosser DM - J. Exp. Med. (1997)

Bottom Line: The results of several different experimental approaches suggest that IL-12 downregulation was due to extracellular calcium influxes that resulted from receptor ligation.Finally, bone marrow-derived macrophages from FcR gamma chain-deficient mice, which fail to flux calcium after receptor ligation, failed to inhibit IL-12(p40) mRNA induction.These results indicate that the calcium influxes that occur as a result of receptor ligation are responsible for inhibiting the induction of IL-12 by LPS.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.

ABSTRACT
Interleukin (IL)-12 is a monocyte- and macrophage-derived cytokine that plays a crucial role in both the innate and the acquired immune response. In this study, we examined the effects that ligating specific macrophage receptors had on the induction of IL-12 by lipopolysaccharide (LPS). We report that ligation of the macrophage Fcgamma, complement, or scavenger receptors inhibited the induction of IL-12 by LPS. Both mRNA synthesis and protein secretion were diminished to near-undetectable levels following receptor ligation. Suppression was specific to IL-12 since IL-10 and tumor necrosis factor-alpha (TNF-alpha) production were not inhibited by ligating macrophage receptors. The results of several different experimental approaches suggest that IL-12 downregulation was due to extracellular calcium influxes that resulted from receptor ligation. First, preventing extracellular calcium influxes, by performing the assays in EGTA, abrogated FcgammaR-mediated IL-12(p40) mRNA suppression. Second, exposure of macrophages to the calcium ionophores, ionomycin or A23187, mimicked receptor ligation and inhibited IL-12(p40) mRNA induction by LPS. Finally, bone marrow-derived macrophages from FcR gamma chain-deficient mice, which fail to flux calcium after receptor ligation, failed to inhibit IL-12(p40) mRNA induction. These results indicate that the calcium influxes that occur as a result of receptor ligation are responsible for inhibiting the induction of IL-12 by LPS. Hence, the ligation of phagocytic receptors on macrophages can lead to a dramatic decrease in IL-12 induction. This downregulation may be a way of limiting proinflammatory responses of macrophages to extracellular pathogens, or suppressing the development of cell-mediated immunity to intracellular pathogens.

Show MeSH

Related in: MedlinePlus

FcγR ligation on BMMφ from FcR γ−/− mice. (A)  BMMφ were preincubated with mAb 2.4G2 before stimulation with the  secondary antibody, goat anti–rat IgG mAb. Relative fluorescence indicative of intracellular Ca2+ is shown for wild-type cells (C57BL/6) and FcR  γ−/− cells. (B) BMMφ from FcR γ−/− or C57BL/6 mice were exposed to LPS alone or LPS in combination with E-IgG. 6 h after the addition of stimuli, total RNA was isolated and used to carry out competitive RT-PCR, using primers for HPRT, or the inducible p40 subunit of  IL-12, as described in Materials and Methods. Results are representative  of three separate experiments.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2196339&req=5

Figure 7: FcγR ligation on BMMφ from FcR γ−/− mice. (A) BMMφ were preincubated with mAb 2.4G2 before stimulation with the secondary antibody, goat anti–rat IgG mAb. Relative fluorescence indicative of intracellular Ca2+ is shown for wild-type cells (C57BL/6) and FcR γ−/− cells. (B) BMMφ from FcR γ−/− or C57BL/6 mice were exposed to LPS alone or LPS in combination with E-IgG. 6 h after the addition of stimuli, total RNA was isolated and used to carry out competitive RT-PCR, using primers for HPRT, or the inducible p40 subunit of IL-12, as described in Materials and Methods. Results are representative of three separate experiments.

Mentions: We next examined IL-12 production in BMMφ from FcR γ−/− mice. Macrophages from these mice bind IgG-opsonized erythrocytes normally (data not shown) but lack the ability to efficiently phagocytose IgG-opsonized erythrocytes (33). The deletion of the FcR γ chain profoundly impairs signaling through the FcγR (33). Unlike wild-type cells, which rapidly flux calcium upon FcγR ligation, BMMφ from FcR γ−/− mice fail to cause calcium fluxes following FcγR ligation (Fig. 7 A). Macrophages from normal and FcR γ−/− mice were incubated in LPS alone or with LPS and E-IgG (Fig. 7 B). Both C57BL/6 and FcR γ−/− BMMφ expressed IL-12(p40) mRNA after LPS treatment. As expected, ligation of FcγR on C57BL/6 BMMφ resulted in a marked suppression of LPS-mediated IL-12(p40) mRNA induction. However, the ligation of FcγR on γ−/− BMMφ did not inhibit the induction of IL-12(p40) mRNA by LPS. These results correlate signaling through the FcγR with the downregulation of IL-12 production after receptor ligation.


Selective suppression of interleukin-12 induction after macrophage receptor ligation.

Sutterwala FS, Noel GJ, Clynes R, Mosser DM - J. Exp. Med. (1997)

FcγR ligation on BMMφ from FcR γ−/− mice. (A)  BMMφ were preincubated with mAb 2.4G2 before stimulation with the  secondary antibody, goat anti–rat IgG mAb. Relative fluorescence indicative of intracellular Ca2+ is shown for wild-type cells (C57BL/6) and FcR  γ−/− cells. (B) BMMφ from FcR γ−/− or C57BL/6 mice were exposed to LPS alone or LPS in combination with E-IgG. 6 h after the addition of stimuli, total RNA was isolated and used to carry out competitive RT-PCR, using primers for HPRT, or the inducible p40 subunit of  IL-12, as described in Materials and Methods. Results are representative  of three separate experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196339&req=5

Figure 7: FcγR ligation on BMMφ from FcR γ−/− mice. (A) BMMφ were preincubated with mAb 2.4G2 before stimulation with the secondary antibody, goat anti–rat IgG mAb. Relative fluorescence indicative of intracellular Ca2+ is shown for wild-type cells (C57BL/6) and FcR γ−/− cells. (B) BMMφ from FcR γ−/− or C57BL/6 mice were exposed to LPS alone or LPS in combination with E-IgG. 6 h after the addition of stimuli, total RNA was isolated and used to carry out competitive RT-PCR, using primers for HPRT, or the inducible p40 subunit of IL-12, as described in Materials and Methods. Results are representative of three separate experiments.
Mentions: We next examined IL-12 production in BMMφ from FcR γ−/− mice. Macrophages from these mice bind IgG-opsonized erythrocytes normally (data not shown) but lack the ability to efficiently phagocytose IgG-opsonized erythrocytes (33). The deletion of the FcR γ chain profoundly impairs signaling through the FcγR (33). Unlike wild-type cells, which rapidly flux calcium upon FcγR ligation, BMMφ from FcR γ−/− mice fail to cause calcium fluxes following FcγR ligation (Fig. 7 A). Macrophages from normal and FcR γ−/− mice were incubated in LPS alone or with LPS and E-IgG (Fig. 7 B). Both C57BL/6 and FcR γ−/− BMMφ expressed IL-12(p40) mRNA after LPS treatment. As expected, ligation of FcγR on C57BL/6 BMMφ resulted in a marked suppression of LPS-mediated IL-12(p40) mRNA induction. However, the ligation of FcγR on γ−/− BMMφ did not inhibit the induction of IL-12(p40) mRNA by LPS. These results correlate signaling through the FcγR with the downregulation of IL-12 production after receptor ligation.

Bottom Line: The results of several different experimental approaches suggest that IL-12 downregulation was due to extracellular calcium influxes that resulted from receptor ligation.Finally, bone marrow-derived macrophages from FcR gamma chain-deficient mice, which fail to flux calcium after receptor ligation, failed to inhibit IL-12(p40) mRNA induction.These results indicate that the calcium influxes that occur as a result of receptor ligation are responsible for inhibiting the induction of IL-12 by LPS.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Temple University School of Medicine, Philadelphia, Pennsylvania 19140, USA.

ABSTRACT
Interleukin (IL)-12 is a monocyte- and macrophage-derived cytokine that plays a crucial role in both the innate and the acquired immune response. In this study, we examined the effects that ligating specific macrophage receptors had on the induction of IL-12 by lipopolysaccharide (LPS). We report that ligation of the macrophage Fcgamma, complement, or scavenger receptors inhibited the induction of IL-12 by LPS. Both mRNA synthesis and protein secretion were diminished to near-undetectable levels following receptor ligation. Suppression was specific to IL-12 since IL-10 and tumor necrosis factor-alpha (TNF-alpha) production were not inhibited by ligating macrophage receptors. The results of several different experimental approaches suggest that IL-12 downregulation was due to extracellular calcium influxes that resulted from receptor ligation. First, preventing extracellular calcium influxes, by performing the assays in EGTA, abrogated FcgammaR-mediated IL-12(p40) mRNA suppression. Second, exposure of macrophages to the calcium ionophores, ionomycin or A23187, mimicked receptor ligation and inhibited IL-12(p40) mRNA induction by LPS. Finally, bone marrow-derived macrophages from FcR gamma chain-deficient mice, which fail to flux calcium after receptor ligation, failed to inhibit IL-12(p40) mRNA induction. These results indicate that the calcium influxes that occur as a result of receptor ligation are responsible for inhibiting the induction of IL-12 by LPS. Hence, the ligation of phagocytic receptors on macrophages can lead to a dramatic decrease in IL-12 induction. This downregulation may be a way of limiting proinflammatory responses of macrophages to extracellular pathogens, or suppressing the development of cell-mediated immunity to intracellular pathogens.

Show MeSH
Related in: MedlinePlus