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A macrophage invasion mechanism for mycobacteria implicating the extracellular domain of CD43.

Fratazzi C, Manjunath N, Arbeit RD, Carini C, Gerken TA, Ardman B, Remold-O'Donnell E, Remold HG - J. Exp. Med. (2000)

Bottom Line: Fluorescence microscopy demonstrated that the associated M. avium had been ingested by the CD43(+/+) M(phi).CD43 expression by the M(phi) was also required for optimal induction by M. avium of tumor necrosis factor (TNF)-alpha production, which likewise could be reconstituted by Galgp.In contrast, interleukin (IL)-10 production by M. avium-infected M(phi) was CD43 independent, demonstrating discordant regulation of TNF-alpha and IL-10.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02111, USA.

ABSTRACT
We studied the role of CD43 (leukosialin/sialophorin), the negatively charged sialoglycoprotein of leukocytes, in the binding of mycobacteria to host cells. CD43-transfected HeLa cells bound Mycobacterium avium, but not Salmonella typhimurium or Shigella flexneri. Quantitative bacteriology showed that macrophages (M(phi)) from wild-type mice (CD43(+/+)) bound M. avium, Mycobacterium bovis (bacillus Calmette-Guérin), and Mycobacterium tuberculosis (strain H37Rv), whereas M(phi) from CD43 knockout mice (CD43(-/)-) did not. Fluorescence microscopy demonstrated that the associated M. avium had been ingested by the CD43(+/+) M(phi). The inability of CD43(-/)- M(phi) to bind M. avium could be restored by addition of galactoglycoprotein (Galgp), the extracellular mucin portion of CD43. The effect of Galgp is not due to opsonization of the bacteria, but required its interaction with the M(phi) other mucins had no effect. CD43 expression by the M(phi) was also required for optimal induction by M. avium of tumor necrosis factor (TNF)-alpha production, which likewise could be reconstituted by Galgp. In contrast, interleukin (IL)-10 production by M. avium-infected M(phi) was CD43 independent, demonstrating discordant regulation of TNF-alpha and IL-10. These findings describe a novel role of CD43 in promoting stable interaction of mycobacteria with receptors on the M(phi) enabling the cells to respond specifically with TNF-alpha production.

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Addition of Galgp restores binding of M. avium by CD43−/− Mφ. (A) M. avium were incubated for varying times with CD43−/− Mφ in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Bacteria/cell ratio was 2:1. The Mφ were harvested and extensively washed, and adherent bacteria were quantified. The binding of M. avium to CD43+/+ Mφ in the absence of Galgp is shown for comparison (•, dashed lines). (B) M. avium at varying bacteria/cell ratios as indicated were incubated with CD43−/− Mφ for 4 h in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Shown are mean values ± SEM for three mice of each group. The number of mycobacteria associated with CD43−/− Mφ in the presence and absence of 100 μg/ml Galgp was significantly different in all conditions (P < 0.0001). Comparable effects of Galgp addition were observed in two additional experiments. Galgp at 200 μg/ml produced similar results (data not shown).
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Figure 4: Addition of Galgp restores binding of M. avium by CD43−/− Mφ. (A) M. avium were incubated for varying times with CD43−/− Mφ in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Bacteria/cell ratio was 2:1. The Mφ were harvested and extensively washed, and adherent bacteria were quantified. The binding of M. avium to CD43+/+ Mφ in the absence of Galgp is shown for comparison (•, dashed lines). (B) M. avium at varying bacteria/cell ratios as indicated were incubated with CD43−/− Mφ for 4 h in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Shown are mean values ± SEM for three mice of each group. The number of mycobacteria associated with CD43−/− Mφ in the presence and absence of 100 μg/ml Galgp was significantly different in all conditions (P < 0.0001). Comparable effects of Galgp addition were observed in two additional experiments. Galgp at 200 μg/ml produced similar results (data not shown).

Mentions: The extracellular mucin region of CD43 was identified as a normal component of human plasma, given the name Galgp, and isolated 17. We reasoned that if CD43 functions as a receptor for mycobacteria, addition of soluble extracellular CD43 to wild-type Mφ might abrogate mycobacterial binding by competing with cell surface CD43. However, addition of Galgp to CD43+/+ Mφ resulted in enhanced mycobacterial binding (not shown). More importantly, addition of 100 μg/ml Galgp to CD43−/− Mφ at the time of infection restored the time-dependent (Fig. 4 A) and bacterial dose–dependent (Fig. 4 B) association of M. avium with CD43−/− Mφ. Restoration of M. avium binding to CD43−/− Mφ by Galgp (100 μg/ml) was also seen when the cells were evaluated by acid-fast staining (data not shown). These findings suggest that Mφ CD43, rather than serving as a receptor for mycobacteria, functions by promoting or stabilizing binding or uptake of mycobacteria.


A macrophage invasion mechanism for mycobacteria implicating the extracellular domain of CD43.

Fratazzi C, Manjunath N, Arbeit RD, Carini C, Gerken TA, Ardman B, Remold-O'Donnell E, Remold HG - J. Exp. Med. (2000)

Addition of Galgp restores binding of M. avium by CD43−/− Mφ. (A) M. avium were incubated for varying times with CD43−/− Mφ in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Bacteria/cell ratio was 2:1. The Mφ were harvested and extensively washed, and adherent bacteria were quantified. The binding of M. avium to CD43+/+ Mφ in the absence of Galgp is shown for comparison (•, dashed lines). (B) M. avium at varying bacteria/cell ratios as indicated were incubated with CD43−/− Mφ for 4 h in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Shown are mean values ± SEM for three mice of each group. The number of mycobacteria associated with CD43−/− Mφ in the presence and absence of 100 μg/ml Galgp was significantly different in all conditions (P < 0.0001). Comparable effects of Galgp addition were observed in two additional experiments. Galgp at 200 μg/ml produced similar results (data not shown).
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Related In: Results  -  Collection

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Figure 4: Addition of Galgp restores binding of M. avium by CD43−/− Mφ. (A) M. avium were incubated for varying times with CD43−/− Mφ in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Bacteria/cell ratio was 2:1. The Mφ were harvested and extensively washed, and adherent bacteria were quantified. The binding of M. avium to CD43+/+ Mφ in the absence of Galgp is shown for comparison (•, dashed lines). (B) M. avium at varying bacteria/cell ratios as indicated were incubated with CD43−/− Mφ for 4 h in the absence (○) and presence (⋄) of 100 μg/ml Galgp. Shown are mean values ± SEM for three mice of each group. The number of mycobacteria associated with CD43−/− Mφ in the presence and absence of 100 μg/ml Galgp was significantly different in all conditions (P < 0.0001). Comparable effects of Galgp addition were observed in two additional experiments. Galgp at 200 μg/ml produced similar results (data not shown).
Mentions: The extracellular mucin region of CD43 was identified as a normal component of human plasma, given the name Galgp, and isolated 17. We reasoned that if CD43 functions as a receptor for mycobacteria, addition of soluble extracellular CD43 to wild-type Mφ might abrogate mycobacterial binding by competing with cell surface CD43. However, addition of Galgp to CD43+/+ Mφ resulted in enhanced mycobacterial binding (not shown). More importantly, addition of 100 μg/ml Galgp to CD43−/− Mφ at the time of infection restored the time-dependent (Fig. 4 A) and bacterial dose–dependent (Fig. 4 B) association of M. avium with CD43−/− Mφ. Restoration of M. avium binding to CD43−/− Mφ by Galgp (100 μg/ml) was also seen when the cells were evaluated by acid-fast staining (data not shown). These findings suggest that Mφ CD43, rather than serving as a receptor for mycobacteria, functions by promoting or stabilizing binding or uptake of mycobacteria.

Bottom Line: Fluorescence microscopy demonstrated that the associated M. avium had been ingested by the CD43(+/+) M(phi).CD43 expression by the M(phi) was also required for optimal induction by M. avium of tumor necrosis factor (TNF)-alpha production, which likewise could be reconstituted by Galgp.In contrast, interleukin (IL)-10 production by M. avium-infected M(phi) was CD43 independent, demonstrating discordant regulation of TNF-alpha and IL-10.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02111, USA.

ABSTRACT
We studied the role of CD43 (leukosialin/sialophorin), the negatively charged sialoglycoprotein of leukocytes, in the binding of mycobacteria to host cells. CD43-transfected HeLa cells bound Mycobacterium avium, but not Salmonella typhimurium or Shigella flexneri. Quantitative bacteriology showed that macrophages (M(phi)) from wild-type mice (CD43(+/+)) bound M. avium, Mycobacterium bovis (bacillus Calmette-Guérin), and Mycobacterium tuberculosis (strain H37Rv), whereas M(phi) from CD43 knockout mice (CD43(-/)-) did not. Fluorescence microscopy demonstrated that the associated M. avium had been ingested by the CD43(+/+) M(phi). The inability of CD43(-/)- M(phi) to bind M. avium could be restored by addition of galactoglycoprotein (Galgp), the extracellular mucin portion of CD43. The effect of Galgp is not due to opsonization of the bacteria, but required its interaction with the M(phi) other mucins had no effect. CD43 expression by the M(phi) was also required for optimal induction by M. avium of tumor necrosis factor (TNF)-alpha production, which likewise could be reconstituted by Galgp. In contrast, interleukin (IL)-10 production by M. avium-infected M(phi) was CD43 independent, demonstrating discordant regulation of TNF-alpha and IL-10. These findings describe a novel role of CD43 in promoting stable interaction of mycobacteria with receptors on the M(phi) enabling the cells to respond specifically with TNF-alpha production.

Show MeSH
Related in: MedlinePlus