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Toxoplasma gondii down modulates cadherin expression in skeletal muscle cells inhibiting myogenesis.

Gomes AF, Guimarães EV, Carvalho L, Correa JR, Mendonça-Lima L, Barbosa HS - BMC Microbiol. (2011)

Bottom Line: Immunofluorescence and immunoblotting analysis of parasite-host cell interaction showed a 54% reduction in cadherin expression at 24 h of infection.Concomitantly, a reduction in M-cadherin mRNA levels was observed after 3 and 24 h of T. gondii-host cell interaction.These data suggest that T. gondii is able to down regulate M-cadherin expression, leading to molecular modifications in the host cell surface that interfere with membrane fusion and consequently affect the myogenesis process.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratório de Biologia Estrutural, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, (Av, Brasil 4365), Rio de Janeiro (21040-361), Brazil.

ABSTRACT

Background: Toxoplasma gondii belongs to a large and diverse group of obligate intracellular parasitic protozoa. Primary culture of mice skeletal muscle cells (SkMC) was employed as a model for experimental toxoplasmosis studies. The myogenesis of SkMC was reproduced in vitro and the ability of T. gondii tachyzoite forms to infect myoblasts and myotubes and its influence on SkMC myogenesis were analyzed.

Results: In this study we show that, after 24 h of interaction, myoblasts (61%) were more infected with T. gondii than myotubes (38%) and inhibition of myogenesis was about 75%. The role of adhesion molecules such as cadherin in this event was investigated. First, we demonstrate that cadherin localization was restricted to the contact areas between myocytes/myocytes and myocytes/myotubes during the myogenesis process. Immunofluorescence and immunoblotting analysis of parasite-host cell interaction showed a 54% reduction in cadherin expression at 24 h of infection. Concomitantly, a reduction in M-cadherin mRNA levels was observed after 3 and 24 h of T. gondii-host cell interaction.

Conclusions: These data suggest that T. gondii is able to down regulate M-cadherin expression, leading to molecular modifications in the host cell surface that interfere with membrane fusion and consequently affect the myogenesis process.

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Cadherin distribution in SkMC after 24 h of T. gondii interaction. Confocal Microscopy analysis showing: (A) In 3-day-old SkMC cultures, after differentiation, myoblasts present intense cadherin labeling at the contact points (arrows). (B and C) In myoblasts after 24 h of interaction with T. gondii (thick arrow), cadherin (thin arrow) becomes disorganized forming aggregates at different sites, around and inside the parasitophorous vacuole (for detail, see inset). (D) Infected myoblasts after 24 h of interaction with T. gondii have little or no labeling for cadherin at points of cell-cell contact (thick arrow). Note that only uninfected cells show strong cadherin expression (thin arrow). Nuclei of cells and parasites labeled with DAPI, in blue. Bars, 20 μm
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Figure 4: Cadherin distribution in SkMC after 24 h of T. gondii interaction. Confocal Microscopy analysis showing: (A) In 3-day-old SkMC cultures, after differentiation, myoblasts present intense cadherin labeling at the contact points (arrows). (B and C) In myoblasts after 24 h of interaction with T. gondii (thick arrow), cadherin (thin arrow) becomes disorganized forming aggregates at different sites, around and inside the parasitophorous vacuole (for detail, see inset). (D) Infected myoblasts after 24 h of interaction with T. gondii have little or no labeling for cadherin at points of cell-cell contact (thick arrow). Note that only uninfected cells show strong cadherin expression (thin arrow). Nuclei of cells and parasites labeled with DAPI, in blue. Bars, 20 μm

Mentions: Indirect immunofluorescence assays were performed in order to localize cadherin, an adhesion molecule involved in homophilic recognition during myoblast and myotube fusion. In SkMC 2-day-old cultures, the myoblasts are still in multiplication and differentiation process. Cadherin is strongly revealed in every cell with higher fluorescence intensity in edges near the membrane and at the point of cell-cell contact (Figure 3A). Apparently, the existence of a single, newly internalized parasite did not lead to any change in the profile of cadherin distribution in host cells (Figure 3B), as demonstrated by immunofluorescence microscopy. The same results were maintained during the first 3 h of interaction (data not shown). After differentiation, myoblasts revealed cadherin highly concentrated at the cell-cell contact point (Figure 4A). However, this profile was not observed after 24 h of T. gondii infection. Besides disorganization, cadherin appeared in aggregates at different points of the SkMC, including around and inside the parasitophorous vacuole (Figure 4B and 4C - inset). Infected myoblasts showed low or no labeling for cadherin at cell-cell contact point (Figure 4B and inset and C). Even in cultures infected for 36 h, only uninfected cells present strong cadherin expression (Figure 4D).


Toxoplasma gondii down modulates cadherin expression in skeletal muscle cells inhibiting myogenesis.

Gomes AF, Guimarães EV, Carvalho L, Correa JR, Mendonça-Lima L, Barbosa HS - BMC Microbiol. (2011)

Cadherin distribution in SkMC after 24 h of T. gondii interaction. Confocal Microscopy analysis showing: (A) In 3-day-old SkMC cultures, after differentiation, myoblasts present intense cadherin labeling at the contact points (arrows). (B and C) In myoblasts after 24 h of interaction with T. gondii (thick arrow), cadherin (thin arrow) becomes disorganized forming aggregates at different sites, around and inside the parasitophorous vacuole (for detail, see inset). (D) Infected myoblasts after 24 h of interaction with T. gondii have little or no labeling for cadherin at points of cell-cell contact (thick arrow). Note that only uninfected cells show strong cadherin expression (thin arrow). Nuclei of cells and parasites labeled with DAPI, in blue. Bars, 20 μm
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3116462&req=5

Figure 4: Cadherin distribution in SkMC after 24 h of T. gondii interaction. Confocal Microscopy analysis showing: (A) In 3-day-old SkMC cultures, after differentiation, myoblasts present intense cadherin labeling at the contact points (arrows). (B and C) In myoblasts after 24 h of interaction with T. gondii (thick arrow), cadherin (thin arrow) becomes disorganized forming aggregates at different sites, around and inside the parasitophorous vacuole (for detail, see inset). (D) Infected myoblasts after 24 h of interaction with T. gondii have little or no labeling for cadherin at points of cell-cell contact (thick arrow). Note that only uninfected cells show strong cadherin expression (thin arrow). Nuclei of cells and parasites labeled with DAPI, in blue. Bars, 20 μm
Mentions: Indirect immunofluorescence assays were performed in order to localize cadherin, an adhesion molecule involved in homophilic recognition during myoblast and myotube fusion. In SkMC 2-day-old cultures, the myoblasts are still in multiplication and differentiation process. Cadherin is strongly revealed in every cell with higher fluorescence intensity in edges near the membrane and at the point of cell-cell contact (Figure 3A). Apparently, the existence of a single, newly internalized parasite did not lead to any change in the profile of cadherin distribution in host cells (Figure 3B), as demonstrated by immunofluorescence microscopy. The same results were maintained during the first 3 h of interaction (data not shown). After differentiation, myoblasts revealed cadherin highly concentrated at the cell-cell contact point (Figure 4A). However, this profile was not observed after 24 h of T. gondii infection. Besides disorganization, cadherin appeared in aggregates at different points of the SkMC, including around and inside the parasitophorous vacuole (Figure 4B and 4C - inset). Infected myoblasts showed low or no labeling for cadherin at cell-cell contact point (Figure 4B and inset and C). Even in cultures infected for 36 h, only uninfected cells present strong cadherin expression (Figure 4D).

Bottom Line: Immunofluorescence and immunoblotting analysis of parasite-host cell interaction showed a 54% reduction in cadherin expression at 24 h of infection.Concomitantly, a reduction in M-cadherin mRNA levels was observed after 3 and 24 h of T. gondii-host cell interaction.These data suggest that T. gondii is able to down regulate M-cadherin expression, leading to molecular modifications in the host cell surface that interfere with membrane fusion and consequently affect the myogenesis process.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratório de Biologia Estrutural, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, (Av, Brasil 4365), Rio de Janeiro (21040-361), Brazil.

ABSTRACT

Background: Toxoplasma gondii belongs to a large and diverse group of obligate intracellular parasitic protozoa. Primary culture of mice skeletal muscle cells (SkMC) was employed as a model for experimental toxoplasmosis studies. The myogenesis of SkMC was reproduced in vitro and the ability of T. gondii tachyzoite forms to infect myoblasts and myotubes and its influence on SkMC myogenesis were analyzed.

Results: In this study we show that, after 24 h of interaction, myoblasts (61%) were more infected with T. gondii than myotubes (38%) and inhibition of myogenesis was about 75%. The role of adhesion molecules such as cadherin in this event was investigated. First, we demonstrate that cadherin localization was restricted to the contact areas between myocytes/myocytes and myocytes/myotubes during the myogenesis process. Immunofluorescence and immunoblotting analysis of parasite-host cell interaction showed a 54% reduction in cadherin expression at 24 h of infection. Concomitantly, a reduction in M-cadherin mRNA levels was observed after 3 and 24 h of T. gondii-host cell interaction.

Conclusions: These data suggest that T. gondii is able to down regulate M-cadherin expression, leading to molecular modifications in the host cell surface that interfere with membrane fusion and consequently affect the myogenesis process.

Show MeSH
Related in: MedlinePlus