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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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Related in: MedlinePlus

Western blot analysis of cadherin protein expression. (A) Percent index variance analysis of the western blot showing cadherin expression: (C1) 2-day old uninfected cultures, (C2) 3-day old uninfected SkMC (control), (I) cultures after 24 h of interaction with T. gondii tachyzoites, and (P) parasites alone (confirming the absence of synthesis of cadherin by T. gondii tachyzoites). Quantitative analysis revealed only 10% reduction in the expression of cadherin between normal cultures, reaching values of more than 50% reduction in T. gondii infected SkMC after 24 h. Results are representative of three independent experiments. Student's T-test (*) p ≤ 0.05.
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Figure 6: Western blot analysis of cadherin protein expression. (A) Percent index variance analysis of the western blot showing cadherin expression: (C1) 2-day old uninfected cultures, (C2) 3-day old uninfected SkMC (control), (I) cultures after 24 h of interaction with T. gondii tachyzoites, and (P) parasites alone (confirming the absence of synthesis of cadherin by T. gondii tachyzoites). Quantitative analysis revealed only 10% reduction in the expression of cadherin between normal cultures, reaching values of more than 50% reduction in T. gondii infected SkMC after 24 h. Results are representative of three independent experiments. Student's T-test (*) p ≤ 0.05.

Mentions: The total cadherin pool was detected using a pan-cadherin-specific antibody, which recognizes the 130 kDa protein [27], since proteins were extracted from 2-3-day-old uninfected cultures (controls) and T. gondii 24 h infected cultures. Quantitative data obtained by densitometric analysis showed that 3-day-old SkMC presented a reduction of only 10% in the synthesis of cadherin when compared to 2-day-old cultures. Regarding the participation of Toxoplasma in the modulation of cadherin synthesis, our data showed a significant decline of cadherin expression after 24 h of T. gondii-SkMC interaction, reaching a 54% reduction. These data demonstrate the variable rate of changes between infected and control SkMC during the analyzed period (Figure 6). For quantitative analysis, western blot signals were normalized against total proteins detected per lane in the corresponding MemCode stained membrane using the QuantityOne software (not shown).


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)

Western blot analysis of cadherin protein expression. (A) Percent index variance analysis of the western blot showing cadherin expression: (C1) 2-day old uninfected cultures, (C2) 3-day old uninfected SkMC (control), (I) cultures after 24 h of interaction with T. gondii tachyzoites, and (P) parasites alone (confirming the absence of synthesis of cadherin by T. gondii tachyzoites). Quantitative analysis revealed only 10% reduction in the expression of cadherin between normal cultures, reaching values of more than 50% reduction in T. gondii infected SkMC after 24 h. Results are representative of three independent experiments. Student's T-test (*) p ≤ 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Figure 6: Western blot analysis of cadherin protein expression. (A) Percent index variance analysis of the western blot showing cadherin expression: (C1) 2-day old uninfected cultures, (C2) 3-day old uninfected SkMC (control), (I) cultures after 24 h of interaction with T. gondii tachyzoites, and (P) parasites alone (confirming the absence of synthesis of cadherin by T. gondii tachyzoites). Quantitative analysis revealed only 10% reduction in the expression of cadherin between normal cultures, reaching values of more than 50% reduction in T. gondii infected SkMC after 24 h. Results are representative of three independent experiments. Student's T-test (*) p ≤ 0.05.
Mentions: The total cadherin pool was detected using a pan-cadherin-specific antibody, which recognizes the 130 kDa protein [27], since proteins were extracted from 2-3-day-old uninfected cultures (controls) and T. gondii 24 h infected cultures. Quantitative data obtained by densitometric analysis showed that 3-day-old SkMC presented a reduction of only 10% in the synthesis of cadherin when compared to 2-day-old cultures. Regarding the participation of Toxoplasma in the modulation of cadherin synthesis, our data showed a significant decline of cadherin expression after 24 h of T. gondii-SkMC interaction, reaching a 54% reduction. These data demonstrate the variable rate of changes between infected and control SkMC during the analyzed period (Figure 6). For quantitative analysis, western blot signals were normalized against total proteins detected per lane in the corresponding MemCode stained membrane using the QuantityOne software (not shown).

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