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Productive infection of human skeletal muscle cells by pandemic and seasonal influenza A(H1N1) viruses.

Desdouits M, Munier S, Prevost MC, Jeannin P, Butler-Browne G, Ozden S, Gessain A, Van Der Werf S, Naffakh N, Ceccaldi PE - PLoS ONE (2013)

Bottom Line: The pathogenesis of these influenza-associated myopathies (IAM) remains unkown, although the direct infection of muscle cells is suspected.Infection led to a cytopathic effect with the lysis of muscle cells, as characterized by the release of lactate dehydrogenase.Our results are compatible with the hypothesis of a direct muscle infection causing rhabdomyolysis in IAM patients.

View Article: PubMed Central - PubMed

Affiliation: Unité Epidémiologie et Physiopathologie des Virus Oncogènes, Institut Pasteur, Paris, France ; UMR 3569, CNRS, Paris, France ; Cellule Pasteur, Université Paris Diderot, Sorbonne Paris Cité, Paris, France.

ABSTRACT
Besides the classical respiratory and systemic symptoms, unusual complications of influenza A infection in humans involve the skeletal muscles. Numerous cases of acute myopathy and/or rhabdomyolysis have been reported, particularly following the outbreak of pandemic influenza A(H1N1) in 2009. The pathogenesis of these influenza-associated myopathies (IAM) remains unkown, although the direct infection of muscle cells is suspected. Here, we studied the susceptibility of cultured human primary muscle cells to a 2009 pandemic and a 2008 seasonal influenza A(H1N1) isolate. Using cells from different donors, we found that differentiated muscle cells (i. e. myotubes) were highly susceptible to infection by both influenza A(H1N1) isolates, whereas undifferentiated cells (i. e. myoblasts) were partially resistant. The receptors for influenza viruses, α2-6 and α2-3 linked sialic acids, were detected on the surface of myotubes and myoblasts. Time line of viral nucleoprotein (NP) expression and nuclear export showed that the first steps of the viral replication cycle could take place in muscle cells. Infected myotubes and myoblasts exhibited budding virions and nuclear inclusions as observed by transmission electron microscopy and correlative light and electron microscopy. Myotubes, but not myoblasts, yielded infectious virus progeny that could further infect naive muscle cells after proteolytic treatment. Infection led to a cytopathic effect with the lysis of muscle cells, as characterized by the release of lactate dehydrogenase. The secretion of proinflammatory cytokines by muscle cells was not affected following infection. Our results are compatible with the hypothesis of a direct muscle infection causing rhabdomyolysis in IAM patients.

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Secretion of 5 cytokines and 1 chemokine by infected muscle cells.The concentrations of IFNα, IFNγ, TNFα, IL-6 and IL‑8 and MCP-1 were simultaneously measured in the culture supernatants of myotubes (left panels) and myoblasts (right panels), at 8h (A, B) and 24h (C, D) after infection with the influenza viruses A/California/7/2009 (blue bars) or A/Paris/1149/2008 (green bars), or in non-infected cells cultured in parallel (white bars). These results are representative of two independent experiments on cells from donor CHQ.
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pone-0079628-g007: Secretion of 5 cytokines and 1 chemokine by infected muscle cells.The concentrations of IFNα, IFNγ, TNFα, IL-6 and IL‑8 and MCP-1 were simultaneously measured in the culture supernatants of myotubes (left panels) and myoblasts (right panels), at 8h (A, B) and 24h (C, D) after infection with the influenza viruses A/California/7/2009 (blue bars) or A/Paris/1149/2008 (green bars), or in non-infected cells cultured in parallel (white bars). These results are representative of two independent experiments on cells from donor CHQ.

Mentions: We assessed the concentration of 5 cytokines (IFNα, IFNγ, TNFα, IL-6, IL-8) and 1 chemokine (MCP-1) in the supernatant of cultured myoblasts or myotubes at 8 hPI (Figure 7A, 7B) and 24 hPI (Figure 7C, 7D). In the supernatants of myotubes, whether infected or not, all proteins were undetectable except IFNα, whose levels were just above the detection threshold in all conditions (Figure 7A, 7C). High levels of MCP-1, IL-6 and moderate levels of IL-8 were detected in the supernatant of mock-infected myoblasts at 8 hPI, and even more so at 24 hPI (Figure 7B, 7D, white bars), which probably corresponded to the accumulation of these proteins over time, in agreement with the literature [48]. IFNα, TNFα and IFNγ remained below or just above the detection threshold. Infection by the pandemic or the seasonal influenza A(H1N1) viruses had no effect on the secretion of the tested proteins (Figure 7B, 7D, blue and green bars) .


Productive infection of human skeletal muscle cells by pandemic and seasonal influenza A(H1N1) viruses.

Desdouits M, Munier S, Prevost MC, Jeannin P, Butler-Browne G, Ozden S, Gessain A, Van Der Werf S, Naffakh N, Ceccaldi PE - PLoS ONE (2013)

Secretion of 5 cytokines and 1 chemokine by infected muscle cells.The concentrations of IFNα, IFNγ, TNFα, IL-6 and IL‑8 and MCP-1 were simultaneously measured in the culture supernatants of myotubes (left panels) and myoblasts (right panels), at 8h (A, B) and 24h (C, D) after infection with the influenza viruses A/California/7/2009 (blue bars) or A/Paris/1149/2008 (green bars), or in non-infected cells cultured in parallel (white bars). These results are representative of two independent experiments on cells from donor CHQ.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3818236&req=5

pone-0079628-g007: Secretion of 5 cytokines and 1 chemokine by infected muscle cells.The concentrations of IFNα, IFNγ, TNFα, IL-6 and IL‑8 and MCP-1 were simultaneously measured in the culture supernatants of myotubes (left panels) and myoblasts (right panels), at 8h (A, B) and 24h (C, D) after infection with the influenza viruses A/California/7/2009 (blue bars) or A/Paris/1149/2008 (green bars), or in non-infected cells cultured in parallel (white bars). These results are representative of two independent experiments on cells from donor CHQ.
Mentions: We assessed the concentration of 5 cytokines (IFNα, IFNγ, TNFα, IL-6, IL-8) and 1 chemokine (MCP-1) in the supernatant of cultured myoblasts or myotubes at 8 hPI (Figure 7A, 7B) and 24 hPI (Figure 7C, 7D). In the supernatants of myotubes, whether infected or not, all proteins were undetectable except IFNα, whose levels were just above the detection threshold in all conditions (Figure 7A, 7C). High levels of MCP-1, IL-6 and moderate levels of IL-8 were detected in the supernatant of mock-infected myoblasts at 8 hPI, and even more so at 24 hPI (Figure 7B, 7D, white bars), which probably corresponded to the accumulation of these proteins over time, in agreement with the literature [48]. IFNα, TNFα and IFNγ remained below or just above the detection threshold. Infection by the pandemic or the seasonal influenza A(H1N1) viruses had no effect on the secretion of the tested proteins (Figure 7B, 7D, blue and green bars) .

Bottom Line: The pathogenesis of these influenza-associated myopathies (IAM) remains unkown, although the direct infection of muscle cells is suspected.Infection led to a cytopathic effect with the lysis of muscle cells, as characterized by the release of lactate dehydrogenase.Our results are compatible with the hypothesis of a direct muscle infection causing rhabdomyolysis in IAM patients.

View Article: PubMed Central - PubMed

Affiliation: Unité Epidémiologie et Physiopathologie des Virus Oncogènes, Institut Pasteur, Paris, France ; UMR 3569, CNRS, Paris, France ; Cellule Pasteur, Université Paris Diderot, Sorbonne Paris Cité, Paris, France.

ABSTRACT
Besides the classical respiratory and systemic symptoms, unusual complications of influenza A infection in humans involve the skeletal muscles. Numerous cases of acute myopathy and/or rhabdomyolysis have been reported, particularly following the outbreak of pandemic influenza A(H1N1) in 2009. The pathogenesis of these influenza-associated myopathies (IAM) remains unkown, although the direct infection of muscle cells is suspected. Here, we studied the susceptibility of cultured human primary muscle cells to a 2009 pandemic and a 2008 seasonal influenza A(H1N1) isolate. Using cells from different donors, we found that differentiated muscle cells (i. e. myotubes) were highly susceptible to infection by both influenza A(H1N1) isolates, whereas undifferentiated cells (i. e. myoblasts) were partially resistant. The receptors for influenza viruses, α2-6 and α2-3 linked sialic acids, were detected on the surface of myotubes and myoblasts. Time line of viral nucleoprotein (NP) expression and nuclear export showed that the first steps of the viral replication cycle could take place in muscle cells. Infected myotubes and myoblasts exhibited budding virions and nuclear inclusions as observed by transmission electron microscopy and correlative light and electron microscopy. Myotubes, but not myoblasts, yielded infectious virus progeny that could further infect naive muscle cells after proteolytic treatment. Infection led to a cytopathic effect with the lysis of muscle cells, as characterized by the release of lactate dehydrogenase. The secretion of proinflammatory cytokines by muscle cells was not affected following infection. Our results are compatible with the hypothesis of a direct muscle infection causing rhabdomyolysis in IAM patients.

Show MeSH
Related in: MedlinePlus