Toxoplasma gondii Impairs Myogenesis in vitro, With Changes in Myogenic Regulatory Factors, Altered Host Cell Proliferation and Secretory Profile

Abstract

Toxoplasma gondii is the causative agent of toxoplasmosis, a parasitic disease with a wide global prevalence. The parasite forms cysts in skeletal muscle cells and neurons, although no evident association with inflammatory infiltrates has been typically found. We studied the impact of T. gondii infection on the myogenic program of mouse skeletal muscle cells (SkMC). The C2C12 murine myoblast cell line was infected with T. gondii tachyzoites (ME49 strain) for 24 h followed by myogenic differentiation induction. T. gondii infection caused a general decrease in myotube differentiation, fusion and maturation, along with decreased expression of myosin heavy chain. The expression of Myogenic Regulatory Factors Myf5, MyoD, Mrf4 and myogenin was modulated by the infection. Infected cultures presented increased proliferation rates, as assessed by Ki67 immunostaining, whereas neither host cell lysis nor apoptosis were significantly augmented in infected dishes. Cytokine Bead Array indicated that IL-6 and MCP-1 were highly increased in the medium from infected cultures, whereas TGF-β1 was consistently decreased. Inhibition of the IL-6 receptor or supplementation with recombinant TGF-β failed to reverse the deleterious effects caused by the infection. However, conditioned medium from infected cultures inhibited myogenesis in C2C12 cells. Activation of the Wnt/β-catenin pathway was impaired in T. gondii-infected cultures. Our data indicate that T. gondii leads SkMCs to a pro-inflammatory phenotype, leaving cells unresponsive to β-catenin activation, and inhibition of the myogenic differentiation program. Such deregulation may suggest muscle atrophy and molecular mechanisms similar to those involved in myositis observed in human patients.

Infection of mouse neural progenitor cells by Toxoplasma gondii affects in vitro proliferation, differentiation and migration

“…This effect was also observed when C17.2 neural stem cell line cells were treated with soluble factors released from T. gondii (Gan et al, 2016), in a process mediated by the Wnt/β-catenin signaling pathway. Our group has also described that direct infection of skeletal muscle precursors (myoblasts) affects Wnt/β-catenin signaling pathway activation, which impairs myogenesis and leaves cells in a proliferative, undifferentiated state (Vieira et al, 2019).…”

“…Contrasting results regarding neuronal and astrocytic differentiation in radial glia and intermediate progenitors due to T. gondii infection may, in fact, reinforce the differences in clinical outcomes when vertical transmission occurs during different gestational periods. Moreover, our group has shown, using two independent systems, that T. gondii decreases the secretion of transforming growth factor beta 1 (TGF-β1), in both radial glia (Marcos et al, 2020) and skeletal muscle cells (Vieira et al, 2019). TGF-β1 is a pleiotropic cytokine involved in organogenesis and pathogenic events in vertebrates (reviewed by Massagué, 1998).…”

Impaired muscle stem cell function and abnormal myogenesis in acquired myopathies

“…Single cell RNA sequencing of the infected or uninfected muscle injured or not with cardiotoxin showed that a large proportion of macrophages remains in the inflammatory state post-injury, which is associated with a reduction in the pro-regenerative macrophage subpopulation [ 151 ]. Another group has studied the direct effect of T. gondii on myoblasts in vitro by infecting C2C12 myoblasts with T. gondii [ 153 ]. Myogenic markers such as MyoD and Myogenin were reduced a few days after the infection.…”

“…These changes were not associated with an increase in cell necrosis or apoptosis. Infected cells also secrete higher levels of pro-inflammatory cytokines such as IL-6 and MCP-1 (monocyte chemoattractant protein-1), and the conditioned medium from infected cells reduces the differentiation of non-infected cells [ 153 ]. Overall, T. gondii infection impairs MuSC regenerative capacity by targeting MuSCs directly and indirectly through the perturbation of the immune response.…”