Maduramicin Inhibits Proliferation and Induces Apoptosis in Myoblast Cells

Abstract

Maduramicin, a polyether ionophore antibiotic derived from the bacterium Actinomadura yumaensis, is currently used as a feed additive against coccidiosis in poultry worldwide. It has been clinically observed that maduramicin can cause skeletal muscle and heart cell damage, resulting in skeletal muscle degeneration, heart failure, and even death in animals and humans, if improperly used. However, the mechanism of its toxic action in myoblasts is not well understood. Using mouse myoblasts (C2C12) and human rhabdomyosarcoma (RD and Rh30) cells as an experimental model for myoblasts, here we found that maduramicin inhibited cell proliferation and induced cell death in a concentration-dependent manner. Further studies revealed that maduramicin induced accumulation of the cells at G0/G1 phase of the cell cycle, and induced apoptosis in the cells. Concurrently, maduramicin downregulated protein expression of cyclin D1, cyclin-dependent kinases (CDK4 and CDK6), and CDC25A, and upregulated expression of the CDK inhibitors (p21Cip1 and p27Kip1), resulting in decreased phosphorylation of Rb. Maduramicin also induced expression of BAK, BAD, DR4, TRADD and TRAIL, leading to activation of caspases 8, 9 and 3 as well as cleavage of poly ADP ribose polymerase (PARP). Taken together, our results suggest that maduramicin executes its toxicity in myoblasts at least by inhibiting cell proliferation and inducing apoptotic cell death.

Maduramicin Rapidly Eliminates Malaria Parasites and Potentiates the Gametocytocidal Activity of the Pyrazoleamide PA21A050

“…Several other mammalian tumor cells, including human rhabdomyosarcomas (RD) and mouse myoblasts (C2C12), have been reported to remain Ͼ80% viable after 72 h of treatment with 1 M maduramicin (29). The reason for the differential maduramicin sensitivities of apicomplexans, like P. falciparum and coccidia, and mammalian cells remains unknown, but it has allowed the use of maduramicin as a coccidiostat in animals.…”

“…The reason for the differential maduramicin sensitivities of apicomplexans, like P. falciparum and coccidia, and mammalian cells remains unknown, but it has allowed the use of maduramicin as a coccidiostat in animals. The standard dose used by the poultry industry for chicken feed is 5 to 7 mg/kg, which has been found to result in levels of maduramicin of ϳ100 nM in muscle, with no toxic effects on chickens or other nontarget species (29)(30)(31). Maduramicin treatment has a serum half-life of 13 h in chickens and does not interfere with vaccination responses, indicating that it did not affect the function of antigen-presenting cells or leukocyte activation and proliferation, which are required for the induction of immunity (31).…”

Maduramicin induces apoptosis and necrosis, and blocks autophagic flux in myocardial H9c2 cells

“…Cell viability was evaluated using CellTiter 96 ® AQ ueous One Solution Cell Proliferation Assay Kit (Promega), as described (Chen, Gu, Singh, Shang, Barzegar, Jiang and Huang 2014). Briefly, H9c2 cells suspended in the growth medium were seeded in a 96-well plate at a density of 5 × 10 3 cells/well (in triplicates) and grown overnight.…”

“…Cell death was evaluated by the trypan blue exclusion assay as described (Chen, Gu, Singh, Shang, Barzegar, Jiang and Huang 2014). Briefly, cells were seeded in 100-mm dishes at a density of 3 × 10 5 cells per dish in the growth medium and grown overnight.…”

“…Western blotting was performed, as described previously (Chen, Gu, Singh, Shang, Barzegar, Jiang and Huang 2014). Briefly, equivalent amounts of proteins (whole cell lysates) were separated on 8–15% sodium dodecyl sulfate-polyacrylamide gels and transferred to polyvinylidene difluoride membranes (Millipore, Bedford, MA, USA).…”