Understanding the influence of trenbolone acetate and polyamines on proliferation of bovine satellite cells

Authors: REICHHARDT, CALEB - Utah State University; AHMADPOUR, AMIR - Utah State University; Christensen, Rachael; INECK, NIKOLE - Utah State University; MURDOCH, GORDON - Idaho State University; THORNTON, KARA - Utah State University

Submitted to: Domestic Animal Endocrinology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/23/2020
Publication Date: 3/31/2020
Citation: Reichhardt, C., Ahmadpour, A., Christensen, R., Ineck, N., Murdoch, G., Thornton, K. 2020. Understanding the influence of trenbolone acetate and polyamines on proliferation of bovine satellite cells. Domestic Animal Endocrinology. 74:106479. https://doi.org/10.1016/j.domaniend.2020.106479.
DOI: https://doi.org/10.1016/j.domaniend.2020.106479

Interpretive Summary: Most beef cattle on feed prior to harvest in the U.S. receive at least one growth promoting implant during production, in order to increase growth, efficiency, and economic return to producers. However, the complete metabolic mechanism at the cellular level through which growth promoting implants function to improve skeletal muscle growth remains unknown. Therefore, a study was planned to investigate the effect of specific protein precursors and steroidal hormone analogs on muscle cell growth. This study had two objectives: (1) determine the effect of polyamines and their precursors on proliferation rate in cultured bovine satellite cells (BSC) and (2) understand whether trenbolone acetate (TBA), a testosterone analog, has an impact on the polyamine biosynthetic pathway in these cells. To address these, BSC were harvested from three different finished steers muscle tissue and grown in culture. Once cultures reached a certain stage of maturity (75% confluency), they were treated in 1% fetal bovine serum (FBS) and/or onw of the following ceoncentrations of a specific nutrient: 10 nM TBA, 10 mM methionine (Met), 8 mM ornithine (Orn), 2 mM putrescine (Put), 1.5 mM spermidine (Spd) or 0.5 mM spermine (Spe). The effects of these treatments were documented, mRNA was isolated from cultures to show which growth factors were stimulated during treatment, and the rate of growth was noted. These results indicate that treatments of BSC cultures with polyamines and their precursors increase BSC proliferation rate as well as abundance of mRNA involved in cell proliferation. Additionally, treatment of BSC cultures with TBA, polyamines, or polyamine precursors effect the expression of genes related to the polyamine growth pathway.

Technical Abstract: Approximately 90% of beef cattle on feed in the U.S. receive at least one anabolic implant during production, which results in increased growth, efficiency, and economic return to producers. However, the complete molecular mechanism through which anabolic implants function to improve skeletal muscle growth remains unknown. This study had two objectives: (1) determine the effect of polyamines and their precursors on proliferation rate in bovine satellite cells (BSC) and (2) understand whether trenbolone acetate (TBA), a testosterone analog, has an impact on the polyamine biosynthetic pathway. To address these, BSC were isolated from three different finished steers and grown in culture. Once cultures reached 75% confluency, they were treated in 1% fetal bovine serum (FBS) and/or 10 nM TBA, 10 mM methionine (Met), 8 mM ornithine (Orn), 2 mM putrescine (Put), 1.5 mM spermidine (Spd) or 0.5 mM spermine (Spe). Initially, a range of physiologically relevant concentrations of Met, Orn, Put, Spd, and Spe were tested to determine appropriate experimental doses to implement the afore mentioned experiments. After 1, 12 or 24 h of treatment, mRNA was isolated from cultures and abundance of paired box transcription factor 7 (Pax7), Sprouty 1 (Spry), mitogen activated protein kinase-1 (Mapk), ornithine decarboxylase (Odc), and S adenosylmethionine (Amd1) were determined, and normalized to 18s. Treatment with 10 nM TBA, Met, Orn, Put, Spd or Spe increased (P < 0.05) BSC proliferation when compared to control cultures. Treatment of cultures with Orn or Met increased (P < 0.01) expression of Odc 1 h post treatment when compared to control cultures. Abundance of Amd1 was increased (P < 0.01) 1 h post treatment in cultures treated with Spd or Spe when compared to 1% FBS controls. Cultures treated with TBA had increased (P < 0.01) abundance of Spry mRNA 12 h post treatment, as well as increased mRNA abundance of Mapk (P < 0.01) 12 h and 24 h post treatment when compared to 1% FBS control cultures. Treatment with Met increased (P < 0.01) mRNA abundance of Pax7 1 h post treatment as compared to 1% FBS controls. These results indicate that treatments of BSC cultures with polyamines and their precursors increase BSC proliferation rate as well as abundance of mRNA involved in cell proliferation. Additionally, treatment of BSC cultures with TBA, polyamines, or polyamine precursors effect the expression of genes related to the polyamine biosynthetic pathway and proliferation.