A Novel Cell‐Based Screening Assay to Assess the Effects of Diverse Protein Sources on Muscle Protein Signaling

Abstract

Protein sources that build or maintain muscle mass are of great interest to athletes, individuals on weight loss regimens and aging adults. Optimizing muscle health benefits derived from high quality proteins such as soy, dairy and egg may identify improved ingredients for sports nutrition and human health. The mammalian target of rapamycin (mTOR) is the central molecular pathway in muscle protein synthesis and is activated by dietary amino acids, specifically leucine. We used a cell‐based screening assay to investigate mTOR activation after incubation with proteins from diverse sources. Intact or partially hydrolyzed proteins were treated with pepsin and pancreatin to simulate gut digestion (SGD). Soluble fractions of SGD proteins were applied (1mg/mL) to C2C12 myotubes for 30 min. C2C12 cell lysates were then collected and mTOR activation was quantified by immunoassay of phosphorylated‐70S6KThr389, a critical downstream molecule in the mTOR signaling pathway. A range of mTOR activation was observed in response to the various proteins. The mean response for conventionally processed soy proteins, dairy, beef, and fish proteins were similar. mTOR activation was significantly higher for soy protein than for egg or pea proteins or soy whey preparations. These results suggest that most high quality proteins stimulate the mTOR pathway to a similar degree. Our data also confirm that insulin and protein stimulate mTOR via separate and additive upstream pathways and that these pathways are rapamycin‐sensitive. Our findings show this in vitro system to be a useful tool for comparing different proteins and other ingredients for their ability to impact muscle protein synthesis.