Whey protein-derived exosomes increase protein synthesis and hypertrophy in C2-C12myotubes.
We sought to examine potential amino acid independent mechanisms whereby hydrolyzed whey protein (WP) affects muscle protein synthesis (MPS) and anabolism in vitro. Specifically, we tested (1) whether 3-h and 6-h treatments of WP, essential amino acids, or l-leucine (Leu) affected MPS, and whether 6-h treatments with low-, medium-, or high doses of WP versus Leu affected MPS; (2) whether knockdown of the primary Leu transporter affected WP- and Leu-mediated changes in MPS, mammalian target of rapamycin (mTOR) signaling responses, or both, following 6-h treatments; (3) whether exosomes isolated from WP (WP-EXO) affected MPS, mTOR signaling responses, or both, compared with untreated (control) myotubes, following 6-h, 12-h, and 24-h treatments, and whether they affected myotube diameter following 24-h and 48-h treatments. For all treatments, 7-d post-differentiated C2C12myotubes were examined. In experiment 1, 6-h WP treatments increased MPS compared with control (+46%), Leu (+24%), and essential amino acids (+25%). Moreover, the 6-h low-, medium-, and high WP treatments increased MPS by approximately 40 to 50% more than corresponding Leu treatments. In experiment 2 (LAT short hairpin RNA-transfected myotubes), 6-h WP treatments increased MPS compared with control (+18%) and Leu (+19%). In experiment 3, WP-EXO treatments increased MPS over controls at 12h (+18%) and 24h (+45%), and myotube diameters increased with 24-h (+24%) and 48-h (+40%) WP-EXO treatments compared with controls. The WP-EXO treatments did not appear to operate through mTOR signaling; instead, they increased mRNA and protein levels o eukaryotic initiation factor 4A. Bovine-specific microRNA following 24-h WP-EXO treatments were enriched in myotubes (chiefly miR-149-3p, miR-2881), but were not related to hypertrophic gene targets. To summarize, hydrolyzed WP-EXO increased skeletal MPS and anabolism in vitro, and this may be related to an unknown mechanism that increases translation initiation factors rather than enhancing mTOR signaling or the involvement of bovine-specific microRNA.
School of Kinesiology, Auburn University, Auburn, AL 36849.
C Brooks
CB
Mobley
School of Kinesiology, Auburn University, Auburn, AL 36849.
Petey W
PW
Mumford
Department of Physiology, University of Kentucky Medical College of Medicine, Lexington 40506.
John J
JJ
McCarthy
Harrison School of Pharmacy, Auburn University, Auburn, AL 36849.
Michael E
ME
Miller
School of Kinesiology, Auburn University, Auburn, AL 36849; Department of Cellular Biology and Physiology, Edward Via College of Osteopathic Medicine, Auburn Campus, Auburn, AL 36849.
Kaelin C
KC
Young
School of Kinesiology, Auburn University, Auburn, AL 36849; Department of Cellular Biology and Physiology, Edward Via College of Osteopathic Medicine, Auburn Campus, Auburn, AL 36849.
Jeffrey S
JS
Martin
School of Kinesiology, Auburn University, Auburn, AL 36849; Department of Cellular Biology and Physiology, Edward Via College of Osteopathic Medicine, Auburn Campus, Auburn, AL 36849.
Darren T
DT
Beck
AP Nutrition LLC, Draper, UT 84020.
Christopher M
CM
Lockwood
School of Kinesiology, Auburn University, Auburn, AL 36849; Department of Cellular Biology and Physiology, Edward Via College of Osteopathic Medicine, Auburn Campus, Auburn, AL 36849. Electronic address: mdr0024@auburn.edu.
Michael D
MD
Roberts
D000818:Animals; D002417:Cattle; D055354:Exosomes; D006984:Hypertrophy; D007930:Leucine / Q000378:metabolism; D018485:Muscle Fibers, Skeletal; D018482:Muscle, Skeletal / Q000378:metabolism; D010766:Phosphorylation; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D000067816:Whey Proteins
D000067816:Whey Proteins; D058570:TOR Serine-Threonine Kinases; D007930:Leucine
exosomes; leucine; muscle protein synthesis; whey protein