Bovine Milk Extracellular Vesicles (EVs) Modification Elicits Skeletal Muscle Growth in Rats.

Hailey A Parry,Paul A Roberson,Michael D Roberts,Andreas N Kavazis,Janos Zempleni,C Brooks Mobley,Arny A Ferrando,John J Mccarthy,Cody T Haun,Ivan J Vechetti,Kaelin C Young,Petey W Mumford,Yufeng Zhang,Matthew A Romero

doi:10.3389/fphys.2019.00436

Abstract

The current study investigated how bovine milk extracellular vesicles (EVs) affected rotarod performance and biomarkers of skeletal muscle physiology in young, growing rats. Twenty-eight-day Fisher 344 rats were provided an AIN-93G-based diet for 4 weeks that either remained unadulterated [EVs and RNA-sufficient (ERS; n = 12)] or was sonicated [EVs and RNA-depleted (ERD; n = 12)]. Prior to (PRE) and on the last day of the intervention (POST), animals were tested for maximal rotarod performance. Following the feeding period, the gastrocnemius muscle was analyzed at the histological, biochemical, and molecular levels and was also used to measure mitochondrial function and reactive oxygen species (ROS) emission. A main effect of time was observed for rotarod time (PRE > POST, p = 0.001). Terminal gastrocnemius mass was unaffected by diet, although gastrocnemius muscle fiber cross sectional area was 11% greater (p = 0.018) and total RNA (a surrogate of ribosome density) was 24% greater (p = 0.001) in ERD. Transcriptomic analysis of the gastrocnemius indicated that 22 mRNAs were significantly greater in ERS versus ERD (p < 0.01), whereas 55 mRNAs were greater in ERD versus ERS (p < 0.01). There were no differences in gastrocnemius citrate synthase activity or mitochondrial coupling (respiratory control ratio), although mitochondrial ROS production was lower in ERD gastrocnemius (p = 0.016), which may be explained by an increase in glutathione peroxidase protein levels (p = 0.020) in ERD gastrocnemius. Dietary EVs profiling confirmed that sonication in the ERD diet reduced EVs content by ∼60%. Our findings demonstrate that bovine milk EVs depletion through sonication elicits anabolic and transcriptomic effects in the gastrocnemius muscle of rapidly maturing rats. While this did not translate into a functional outcome between diets (i.e., rotarod performance), longer feeding periods may be needed to observe such functional effects.

Highlights

Extracellular vesicles (EVs) are nanoparticle-sized lipid bilayercoated vesicles that are secreted from all cells (Colombo et al, 2013, 2014; Yanez-Mo et al, 2015; Zempleni, 2017)
While dietary milk EVs have been shown to be less enriched in skeletal muscle following oral gavage compared to other tissues (Leiferman et al, 2018; Manca et al, 2018), we have previously demonstrated that whey protein-derived EVs elicit an anabolic effect on C2C12 myotubes in vitro (Mobley et al, 2017c)
It is interesting that more gastrocnemius transcripts were up-regulated in gastrocnemius muscles from EVs and ribonucleic acid (RNA)-depleted (ERD) versus EVs and RNA-sufficient (ERS) rats, and bioinformatics showed processes related to cadherin binding, fatty acid biosynthesis, and protein kinase activity were upregulated in ERD-fed rats

Summary

Introduction

Extracellular vesicles (EVs) are nanoparticle-sized lipid bilayercoated vesicles that are secreted from all cells (Colombo et al, 2013, 2014; Yanez-Mo et al, 2015; Zempleni, 2017). In 2012, a novel observation was made in which exogenous rice-borne miRNAs were present in serum and tissues of various animals which consumed a rice-based diet (Zhang et al, 2012). MIR168a was shown to bind to the low-density lipoprotein receptor adapter protein 1 (ldlrap1) mRNA, inhibit ldlrap expression in the liver of mice, and decrease LDL levels in mouse plasma (Zhang et al, 2012). This was the first in vivo observation of physiological alterations occurring due to miRNAs obtained through the diet

Objectives

Methods

Results

Conclusion