Blueberry (BB) consumption is linked to improved health. The bioconversion of the polyphenolic content of BB by fermentative bacteria in the large intestine may be a necessary step for the health benefits attributed to BB consumption. The identification of specific gut microbiota taxa that respond to BB consumption and that mediate the bioconversion of consumed polyphenolic compounds into bioactive forms is required to improve our understanding of how polyphenols impact human health. We tested the ability of polyphenol-rich fractions purified from whole BB—namely, anthocyanins/flavonol glycosides (ANTH/FLAV), proanthocyanidins (PACs), the sugar/acid fraction (S/A), and total polyphenols (TPP)—to modulate the fecal microbiota composition of healthy adults in an in vitro colon system. In a parallel pilot study, we tested the effect of consuming 38 g of freeze-dried BB powder per day for 6 weeks on the fecal microbiota of 17 women in two age groups (i.e., young and older). The BB ingredients had a distinct effect on the fecal microbiota composition in the artificial colon model. The ANTH/FLAV and PAC fractions were more effective in promoting microbiome alpha diversity compared to S/A and TPP, and these effects were attributed to differentially responsive taxa. Dietary enrichment with BB resulted in a moderate increase in the diversity of the microbiota of the older subjects but not in younger subjects, and certain health-relevant taxa were significantly associated with BB consumption. Alterations in the abundance of some gut bacteria correlated not only with BB consumption but also with increased antioxidant activity in blood. Collectively, these pilot data support the notion that BB consumption is associated with gut microbiota changes and health benefits.

The accumulation of damaged mitochondria due to insufficient autophagy has been implicated in the pathophysiology of skeletal muscle aging. Ulk1 is an autophagy-related kinase that initiates autophagosome assembly and may also play a role in autophagosome degradation (i.e., autophagy flux), but the contribution of Ulk1 to healthy muscle aging is unclear. Therefore, the purpose of this study was to investigate the role of Ulk1-mediated autophagy in skeletal muscle aging. At age 22 months (80% survival rate), muscle contractile and metabolic function were assessed using electrophysiology in muscle-specific Ulk1 knockout mice (MKO) and their littermate controls (LM). Specific peak-isometric torque of the ankle dorsiflexors (normalized by tibialis anterior muscle cross-sectional area) and specific force of the fast-twitch extensor digitorum longus muscles was reduced in MKO mice compared to LM mice (p < 0.03). Permeabilized muscle fibers from MKO mice had greater mitochondrial content, yet lower mitochondrial oxygen consumption and greater reactive oxygen species production compared to fibers from LM mice (p ≤ 0.04). Alterations in neuromuscular junction innervation patterns as well as changes to autophagosome assembly and flux were explored as possible contributors to the pathological features in Ulk1 deficiency. Of primary interest, we found that Ulk1 phosphorylation (activation) to total Ulk1 protein content was reduced in older muscles compared to young muscles from both human and mouse, which may contribute to decreased autophagy flux and an accumulation of dysfunctional mitochondria. Results from this study support the role of Ulk1-mediated autophagy in aging skeletal muscle, reflecting Ulk1′s dual role in maintaining mitochondrial integrity through autophagosome assembly and degradation.

Skeletal muscle progenitor cells (MPCs) repair damaged muscle postinjury. Pyruvate kinase M2 (PKM2) is a glycolytic enzyme (canonical activity) that can also interact with other proteins (noncanonical activity) to modify diverse cellular processes. Recent evidence links PKM2 to MPC proliferation. This study aimed to understand cellular roles for PKM2 in MPCs and the necessity of PKM2 in MPCs for muscle regeneration postinjury. Cultured, proliferating MPCs (C2C12 cells) were treated with a short hairpin RNA targeting PKM2 or small molecules that selectively affect canonical and noncanonical PKM2 activity (shikonin and TEPP-46). Cell number was measured, and RNA-sequencing and metabolic assays were used in follow-up experiments. Immunoprecipitation coupled to proteomics was used to identify binding partners of PKM2. Lastly, an MPC-specific PKM2 knockout mouse was generated and challenged with a muscle injury to determine the impact of PKM2 on regeneration. When the noncanonical activity of PKM2 was blocked or impaired, there was an increase in reactive oxygen species concentrations (1.6–2.0-fold, P < 0.01). Blocking noncanonical PKM2 activity also increased lactate excretion (1.2–1.6-fold, P < 0.05) and suppressed mitochondrial oxygen consumption (1.3–1.6-fold, P < 0.01). Glutamate dehydrogenase 1 (GLUD1) was identified as a PKM2 binding partner and blocking noncanonical PKM2 activity increased GLUD activity (1.5–1.6-fold, P < 0.05). Mice with an MPC-specific PKM2 deletion did not demonstrate impaired muscle regeneration. The results suggest that the noncanonical activity of PKM2 is important for MPC proliferation in vitro and demonstrate GLUD1 as a PKM2 binding partner. Because no impairments in muscle regeneration were detected in a mouse model, the endogenous environment may compensate for loss of PKM2.

The use of primary human tissue and cells is ideal for the investigation of biological and physiological processes such as the skeletal muscle regenerative process. There are recognized challenges to working with human primary adult stem cells, particularly human muscle progenitor cells (hMPCs) derived from skeletal muscle biopsies, including low cell yield from collected tissue and a large degree of donor heterogeneity of growth and death parameters among cultures. While incorporating heterogeneity into experimental design requires a larger sample size to detect significant effects, it also allows us to identify mechanisms that underlie variability in hMPC expansion capacity, and thus allows us to better understand heterogeneity in skeletal muscle regeneration. Novel mechanisms that distinguish the expansion capacity of cultures have the potential to lead to the development of therapies to improve skeletal muscle regeneration.

Peptide YY (PYY) is considered a gut peptide with roles in post-prandial appetite and glucose regulation. Circulating PYY protein levels increase during aerobic exercise. Furthermore, people who have greater increases in muscle progenitor cells (hMPCs), the adult stem cell population responsible for skeletal muscle (SkM) repair, after resistance training have higher PYY transcript levels in SkM prior to training. Currently, examination of PYY expression patterns in SkM and/or hMPCs is lacking. Our objective was to identify the expression patterns of PYY in SkM and hMPCs. PYY and the associated Y receptors were analyzed in SkM biopsy tissue and cultured hMPCs from young and old human participants. Additional experiments to assess the role and regulation of PYY in hMPCs were performed. In SkM, PYY and one of the three Y receptors (Y1r) were detectable, but expression patterns were not affected by age. In expanding hMPCs, PYY and all three Y receptor (Y1r, Y2r, and Y5r) proteins were expressed in a temporal fashion with young hMPCs having greater levels of Y receptors at various time points. Exogenous PYY did not affect hMPC population expansion. hMPC PYY levels increased following the metabolic stimulus, 5-Aminoimidazole-4-carboxamide ribonucleotide (AICAR), but were not affected by the inflammatory stimulus, tumor necrosis factor alpha (TNFα). In conclusion, PYY and Y receptor expression are not impacted by age in SkM tissue but are reduced in old vs. young expanding hMPCs. Furthermore, endogenous PYY production is stimulated by low energy states and thus may be integral for skeletal muscle and hMPC responses to metabolic stimuli.

Food antigens elicit immune tolerance through the action of intestinal regulatory T (T _reg ) cells. Unlike food allergens, the proteins that mediate tolerance are mostly undescribed. Here, we found that epitopes derived from seed storage proteins are targets of murine intestinal T _reg cells, with the most frequent response targeting the C terminus of the maize protein alpha-zein. A major histocompatibility complex (MHC) tetramer loaded with this antigen revealed that zein-specific T cells are predominantly intestinal T _reg cells, develop concurrently with weaning, and constitute up to 2% of the peripheral T _reg cell pool. Zein-responsive T _reg cells repressed naïve T cell proliferation ex vivo, and prior dietary exposure resulted in a constrained response upon diverse inflammatory challenges in vivo, supporting a specific role for gut-resident T _reg cells in suppressing systemic immune responses. Our work reveals the development, immune-suppressive characteristics, and function of naturally occurring T _reg cells that recognize dietary seed storage proteins, a previously undescribed class of antigens in oral tolerance.