Biology subjects

Norton, S. C.

Publications and source records attributed to Norton, S. C..

3 recordsLinked to original sources

bioRxiv2026

Exogenous L6 myotube mitochondrial transplantation attenuates hypertrophy and elicits a unique proteomic signature in phenylephrine-treated H9C2 cardiomyocytes

Mitochondrial transplantation has recently emerged as an alternative treatment for cardiovascular disease (CVD), aimed at increasing mitochondrial number and improving mitochondrial function. Though mitochondrial dysfunction is a key factor in the development of right ventricular hypertrophy/failure, the effects of mitochondrial transplantation on disease mitigation have been largely unexplored. Therefore, this in vitro study aimed to determine whether the transplantation of exogenous L6 myotube mitochondria mitigates negative outcomes in H9C2 cardiomyocytes that were stimulated to hypertrophy with phenylephrine. Control (CTL), control with mitochondrial transplantation (MitoTx), Phenylephrine only (Phe-only), and phenylephrine with mitochondrial transplantation (Phe+MitoTx) treatments were evaluated. Pilot experiments indicated mitochondrial transplantation into healthy cardiomyocytes acutely increased Complex I-linked oxidative phosphorylation (OXPHOS) capacity (p=0.005) and maximal respiratory capacity (p=0.022) within 24 hours, and these data alongside microscopic evidence of fluorescently labeled L6 mitochondria in H9C2 cardiomyocytes suggested successful transplantation. Regarding treatment comparisons, Phe-only showed a significant increase in cell area (p<0.05), while Phe+MitoTx blunted the hypertrophic cardiomyocyte response. Consistent with these results, proteomic analysis of 4,806 proteins showed that transplantation enriched the mitochondrial proteome, impacting pathways including OXPHOS, respiration, fatty acid and amino acid metabolism, while suppressing extracellular matrix remodeling and de-differentiation signatures. This response was observed in healthy and phenylephrine-stressed cardiomyocytes. In conclusion, our in vitro data indicates that transplantation of L6 skeletal muscle mitochondria mitigates negative effects induced by phenylephrine in H9C2 cardiomyocytes. However, more rigorous in vivo studies are needed to determine if this is a suitable approach for disease mitigation.

cell biology↗

bioRxiv2026

Aging concomitantly reduces skeletal muscle proteome plasticity and hypertrophic responses to resistance training

Skeletal muscle mass and training adaptations decline with aging, yet the proteomic basis of these attenuated responses remains unclear. We hypothesized that aging is accompanied by diminished proteome plasticity in response to resistance training (RT). The soluble proteome of VL biopsies was profiled in 17 younger (21.9 {+/-} 2.5 yr) and 15 older (57.5 {+/-} 6.9 yr) untrained males before and after 10-12 weeks of supervised RT using data-independent acquisition mass spectrometry (2,113 quantified proteins). At baseline, we detected 196 differentially expressed proteins (DEPs) significantly differed between age groups by {Pi}-score (278 by FDR). A 5.6-fold difference in training-responsive was observed in younger vs. older adults (100 vs. 18 {Pi}-score DEPs; 134 vs. 0 FDR-significant). Despite this quantitative attenuation, 61.6% of proteins changed in the same direction in both age groups (Spearman {rho} = 0.284, p = 3.46 x 10-), indicating conserved but amplitude-compressed training responses (median |log2FC|: 0.13 young vs. 0.09 old). RT in older adults partially reversed the aging proteome in that directionally different changes were observed in 75.2% of aging- or training-significant proteins in aging and training contrasts, with ribosomal and translational machinery showing the strongest reversal (cytoplasmic translation NES: -2.90 with aging, +2.60 with training). Ten WGCNA co-expression modules were identified, with age emerging as the dominant organizing principle (Turquoise module r-equiv = +0.59, p < 0.001). Module eigengenes discriminated age groups at the univariate level (Turquoise/Lipid Catabolism AUC = 0.96, q < 0.012), and training-induced module changes correlated with hypertrophic outcomes. Aging markedly attenuates but does not qualitatively alter skeletal muscle proteome plasticity. RT partially reverses aging proteome signatures, with translational machinery being the most responsive and mitochondrial programs the least responsive. Baseline proteomic state constrains adaptive capacity, suggesting that the molecular features distinguishing aging muscle directly may limit its hypertrophic response to RT.

physiology↗

bioRxiv2025

Proteomic profiling of skeletal muscle ribosomes from higher versus lower responders to 10 weeks of resistance training

Ribosome biogenesis is a key driver of resistance training (RT)-induced skeletal muscle hypertrophy in humans. However, high resolution insight into RT-induced compositional alterations in ribosomes remain unexplored. Therefore, the purpose of this study was twofold: 1) develop a protocol sufficient to enrich and examine the ribosomal proteome from a small amount of human muscle tissue, and 2) determine if ribosomal protein composition differs between higher and lower responders to RT. Fourteen participants completed 10 weeks of RT (23 sessions) and were stratified into higher (n=7) and lower (n=7) responders based on changes in vastus lateralis muscle cross-sectional area (VL mCSA) and mean myofiber cross-sectional area (fCSA) from baseline (PRE) to after the 23rd RT session (POST). Participants then performed a twenty fourth RT session and biopsies were collected 24 hours post-RT (POST-24h) to examine acute bout (POST to POST-24h) ribosomal proteome alterations to RT. Ribosome enrichment and analysis included ultracentrifugation (100,000 g, 3h, 2{degrees}C) through 20% sucrose gradients followed by shotgun proteomics to quantify ribosomal protein composition. Our protocol produced exceptional ribosome enrichment, with 74 distinct ribosomal proteins being detected (92% coverage of the putative 80 cytosolic ribosomal proteins) as well as [~]164-fold and [~]71-fold increases in large (RPL) and small (RPS) protein subunit abundances, respectively, compared to conventional tissue homogenization. Despite robust phenotypic differences in hypertrophy between responder groups to the 10-week RT protocol (mCSA: +31% versus +3%; fCSA: +29% versus -2%, P<0.05 for both outcomes), ribosomal protein composition was not significantly different between groups (higher vs. lower responders), nor was there a significant time (POST to POST-24h) or a group*time interaction (P>0.028 for all comparisons). In conclusion, we present a highly effective ribosome enrichment protocol requiring minimal human muscle tissue. Though preliminary analyses indicate that ribosomal protein composition was similar between hypertrophic phenotypes in response to an acute RT session after a 10-week RT intervention, future research leveraging our techniques with more sampling time points are needed to provide more definitive conclusions.

molecular biology↗