Schmitt, Rebecca E

Cancer cachexia, and its associated complications, represent a large and currently untreatable roadblock to effective cancer management. Many potential therapies have been proposed and tested—including appetite stimulants, targeted cytokine blockers, and nutritional supplementation—yet highly effective therapies are lacking. Innovative approaches to treating cancer cachexia are needed. Members of the Kruppel-like factor (KLF) family play wide-ranging and important roles in the development, maintenance, and metabolism of skeletal muscle. Within the KLF family, we identified KLF10 upregulation in a multitude of wasting contexts—including in pancreatic, lung, and colon cancer mouse models as well as in human patients. We subsequently interrogated loss-of-function of KLF10 as a potential strategy to mitigate cancer associated muscle wasting. In vivo studies leveraging orthotopic implantation of pancreas cancer cells into wild-type and KLF10 KO mice revealed significant preservation of lean mass and robust suppression of pro-atrophy muscle-specific ubiquitin ligases Trim63 and Fbxo32, as well as other factors implicated in atrophy, calcium signaling, and autophagy. Bioinformatics analyses identified Transforming growth factor beta (TGF-β), a known inducer of KLF10 and cachexia promoting factor, as a key upstream regulator of KLF10. We provide direct in vivo evidence that KLF10 KO mice are resistant to the atrophic effects of TGF-β. ChIP-based binding studies demonstrated direct binding to Trim63 , a known wasting-associated atrogene. Taken together, we report a critical role for the TGF-β/KLF10 axis in the etiology of pancreatic cancer–associated muscle wasting and highlight the utility of targeting KLF10 as a strategy to prevent muscle wasting and limit cancer-associated cachexia.

AbstractBackgroundOver the past decade, advances in sepsis identification and management have resulted in decreased sepsis mortality. This increase in survivorship has highlighted a new clinical obstacle: chronic critical illness (CCI), for which there are no effective treatment options. Up to half of sepsis survivors suffer from CCI, which can include multi‐organ dysfunction, chronic inflammation, muscle wasting, physical and mental disabilities, and enhanced frailty. These symptoms prevent survivors from returning to regular day‐to‐day activities and are directly associated with poor quality of life.MethodsMice were subjected to cecal ligation and puncture (CLP) with daily chronic stress (DCS) as an in vivo model to study sepsis late‐effects/sequelae on skeletal muscle components. Longitudinal monitoring was performed via magnetic resonance imaging, skeletal muscle and/or muscle stem cell (MuSCs) assays (e.g., post‐necropsy wet muscle weights, minimum Feret diameter measurements, in vitro MuSC proliferation and differentiation, number of regenerating myofibres and numbers of Pax7‐positive nuclei per myofibre), post‐sepsis whole muscle metabolomics and MuSC isolation and high‐content transcriptional profiling.ResultsWe report several findings supporting the hypothesis that MuSCs/muscle regeneration are critically involved in post‐sepsis muscle recovery. First, we show that genetic ablation of muscle stem cells (MuSCs) impairs post‐sepsis muscle recovery (maintenance of 5–8% average lean mass loss compared with controls). Second, we observe impaired MuSCs expansion capacity and morphological defects at 26 days post‐sepsis compared with control MuSCs (P < 0.001). Third, when subjected to an experimental muscle injury, sepsis‐recovered mice exhibited evidence of impaired muscle regeneration compared with non‐septic mice receiving the same muscle injury (CLP/DCS injured mean minimum Feret is 92.1% of control injured, P < 0.01). Fourth, we performed a longitudinal RNA sequencing study on MuSCs isolated from post‐sepsis mice and found clear transcriptional differences in all post‐sepsis samples compared with controls. At Day 28, CLP/DCS mice satellite cells have multiple altered metabolic pathways, such as oxidative phosphorylation, mitochondrial dysfunction, sirtuin signalling and oestrogen receptor signalling, compared with controls (P < 0.001).ConclusionsOur data show that MuSCs and muscle regeneration are required for effective post‐sepsis muscle recovery and that sepsis triggers morphological, functional, and transcriptional changes in MuSCs. Moving forward, we strive to leverage a more complete understanding of post‐sepsis MuSC/regenerative defects to identify and test novel therapies that promote muscle recovery and improve quality of life in sepsis survivors.

AbstractBackgroundPersistent loss of skeletal muscle mass and function as well as altered fat metabolism are frequently observed in severe sepsis survivors. Studies examining sepsis‐associated tissue dysfunction from the perspective of the tissue microenvironment are scarce. In this study, we comprehensively assessed transcriptional changes in muscle and fat at single‐cell resolution following experimental sepsis induction.MethodsSkeletal muscle and visceral white adipose tissue from control mice or mice 1 day or 1 month following faecal slurry‐induced sepsis were used. Single cells were mechanically and enzymatically prepared from whole tissue, and viable cells were further isolated by fluorescence activated cell sorting. Droplet‐based single‐cell RNA‐sequencing (scRNA‐seq; 10× Genomics) was used to generate single‐cell gene expression profiles of thousands of muscle and fat‐resident cells. Bioinformatics analyses were performed to identify and compare individual cell populations in both tissues.ResultsIn skeletal muscle, scRNA‐seq analysis classified 1438 single cells into myocytes, endothelial cells, fibroblasts, mesenchymal stem cells, macrophages, neutrophils, T‐cells, B‐cells, and dendritic cells. In adipose tissue, scRNA‐seq analysis classified 2281 single cells into adipose stem cells, preadipocytes, endothelial cells, fibroblasts, macrophages, dendritic cells, B‐cells, T‐cells, NK cells, and gamma delta T‐cells. One day post‐sepsis, the proportion of most non‐immune cell populations was decreased, while immune cell populations, particularly neutrophils and macrophages, were highly enriched. Proportional changes of endothelial cells, neutrophils, and macrophages were validated using faecal slurry and cecal ligation and puncture models. At 1 month post‐sepsis, we observed persistent enrichment/depletion of cell populations and further uncovered a cell‐type and tissue‐specific ability to return to a baseline transcriptomic state. Differential gene expression analyses revealed key genes and pathways altered in post‐sepsis muscle and fat and highlighted the engagement of infection/inflammation and tissue damage signalling. Finally, regulator analysis identified gonadotropin‐releasing hormone and Bay 11‐7082 as targets/compounds that we show can reduce sepsis‐associated loss of lean or fat mass.ConclusionsThese data demonstrate persistent post‐sepsis muscle and adipose tissue disruption at the single‐cell level and highlight opportunities to combat long‐term post‐sepsis tissue wasting using bioinformatics‐guided therapeutic interventions.