Activated mast cells in skeletal muscle can be a potential mediator for cancer‐associated cachexia

Abstract

Background Eighty per cent of United States advanced cancer patients faces a worsened prognosis due to cancer‐associated cachexia. Inflammation is one driver of muscle atrophy in cachexia, and skeletal muscle‐resident immune cells could be a source of inflammation. This study explores the efficacy of cancer activated skeletal muscle‐resident mast cells as a biomarker and mediator of cachexia. Methods Individual gene markers for immune cells were assessed in a publicly available colon carcinoma cohort of normal (n = 3), moderate cachexia (n = 3), and severe cachexia (n = 4) mice. Lewis lung carcinoma (LL/2) cells induced cachexia in C57BL/6 mice, and a combination of toluidine blue staining, immunofluorescence, quantitative polymerase chain reaction, and western blots measured innate immune cell expression in hind limb muscles. In vitro measurements included C2C12 myotube diameter before and after treatment with media from primary murine mast cells activated with LL/2 conditioned media. To assess translational potential in human samples, innate immune cell signatures were assessed for correlation with skeletal muscle atrophy and apoptosis, dietary excess, and cachexia signatures in normal skeletal muscle tissue. Gene set enrichment analysis was performed with innate immune cell signatures in publicly available cohorts for upper gastrointestinal (GI) cancer and pancreatic ductal adenocarcinoma (PDAC) patients (accession: GSE34111 and GSE130563, respectively). Results Individual innate immunity genes (TPSAB1 and CD68) showed significant increases in severe cachexia (weight loss > 15%) mice in a C26 cohort (GSE24112). Induction of cachexia in C57BL/6 mice with LL/2 subcutaneous injection significantly increased the number of activated skeletal muscle‐resident degranulating mast cells. Murine mast cells activated with LL/2 conditioned media decreased C2C12 myotube diameter (P ≤ 0.05). Normal human skeletal muscle showed significant positive correlations between innate immune cell signatures and muscle apoptosis and atrophy, dietary excess, and cachexia signatures. The mast cell signature was up‐regulated (positive normalized enrichment score and false discovery rate ≤ 0.1) in upper GI cachectic patients (n = 12) compared with control (n = 6), as well as in cachectic PDAC patients (n = 17) compared with control patients (n = 16). Conclusions Activated skeletal muscle‐resident mast cells are enriched in cachectic muscles, suggesting skeletal‐muscle resident mast cells may serve as a biomarker and mediator for cachexia development to improve patient diagnosis and prognosis.

Oral Exposure to Polystyrene Microplastics of Mice on a Normal or High-Fat Diet and Intestinal and Metabolic Outcomes

“…For immunochemistry, sections of the small intestine were prepared and blocked with Blocking One Histo (Nacalai Tesque, Inc.) for 30 min. Subsequently, the samples were stained with monoclonal primary anti-Muc2 (1/500 dilution; ab272692, Abcam) 59 at 4°C overnight. After washing for 5 min in PBS, the samples were subsequently stained with a Texas-red-conjugated antimouse secondary antibody (1/1,000 dilution; Jackson ImmunoResearch) in the dark at room temperature for 60 min.…”

A protocol for single nucleus RNA-seq from frozen skeletal muscle

“…In our samples this cell type was present in two of four pathologically affected muscles, but in higher proportions than controls (Fig S3). There is some evidence that mast cells are enriched in inflammatory myopathies (Yokota et al, 2014), Duchenne muscular dystrophy (Gorospe et al, 1994) and cachectic muscle (Widner et al, 2021). These results emphasize this method's ability to detect rare cell types and enable further investigation into their behaviour in pathological conditions.…”

ACSL4 contributes to ferroptosis‐mediated rhabdomyolysis in exertional heat stroke

“…Furthermore, the iron accumulation in RM tissue was confirmed by the Prussia blue staining ( Figure S3A ), and this process is accompanied by significant degradation of heme ( Figure S3B ). Based on our KEGG pathway and GO enrichment analysis ( Figure S2 ), we found that the expression of the iron homoeostasis‐related genes in Gas muscle tissue, such as the transferrin receptor (TFR), 5 SLC39A14, 22 and heme oxygenase 1 (Hmox1), 21 were dysregulated, and we performed the western blotting to determine the expression of the iron homoeostasis‐related genes in Gas muscle tissue, such as TFR, nuclear receptor coactivator 4 (NCOA4), 23 ferroportin1 (FPN), 24 SLC39A14, Hmox1, and ferritin heavy chain 1 (FTH1) 23 ( Figure S3C ). Our analysis demonstrate that the levels of TFR reached to the peak levels once EHS onset, the levels of NCOA4 and SLC39A14 significantly increased once EHS onset, and reached to the peak level at 6 h, then slightly decreased at 24 h after EHS.…”

“…Massive degenerative changes of skeletal muscle cells, including cell death, are the most common pathological change of EHS. 3 , 4 Apoptosis, 5 necroptosis, 6 and autophagy 7 , 8 represent extremes of the response of the muscle to injury and often coexisted in experimental models of acute muscle injury. 9 However, the above‐mentioned various deaths cannot clarify the mechanism of RM development following EHS.…”