Skeletal muscle cells possess a ‘memory’ of acute early life TNF-α exposure: role of epigenetic adaptation

Abstract

Sufficient quantity and quality of skeletal muscle is required to maintain lifespan and healthspan into older age. The concept of skeletal muscle programming or memory has been suggested to contribute to accelerated muscle decline in the elderly in association with early life stress such as fetal malnutrition. Further, muscle cells in vitro appear to remember the in vivo environments from which they are derived (e.g. cancer, obesity, type II diabetes, physical inactivity and nutrient restriction). Tumour-necrosis factor alpha (TNF-α) is a pleiotropic cytokine that is chronically elevated in sarcopenia and cancer cachexia. Higher TNF-α levels are strongly correlated with muscle loss, reduced strength and therefore morbidity and earlier mortality. We have extensively shown that TNF-α impairs regenerative capacity in mouse and human muscle derived stem cells (Meadows 2000;Foulstone 2001Foulstone , 2004Al-Shanti 2008;Saini 2008;Sharples 2010). We have also recently established an epigenetically mediated mechanism (SIRT1-histone deacetylase) regulating survival of myoblasts in the presence of TNF-α (Saini 2012). We therefore wished to extend this work in relation to muscle memory of catabolic stimuli and the potential underlying epigenetic modulation of muscle loss. To enable this aim; C2C12 myoblasts were cultured in the absence or presence of early TNF-α (early proliferative lifespan) followed by 30 population doublings in the absence of TNF-α, prior to the induction of differentiation in low serum media (LSM) in the absence or presence of late TNF-α (late proliferative lifespan). The cells that received an early plus late lifespan dose of TNF-α exhibited reduced morphological (myotube number) and biochemical (creatine kinase activity) differentiation vs. control cells that underwent the same number of proliferative divisions but only a later life encounter with TNF-α. This suggested that muscle cells had a morphological memory of the acute early lifespan TNF-α encounter. Importantly, methylation of myoD CpG islands were increased in the early TNF-α cells, 30 population doublings later, suggesting that even after an acute encounter with TNF-α, the cells have the capability of retaining elevated methylation for at least 30 cellular divisions. Despite these fascinating findings, there were no further increases in myoD methylation or changes in its gene expression when these cells were exposed to a later TNF-α dose suggesting that this was not directly responsible for the decline in differentiation observed. In conclusion, data suggest that elevated myoD methylation is retained throughout muscle cells proliferative lifespan as result of early life TNF-α treatment and has implications for the epigenetic control of muscle loss.

Long-lasting effect of obesity on skeletal muscle transcriptome

“…Earlier studies reported that high-fat diet increases [70] and CR reduces TNFα expression in SM [71], suggesting that CR may circumvent the apoptotic effect of TNFα in SM [72]. Acute early life exposures to TNFα renders muscle cells more susceptible to impaired regeneration when inflammation is encountered in later proliferative life [73], which supports the role TNFα in muscle degeneration. Interestingly, the TNFRSF12A gene was significantly upregulated by WSD and downregulated by CR (Additional file 2: Table S1 and Table 2), while the transcriptional activation of this gene has been previously linked to diffuse muscle atrophy [74].…”

Transcriptomic and epigenetic regulation of disuse atrophy and the return to activity in skeletal muscle

“…HRM-PCR for total DNA methylation HRM-PCR for CpG methylation was performed as previously described (24). In brief, 20 ng DNA was subjected to HRM-PCR using EpiTect HRM-PCR kits and Rotorgene 3000Q (Qiagen) with Rotorgene software.…”

“…This relationship was subsequently used to quantify the percentage methylation from the integrated raw melt curves of the experimental samples. Calculations were performed by using a Python-based program, MethylCal, that was developed for this purpose in-house [used previously (24)].…”

“…It has recently been suggested that denervationinduced atrophy results in the differential expression of genes that are associated with chromatin remodeling (23). Recent in vitro evidence also suggests that epigenetic mechanisms may influence regeneration and myotube atrophy (24). Skeletal muscle cells that have encountered the atrophic stimulus of the inflammatory cytokine, TNF-a, during their early proliferative life are more susceptible to TNF-a later in their proliferative life, and demonstrate impaired differentiation and regeneration compared with matched, untreated controls (24).…”

Transcriptional memory in skeletal muscle. Don't forget (to) exercise

“…A general trend towards reduced promoter methylation levels of genes involved in energy metabolism, myogenesis, contractile activity, and oxidative stress resistance became apparent when global methylation patterns in skeletal muscle of healthy aged men with a lifelong history of physical activity were compared to “couch potatoes” of the same age group (Sailani et al, 2019). Reciprocally, also “bad memories” inflicted by physical inactivity, unhealthy diet, and prenatal stress have been reported to become reflected by differential DNA methylation patterns in skeletal muscle tissue (Alibegovic et al, 2009; Jacobsen et al, 2012; Jacobsen et al, 2014; Nilsson & Ling, 2017; Nitert et al, 2012; Sharples, Polydorou et al, 2016; Sheppard et al, 2017).…”