Obesity influences Muscle Stem Cells
Obesity has become epidemic, it affects almost 1 out of 3 people and occurs when the Body Mass Index (B.M.I) is higher than 30 kg/m2. Obesity causes reduced muscle growth and various metabolic disorder, like type 2 diabetes. According to a new research from Lund University in Sweden, this disease could bring epigenetic changes during the formation of new muscle cells [1]. In this study the same changes in obese and non-obese subjects were analysed to detect how much this difference could be appreciable, to better understand obesity causes and effects. In obese subjects, DNA methylation after differentiation appeared to be modified almost 4 times more than in non-obese. The transcriptome analysis has been made in the same condition to see if changes in methylome caused different genes regulation. This study showed both up- and down-regulation of already known factors related to myogenesis that provoke a decrease in muscular tone and mass. Importantly, they discovered three genes whose involvement in myogenesis was never seen before. These new myogenesis-related genes were Interleukin 32 (a pro-inflammatory cytokine) and two gene related with pregnancy: Pregnancy-Specific beta-1-Glycoproteins (PSGs) and Chorionic Gonadotropin Beta-polypeptides (CGBs). IL-32 was analysed in detail and evidences showed its negative role in myogenesis.
Myogenesis is the differentiation process of muscle cells and in adulthood it can happen only after a trauma or a damage of the muscular tissues. The mesodermal progenitor cells after these events start to produce the myocyte enhancer factors 2 (MEF2s) and the myogenic regulatory factors (MRFs), such as MyoD, Myf5 or Myogenin, which are helix-loop-helix transcription factors, which take part in different steps of differentiation [2].
The next step is the proliferation of myoblasts which will eventually end up fusing together to generate multi-nucleated myotubes. It is already known that during differentiation many changes can occur in the methylome of muscle skeletal cells [3, 4] suggesting that the degree of methylation plays an important role on the myogenesis. Cajsa Davegårdh and collaborators studied how DNA methylation is modified in muscle stem cells of healthy and obese individuals. DNA methylation is an epigenetic modification by which methyl groups (-CH3) are introduced on the DNA, mainly but not exclusively, on specific regions called CpG Islands, which are very rich in Cytosine and Guanine. These modifications are used to modulate the activity of the genes [5] .
It is well known that high fat diet, lack of exercise, aging, and other environmental factors can modify DNA methylation in human skeletal muscle cells [6-9]. Starting from these knowledges, Davegårdh et al. analysed the whole methylome on both myoblasts and myotubes of 28 people: 14 healthy and 14 obese subjects. DNA methylation analysis was conducted using Infinium Human-Methylation450 BeadChip by Illumina™. This microarray is able to determine methylation level of more than 450,000 loci in the genome, even those in areas with low density of CpG islands.
By these experiments, they found that the average degree of DNA methylation was not much different between the two groups. Nevertheless, obese subjects had a larger number of individual CpG island, that showed a different level of methylation after differentiation, with respect to healthy subjects. They also observed that cells from the obese subject maintained the altered epigenetic pattern, even after being cultured in the same conditions of non-obese cells. Thus, obesity influences the epigenetic memory of muscle stem cells during myogenesis. The modified methylation levels were partially confirmed by a quantitative PCR test on the three catalytically active DNA methyl-transferases (DNMTs): their expression was slightly higher in obese subjects than in non-obese.The transcriptome analysis was made using HumanHT-12 Expression BeadChip by Illumina™ on myoblast and myotubes of both healthy and obese subjects. These microarrays allowed them to profile the expression pattern of almost the whole genome.The gene sets obtained were then compared through GSEA (Gene Set Enrichment Analysis) in which gene sets are compared to discover differences in expression pattern. As expected, MRFs and metabolism-related gene expression varied according to the state of cell differentiation both in obese and non-obese subjects. However, they did not find significant differences in expression of genes differentially methylated between obese and non-obese subjects in myoblasts nor in myotubes. The transcriptomic analysis pointed out the up-regulation of IL-32 and a down-regulation of PSGs and CGBs, after differentiation. This was the first time that those genes were linked to myogenesis. Between all of them, Interleukin 32 was selected to follow up experiments: with siRNA in human cells and heterologous expression in mouse, the authors pointed out that IL-32 expression has a negative role on MRFs expression, like MYOD1, and it can lead to insulin insensitivity, a very usual consequence of obesity. This novel role of IL-32 in myogenesis could be the first step for future study regarding IL-32 inhibitors to improve muscle mass or to further explore its function in muscle biology.
In conclusion, although the data from DNA methylation analysis highlighted a connection between altered methylation and obesity, its function remained unclear. In fact, Davegårdh and collaborators could not explain the cause and effect mechanism, whether the methylations are responsible of increasing obesity risk or it is obesity to enhance DNA methylation. As the author stated: “While approximately three times more methylation changes took place in the obese during differentiation, only a modest number of differences were observed in myoblasts of non-obese subjects. It may be explained by large variances within groups” [1]. This aspect could also be elaborated in future studies, to better understand obesity and to explore what happens on methylome while losing weight.
References
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