AMPKα1-LDHA, a new metabolic pathway, regulating stem cell fate

At resting state, muscle stem cells (MuSCs, also called satellite cells) remain quiescent in their niche, hanging to the myofiber. After injury, MuSCs activate, proliferate and fully differentiate to reform myofibers in order to repair the tissue. Also, MuSCs are able to return to quiescence and replenish the stem cell pool even after multiple injuries [1]. Although the control of the return into quiescence, also named self-renewal, is crucial for skeletal muscle homeostasis, few is known about its regulation. It is claimed that regulation of stem cell fate (embryonic stem cells, cancer stem cells, neural stem cells and hematopoietic stem cells and during aging) is dependent of the metabolic state of the cell and the surrounding stem cell niche. Thus, tailoring metabolic pathways might aid to regulate stem cell fate choice between self-renewal and differentiation. Recently, different studies demonstrate that MuSC activation is modulated by NAD⁺ metabolism, mTORC activity and also autophagy, supporting the link between metabolism and stem cell fate [2].

AMP Activated Kinase (AMPK) is the master regulator of cell metabolism. Composed of 3 different subunits (α, β and γ), each one having up to 3 isoforms, AMPK activity and targets can differ between cells and tissues. AMPK has been shown to have many roles in skeletal muscle homeostasis [3,4].

Recently, our work demonstrates for the first time that tailoring the metabolism can control MuSC fate and then that AMPKα1 (and not AMPKα2) has a crucial function in the Warburg-like effect [5]. Indeed, based on in vitro (freshly isolated MuSCs), ex vivo (isolated fibers) and in vivo (regenerating muscles) experiments, our study highlights that deleting AMPKα1 gene forces MuSCs to self-renew and delays their differentiation, inducing impairment of skeletal muscle regeneration (i.e. reduction of weight, myofiber size and number of nuclei per fiber). Furthermore, AMPKα1^−/− muscle precursor cells exhibit in vitro a more glycolytic metabolism showed by an increase in lactate release and a higher activity of the key enzyme of the glycolysis: the Lactate Dehydrogenase (LDH). Mitochondria content of these cells seems to be altered. Indeed, a decrease of the expression of mitochondrial biogenesis genes (PGC1α and PGC1β), a decrease of the citrate synthase activity (a key enzyme of the OXPHOS), a decrease of the TOM22 expression (a transporter involved in mitochondrial function) and a reduced maximal Oxygen Consumption Rate are evidenced in absence of AMPKα1. Altogether, these data suggest that MuSCs need to switch their metabolism from glycolysis to oxidative phosphorylation in order to fully differentiate.

LDH has been described as a non-limiting enzyme of the glycolysis. However, our study showed that LDH activity is strongly modulated in absence of AMPKα1 in non-differentiated cells such as hematopoietic stem cells and MuSCs, as compared with differentiated and post-mitotic cells such as myofibers.

In parallel, by using different type of media (glucose supplemented media or galactose supplemented media), we demonstrated that pushing cells to perform only oxidative phosphorylation (using the galactose media) tailors their fate by inhibiting their self-renewal. Interestingly, AMPKα1^−/− MuSCs do not modify their self-renewal, whatever the media used, showing that AMPKα1 is the key regulator allowing MuSCs to skew their metabolism in order to differentiate.

Moreover, specific inhibition of LDH activity (using Oxamate, an allosteric inhibitor of the LDH) induces a decrease of AMPKα1 MuSC self-renewal. Inversely, activation of AMPKα1 (using the compound 991, a potent and specific AMPK activator) induces a decrease of MuSC self-renewal, associated with a decrease of LDH activity.

Finally, in vivo experiments, by electroporation of a plasmid coding for the LDH, demonstrate that MuSC self-renewal can be modulated by increasing only LDH activity, validating our previous in vitro experiments. Altogether, these data strongly support the crucial function of AMPKα1-LDH pathway in MuSC fate and that tailoring MuSC metabolism can deeply affect their fate.

Nothing is known about the possible function of the second isoform of the catalytic subunit (AMPKα2). Lantier and colleagues have shown an increase of the expression of AMPKα2 during myoblast differentiation [6]. These data suggest that AMPKα2 is also involved in muscle cell behaviour and raise new questions about its role in the regulation of muscle cell fate. It has been shown that some of the metabolic adaptations seem to preferentially occur through the regulation of AMPKα2, suggesting distinct physiological roles between AMPKα1 and AMPKα2. Indeed, a lack of AMPKα2 was associated with a lower ATP amount in skeletal muscle [7], indicating that the remaining AMPKα1 isoform was not able to compensate for the loss of AMPKα2 to maintain muscle energy balance. These results suggest a regulated balance between AMPKα1 and AMPKα2 activities during myogenesis. AMPKα1 tailors early activation of MuSCs and their self-renewal, through the regulation of the LDH, while AMPKα2 may modulate myogenesis (i.e. differentiation and fusion).

Figure 1.

After damage, MuSCs activate, proliferate and differentiate or self-renew. While inhibition of AMPKα1 triggers the cells to self-renew by up-regulating the Lactate Dehydrogenase (LDH) activity and glycolysis; down-regulation of the OXPHOS and in general the mitochondrial activity, activate AMPKα1 and leads MuSC to differentiate.

Funding Statement

European Commission [LSHM-CT-2004-005272], Seventh Framework Programme (FP7) [241440], Association Française contre les Myopathies [19046]

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.