ABSTRACT
Background Muscle stem cells (MuSCs) are requisite for skeletal muscle regeneration and homeostasis. Proper functioning of MuSCs, including activation, proliferation, and fate decision, is determined by an orchestrated series of events and communication between MuSCs and their niche consisting of the host myofiber and neighbouring cells. A multitude of biochemical stimuli are known to regulate fate and function of MuSCs. However, in addition to biochemical factors, it is conceivable that MuSCs residing between basal lamina and sarcolemma of myofibers are subjected to mechanical forces during muscle stretch-shortening cycles due to myofascial connections between MuSCs and myofibers. MuSCs have been shown to respond to mechanical forces in vitro but it remains to be proven whether physical forces are also exerted on MuSCs in their native niche and whether they contribute to the functioning and fate of MuSCs.
Methods MuSCs deformation in their native niche resulting from mechanical loading of ex vivo myofiber bundles were visualized utilizing mT/mG double-fluorescent Cre-reporter mouse and multiphoton microscopy. MuSCs were subjected to 1 hour pulsating fluid shear stress with a peak shear stress rate of 8.8 Pa/s. After treatment, nitric oxide and mRNA expression levels of genes involved in regulation of MuSC proliferation and differentiation were determined.
Results Ex vivo stretching of extensor digitorum longus and soleus myofiber bundles caused compression as well as tensile and shear deformation of MuSCs in their niche. MuSCs responded to pulsating fluid shear stress in vitro with increased nitric oxide production and an upward trend in iNOS mRNA levels, while nNOS expression was unaltered. Pulsating fluid shear stress enhanced gene expression of c-Fos, Cdk4, and IL-6, while expression of Wnt1, MyoD, Myog, Wnt5a, COX2, Rspo1, Vangl2, Wnt10b, and MGF remained unchanged.
Conclusions We conclude that MuSCs in their native niche are subjected to force-induced deformations due to myofiber stretch-shortening. Moreover, MuSCs are mechanoresponsive as evident by pulsating fluid shear stress-mediated expression of factors by MuSCs known to promote proliferation.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
E-mail addresses of all authors M. Haroon n.i.l.haroon{at}vu.nl, J. Klein-Nulend j.kleinnulend{at}acta.nl, A.D. Bakker a.bakker{at}acta.nl, J. Jin j.jin{at}acta.nl, C. Offringa c.offringa{at}vu.nl, F. Le Grand fabien.le-grand{at}inserm.fr, L. Giordani lorenzo.giordani{at}courriel.upmc.fr, K.J. Liu karen.liu{at}kcl.ac.uk, R.D. Knight robert.knight{at}kcl.ac.uk, R.T. Jaspers r.t.jaspers{at}vu.nl
ABREVIATIONS
- AP-1
- Activator protein-1
- Ca2+
- Calcium
- Cdk4
- Cyclin-dependent kinase 4
- COX2
- Cyclooxygenase-2
- Dkk1
- Dickkopf-related protein 1
- ECM
- Extracellular matrix
- EDL
- Extensor digitorum longus
- eNOS
- Endothelial nitric oxide synthase
- FBS
- Fetal bovine serum
- HGF
- Hepatocyte growth factor
- IL-6
- Interleukin-6
- iNOS
- Inducible nitric oxide synthase
- mG
- membrane targeted green fluorescent protein
- MGF
- Mechano growth factor
- MITK
- Medical Imaging Interaction Toolkit
- mT
- Membrane-targeted tandem dimer Tomato
- MuSCs
- Muscle stem cells
- MyoD
- Myogenic differenciation
- Myog
- Myogenin
- nNOS
- Neuronal nitric oxide synthase
- NO
- Nitric oxide
- NO2−
- Nitrite
- NOS
- nitric oxide synthase
- Pax7
- Paired box 7
- PCP
- Planar cell polarity
- PFSS
- Pulsating fluid shear stress
- Rspo1
- R-spondin-1
- SO
- Soleus
- Vangl2
- Vang-like protein 2