RSPO3 is a novel contraction-inducible factor identified in an “in vitro exercise model” using primary human myotubes

Abstract

The physiological significance of skeletal muscle as a secretory organ is now well known but we can only speculate as to the existence of as-yet-unidentified myokines, especially those upregulated in response to muscle contractile activity. We first attempted to establish an “insert-chamber based in vitro exercise model” allowing the miniature but high cell-density culture state enabling highly developed contractile human myotubes to be readily obtained by applying electric pulse stimulation (EPS). By employing this in vitro exercise model, we identified R-spondin 3 (RSPO3) as a novel contraction-inducible myokine produced by cultured human myotubes. Contraction-dependent muscular RSPO3 mRNA upregulation was confirmed in skeletal muscles of mice subjected to sciatic nerve mediated in situ contraction as well as those of mice after 2 h of running. Pharmacological in vitro experiments demonstrated a relatively high concentration of metformin (millimolar range) to suppress the contraction-inducible mRNA upregulation of human myokines including RSPO3, interleukin (IL)-6, IL-8 and CXCL1. Our data also suggest human RSPO3 to be a paracrine factor that may positively participate in the myogenesis processes of myoblasts and satellite cells. Thus, the “insert chamber-based in vitro exercise model” is a potentially valuable research tool for investigating contraction-inducible biological responses of human myotubes usually exhibiting poorer contractility development even in the setting of EPS treatment.

Identification of Clinically Relevant Brain Endothelial Cell Biomarkers in Plasma

“…As RSPO3 regulates vascular network remodeling and activates inflammatory gene expression that can exert synergistic effects on BBB permeability, [36][37][38] the adverse neuroimaging and clinical outcomes associated with RSPO3 may reflect secondary consequences of aberrant inflammatory signaling (a postulation supported by its positive relationship with plasma GFAP), compromised BBB integrity or systemic drivers of angiogenesis, such as differential metabolic demands. 39 With the exception of RSPO3, our candidate BEC proteins displayed unexpected associations with ADRD biomarkers, including Aβ 42/40 and pTau-181. Given that BBB integrity likely influences the extent to which biomarkers originating from the central nervous system can migrate into the periphery, the observed relationships may be attributed to neurobiological mechanisms that are distinct from coevolving neurodegenerative processes.…”

Increased R‐spondin 3 contributes to aerobic exercise‐induced protection against renal vascular endothelial hyperpermeability and acute kidney injury

“…A previous study has reported that a 2 h of running exercise training per se can increase RSPO3 mRNA expression in murine skeletal muscles. 25 We then observed the effect of exercise training on RSPO3 mRNA expression in the soleus muscle, liver, and myocardium. The serum levels of RSPO3 were also examined.…”

“…In particular, RSPO1 and RSPO3 protect against endothelial dysfunction in murine models of mesenteric ischaemia/reperfusion injury 23 and mechanical stretch‐induced lung injury, 24 respectively. Intriguingly, a recent study indicates that muscular RSPO3 expression is significantly upregulated in mice after 2 h of wheel running 25 . These studies prompt us to investigate whether exercise training exerts protection against sepsis‐associated AKI via modulating RSPO3 expression.…”

“…Intriguingly, a recent study indicates that muscular RSPO3 expression is significantly upregulated in mice after 2 h of wheel running. 25 These studies prompt us to investigate whether exercise training exerts protection against sepsisassociated AKI via modulating RSPO3 expression.…”