A tumour
necrosis factor (TNF)-α-induced
atrophy model was used to examine
handelin activity in cultured C2C12 myotubes in vitro.
Lipopolysaccharide (LPS)-treated 8-week-old model mice and 23-month-old (aged) mice were used to examine the
therapeutic effects of
handelin on
cachexia- and aging-induced
muscle atrophy, respectively, in vivo.
Protein and
mRNA expressions were analysed by Western blotting, ELISA and quantitative PCR, respectively. Skeletal muscle mass was measured by histological analysis.
RESULTS:
Handelin treatment resulted in an upregulation of
protein levels of early (MyoD and
myogenin) and late (
myosin heavy chain, MyHC)
differentiation markers in C2C12 myotubes (P < 0.05), and enhanced mitochondrial respiratory (P < 0.05). In TNF-α-induced myotube
atrophy model,
handelin maintained MyHC
protein levels, increased
insulin-like growth factor (Igf1)
mRNA expression and phosphorylated
protein kinase B protein levels (P < 0.05).
Handelin also reduced atrogin-1 expression, inhibited nuclear factor-κB activation and reduced
mRNA levels of
interleukin (Il)6, Il1b and
chemokine ligand 1 (Cxcl1) (P < 0.05). In LPS-treated mice,
handelin increased
body weight (P < 0.05), the weight (P < 0.01) and cross-sectional area (CSA) of the soleus muscle (P < 0.0001) and improved motor function (P < 0.05). In aged mice,
handelin slightly increased the weight of the tibialis anterior muscle (P = 0.06) and CSA of the tibialis anterior and gastrocnemius muscles (P < 0.0001). In the tibialis anterior muscle of aged mice,
handelin upregulated
mRNA levels of Igf1 (P < 0.01), anti-inflammatory
cytokine Il10 (P < 0.01), mitochondrial biogenesis genes (P < 0.05) and
antioxidant-related
enzymes (P < 0.05) and strengthened Sod and
Cat enzyme activity (P < 0.05).
Handelin also reduced lipid peroxidation and protein carbonylation, downregulated
mRNA levels of Fbxo32, Mstn, Cxcl1, Il1b and Tnf (P < 0.05), and decreased IL-1β levels in serum (P < 0.05). Knockdown of Hsp70 or using an Hsp70 inhibitor abolished the ameliorating effects of
handelin on myotube
atrophy.
CONCLUSIONS:
Handelin ameliorated
cachexia- and aging-induced skeletal muscle
atrophy in vitro and in vivo, by maintaining homeostasis of
protein synthesis and degradation, possibly by inhibiting
inflammation.
Handelin is a potentially promising
drug candidate for the treatment of muscle wasting.