Glycyl-l-histidyl-l-lysine-Cu2+ rescues cigarette smoking-induced skeletal muscle dysfunction via a sirtuin 1-dependent pathway.

Abstract

Skeletal muscle dysfunction is an important co-morbidity in patients with chronic obstructive pulmonary disease (COPD) and is significantly associated with increased mortality. Oxidative stress has been demonstrated an important trigger for COPD-related skeletal muscle dysfunction. Glycine-histidine-lysine (GHK) is an active tripeptide, which is a normal component of human plasma, saliva, and urine; promotes tissue regeneration; and acts as an anti-inflammatory and antioxidant properties. The purpose of this study was to determine whether GHK is involved in COPD-related skeletal muscle dysfunction. The plasma GHK level in patients with COPD (n=9) and age-paired healthy subjects (n=11) were detected using reversed-phase high-performance liquid chromatography. The complex GHK with Cu (GHK-Cu) was used in in vitro (C2C12 myotubes) and in vivo experiments (cigarette smoking [CS]-exposure mouse model) to explore the involvement of GHK in CS-induced skeletal muscle dysfunction. Compared with healthy control, plasma GHK levels were decreased in patients with COPD (70.27±38.87ng/mL vs. 133.0±54.54ng/mL, P=0.009). And plasma GHK levels in patients with COPD were associated with pectoralis muscle area (R=0.684, P=0.042), inflammatory factor TNF-α (R=-0.696, P=0.037), and antioxidative stress factor SOD2 (R=0.721, P=0.029). GHK-Cu was found to rescue CSE-induced skeletal muscle dysfunction in C2C12 myotubes, as evidenced by increased expression of myosin heavy chain, reduced expression of MuRF1 and atrogin-1, elevated mitochondrial content, and enhanced resistance to oxidative stress. In CS-induced muscle dysfunction C57BL/6 mice, GHK-Cu treatment (0.2 and 2mg/kg) reduces CS-induced muscle mass loss (skeletal muscle weight (1.19±0.09% vs. 1.29±0.06%, 1.40±0.05%; P<0.05) and muscle cross-sectional area elevated (1055±552.4μm2 vs. 1797±620.9μm2 , 2252±534.0μm2 ; P<0.001), and also rescues CS-induced muscle weakness, indicated by improved grip strength (175.5±36.15g vs. 257.6±37.98g, 339.1±72.22g; P<0.01). Mechanistically, GHK-Cu directly binds and activates SIRT1(the binding energy was -6.1kcal/mol). Through activating SIRT1 deacetylation, GHK-Cu inhibits FoxO3a transcriptional activity to reduce protein degradation, deacetylates Nrf2 and contribute to its action of reducing oxidative stress by generation of anti-oxidant enzymes, increases PGC-1α expression to promote mitochondrial function. Finally, GHK-Cu could protect mice against CS-induced skeletal muscle dysfunction via SIRT1. Plasma glycyl-l-histidyl-l-lysine level in patients with chronic obstructive pulmonary disease was significantly decreased and was significantly associated with skeletal muscle mass. Exogenous administration of glycyl-l-histidyl-l-lysine-Cu2+ could protect against cigarette smoking-induced skeletal muscle dysfunction via sirtuin 1.