Effects of GHK-Cu on MMP and TIMP Expression, Collagen and Elastin Production, and Facial Wrinkle Parameters

Abstract

Background: Glycyl-L-histidyl-L-lysine-copper (GHK-Cu) is an endogenous tripeptide-copper complex involved in collagen synthesis and is used topically as a skin anti-aging and wound healing agent. However, its biological effects are yet to be fully elucidated. Objectives: To investigate the effects of GHK-Cu on gene expression of metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), and on production of collagen and elastin by human adult dermal fibroblasts (HDFa); and to investigate the effectiveness of topical application of GHK-Cu on wrinkle parameters in volunteers. Methods: Cultured HDFa were incubated with GHK-Cu at 0.01, 1 and 100 nM in cell culture medium. Gene expression (mRNA) for MMP1, MMP2, TIMP1 and TIMP2 in treated and control HDFa was measured by RT-PCR. Cellular production of collagen and elastin was measured colourmetrically using commercial assay kits. Correlations between gene expression and collagen and elastin production were determined. A randomised, double-blind clinical trial involving twice daily application of GHK-Cu, encapsulated in lipid-based nano-carrier, to facial skin of female subjects (n= 40, aged 40 to 65) was run over 8 weeks. The formulation vehicle (a serum) and a commercial cosmetic product containing Matrixyl® 3000, a lipophilic GHK derivative, were used as controls. Results: GHK-Cu significantly increased gene expression of MMP1 and MMP2 at the lowest concentration whilst simultaneously increasing the expression of TIMP1 at all the tested concentrations. All examined concentrations of GHK-Cu increased both collagen and elastin production. An increase of the mRNA expression ratio of TIMPs to MMPs was associated with an increase in collagen/elastin production. Application of GHK-Cu in nano-carriers to facial skin of volunteers significantly reduced wrinkle volume (31.6%; p=0.004) compared to Matrixyl® 3000, and significantly reduced wrinkle volume (55.8%; p<0.001) and wrinkle depth (32.8%; p=0.012) compared to control serum. Conclusions: GHK-Cu significantly increased collagen and elastin production by HDFa cells depending on the relative mRNA expression of their TIMP(s) over MMP. Topical application of GHK-Cu with the aid of nano-carriers reduced wrinkle volume to a significantly greater extent than the vehicle alone or a commercial product containing Matrixyl 3000®, a GHK lipophilic derivative.