Collagen Fragmentation Promotes Oxidative Stress and Elevates Matrix Metalloproteinase-1 in Fibroblasts in Aged Human Skin

Abstract

Aged human skin is fragile because of fragmentation and loss of type I collagen fibrils, which confer strength and resiliency. We report here that dermal fibroblasts express increased levels of collagen-degrading matrix metalloproteinases-1 (MMP-1) in aged (>80 years old) compared with young (21 to 30 years old) human skin in vivo. Transcription factor AP-1 and ␣2␤1 integrin, which are key regulators of MMP-1 expression, are also elevated in fibroblasts in aged human skin in vivo. MMP-1 treatment of young skin in organ culture causes fragmentation of collagen fibrils and reduces fibroblast stretch, consistent with reduced mechanical tension , as observed in aged human skin. Limited fragmentation of three-dimensional collagen lattices with exogenous MMP-1 also reduces fibroblast stretch and mechanical tension. Furthermore, fibroblasts cultured in fragmented collagen lattices express elevated levels of MMP-1, AP-1, and ␣2␤1 integrin. Importantly, culture in fragmented collagen raises intracellular oxidant levels and treatment with antioxidant MitoQ 10 significantly reduces MMP-1 expression. These data identify positive feedback regulation that couples age-dependent MMP-1-catalyzed collagen fragmentation and oxidative stress. We propose that this self perpetuating cycle promotes human skin aging. These data extend the current understanding of the oxidative theory of aging beyond a cellular-centric view to include extracellular matrix and the critical role that connective tissue microenvironment plays in the biology of aging. Skin connective tissue (dermis) provides structural support for the skin's vasculature, appendages, and epidermis, which are vital to the function of skin. Structural integrity and function of the dermis are primarily dependent on its extracellular matrix, which is primarily composed of type I collagen fibrils. Type I collagen is the most abundant structural protein in skin, 1 and fragmented collagen fibrils are prominent, characteristic features of aged human skin in vivo.2-4 This fragmentation seriously impairs both the mechanical properties of skin, and the functions of cells that reside within the dermis. Clinically, this impairment manifests as delayed wound healing, reduced vascularization, propensity to bruise, and thin skin. Failure of normal functional interactions among dermal cells and their extracellular matrix microenvironment underlie these age-dependent phenotypic alterations. 6Damage to the collagenous extracellular matrix of the dermis can be observed at both the histological and ultrastructural level. 5,[7][8][9] In young dermis, intact, tightly packed, well-organized, long collagen fibrils are abundant. In contrast, in aged dermis, collagen fibrils are fragmented, disorganized, and sparse, resulting in the appearance of amorphous open space. Quantitative biochemical analysis reveals that the amount of fragmented collagen is 4.3-fold greater in aged (Ͼ80 years old) compared with young (21 to 30 years old) human dermis in vivo.

Reduction of fibroblast size/mechanical force down‐regulates TGF ‐β type II receptor: implications for human skin aging

“…These enlarged fibroblasts produce increased levels of collagen and CTGF/CCN2. Interestingly, we recently reported that reduced fibroblast size/mechanical force also up‐regulates matrix metalloproteinase‐1 expression and thus causes collagen fibril fragmentation (Qin et al ., 2014), as observed in aged human skin in vivo (Fisher et al ., 2008, 2009). These data combined with our current findings provide a foundation for our understanding the cellular and molecular basis of age‐related loss of collagen production and increased collagen fragmentation in aged human skin.…”

“…Cells between passage 3 and 10 were used for all experiments. 3D collagen lattices were prepared based on previous publication with minor modification (Fisher et al ., 2009). Briefly, neutralized rat tail type I collagen (2 mg mL −1 , BD, Biosciences) was suspended in medium cocktail [DMEM, NaHCO 3 (44 m m ), L‐glutamine (4 m m ), Folic Acid (9 m m ), and neutralized with 1N NaOH to pH 7.2].…”

“…During aging, the dermal ECM undergoes progressive fragmentation that impairs fibroblast attachment with consequent reduction of size (Varani et al ., 2004; Fisher et al ., 2008, 2009; Quan et al ., 2013a). In aged skin, reduced size of fibroblasts is accompanied by their decreased production of key ECM components, such as type I collagen, fibronectin, and connective tissue growth factor (CTGF/CCN2) (Varani et al ., 2004, 2006; Fisher et al ., 2008; Quan et al ., 2010).…”

Wpływ L-argininy i kwasu askorbinowego na zawartość tłuszczu trzewnego oraz stężenia metaloproteinaz 2 i 9 u szczurów karmionych dietą wysokotłuszczową

“…In pigs fed a high-cholesterol diet [31] the supplementation of vitamins C and E markedly prevents lipid peroxidation, increases neointimal collagen content, and reduces hypercholesterolaemiainduced changes in vascular MMP-1. The fibroblasts of aged human skin cultured in fragments, manifest raised intracellular oxidant levels; conversely, treatment with antioxidant MitoQ (10) significantly reduces MMP-1 expression [32]. The underlying mechanisms that have been discussed include: upregulation of the expression of genes involved in cell proliferation, regulation of transcription, as well as downregulation of genes involved in lipid metabolism, cell adhesion, and differentiation.…”

The decreased secretion of hyaluronan by older human fibroblasts under physiological conditions is mainly associated with the down-regulated expression of hyaluronan synthases but not with the expression levels of hyaluronidases

“…In photoShuko Terazawa and Hiroaki Nakajima have contributed equally to this work. aged skin, there is a marked deficiency of HA (Margelin et al 1996;Takahashi et al 1995Takahashi et al , 1996 in addition to the fragmentation and loss of type I collagen fibrils due to the up-regulated activity of matrix-metalloproteinase (MMP)-1 (Fisher et al 2008(Fisher et al , 2009). Available evidence indicates that naturally aged, sun-protected skin and photoaged skin share important molecular features, including connective tissue damage, elevated matrix metalloproteinase levels and reduced collagen production (Varani et al 2001(Varani et al , 2002.…”