Matrix-degrading metalloproteinases in photoaging

Abstract

UV radiation from the sun impacts skin health adversely through complex, multiple molecular pathways. Premature skin aging (photoaging) is among the most widely appreciated harmful effects of chronic exposure to solar UV radiation. Extensive damage to the dermal connective tissue is a hallmark of photoaged skin. Disruption of the normal architecture of skin connective tissue impairs skin function and causes it to look aged. UV irradiation induces expression of certain members of the matrix metalloproteinase (MMP) family, which degrade collagen and other extracellular matrix proteins that comprise the dermal connective tissue. Although the critical role of MMPs in photoaging is undeniable, important questions remain. This article summarizes our current understanding of the role of MMPs in the photoaging process and presents new data that (1) describe the expression and regulation by UV irradiation of all members of the MMP family in human skin in vivo and (2) quantify the relative contributions of epidermis and dermis to the expression of UV irradiation-induced MMPs in human skin in vivo.Journal of Investigative Dermatology Symposium Proceedings (2009) 14, 20-24; doi:10.1038/jidsymp.2009.8.

Figures

Figure 1

Basal gene expression of MMP family members in human skin in vivo. Full thickness (4mm) sun-protected buttock human skin was obtained from healthy adult human volunteers (eight subjects, average 36 years age), as previously described (Fisher, G.J, 1991. J Invest Dermatol 96:699–707; Quan, 2004, Amer J Pathol 169:482–490). Total RNA was extracted, using a commercial kit (RNeasy midikit, Qiagen, Chatsworth, CA) according to the manufacturer’s protocol. Reverse transcription was performed using Taqman Reverse Transcription kit (Applied Biosystems, Foster City, CA). Real-time RT-PCR was performed using a Taqman Universal PCR Master Mix kit (Applied Biosystems, Foster City, CA) and 7500 Real-Time PCR System (Applied Biosystems, Foster City, CA). All primers and probes were purchased from Applied Biosystems (Assays-on-Demand™ Gene Expression Products). Results are means±SEM of MMPs mRNA normalized to 36B4 (internal control) mRNA. All procedures involving human subjects were conducted in accord with the regulations set forth by the University of Michigan Institutional Review Board, and all subjects provided written informed consent.

Figure 2

UV irradiation induces MMP-1, MMP-3, and MMP-9 in human skin in vivo. Sun-protected buttock skin of healthy adult human volunteers was exposed to twice the minimum erythema dose of solar-simulated UV irradiation (SPEC 450W xenon arc solar simulator). Full thickness (4mm) biopsies were obtained 8 and 24 hours post irradiation. Total RNA was extracted and MMP-1, MMP-3, and MMP-9 mRNA levels were determined by real-time RT-PCR, as described in figure 1 legend. Results are means±SEM of MMPs mRNA normalized to 36B4 (internal control) mRNA. N=8, p

Figure 3

Laser capture microdisection (LCM) coupled real-time RT-PCR. Human skin punch biopsies were embedded in OCT, sectioned, and stained with H&E. Epidermis and dermis were captured using LCM (Leica ASLMD System, Leica Microsystems, Germany). Arrows indicate locations where epidermis and dermis were separated by laser. Total RNA was extracted from captured epidermis and dermis, and quantitative real-time RT-PCR was performed as described in figure 1 legend.

Figure 4

UV irradiation induces gene expression of MMP-1, MMP-3, and MMP-9 primarily in human epidermis in vivo. Sun-protected buttock skin of healthy adult human volunteers was exposed to twice the minimum erythema dose of solar-simulated UV irradiation (SPEC 450W xenon arc solar simulator). Full thickness (4mm) biopsies were obtained 24 hours post irradiation as described in Figure 1 legend. Total RNA were extracted from epidermis and dermis, which were obtained by laser capture microdissection. MMP-1, MMP-3, and MMP-9 mRNA levels were determined by real-time RT-PCR, as described in Figure 1 legend. Results are means±SEM of MMPs mRNA normalized to 36B4 (internal control) mRNA. N=6, p

Figure 5

Collagenase activity induced by solar-simulated UV irradiation in human skin in vivo. Sun-protected buttock skin of healthy adult human volunteers was exposed to twice the minimum erythema dose of solar-simulated UV irradiation (SPEC 450W xenon arc solar simulator). Full thickness (4mm) biopsies were obtained 24 hours post irradiation as described in Figure 1 legend. Collagenase activity was detected by in situ zymography, using FITC-labeled type I collagen as substrate (green fluorescence). Collagenease-catalyzed collagen cleavage causes loss of green fluorescence, resulting in darkened areas, which are most noticeable in the epidermis and upper dermis of UV-irradiated human skin. Images are representative of five experiments.