Diabetes mellitus and the skin.

Diabetes is a debilitating, life-threatening disease accounting in 2015 for the death of 5 million people worldwide. According to new estimations, 415 million adults currently suffer from the disease, and this number is expected to rise to 642 million by 2040. High glucose blood levels also affect the skin among systemic organs, and skin disorders can often predict the onset of this metabolic disorder. In this review, we address the pathomechanistic effects of diabetes on the skin and give an overview on the most common skin diseases associated with diabetes.

[Wound healing in the elderly]. / Wundheilung im Alter.

Restoration of tissue integrity is essential for host defense and protection of the organism. The efficacy and quality of skin repair varies significantly over a person's lifetime. Whereas prenatal wound healing is characterized by regeneration and scarless healing, scarring, fibrosis, and loss of function are features of postnatal repair. In fact, aging is the prominent risk factor for chronic wounds, skin fragility, infections, comorbidities, and decreased quality of life. Current strategies for restoration of tissue integrity and wound therapy are not sufficient and require further investigation of the underlying pathomechanisms and the development of causal-based concepts.

Interplay of H2A deubiquitinase 2A-DUB/Mysm1 and the p19(ARF)/p53 axis in hematopoiesis, early T-cell development and tissue differentiation.

Monoubiquitination of core histone 2A (H2A-K119u) has a critical role in gene regulation in hematopoietic differentiation and other developmental processes. To explore the interplay of histone H2A deubiquitinase Myb-like SWIRM and MPN domain containing1 (2A-DUB/Mysm1) with the p53 axis in the sequential differentiation of mature lymphocytes from progenitors, we systematically analyzed hematopoiesis and early T-cell development using Mysm1(-/-) and p53(-/-)Mysm1(-/-) mice. Mysm1(-/-) thymi were severely hypoplastic with <10% of wild-type cell numbers as a result of a reduction of early thymocyte progenitors in context with defective hematopoietic stem cells, a partial block at the double-negative (DN)1-DN2 transition and increased apoptosis of double-positive thymocytes. Increased rates of apoptosis were also detected in other tissues affected by Mysm1 deficiency, including the developing brain and the skin. By quantitative PCR and chromatin immunoprecipitation analyses, we identified p19(ARF), an important regulator of p53 tumor suppressor protein levels, as a potential Mysm1 target gene. In newly generated p53(-/-)Mysm1(-/-) double-deficient mice, anomalies of Mysm1(-/-) mice including reduction of lymphoid-primed multipotent progenitors, reduced thymocyte numbers and viability, and interestingly defective B-cell development, growth retardation, neurological defects, skin atrophy, and tail malformation were almost completely restored as well, substantiating the involvement of the p53 pathway in the alterations caused by Mysm1 deficiency. In conclusion, this investigation uncovers a novel link between H2A deubiquitinase 2A-DUB/Mysm1 and suppression of p53-mediated apoptotic programs during early lymphoid development and other developmental processes.

Photosensitization of uroporphyrin augments the ultraviolet A-induced synthesis of matrix metalloproteinases in human dermal fibroblasts.

Porphyria cutanea tarda is characterized by severe connective tissue damage in sun-exposed skin. The regulated synthesis and degradation of the extracellular matrix by various matrix metalloproteinases (MMPs) determine its amount and composition within the skin. In this study, we therefore asked whether long-wave ultraviolet irradiation (340-450 nm) in conjunction with uroporphyrin I could modulate the synthesis of MMPs with substrate specificities for dermal (collagens I, III, V; proteoglycans) and basement membrane components (collagens IV, VII; fibronectin; laminin) and whether synthesis of the counteracting tissue inhibitor of metalloproteinases is also affected. After irradiation of uroporphyrin-pretreated fibroblasts, specific mRNAs of MMP-1 and MMP-3 increased concomitantly up to 2.7-fold compared with ultraviolet-irradiated cells and up to 10-fold compared with mock-irradiated or uroporphyrin I-treated controls. In contrast, mRNA levels of tissue inhibitor of metalloproteinases remained unaltered. Similar results were obtained by immunoprecipitation. Gelatin and casein zymography revealed increased proteolytic activity of MMP-2 and MMP-3 in blister fluids of patients with porphyria cutanea tarda, indicating that similar events may occur in vivo. Using deuterium oxide as enhancer and sodium azide as quencher of singlet oxygen, we could increase or reduce MMP synthesis, suggesting that singlet oxygen is the major intermediate in the upregulation of MMPs after irradiation of uroporphyrin-pretreated fibroblasts. Taken together, our results show that ultraviolet irradiation alone, and to a greater extent in conjunction with uroporphyrin I, results in an unbalanced synthesis of MMPs that may contribute to the destruction of the dermis and basement membrane, leading to blistering and accelerated photoaging in porphyria cutanea tarda patients.

Isolation and identification of psoralen plus ultraviolet A (PUVA)-induced genes in human dermal fibroblasts by polymerase chain reaction-based subtractive hybridization.

Premature aging of the skin is a prominent side-effect of psoralen photoactivation, a therapy used for a variety of skin disorders. Recently, we demonstrated that treatment of human dermal fibroblasts with 8-methoxypsoralen and ultraviolet A irradiation resulted in a permanent growth arrest with a switch of mitotic to postmitotic fibroblasts. Furthermore, an upregulation of matrix-degrading metalloproteinases and a high level of de novo expression of the senescence-associated beta-galactosidase was detected in the PUVA-treated postmitotic fibroblasts. The molecular basis for this PUVA-induced change in the functional and morphologic phenotype of fibroblasts resembling or mimicking replicative senescence is, however, unknown. Herein after, we have used a polymerase chain reaction-based subtractive hybridization protocol to identify human genes that are induced by PUVA treatment. Application of polymerase chain reaction-Select resulted in the cloning of four PUVA genes. Sequence analysis and homology searches identified three cDNA clones of known genes related to cell cycle regulation (p21waf1/cip1), stress response (ferritin H) and connective tissue metabolism (tissue inhibitor of metalloproteinases-3), whereas one cDNA clone represented a novel gene (no. 478). Northern blot analyses were performed to confirm a PUVA-dependent increase in specific mRNA levels in human dermal fibroblasts in vitro. This report on the identification of growth arrest related genes in PUVA-treated fibroblasts may stimulate further research addressing the causal role of these known and novel genes in extrinsic and intrinsic aging processes on a molecular and cellular level.

Singlet oxygen may mediate the ultraviolet A-induced synthesis of interstitial collagenase.

Singlet oxygen has been postulated to be generated by Ultraviolet (UV) A irradiation of mammalian cells. We studied the role of singlet oxygen in the downstream signaling of the complex UV response leading to the induction of matrix-metalloproteinase-1 (interstitial collagenase/MMP-1). Exposure of cultured human fibroblasts to singlet oxygen, generated in a dark reaction by thermodissociation of the endoperoxide of the disodium salt of 3,3'-(1,4-naphthylidene) dipropionate (NDPO2) induced collagenase mRNA steady state levels in a dose dependent manner. The increase in collagenase expression after singlet-oxygen exposure generated with 3 mM NDPO2 was equivalent to that observed with UVA at a dose rate of 200-300 kJ/m2 and developed in a similar time course. In contrast, mRNA levels of TIMP-1, the specific tissue inhibitor of metalloproteinases, remained unchanged. Indirect evidence for the role of singlet oxygen in the UVA induction of collagenase comes from studies using singlet oxygen enhancer or quencher. Accordingly, incubation in deuterium oxide, an enhancer of singlet-oxygen lifetime, led to an additional increase in steady-state levels of collagenase mRNA after exposure to NDPO2 or to UVA irradiation. In contrast, sodium azide, a potent quencher of singlet oxygen, almost totally abrogated the induction of collagenase after exposure of fibroblasts to NDPO2 or to UVA irradiation. Similar results were obtained in studies of the proteins by radioimmunoprecipitation of MMP-1 and TIMP-1 using specific antibodies. Collectively, our data provide circumstantial evidence that singlet oxygen mediates the UVA induction of collagenase in vitro, whereas it does not exert any effect on TIMP-1 synthesis. The unbalanced synthesis of interstitial collagenase may contribute to the connective tissue damage in vivo related to photoaging and other photocutaneous disorders.

UVA-induced autocrine stimulation of fibroblast-derived-collagenase by IL-6: a possible mechanism in dermal photodamage?

Like other cytokines, IL-6 has been reported to stimulate collagenase. In this study we were interested in whether IL-6 is involved in the ultraviolet (UV) mediated up-regulation of fibroblast-derived collagenase. Confluent fibroblast monolayers were irradiated under standardized conditions. Following UVA irradiation the bioactivity of IL-6 increased up to fiftyfold in the supernatants of irradiated compared to mock-irradiated fibroblasts. As determined by Northern blot analysis this was also reflected on the pre-translational level by a tenfold increase of IL-6-specific mRNA following UVA irradiation. Induction of IL-6-specific mRNA was maximal at 6 h post-irradiation, thus clearly preceding the maximal induction of collagenase mRNA at 24 h post-irradiation. To elucidate the regulatory role of IL-6 in the UVA induction of fibroblast-derived collagenase, monospecific polyclonal neutralizing antibodies directed against recombinant human IL-6 and antisense oligonucleotides specifically inhibiting the translation of IL-6 mRNA were used at various concentrations. The amount of UVA-induced collagenase mRNA was reduced in a dose-dependent manner when antibodies or specific antisense oligonucleotides were present during and after irradiation. Taken together our data provide first evidence that UVA enhances IL-6 synthesis and secretion in fibroblasts. IL-6 induces via an autocrine mechanism collagenase and may thus contribute to the actinic damage of the dermis.

Adaptive antioxidant response of manganese-superoxide dismutase following repetitive UVA irradiation.

In response to the attack of reactive oxygen species, the skin has developed a complex antioxidant defense system including among others the manganese-superoxide dismutase (MnSOD). MnSOD dismutates the superoxide anion (O2*-) derived from the reduction of molecular oxygen to hydrogen peroxide (H2O2), which is detoxified by glutathione peroxidase to water and molecular oxygen. We have addressed the question whether MnSOD is inducible upon UVA irradiation and whether repetitive UV exposure, as practiced for the light-hardening during phototherapy of various photodermatoses, can even enhance the adaptive antioxidant response. Single exposure of four different strains of fibroblasts to UVA irradiation resulted in a dose- and time-dependent increase in specific MnSOD mRNA levels. Interestingly, repetitive UVA exposure at days 1, 2, and 3 at a dose rate of 200 kJ per m2 resulted in a 5-fold induction of specific MnSOD mRNA levels following the third UVA exposure. Similar results were obtained for MnSOD activity. This adaptive response in terms of upregulation of the antioxidant enzyme MnSOD correlates with the protection against high UV doses, if cells were preexposed to sublethal UV doses. Importantly, MnSOD substantially differed between the tested individuals in both mRNA and activity levels. Taken together, we here provide evidence for the increasing induction of MnSOD upon repetitive UVA irradiation that may contribute to the effective adaptive UVA response of the skin during light hardening in phototherapy. Interindividual differences in the inducibility of MnSOD might account for differences in the susceptibility to develop photodermatologic disorders related to photosensitivity, photoaging, and skin cancer. The molecular basis for interindividual differences in the inducibility of antioxidant enzymes remains to be elucidated.

Selective pick-up of increased iron by deferoxamine-coupled cellulose abrogates the iron-driven induction of matrix-degrading metalloproteinase 1 and lipid peroxidation in human dermal fibroblasts in vitro: a new dressing concept.

Using atomic absorption spectrum analysis, we found iron levels in exudates from chronic wounds to be significantly increased (3.71 +/- 1.56 micromol per g protein) compared to wound fluids from acute wounds derived from blister fluids (1.15 +/- 0.62 micromol per g protein, p < 0.02), drainage fluids of acute wounds (0.87 +/- 0.34 micromol per g protein, p < 0.002), and pooled human plasma of 50 volunteers (0.42 micromol per g protein). Increased free iron and an increase in reactive oxygen species released from neutrophils represent pathogenic key steps that --via the Fenton reaction - are thought to be responsible for the persistent inflammation, increased connective tissue degradation, and lipid peroxidation contributing to the prooxidant hostile microenvironment of chronic venous leg ulcers. We herein designed a selective pick-up dressing for iron ions by covalently binding deferoxamine to cellulose. No leakage occurred following gamma sterilization of the dressing and, more importantly, the deferoxamine-coupled cellulose dressing retained its iron complexing properties sufficient to reduce iron levels found in chronic venous ulcers to levels comparable to those found in acute wounds. In order to study the functionality of the dressing, human dermal fibroblasts were exposed to a Fenton reaction mimicking combination of 220 microM Fe(III) citrate and 1 mM ascorbate resulting in a 4-fold induction of matrix-degrading metalloproteinase 1 as determined by a matrix-degrading metalloproteinase 1 specific enzyme-linked immunosorbent assay. This induction was completely suppressed by dissolved deferoxamine at a concentration of 220 microM or by an equimolar amount of deferoxamine immobilized to cellulose. In addition, the Fe(III) citrate and ascorbate driven Fenton reaction resulted in an 8-fold increase in malondialdehyde, the major product of lipid peroxidation, as determined by high pressure liquid chromatography. This increase in malondialdehyde levels could be significantly reduced in the presence of the selective pick-up dressing coupled with deferoxamine suggesting that the deferoxamine dressing, in fact, prevents the development of a damaging prooxidant microenvironment and also protects from unfavorable consequences like matrix-degrading metalloproteinase 1 and lipid peroxide induction.

Hydrogen peroxide (H2O2) increases the steady-state mRNA levels of collagenase/MMP-1 in human dermal fibroblasts.

Reactive oxygen species (ROS) have been shown to be important messenger molecules in the induction of several genes. In human dermal fibroblasts the herbicide paraquat (PQ2+) was used to induce intracellular oxidative stress that was modulated by the inhibition of copper, zinc superoxide dismutase (Cu,ZnSOD), glutathione peroxidase (GSHPx), catalase, and blocking of the Fenton reaction. Interstitial collagenase (MMP-1) mRNA increased time dependently for up to 72 h following paraquat treatment. A correlation with the translation of MMP-1 could, however, only be detected up to 24 h, indicating an uncoupling of transcription and translation. Interleukin-1 alpha and beta mRNA showed two peaks at 6 h and 72 h. The inhibition of catalase by aminotriazol (ATZ), inhibition of GSHPx by buthionine sulfoximine (BSO), and blocking the Fenton reaction by the iron chelator desferrioxamine (DFO) in concert led to an increase in steady-state MMP-1 mRNA levels, possibly dependent on intracellular H2O2 increase. This combined treatment potentiated MMP-1 mRNA induction up to 6.5-fold compared to paraquat treated controls. Furthermore, exogenously added H2O2 caused an increase in MMP-1 mRNA levels. In contrast, inhibition of Cu,ZnSOD by diethyldithiocarbamate (DDC), leading to diminished H2O2 production from O2.-, decreased MMP-1 mRNA induction. Collectively, our data provide evidence that H2O2 is an important intermediate in the downstream signalling pathway finally leading to the induction of increased steady state MMP-1 mRNA levels. The synthesis of MMPs may contribute to connective tissue damage in vivo related to photoaging, inflammatory diseases, and tumor invasion.

The first peak of the UVB irradiation-dependent biphasic induction of vascular endothelial growth factor (VEGF) is due to phosphorylation of the epidermal growth factor receptor and independent of autocrine transforming growth factor alpha.

Ultraviolet B (UVB) irradiation, the major damaging component of sunlight, has earlier been reported to enhance cutaneous angiogenesis in chronically sun-exposed skin. We herein provide first evidence for a biphasic induction of the vascular endothelial growth factor (VEGF) following UVB irradiation of the human epidermal cell line HaCaT. The first VEGF peak occurred on mRNA level at 1 h and on protein level at 4 h postirradiation and is fully mediated by the UVB-dependent phosphorylation of the epidermal growth factor receptor, which subsequent to its phosphorylation also initiates at least in part the synthesis of transforming growth factor alpha that confers as shown previously the second late VEGF peak at 8 h on mRNA and at 24 h on protein level.

Singlet oxygen induces collagenase expression in human skin fibroblasts.

Singlet oxygen generated in a dark reaction by thermodissociation of an endoperoxide (NDPO2) elicits an increase in mRNA of interstitial collagenase (MMP-1) in cultured human fibroblasts. The effect is enhanced in deuterium oxide-based medium and is abolished in the presence of non-toxic doses of sodium azide. In contrast, the mRNA level of the tissue inhibitor of metalloproteinases (TIMP-1) remains unaltered under these experimental conditions. These observations support the suggestion that an unbalanced synthesis of collagenase and TIMP reported to occur following UV-A irradiation or during inflammatory conditions may be mediated by singlet oxygen.

Singlet oxygen is an early intermediate in cytokine-dependent ultraviolet-A induction of interstitial collagenase in human dermal fibroblasts in vitro.

Ultraviolet (UV) A irradiation of human dermal fibroblasts elicits an increase in specific mRNA amounts and bioactivities of the cytokines IL-1alpha, IL-1beta, and IL-6. These effects are enhanced in deuterium oxide-based medium and are diminished in the presence of non-toxic concentrations of sodium azide. Furthermore, generating singlet oxygen outside the cells by irradiation of rose bengal-coated resin particles with visible light (lambda > 450 nm) results in the induction of interstitial collagenase, IL-1 and IL-6, similar to the response observed with UVA irradiation. These observations suggest that singlet oxygen is an early intermediate in the signaling pathway of IL-1 and IL-6 mediating UVA induction of interstitial collagenase (E.C. 3.4.24.7). Furthermore, singlet oxygen appears to initiate this complex UV response at the cell membrane.

Ultraviolet-B induction of interstitial collagenase and stromelyin-1 occurs in human dermal fibroblasts via an autocrine interleukin-6-dependent loop.

Ultraviolet-B irradiation of human dermal fibroblasts has earlier been shown to induce matrix-degrading metalloproteinases, thus driving connective tissue degradation in photoaging and photocarcinogenesis. Herein, we report that Ultraviolet-B irradiation led to a dramatic increase in specific mRNA and protein levels of interstitial collagenase, stromelysin and interleukin-6. By contrast, the major tissue inhibitor of matrix-degrading metalloproteinases, TIMP-1, was unaffected. Monospecific neutralizing antibodies directed against human interleukin-6 significantly reduced the interstitial collagenase and stromelysin-1 protein levels. Taken together, our data provide the first evidence that Ultraviolet-B induction of interstitial collagenase and stromelysin-1 occurs via the synthesis and release of interleukin-6. Hence, this newly identified autocrine mechanism may contribute to dermal photodamage.

PCR-based subtractive hybridization identifies repressed genes in growth-arrested human dermal fibroblasts following combined treatment with 8-methoxypsoralen and UVA irradiation (PUVA).

To identify genes which are repressed in growth-arrested human dermal fibroblasts upon a single treatment with 8-methoxypsoralen and UVA irradiation (PUVA) we have used a PCR-based subtractive hybridization protocol resulting in cloning of four PUVA-repressed genes. Sequence analysis and homology searches identified three known genes related to growth control, lipid and connective tissue metabolism. One cDNA clone represented a novel gene. Northern blot analyses confirmed a PUVA-dependent reduction in mRNA expression in fibroblasts in vitro. The identification of growth arrest related repressed genes in PUVA-treated fibroblasts may stimulate further research addressing the causal role of these genes in the control and regulation of the postmitotic phenotype of fibroblasts on a molecular and cellular level.

Photoaging of the skin from phenotype to mechanisms.

The skin is increasingly exposed to ambient UV-irradiation thus increasing its risk for photooxidative damage with longterm detrimental effects like photoaging, which is characterized by wrinkles, loss of skin tone, and resilience. Photoaged skin displays prominent alterations in the cellular component and the extracellular matrix of the connective tissue with an accumulation of disorganized elastin and its microfibrillar component fibrillin in the deep dermis and a severe loss of interstitial collagens, the major structural proteins of the dermal connective tissue. The unifying pathogenic agents for these changes are UV-generated reactive oxygen species (ROS) that deplete and damage non-enzymatic and enzymatic antioxidant defense systems of the skin. As well as causing permanent genetic changes, ROS activate cytoplasmic signal transduction pathways in resident fibroblasts that are related to growth, differentiation, senescence, and connective tissue degradation. This review focuses on the role of UV-induced ROS in the photodamage of the skin resulting in biochemical and clinical characteristics of photoaging. In addition, the relationship of photoaging to intrinsic aging of the skin will be discussed. A decrease in the overall ROS load by efficient sunscreens or other protective agents may represent promising strategies to prevent or at least minimize ROS induced photoaging.

A newly adapted pulsed-field gel electrophoresis technique allows to detect distinct types of DNA damage at low frequencies in human dermal fibroblasts upon exposure to non-toxic H2O2 concentrations.

Reactive oxygen species (ROS) comprise several oxygen containing compounds, among them hydrogen peroxide (H2O2), which are generated by internal and external sources and play pleiotropic roles in physiological and pathological states. Skin cells as well as cells from other tissues have developed antioxidant defense mechanisms to protect themselves from high concentrations of ROS. Although biological and pathological roles of ROS have previously been elucidated, so far only limited knowledge exists regarding ROS-mediated generation of DNA breaks and base lesions occurring at low frequency in intact skin cells. This study was therefore designed to probe a newly adapted pulsed-field gel electrophoresis technique for the adequate measurement of high molecular weight DNA fragments as well as to investigate the protective role of the antioxidant enzyme catalase against H2O2-mediated damage in human dermal fibroblasts. We stably transfected and overexpressed the full-length catalase cDNA in the human dermal fibroblast cell line 1306 in culture and found that these cells are significantly more protected from cytotoxicity, overall DNA strand breaks, and 8-oxodeoxyguanine base lesions resulting from H2O2-triggered oxidative stress compared to vector-transfected 1306 cells or secondary dermal fibroblasts. This work has outlined the importance of catalase in the protection from H2O2-mediated cytotoxicity and DNA damage which--if unbalanced--even when occurring at low frequency are known to lead to genomic instability, a hallmark in carcinogenesis and premature aging.

UVA-induced autocrine stimulation of fibroblast-derived collagenase/MMP-1 by interrelated loops of interleukin-1 and interleukin-6.

Previous work has shown that fibroblast-derived collagenase/matrix-metalloproteinase-1 (MMP-1), responsible for the breakdown of dermal interstitial collagen, was dose-dependently induced in vitro and in vivo by UVA irradiation and this induction was at least partly mediated by interleukin-6 (IL-6). We here provide evidence that UVA-induced IL-1 alpha and IL-1 beta play a central role in the induction of the synthesis both of IL-6 and collagenase/MMP-1. In contrast to the late increase of IL-1 alpha and IL-1 beta mRNA levels at 6 h postirradiation, bioactivity of IL-1 is already detectable at 1 h postirradiation. This early peak of IL-1 bioactivity appears to be responsible for the induction of IL-6 synthesis and together with IL-6 lead to an increase of the steady-state mRNA level of collagenase/MMP-1 as deduced from studies using IL-1 alpha and IL-1 beta antisense oligonucleotides or neutralizing antibodies against IL-1 alpha and IL-1 beta. Besides the early posttranslationally controlled release of intracellular IL-1, a latter pretranslationally controlled synthesis and release of IL-1 perpetuates the UV response. From these data we suggest a UV-induced cytokine network consisting of IL-1 alpha, IL-1 beta and IL-6, which via interrelated autocrine loops induce collagenase/MMP-1 and thus may contribute to the loss of interstitial collagen in cutaneous photoaging.

Ultraviolet B wavelength dependence for the regulation of two major matrix-metalloproteinases and their inhibitor TIMP-1 in human dermal fibroblasts.

The wavelength dependence for the regulation of two major matrix-metalloproteinases, interstitial collagenase (MMP-1) and stromelysin-1 (MMP-3), and their major inhibitor, tissue inhibitor of metalloproteinases (TIMP-1), was studied in human dermal fibroblasts in vitro. Monochromatic irradiation at 302, 307, 312 and 317 nm with intensities ranging from 20 to 300 J/m2 increased MMP-1 and MMP-3 mRNA steady-state levels and the secretion of the corresponding proteins up to 4.4-fold, whereas almost no increase was observed at wavelengths < 290 nm. In contrast, the synthesis of TIMP-1 increased only marginally. This imbalance may contribute to the severe connective tissue damage related to photoaging of the skin. The wavelengths responsible for MMP-1 and MMP-3 induction reported here are distinct from the absorption spectrum of DNA and are different from results previously reported in the literature. Importantly, they overlap with wavelengths whose intensity is predicted to increase on the earth's surface upon ozone depletion. Intensities and particular wavelengths used in our studies in vitro can be absorbed readily by fibroblasts within the skin in vivo and, thus, are relevant for risk assessment and development of protective agents.

Ultraviolet B wavelength dependence for the regulation of two major matrix-metalloproteinases and their inhibitor TIMP-1 in human dermal fibroblasts.

The wavelength dependence for the regulation of two major matrix-metalloproteinases, interstitial collagenase (MMP-1) and stromelysin-1 (MMP-3), and their major inhibitor, tissue inhibitor of metalloproteinases (TIMP-1), was studied in human dermal fibroblasts in vitro. Monochromatic irradiation at 302, 307, 312 and 317 nm with intensities ranging from 20 to 300 J/m2 increased MMP-1 and MMP-3 mRNA steady-state levels and the secretion of the corresponding proteins up to 4.4-fold, whereas almost no increase was observed at wavelengths < 290 nm. In contrast, the synthesis of TIMP-1 increased only marginally. This imbalance may contribute to the severe connective tissue damage related to photoaging of the skin. The wavelengths responsible for MMP-1 and MMP-3 induction reported here are distinct from the absorption spectrum of DNA and are different from results previously reported in the literature. Importantly, they overlap with wavelengths whose intensity is predicted to increase on the earth's surface upon ozone depletion. Intensities and particular wavelengths used in our studies in vitro can be absorbed readily by fibroblasts within the skin in vivo and, thus, are relevant for risk assessment and development of protective agents.