Gebbers Narcisa, Hirt-Burri Nathalie, Scaletta Corinne, Hoffmann Gerd, Applegate Lee Ann
Orthopedic Cell Therapy Unit, University Hospital, Lausanne, Switzerland.
Ger Med Sci. 2007 Nov 14;5:Doc08.
Excessive exposure to solar ultraviolet radiation is involved in the complex biologic process of cutaneous aging. Wavelengths in the ultraviolet-A and -B range (UV-A and UV-B) have been shown to be responsible for the induction of proteases, e. g. the collagenase matrix metalloproteinase 1 (MMP-1), which are related to cell aging. As devices emitting longer wavelengths are widely used in therapeutic and cosmetic interventions and as the induction of MMP-1 by water-filtered infrared-A (wIRA) had been discussed, it was of interest to assess effects of wIRA on the cellular and molecular level known to be possibly involved in cutaneous degeneration.
Investigation of the biological implications of widely used water-filtered infrared-A (wIRA) radiators for clinical use on human skin fibroblasts assessed by MMP-1 gene expression (MMP-1 messenger ribonucleic acid (mRNA) expression).
Human skin fibroblasts were irradiated with approximately 88% wIRA (780-1400 nm) and 12% red light (RL, 665-780 nm) with 380 mW/cm(2) wIRA(+RL) (333 mW/cm(2) wIRA) on the one hand and for comparison with UV-A (330-400 nm, mainly UV-A1) and a small amount of blue light (BL, 400-450 nm) with 28 mW/cm(2) UV-A(+BL) on the other hand. Survival curves were established by colony forming ability after single exposures between 15 minutes and 8 hours to wIRA(+RL) (340-10880 J/cm(2) wIRA(+RL), 300-9600 J/cm(2) wIRA) or 15-45 minutes to UV-A(+BL) (25-75 J/cm(2) UV-A(+BL)). Both conventional Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) and quantitative real-time RT-PCR techniques were used to determine the induction of MMP-1 mRNA at two physiologic temperatures for skin fibroblasts (30 degrees C and 37 degrees C) in single exposure regimens (15-60 minutes wIRA(+RL), 340-1360 J/cm(2) wIRA(+RL), 300-1200 J/cm(2) wIRA; 30 minutes UV-A(+BL), 50 J/cm(2) UV-A(+BL)) and in addition at 30 degrees C in a repeated exposure protocol (up to 10 times 15 minutes wIRA(+RL) with 340 J/cm(2) wIRA(+RL), 300 J/cm(2) wIRA at each time).
Single exposure of cultured human dermal fibroblasts to UV-A(+BL) radiation yielded a very high increase in MMP-1 mRNA expression (11 +/-1 fold expression for RT-PCR and 76 +/-2 fold expression for real-time RT-PCR both at 30 degrees C, 75 +/-1 fold expression for real-time RT-PCR at 37 degrees C) and a dose-dependent decrease in cell survival. In contrast, wIRA(+RL) did not produce cell death and did not induce a systematic increase in MMP-1 mRNA expression (less than twofold expression, within the laboratory range of fluctuation) detectable with the sensitive methods applied. Additionally, repeated exposure of human skin fibroblasts to wIRA(+RL) did not induce MMP-1 mRNA expression systematically (less than twofold expression by up to 10 consecutive wIRA(+RL) exposures and analysis with real-time RT-PCR).
wIRA(+RL) even at the investigated disproportionally high irradiances does not induce cell death or a systematic increase of MMP-1 mRNA expression, both of which can be easily induced by UV-A radiation. Furthermore, these results support previous findings of in vivo investigations on collagenase induction by UV-A but not wIRA and show that infrared-A with appropriate irradiances does not seem to be involved in MMP-1 mediated photoaging of the skin. As suggested by previously published studies wIRA could even be implicated in a protective manner.
过度暴露于太阳紫外线辐射参与皮肤老化的复杂生物学过程。已证明紫外线-A和紫外线-B范围内的波长(UV-A和UV-B)可诱导蛋白酶,例如与细胞老化相关的胶原酶基质金属蛋白酶1(MMP-1)。由于发射较长波长的设备广泛用于治疗和美容干预,并且已讨论了水过滤红外-A(wIRA)对MMP-1的诱导作用,因此评估wIRA在已知可能参与皮肤退化的细胞和分子水平上的作用具有重要意义。
通过MMP-1基因表达(MMP-1信使核糖核酸(mRNA)表达)评估临床广泛使用的水过滤红外-A(wIRA)辐射器对人皮肤成纤维细胞的生物学影响。
一方面,用约88%的wIRA(780 - 1400 nm)和12%的红光(RL,665 - 780 nm)以380 mW/cm²的wIRA(+RL)(333 mW/cm²的wIRA)照射人皮肤成纤维细胞;另一方面,为作比较,用28 mW/cm²的UV-A(+BL)(330 - 400 nm,主要是UV-A1)和少量蓝光(BL,400 - 450 nm)照射。通过在15分钟至8小时内单次暴露于wIRA(+RL)(340 - 10880 J/cm²的wIRA(+RL),300 - 9600 J/cm²的wIRA)或15 - 45分钟内暴露于UV-A(+BL)(25 - 75 J/cm²的UV-A(+BL))后的集落形成能力建立存活曲线。使用传统逆转录聚合酶链反应(RT-PCR)和定量实时RT-PCR技术,在皮肤成纤维细胞的两个生理温度(30℃和37℃)下,通过单次暴露方案(15 - 60分钟的wIRA(+RL),340 - 1360 J/cm²的wIRA(+RL),300 - 1200 J/cm²的wIRA;30分钟的UV-A(+BL),50 J/cm²的UV-A(+BL))以及在30℃下的重复暴露方案(高达10次15分钟的wIRA(+RL),每次340 J/cm²的wIRA(+RL),300 J/cm²的wIRA)来测定MMP-1 mRNA的诱导情况。
培养的人真皮成纤维细胞单次暴露于UV-A(+BL)辐射后,MMP-1 mRNA表达大幅增加(30℃时RT-PCR为11±1倍表达,实时RT-PCR为76±2倍表达;37℃时实时RT-PCR为75±1倍表达),且细胞存活率呈剂量依赖性下降。相比之下,wIRA(+RL)未导致细胞死亡,也未诱导MMP-1 mRNA表达系统性增加(小于两倍表达,在实验室波动范围内),采用的敏感方法可检测到这一点。此外,人皮肤成纤维细胞重复暴露于wIRA(+RL)也未系统性诱导MMP-1 mRNA表达(通过实时RT-PCR分析,连续高达10次wIRA(+RL)暴露后小于两倍表达)。
即使在研究的不成比例的高辐照度下,wIRA(+RL)也不会诱导细胞死亡或MMP-1 mRNA表达系统性增加,而UV-A辐射很容易诱导这两种情况。此外,这些结果支持先前关于UV-A而非wIRA诱导胶原酶的体内研究结果,表明适当辐照度的红外-A似乎不参与MMP-1介导的皮肤光老化。如先前发表的研究所暗示,wIRA甚至可能具有保护作用。