紫外线对光化性角化病和正常皮肤p53、基质金属蛋白酶2、9表达的影响

目的 探讨不同剂量紫外线对光化性角化病（AK）与正常皮肤组织中增殖与凋亡、及p53、基质金属蛋白酶（MMP）2和MMP9表达的影响。方法 AK与正常皮肤组织分别分为4个组，对照组（照射剂量为0 J/cm2）、5 J/cm2组、10 J/cm2组、20 J/cm2组。每块组织连续照射4 d，继续培养24 h后取材。TUNEL检测细胞凋亡、Ki-67检测细胞增殖情况，定量PCR及免疫组化法检测p53、MMP2及MMP9表达。 结果 紫外线照射后，与对照组相比，凋亡细胞百分比在正常皮肤组织 10、20 J/cm2组（46.8 ± 2.1，56.7 ± 2.4）增加，在AK 20 J/cm2（43.5 ± 1.5）组增加，正常皮肤组织10、20 J/cm2组比AK同剂量组增加（P < 0.05）；Ki-67阳性细胞百分比在正常皮肤组织20 J/cm2组（3.34 ± 0.76）表达减少（P < 0.05），在AK中无明显变化（P > 0.05）；p53 mRNA（5 J/cm2：1.106 ± 0.025；10 J/cm2：1.259 ± 0.045；20 J/cm2：1.425 ± 0.053）及蛋白（10 J/cm2：0.1169 ± 0.0032；20 J/cm2：0.1454 ± 0.0047）在正常皮肤组织中表达增加，在AK中，mRNA（10 J/cm2：0.611 ± 0.050；20 J/cm2：0.578 ± 0.070）及蛋白（20 J/cm2：0.0404 ± 0.0027）表达减少（P < 0.05）；正常皮肤组织中MMP2 mRNA及蛋白（10 J/cm2：1.086 ± 0.013，0.0843 ± 0.0024；20 J/cm2：1.417 ± 0.036，0.1236 ± 0.0042）、MMP9 mRNA及蛋白（20 J/cm2：1.395 ± 0.026，0.3065 ± 0.0162）表达增加，AK中MMP2 mRNA及蛋白（20 J/cm2：1.296 ± 0.028，0.0744 ± 0.0032）、MMP9 mRNA及蛋白（10 J/cm2：1.298 ± 0.035，0.0992 ± 0.0053；20 J/cm2：1.286 ± 0.032，0.1010 ± 0.0063）表达增加（P < 0.05）。结论 紫外线促进AK进展可能与其下调p53表达，上调MMP2和MMP9有关。

Effect of ultraviolet radiation on the exp ression of p53, matrix metalloproteinase-2 and-9 in actinic keratosis lesions and normal human skin

XU Dan*, LIU Tong-yun, YUAN Rui-hong, TU Ying, GU Hua, HE Li. *Department of Dermatology, First Affiliated Hospital of Kunming Medical University, Kunming 650032, ChinaCorresponding author: HE Li, Email: heli2662@yahoo.com.cn 【Abstract】 Objective To estimate the effect of different doses of ultraviolet （UV） radiation on the proliferation of and apoptosis in kertatinocytes, as well as on the expression of p53, matrix metalloproteinase-2 （MMP2） and -9 （MMP9） in actinic keratosis（AK） lesions and normal human skin. Methods Tissue specimens were obtained from the lesions of 20 patients with AK and sun-exposed normal skin of 20 healthy human subjects, and subjected to an air-exposed culture. Each of the specimens was divided into 4 areas to remain untreated （control area） or be irradiated with UV of 5, 10 and 20 J/cm2 （irradiated areas） for 4 consecutive days. After another 24-hour culture, the tissue cultures were collected followed by the evaluation of apoptosis in and proliferation of keratinocytes by using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling （TUNEL） and Ki-67 staining, and determination of mRNA and protein expressions of p53, MMP2 and MMP9 by using real time PCR and immunohistochemistry respectively. Results A statistical increase was observed in the percentage of apoptotic cells in the normal skin irradiated with UV of 10 and 20 J/cm2 （46.8% ± 2.1% and 56.7% ± 2.4%, both P < 0.05） and in the AK lesions irradiated with UV of 20 J/cm2（43.5% ± 1.5%, P < 0.05） compared with the corresponding unirradiated tissues. The normal skin showed a higher percentage of apoptotic cells than the lesional skin after irradiation with UV of 10 and 20 J/cm2 （both P < 0.05）. The percentage of Ki-67-positive cells was significantly decreased in the normal skin after irradiation with UV of 20 J/cm2 （3.34% ± 0.76%, P < 0.05）, but experienced no statistical changes in the lesional skin after different doses of UV irradiation （all P > 0.05）. There was a statistical elevation in the expression of p53 mRNA （5 J/cm2: 1.106 ± 0.025, 10 J/cm2: 1.259 ± 0.045, 20 J/cm2: 1.425 ± 0.053, all P < 0.05） and protein（10 J/cm2: 0.1169 ± 0.0032, 20 J/cm2: 0.1454 ± 0.0047, both P < 0.05） in the normal skin, but a statistical reduction in the expression of p53 mRNA（10 J/cm2: 0.611 ± 0.050, 20 J/cm2: 0.578 ± 0.070, both P < 0.05） and protein（20 J/cm2: 0.0404 ± 0.0027, P< 0.05） in the lesional skin after irradiation compared with the corresponding unirradiated skin tissues. Further more, a statistical increment was observed in MMP2 mRNA and protein expression in normal skin irradiated with UV of 10 J/cm2 （1.086 ± 0.013, 0.0843 ± 0.0024, respectively, both P < 0.05） and 20 J/cm2 （1.417 ± 0.036, 0.1236 ± 0.0042, respectively, both P < 0.05） and in lesional skin irradiated with UV of 20 J/cm2 （1.296 ± 0.028, 0.0744 ± 0.0032, respectively, both P < 0.05）, as well as in MMP9 mRNA and protein expression in normal skin irradiated with UV of 20 J/cm2 （1.395 ± 0.026, 0.3065 ± 0.0162, respectively, both P < 0.05） and in lesional skin irradiated with UV of 10 J/cm2 （1.298 ± 0.035, 0.0992 ± 0.0053, respectively, both P < 0.05） and 20 J/cm2 （1.286 ± 0.032, 0.1010 ± 0.0063, respectively, both P < 0.05） compared with the corresponding unirradiated tissues. Conclusion Ultraviolet may accelerate the progression of AK by down-regulating p53 expression but up-regulating MMP2 and MMP9 expression. 【Key words】 Actinic keratosis; Ultraviolet rays; Genes，p53; Matrix metalloproteinase 2; Matrix metalloproteinase 9