中华皮肤科杂志 ›› 2018, Vol. 51 ›› Issue (6): 429-433.doi: 10.3760/cma.j.issn.0412-4030.2018.06.007

• 论著 • 上一篇    下一篇

脂溢性角化病皮损组织蛋白酶L2表达及活性对黑素小体降解的影响

苏梦云1,雷铁池1,易文娟1,苗芳1,江珊2,徐世正3   

  1. 1. 武汉大学人民医院
    2.
    3. 武汉大学人民医院皮肤科
  • 收稿日期:2017-06-05 修回日期:2017-11-20 发布日期:2018-05-30
  • 通讯作者: 雷铁池 E-mail:tiechilei@126.com
  • 基金资助:
    国家自然科学基金

Expression and activity of cathepsin L2 and its effect on the degradation of melanosomes in skin lesions of seborrheic keratosis

  • Received:2017-06-05 Revised:2017-11-20 Published:2018-05-30
  • Contact: Tie-Chi LEI E-mail:tiechilei@126.com
  • Supported by:
    National Natural Science Foundation of China

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摘要： 目的 检测脂溢性角化病（SK）皮损中组织蛋白酶L2（CTSL2）表达及活性，观察SK皮损中黑素小体超微结构的变化，研究CTSL2对黑素小体降解的影响。方法 2016年1 - 8月招募武汉大学人民医院皮肤科门诊SK患者20例，取患者皮损和周围正常皮肤组织，其中15例用HE染色、Fontana?Masson嗜银染色观察黑素颗粒分布，透射电镜（TEM）观察黑素小体超微结构变化，Ki67免疫组化染色检测细胞增殖状态。5例用RT?PCR和荧光底物裂解法分别检测CTSL2 mRNA表达水平及酶活性；用蔗糖梯度离心法从废弃人眼球视网膜色素上皮组织分离纯化黑素小体，并与SK皮损表皮裂解物共孵育，TEM观察孵育后黑素小体膜结构的变化。两组间均数比较采用配对t检验，P < 0.05 为差异有统计学意义。结果 SK皮损中可见大量黑素颗粒沉积，而正常皮肤仅在基底层见线形黑素颗粒沉积。TEM观察显示，SK皮损中黑素小体破损率为24.33% ± 3.06%，正常皮肤为49.00% ± 4.00%，两组比较，t = 8.49，P < 0.05。RT?PCR显示，5例SK皮损中CTSL2 mRNA相对表达水平为0.35 ± 0.09，正常皮肤组织为0.43 ± 0.08，两组比较，t = 3.17，P < 0.05。SK皮损中CTSL2酶活性（17.46 ± 0.45）也低于正常皮肤组织（28.78 ± 0.58），t = 34.29，P < 0.05。TEM还观察到，SK皮损裂解物处理的黑素小体破损率为32.33% ± 4.93%，低于正常皮肤43.00% ± 2.65%，两组比较，t = 3.30，P < 0.05。结论 SK皮损中CTSL2表达水平减低影响角质形成细胞对黑素小体的降解，是否直接参与SK皮损的病理发生有待进一步证实。

关键词: 角化病, 脂溢性, 黑素小体, 组织蛋白酶L

Abstract: Su Mengyun, Lei Tiechi, Yi Wenjuan, Miao Fang, Jiang Shan, Xu Shizheng Department of Dermatology, Renmin Hospital of Wuhan University, Wuhan 430060, China Corresponding author: Lei Tiechi, Email: tchlei@whu.edu.cn 【Abstarct】 Objective To determine the of cathepsin L2（CTSL2）and evaluate its activity in skin lesions of seborrheic keratosis（SK）, to observe the ultrastructural changes of melanosomes in the skin lesions of SK, and to estimate the effect of CTSL2 on the degradation of melanosomes. Methods Twenty patients with SK were enrolled from the Department of Dermatology, Renmin Hospital of Wuhan University. The lesional tissue and the perilesional normal skin were biopsied from each patient. Among 15 of the 20 patients, hematoxylin and eosin（HE）staining and Fontana-Masson silver staining were performed to observe the distribution of melanin granules, transmission electron microscopy（TEM）was conducted to observe the ultrastructural changes of melanosomes, and immunohistochemical staining was performed to estimate the cellular proliferative activity. RT-PCR and fluorogenic substrate cleavage assay were performed in the other 5 patients to determine the mRNA of CTSL2 and evaluate its activity, respectively. Sucrose density gradient ultracentrifugation was performed to isolate and purify melanosomes from the retinal pigment epithelium （RPE） harvested from a discarded eyeball of a 35-year old male patient with informed consent. The purified melanosomes were incubated with epidermal lysates of SK lesions, and TEM was used to observe the changes in the membrane structure of melanosomes. Statistical analysis was carried out by paired t test, and a P value < 0.05 was considered statistically significant. Results A large number of melanin granules were deposited in SK lesions, while the linear deposition of melanin granules was only seen in the basal layer of the normal skin. TEM showed that the percentage of damaged melanosomes was much higher in the normal skin （49.00% ± 4.00%） than in the SK lesions （24.33% ± 3.06%）（t = 8.49, P < 0.05）. RT-PCR revealed that the mRNA and activity of CTSL2 were both significantly lower in the SK lesions than in the normal skin （mRNA: 0.35 ± 0.09 vs. 0.43 ± 0.08, t = 3.17, P < 0.05; activity: 17.46 ± 0.45 vs. 28.78 ± 0.58, t = 34.29, P < 0.05）. Moreover, TEM also showed that the percentage of damaged melanosome was lower in the SK lesion lysate-treated group （32.33% ± 4.93%） than in the normal skin lysate-treated group （43.00% ± 2.65%, t = 3.30, P < 0.05）. Conclusion Decreased of CTSL2 in the SK lesions can affect the degradation of melanosomes by keratinocytes. However, whether CTSL2 directly takes part in the pathogenesis of SK or not is still needed to be further confirmed.

Key words: Keratosis, seborrheic, Melanosomes, Cathepsin L

引用本文

苏梦云 雷铁池 易文娟 苗芳 江珊 徐世正. 脂溢性角化病皮损组织蛋白酶L2表达及活性对黑素小体降解的影响[J]. 中华皮肤科杂志, 2018,51(6):429-433. doi:10.3760/cma.j.issn.0412-4030.2018.06.007