Chinese Medical Sciences Journal ›› 2012, Vol. 27 ›› Issue (2): 73-79.

• Original Article • 上一篇    下一篇

Role of ADAM10 and ADAM17 in CD16b Shedding Mediated by Different Stimulators△

Sha Guo, Min Peng, Qing Zhao, and Wei Zhang*   

  1. Department of Immunology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China
  • 收稿日期:2012-07-05 修回日期:2012-07-05 出版日期:2012-07-05 发布日期:2012-07-05
  • 作者简介:*Corresponding author Tel: 86-10-69156417, Fax: 86-10-82282687, E-mail: wzhang@pumc.edu.cn

Role of ADAM10 and ADAM17 in CD16b Shedding Mediated by Different Stimulators△

Sha Guo, Min Peng, Qing Zhao, and Wei Zhang*   

  1. Department of Immunology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China
  • Received:2012-07-05 Revised:2012-07-05 Published:2012-07-05 Online:2012-07-05

摘要: Objective To investigate the main proteinases responsible for CD16b shedding under different stimulators.
Methods HEK293 cell line stably expressing CD16b was constructed by lentivirus system. The cell line was then overexpressed with a disintegrin and metalloproteinase 10 (ADAM10) or ADAM17, suppressed with short hairpin RNA of ADAM10 or ADAM17, and reconstituted with ADAM10 or ADAM17, respectively. After each treatment, the cell line was stimulated with ionomycin or phorbol 12-myristate- 13-acetate (PMA) for 12 hours. The soluble CD16b released from cell membrane was detected by immunoprecipition and immunoblot. Quantitation was then implemented to compare the amount of soluble CD16b in cell supernatant after stimulation.
Results HEK293 cell line stably expressing CD16b was successfully established. When CD16b expressing cell line was overexpressed with ADAM10, shedding of CD16b was increased after stimulation with ionomycin but not PMA; when the cell line overexpressed with ADAM17, shedding of CD16b was increased after stimulation with PMA but not ionomycin. Similarly, when ADAM10 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with ionomycin; when ADAM17 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with PMA. The shedding of CD16b was increased again when CD16b expressing cell line was reconstituted with ADAM10 and stimulated by ionomycin or reconstituted with ADAM17 and stimulated by PMA.
Conclusions Both ADAM10 and ADAM17 could shed CD16b, but they possess differed preferences. ADAM10 is the main sheddase under stimulation of ionomycin, while ADAM17 is the main sheddase under stimulation of PMA.

关键词: a disintegrin and metalloproteinase 10, a disintegrin and metalloproteinase 17, CD16b, shedding

Abstract: Objective To investigate the main proteinases responsible for CD16b shedding under different stimulators.
Methods HEK293 cell line stably expressing CD16b was constructed by lentivirus system. The cell line was then overexpressed with a disintegrin and metalloproteinase 10 (ADAM10) or ADAM17, suppressed with short hairpin RNA of ADAM10 or ADAM17, and reconstituted with ADAM10 or ADAM17, respectively. After each treatment, the cell line was stimulated with ionomycin or phorbol 12-myristate- 13-acetate (PMA) for 12 hours. The soluble CD16b released from cell membrane was detected by immunoprecipition and immunoblot. Quantitation was then implemented to compare the amount of soluble CD16b in cell supernatant after stimulation.
Results HEK293 cell line stably expressing CD16b was successfully established. When CD16b expressing cell line was overexpressed with ADAM10, shedding of CD16b was increased after stimulation with ionomycin but not PMA; when the cell line overexpressed with ADAM17, shedding of CD16b was increased after stimulation with PMA but not ionomycin. Similarly, when ADAM10 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with ionomycin; when ADAM17 was suppressed by short hairpin RNA, CD16b shedding was decreased after stimulation with PMA. The shedding of CD16b was increased again when CD16b expressing cell line was reconstituted with ADAM10 and stimulated by ionomycin or reconstituted with ADAM17 and stimulated by PMA.
Conclusions Both ADAM10 and ADAM17 could shed CD16b, but they possess differed preferences. ADAM10 is the main sheddase under stimulation of ionomycin, while ADAM17 is the main sheddase under stimulation of PMA.

Key words: a disintegrin and metalloproteinase 10, a disintegrin and metalloproteinase 17, CD16b, shedding

基金资助: △Supported by the National Natural Science Foundation of China (30872287).

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