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.