Effect of (E)-2-(4-bromophenyl)-1-(3,4-dihydroxyphenyl) Ethanone Oxime on Proliferation and Activation of Mice Lymphocytes

Abstract

To study the effects of (E)-2-(4-bromophenyl)-1-(3,4-dihydroxyphenyl) ethanone oxime (BDEO) on the proliferation and activation of the mice's splenic lymphocytes and the peripheral blood lymphocytes induced by Concanavalin A (Con A) in vitro and in vivo, and its molecular mechanism. Methods During the lymphocyte proliferation and activation induced by Con A in vitro, MTT and cell counting were used to detect the transformation rates and survival rates of lymphocytes, and ELISA was used to measure the activity of caspase-9; moreover, the levels of Bax, Bcl-2 and caspase-3 were determined by Western blot, in order to observe the effects of BDEO on cell proliferation and activation. The effects of administration of Con A 〔15 mg/(kg·d)〕 and BDEO 〔(3, 6 mg/(kg·d)〕 by intraperitoneal injection on transformation rates of spleen cells and peripheral blood lymphocyte, as well as phagocytosis rate of peritoneal macrophages in mice were also observed in vivo. Results 0.3-1 μmol/L BDEO significantly inhibited the transformation rates and growth of mice lymphocyte (P<0.05). The activity of caspase-9 and the levels of mitochondrial pro-apoptotic protein Bax and Bak gradually increased, then decreased as the BDEO continually accumulated. Anti-apoptotic protein Bcl-2 as well as mitochondrial Cyt C levels first decreased then increased gradually, and cytoplasmic Cyt C, cleaved caspase-9 and cleaved caspase-3 levels showed firstly a increase, then decrease gradually. Additionally, administration of BDEO by intraperitoneal injection significantly inhibited proliferation of spleen lymphocytes and peripheral blood lymphocyte, as well as phagocytosis of peritoneal macrophagesin in mice. Conclusion BDEO might regulate the proliferation and activation of lymphocytes through activation of caspase-3 mainly via a mitochondrial intrinsic pathway; the inhibiting [CM(155mm]effect on the proliferation and transformation rate of lymphocytes was significant when the concentration of BDEO was relatively low; as the concentration accumulated increasingly, the inhibiting effect reduced. The results indicated that BDEO has immunosuppressive activity.

Keywords: Lymphocytes, BDEO, Proliferation, Activation, Cyt C, Caspase-9