[Effect of koumine on proliferation of murine CD4+ T cells purified by magnetic-activated cell sorting in vitro].
OBJECTIVE
To assess the separation efficiency of magnetic-activated cell sorting in the purification of CD4+ T cells from murine spleen, and observe the effects of koumine on the proliferation of the separated cells.


METHODS
CD4+ T cells were isolated from murine spleen by magnetic-activated cell sorting (MiniMACS). Fluorescence-activated cell sortering was employed to determine the purity of CD4+ T cells before and after the separation procedure followed by evaluation of the cell viability using trypan blue staining. Concanavalin A- (ConA, 5 microg/ml) or phytahematoagglutinin (PHA,1 mg/ml)-induced murine T cells were treated with different concentrations of koumine (10-320 microg/ml), and their proliferation was determined by MTT colorimetry, and enzyme-linked immunosorbent assay was used to measure IL-2 level in the cell culture supernatant.


RESULTS
The purity of CD4+ T cells reached (90.3+/-5.8)% after the purification with a cell viability of (94.9+/-3.6)%. Koumine (20-320 microg/ml) dose-dependently inhibited ConA- or PHA-induced proliferation of murine lymphocytes as compared with the controls (P<0.05). Koumine (20, 100, and 200 microg/ml) significantly decreased the level of IL-2 in comparison with the control group (P<0.05).


CONCLUSIONS
CD4+ T cells of high purity can be obtained from murine spleen using MiniMACS without impairing the viability of the cells. Koumine significantly inhibits the proliferation of murine CD4+ T cells due to its immunosuppressive effect and inhibition of IL-2 secretion.
Wang
Zhi-rui
ZR
Department of Pharmacy, Zhujiang Hospital, Southern Medical University, Guangzhou 510282, China. wangzr@fimmu.com
Huang
Chang-quan
CQ
Zhang
Zhong-yi
ZY
Zhang
Lan-lan
LL
Lin
Jing-ming
JM
D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology*; D049109:Cell Proliferation / Q000187:drug effects*; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D005260:Female; D031450:Gelsemium / Q000737:chemistry*; D026121:Indole Alkaloids / Q000494:pharmacology*; D008280:Magnetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C
D026121:Indole Alkaloids; C515085:koumine