Effect of artesunate on the expression of Fas/FasL and Bcl-2/Bax in synoviocytes of rats with adjuvant arthritis

Xingfu SONG; Honggang YUAN; Xianguo CHEN

ABSTRACT

BACKGROUND:

The morbility of rheumatoid arthritis is closely associated with cell apoptosis process. Hyperplasia of synoviocyte lies in the relatively insufficiency of synoviocyte apoptosis. Therefore, induction of synoviocyte apoptosis has clinical significance in the treatment of rheumatoid arthritis. Arteannuin and its derivatives can induce the apoptosis of various cells.

OBJECTIVE:

To observe the effect of artesunate on the expression of Fas/FasL and Bcl-2/Bax, which are correlated with apoptosis in synoviocyte of adjuvant arthritis.

DESIGN:

Randomized and controlled observation.

SETTING:

First College of Clinical Medical Science, Three Gorges UniversityMATERIALS The experiment was conducted in the laboratories of Immunology and Morphology, Three Gorges University from September 2005 to November 2005. Fifty 8-week-old male Wistar rats of clean grade and (150±21) g were provided by the Animal Experimental Center of Tongji Medical College, Huazhong University of Science and Technology; Complete Freud adjuvant by SIGMA, U.S., and artesunate solution by Guilin Pharmaceutical Corporation.rabbit anti-mouse Fas (SC-716), rabbit anti-mouse FasL and Technology (No. SCXK 2004-2007).Complete Freud adjuvant (SIGMA, U.S. No. 093K8932); artesunate solution (Guilin Pharmaceutical Cor); rabbit anti-mouse Fas (SC-716), rabbit anti-mouse FasL multi-antibody (SC-834), goat anti-mouse IgG-HRP multimer, rabbit anti-mouse P53 (M3566) multi-antibody, Bcl-2 (sc-7382) multi-antibody, rabbit anti-mouse Bax (sc-7480) multi-antibody, ABC compound reagent kit and DAB coloring reagent all acquired from Santa Cruz biotechnology (Santa Cruz, USA); methotrexate (MTX) purchased from Shanghai Hualian Pharmaceutical Co., Ltd.; microscope and image analysis system obtained from Leica (Leica,Germany); microtome (Leica, Germany).

METHODS:

Fifty rats were randomly divided into six groups normal group (n =8), model group (n =8), high dose artesunate group (n =8), low dose artesunate group (n =8), artesunate plus methotrexate (MTX) group (n =8), and MTX group (n =8). ①Model construction was referred to literature Except the normal group with 0.1 mL normal saline, all rats were injected with 0.1 mL complete Freund's adjuvant into the right voix pedis to establish the models of adjuvant arthritis. Since the 13th day after modeling, high and low dose artesunate groups were intraperitoneally injected with 40 and 20 mg/kg artesunate, respectively everyday; artesunate plus MTX group was intraperitoneally injected with 20 mg/kg artesunate everyday and 0.4 mg/kg MTX every three days; MTX group with 0.4 mg/kg MTX solution every three days;model group with 1 mL normal group everyday. ②The arthrosis index (Al) of each group was evaluated before and 13 days after administration; the expressions of Fas/FasL, Bcl-2, and Bax in synovial membrane tissue were examined by immunohistochemistry.MAIN OUTCOME

MEASURES:

Al and the expression of Fas/FasL, Bcl-2, and Bax in synovial membrane tissue of each group.

RESULTS:

Fifty rats were involved in the result analysis. ①Thirteen days after administration, the Al of each experiment group (including model control) was remarkably lower than that before treating (P ＜ 0.01). The Al of eachexperiment group was significantly lower than that in model control group (P ＜ 0.01). ②The expression of Fas/FasL in high and low dose artesunate groups and artesunate plus MTX group was up-regulated significantly compared with that in model group (P ＜ 0.01). There was no statistically significant difference in the expression between MTX group and model group (P ＞ 0.05); the expression of Bcl-2 was significantly down-regulated but Bax up-regulated in two artesunate groups,artesunate plus MTX group and MTX group compared with that in model group (P ＜ 0.05).

CONCLUSION:

The findings suggest that artesunate could alleviate adjuvant arthritis, up-regulate the expression of Fas/FasL and Bax, but down-regulate that of Bcl-2, in which the induction of synoviocyte apoptosis may be one of the mechanisms.