原苏木素A调控树突状细胞成熟和功能状态的实验研究

Experimental study on regulating effect of protosappanin A on dendritic cell maturation and functional state

目的:探讨原苏木素A(PrA)对树突状细胞(DC)成熟的调控作用。方法:选择SPF级雄性Wistar大鼠,SD大鼠作为实验对象。在脂多糖(LPS)诱导DC成熟过程中,使用甲基噻唑基四唑(MTT)比色法筛选PrA的作用浓度。使用不同浓度PrA预处理LPS诱导的DC,分析DC表面分子CD80、CD86的表达差异;检测DC激活T细胞增殖的能力的改变,以及调节性T细胞(Treg)表面分子CD4、CD25和Foxp3的表达水平,同时检测上清液中DC分泌的细胞因子白介素(IL)-10、IL-12水平。结果:与未成熟DC(imDC)组比较,LPS-DC组DC表面分子CD80[(31.50±29.04)%比(63.80±14.03)%]、CD86[(36.10±27.21)%比(62.60±12.37)%]表达明显升高,P均<0.01;与LPS-DC组比较,20-DC组(即加入20nmol/L PrA的DC组)CD80[(63.80±14.03)%比(39.70±26.60)]、CD86[(62.60±12.37)%比((37.90±26.93)]表达显著降低,P均<0.05。与LPS-DC组比较,5-DC组、20-DC组DC激活的T细胞增殖能力[(0.39±0.06)比(0.32±0.46)比(0.28±0.08)]、IL-12水平[(250.00±89.81)pg/ml比(176.80±49.89)pg/ml比(134.30±60.64)pg/ml]显著降低,Treg表达[(0.42±0.23)比(0.76±0.20)比(0.93±0.52)]、IL-10水平[(145.80±70.28)pg/ml比(274.00±131.93)pg/ml比(354.00±146.22)pg/ml]显著升高(5-DC组:P均<0.05,20-DC组:P均<0.01)。结论:原苏木素A能抑制LPS诱导的DC成熟,包括降低表面分子CD80和CD86表达、抑制激活异基因T淋巴细胞增殖的能力、诱导Treg增殖、并影响相关细胞因子的水平。

Abstract： Objective： To explore regulating effect of protosappanin A （PrA） on dendritic cell （DC） maturation. Methods： SPF male Wistar rats and SD rats were selected as subjects. During DC maturation induced by lipopolysac- charide （LPS）, methyl thiazolyl tetrazolium （MTT） method was used to screen proper concentrations of PrA. Dif- ferent concentrations of PrA were used to pretreat LPS-induced DC, difference of DC surface molecule CD80 and CD86 expressions were analyzed; DC＇s ability in activating T cell proliferation, expression levels of CD4, CD25 and Foxp3 on surface of regulatory T cells （Treg）, and levels of interleukin （IL） -10 and IL-12 secreted by DC in supernatant were measured. Results： Compared with immature DC （imDC） group, there were significant rise in expressions of DC surface molecule CD80 [ （31.50±29.04）% vs. （63.80± 14.03）%] and CD86 [ （36.10±27.21）% vs. （62.60 ± 12.37）%] in LPS-DC group, P（0.01 both; compared with LPS-DC group, there were significant reduc- tionsin expressions of CD80 [（63.80± 14.03）% vs. （39.70±6.60）] and CD86 [（62.60±12.37）% vs. （ （37.90 ±26.93）] in 20-DC group （DC cultured with 20nmol/L PrA）, P〈0.05 both. Compared with LPS-DC group, there were significant reductions in DC-activated T cell proliferation capacity [ （0.39 ± 0.06） vs. （0.32 ±0.46） vs. （0.28 ± 0.08）] and IL-12 level [ （250.00 ±89.81） pg/ml vs. （176.80 ±49.89） pg/ml vs. （134.30 ±60.64） pg/ml], and significant rise in expression of Treg [ （0.42 ± 0.23） vs. （0.76 ±0.20） vs. （0.93 ± 0.52）] and IL-10 level [（145.80 ± 70.28） pg/ml vs. （274. 00 ± 131.93） pg/ml vs. （354.00 ±146.22） pg/ml] in 5-DC group and 20-DC group （P〈0.05 all for 5-DC group, P〈0.01 all for 20-DC group）. Conclusion.. PrA can inhibit LPS-induced DC maturation, including reducing expressions of surface molecule CD80 and CD86, suppressing the capacity to activate allogeneic T lymphocyte proliferation, inducing Treg proliferation and affecting levels of relative cytokines.