摘要
目的探讨LPS刺激后小鼠腹腔巨噬细胞表面TLR2、TLR4表达的动态变化。方法常规方法从BALB/C小鼠腹腔中分离出巨噬细胞,调整细胞数为2×106/孔,分为6组,每组设3个复孔,加入LPS使其终浓度为1μg/ml,在终浓度为1μg/mlLPS刺激下,5%CO2,37℃孵育0、1.5、3、6、16、24h。用异硫氢酸荧光素(FITC)标记的大鼠抗小鼠TLR2抗体和藻红蛋白(PE)标记的大鼠抗小鼠TLR4抗体对小鼠腹腔巨噬细胞进行免疫荧光染色,流式细胞仪(FCM)测定FITC、PE阳性细胞数及其平均荧光强度(MFI)。结果1.随着LPS刺激时间的延长,TLR2阳性率逐步上升,6h达到高峰后出现缓慢下降趋势,各刺激组阳性率与对照组组相比,P<0.01。TLR2MFI则随着LPS刺激时间的延长而缓慢下降,各刺激组与对照组相比,P<0.05。2.随着LPS刺激时间的延长,TLR4阳性率逐步上升,刺激达24h时,阳性率为14.02%±1.23%,与对照组(8.98%±1.06%)相比,P<0.01。TLR4MFI也随着刺激时间的延长而上升,各刺激组与对照组相比,P<0.05。结论LPS能够引起巨噬细胞TLR2/4表达和分布发生变化,说明TLR2与TLR4均参与针对LPS的反应。
Objective To investigate the effects of increasing stimulating time on expression of TLR2 and TLR4 in peritoneal macrophages of mice ( M induced by LPS in vitro. Methods M were obtained from BALB/C mice in routine. All the cells were divided to six groups, there were 3 repetitions in each group, in which M were incubated with 1靏/ml of LPS(final concentrations) at 37℃ in 5%CO2/95% air for 0, 1.5, 3, 6, 16, 24h respectively and expression of TLR2 and TLR4 were measured by flow cytometry, monoclonal antibodies against TLR2 and TLR4, which were marked by FITC and PE respectively,were used. Results 1.With the stimulating time extending the TLR2 positive percentage went up and the fastigium was at 6h and then it went down slowly. The positive percentage of stimulated groups significantly higher than that of unstimulated group (P<0.05). And the MFIs of TLR2 went down with the process of time significantly. 2.With the stimulating time extending the TLR4 positive percentage went up gradually. The positive percentage 24h group was 14.02%±1.23% significantly higher than that of 0h group(8.98%±1.06%, P<0.01). The MFI of TLR4 also increased with time extending. The MFIs of stimulated groups were all significantly higher than that of 0h group. Conclusions LPS can induce different expression and distribution of TLR2 and TLR4 on M surface, which indicated that both TLR2 and TLR4 take part in responding actions against LPS.
出处
《江西医学检验》
2004年第6期483-486,共4页
Jiangxi Journal of Medical Laboratory Sciences
基金
国家自然科学基金资助项目(30260107)
江西省自然科学基金资助项目(024008)
