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纸质出版:2014
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郭露,张莉莉,曹小洁,刘赟,廖少琼,黎炳护,皮燕,尹延伟,李敬诚.TLR4通过下调ABCA1表达促进平滑肌源性泡沫细胞形成[J].陆军军医大学学报(原第三军医大学学报),2014,36(07):625-630.
Guo Lu,Zhang Lili,Cao Xiaojie,Liu Yun,Liao Shaoqiong,Li Binghu,Pi Yan,Yin Yanwei,Li Jingcheng.TLR4 promotes VSMCs-derived foam cells formation via down-regulating ABCA1 expression[J].J Amry Med Univ (J Third Mil Med Univ),2014,36(07):625-630.
郭露,张莉莉,曹小洁,刘赟,廖少琼,黎炳护,皮燕,尹延伟,李敬诚.TLR4通过下调ABCA1表达促进平滑肌源性泡沫细胞形成[J].陆军军医大学学报(原第三军医大学学报),2014,36(07):625-630. DOI:
Guo Lu,Zhang Lili,Cao Xiaojie,Liu Yun,Liao Shaoqiong,Li Binghu,Pi Yan,Yin Yanwei,Li Jingcheng.TLR4 promotes VSMCs-derived foam cells formation via down-regulating ABCA1 expression[J].J Amry Med Univ (J Third Mil Med Univ),2014,36(07):625-630. DOI:
目的 探讨Toll样受体4(Toll-like receptor 4,TLR4)促进平滑肌源性泡沫细胞的形成是否与三磷酸腺苷-结合盒转运子A1(ATP-binding cassette transporter,ABCA1)表达异常相关。 方法 组织贴块法培养原代血管平滑肌细胞(vascular smooth muscle cells,VSMCs);细胞分为野生型(Wild type)空白对照组、Wild type氧化低密度脂蛋白(oxLDL)组、TLR4敲除(TLR4
-/-
)空白对照组和TLR4
-/-
oxLDL组;油红O染色观察VSMCs内脂质聚集,并用异丙醇萃取油红O测定光密度值;RT-qPCR、Western blot检测VSMCs中TLR4、ABCA1、IL-1β及TNF-α mRNA和蛋白表达水平;细胞免疫荧光染色鉴定原代VSMCs及观察核转录因子-kappa B(NF-κB)核转位。 结果 ①Wild type oxLDL组中见细胞内大量脂滴形成,油红O光密度值为Wild type空白对照组的2.3倍(P0.01);TLR4
-/-
oxLDL组中只见少许脂质沉积于细胞内,油红O光密度值与TLR4
-/-
空白对照组相比无明显变化(P0.05)。②Wild type oxLDL组ABCA1 mRNA及蛋白表达水平较Wild type空白对照组分别下降61%(P0.05),44%(P0.01);TLR4
-/-
oxLDL组ABCA1 mRNA及蛋白表达水平分别为TLR4
-/-
空白对照组的2.3倍(P0.05),1.7倍(P0.05)。③Wild type oxLDL 组VSMCs中NF-κB核转位明显增多,IL-1β 和TNF-α mRNA表达较Wild type空白对照组分别升高3.9倍(P0.05),1.7倍(P0.05);TLR4
-/-
oxLDL组VSMCs中无明显NF-κB核转位,仅IL-1β mRNA表达较TLR4
-/-
空白对照组升高1.2倍(P0.05)。 结论 TLR4及其调节的炎性反应参与平滑肌源性泡沫细胞形成过程中ABCA1表达异常。
Objective To determine whether Toll-like receptor 4 (TLR4) promoting VSMCs-deriv
ed foam cells formation is related with the abnormal expression of ATP-binding cassette transporter 1 (ABCA1). Methods Vascular smooth muscle cells (VSMCs) were isolated from male C57BL/6J (TLR4
-/-
) and C57BL/10ScNj (TLR4
-/-
) mice respectively
and primary cultured and identified. The obtained VSMCs were divided into wild type blank control cells
oxLDL-treated wild type cells (50 μg/mL oxLDL)
TLR4
-/-
blank control cells and oxLDL-treated TLR4
-/-
cells. Neutral lipid inclusions in the VSMCs induced by oxLDL were assessed by Oil Red O staining. Lipid accumulation was quantitated by isopropanol extraction of Oil Red O from stained cells and optical density determinations at 518 nm. The mRNA and/or protein levels of TLR4
ABCA1
IL-1β and TNF-α were determined by Western blotting and RT-qPCR. The NF-κB translocation was identified by immunofluorescence assay. Results Oil red O staining indicated that a large amount of lipid was accumulated in oxLDL-treated wild type VSMCs
with the OD value increased by 2.3 folds compared with that of blank control (
P
0.01). Only few lipids were found in oxLDL-treated TLR4
-/-
VSMCs
with no significant difference with the untreated cells (
P
0.05). The ABCA1 expression at mRNA and protein levels was reduced by 61% (P0.05) and 44% (
P
0.01)
respectively
in oxLDL-treated wild type VSMCs than in untreated cells. However
in the TLR4
-/-
VSMCs stimulated by oxLDL
the ABCA1 mRNA and protein expression were 2.3-fold (
P
0.05) and 1.7-fold higher (
P
0.05) than the TLR4
-/-
blank control
respectively. The translocation of NF-κB was activated by TLR4 in oxLDL-treated wild type VSMCs
and the IL-1β and TNF-α mRNA expression were 3.9 folds (
P
0.05) and 1.7 folds (
P
0.05) respectively of wild type blank controls. However
in oxLDL-treated TLR4 knockout VSMCs
NF-κB translocation and the expression of TNF-α had no significant change
but the expression
of IL-1β was enhanced by 1.2 times (
P
0.05) when compared with untreated TLR4
-/-
VSMCs. Conclusion TLR4 and its mediating inflammation are involved in the abnormal expression of ABCA1 during VSMCs-derived foam cells formation.
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