Impact of GdCl3 on Toll-Like Receptors Expression Induced by Lipopolysaccharide in Murine Macrophage Cell Line RAW264.7_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Authors：

XU Jianbo ¹ , WU Heshui ¹ , ZHANG Jinxiang ² , WANG Lin ³ , JIN Qinwen ¹ , TIAN Yuan ¹ , ZHANG Jinghui ¹ , ZHANG Lei. ¹

Department of General Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China;

Corresponding author：

Keywords：

Gadolinium chloride; Toll-like receptor; Macrophage; Endotoxin

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Objective　To observe the expression of GdCl3 on Toll-like receptors （TLRs） of RAW264.7 from murine macrophage cell line induced by lipopolysaccharide (LPS) stimulation. Methods　Cells were divided into 3 groups: blank group, LPS group and GdCl3 group. And these cells dyed by goat anti-mouse TLR2/4 poly-antibody and anti-goat IgG labelled with fluorescein isothiocyanate (FITC). The synthesis of TLR2/4 protein were determined by flow cytometry (FCM) analysis and reverse transcription polymerase chain reaction (RT-PCR) analyzed their gene expression. Cell supernatants were taken to measure TNF-α production following the ELISA (enzyme-linked immunosorbent assay) protocol. Results　The expressions of TLR2/4 protein and mRNA in GdCl3 group under action of different concentration of GdCl3〔TLR2/4 protein, 200 μmol/L: (70.2±1.28）％/(66.7±2.59)％, 400 μmol/L: (64.9±1.43)％/(60.4±1.25)％, 2 000 μmol/L: (47.4±0.98)％/(32.1±0.74)％; TLR2/4 mRNA (the value of absorbance), 200 μmol/L: (76.42±2.76)/(101.72±3.14), 400 μmol/L: (75.60±3.76)/(89.65±5.17), 2 000 μmol/L: (64.22±4.67)/(78.44±4.88)〕 were significantly lower than those of in LPS group 〔TLR2/4 protein: (94.4±1.76)％/(95.7±0.87)％, P＜0.01; TLR2/4 mRNA: (127.64±3.25)/(119.82±5.59), P＜0.05, P＜0.01〕. The expression of TNF-α in GdCl3 group under action of different concentration of GdCl3〔200 μmol/L: (2 540±77) pg/ml, 400 μmol/L: (2 041±106) pg/ml, 2 000 μmol/L: (1 020±220) pg/ml〕 was also significantly lower that that of in LPS group 〔(4 688±127) pg/ml, P＜0.01)〕. Conclusion　 GdCl3 significantly inhibits TLR expression and secretion of TNF-α under the condition of LPS stimulation in vivo.

Citation： XU Jianbo,WU Heshui,ZHANG Jinxiang,WANG Lin,JIN Qinwen,TIAN Yuan,ZHANG Jinghui,ZHANG Lei.. Impact of GdCl3 on Toll-Like Receptors Expression Induced by Lipopolysaccharide in Murine Macrophage Cell Line RAW264.7. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2006, 13(6): 681-684. doi: Copy

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1. 　Akira S, Sato S. Toll-like receptors and their signaling mechanisms [Ｊ]. Scand J Infect Dis， 2003； 35(9)∶ 555.
2. 　Wu HS, Zhang JX, Wang L, et al. Toll-like receptor 4 inv-olvement in hepatic ischemia/reperfusion injury in mice [Ｊ]. Hepatobiliary Pancreat Dis Int， 2004; 3(2)∶250.
3. 　Medzhitov R. Toll-like receptors and innate immunity [Ｊ]. Nat Rev Immunol， 2001; 1(2)∶135.
4. 　Schauer RJ, Bilzer M, Kalmuk S, et al. Microcirculatory failure after rat liver transplantation is related to Kupffer cell-derived oxidant stress but not involved in early graft dysfunction [Ｊ]. Transplantation， 2001; 72(10)∶1692.
5. 　Shibuya H, Ohkohchi N, Seya K, et al. Kupffer cells generate superoxide anions and modulate reperfusion injury in rat livers after cold preservation [Ｊ]. Hepatology， 1997； 25(2)∶35.
6. 　Dekai Zhang, Guolong Zhang, Matthew S, et al. A toll-like receptor that prevents infection by uropathogenic bacteria[Ｊ]. Science， 2004; 303（5633）∶1522.
7. 　Aliprantis AO, Yang RB, Mark MR, et al. Cell activation and apoptosis by bacterial lipoproteins through toll-like receptor-2 [Ｊ]. Science， 1999； 285 (5428)∶736.
8. 　吴河水，王　峰，王　琳，等. 小鼠肝缺血/再灌注损伤时肝脏Kupffer细胞中TLR2的表达 [Ｊ]. 中国普通外科杂志, 2004； 13(8)∶591.
9. 　Musikacharoen T, Matsuguchi T, Kikuchi T, et al. NF-kappa B and STAT5 play important roles in the regulation of mouse Toll-like receptor 2 gene expression [Ｊ]. J Immunol, 2001； 166（7）∶4516.
10. 　Muzio M, Polntarutti N, Bosisio D, et al. Toll like receptor family (TLT) and signalling pathway [Ｊ]. Eur Cytokine Netw， 2000； 11(3)∶489.
11. 　Lee CM, Yeoh GC, Olynyk JK. Differential effects of gadolinium chloride on Kupffer cells in vivo and in vitro [Ｊ]. Int J Biochem Cell Biol， 2004; 36(3)∶481.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Impact of GdCl3 on Toll-Like Receptors Expression Induced by Lipopolysaccharide in Murine Macrophage Cell Line RAW264.7

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