Fu Ying-jun, Xie Yong, Shi Jun-qiang, Zhou Nan-jin, Guo Guang-hua
Department of Pharmacology, Nanchang University Medical College, Nanchang 330006, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2007 Jun;23(6):523-6.
To investigate the expression of Toll like receptor 4(TLR4) on splenocytes and TNF-alpha mRNA, the dynamic changes of CD4(+) CD25(+) regulatory T cell(Treg) and endotoxin(ET) in peripheral blood of burnt rats during the early phase, and to explore the mechanism against inflammatory reaction in intestine-derived infection after burn.
64 SD male rats were randomly separated into control group and burn model groups. Rats of burn model group were burnt with vapor under 3 mPa pressure and 108 degrees Celsius temperature for 8 seconds to achieve deep partial-thickness burn, and a 30% total body surface area (TBSA) burn model was made. Rats were sacrificed before and 2, 5, 8, 12, 24, 48 and 72 h after burn, and the TLR4 and TNF-alpha mRNA expression in splenocytes were measured at various intervals by RT-PCR. The expression of TLR4 protein was measured by Western blot, and the percentage of Treg cells in CD4(+) T cells was detected by flow cytometry (FCM), and the LPS concentration of plasma was detected by limulus lysate test.
The expression of TLR4 mRNA, TNF-alpha mRNA, TLR4 protein and the levels of Treg, ET were significantly increased at some times points after burn. The expression of TLR4 mRNA and protein reached the peak at 8 h, whereas the TNF-alpha mRNA at 12 h, and Treg and ET at 8 h. The peak values of them were 3.66+/-0.51, 2.27+/-0.19, 1.65+/-0.23, 63.19+/-12.65% and 11.68+/-1.71 Eu/mL respectively, which were of significant difference when compared with the control group (P<0.01). The expression of TLR4 mRNA showed a positive correlation with that of Treg, ET and TNF-alpha mRNA (r=0.898, 0.811, 0.462, P<0.01).
Treg might play a major role in the process of immune regulation in rats after burn, the mechanism of which may be correlated with the increase of LPS-TLR4 signal transduced by intestine-derived infection.
探讨烧伤大鼠早期脾细胞Toll样受体4（TLR4）及肿瘤坏死因子-α（TNF-α）mRNA的表达，外周血CD4(+)CD25(+)调节性T细胞（Treg）及内毒素（ET）的动态变化，以探讨烧伤后肠源性感染抗炎反应机制。
64只雄性SD大鼠随机分为对照组和烧伤模型组。烧伤模型组大鼠在3 mPa压力、108℃温度下用蒸汽烫伤8秒，制成30%总体表面积（TBSA）深Ⅱ度烧伤模型。于烧伤前及烧伤后2、5、8、12、24、48、72小时处死大鼠，不同时间点采用逆转录聚合酶链反应（RT-PCR）检测脾细胞TLR4及TNF-α mRNA表达。采用蛋白质印迹法检测TLR4蛋白表达，流式细胞术（FCM）检测CD4(+)T细胞中Treg细胞百分比，鲎试剂法检测血浆LPS浓度。
烧伤后部分时间点TLR4 mRNA、TNF-α mRNA、TLR4蛋白表达及Treg、ET水平显著升高。TLR4 mRNA和蛋白表达于8小时达峰值，TNF-α mRNA于12小时达峰值，Treg和ET于8小时达峰值。其峰值分别为3.66±0.51、2.27±0.19、1.65±0.23、63.19±12.65%和11.68±1.71 Eu/mL，与对照组比较差异有统计学意义（P<0.01）。TLR4 mRNA表达与Treg、ET及TNF-α mRNA表达呈正相关（r=0.898、0.811、0.462，P<0.01）。
Treg可能在烧伤大鼠免疫调节过程中起主要作用，其机制可能与肠源性感染介导的LPS-TLR4信号转导增加有关。
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