The effects of lymph after intestinal ischemia/reperfusion on the inflammatory factors and Toll-like receptor 4 ligand high mobility group box-1 in Toll-like receptor 4 deficient mice

Abstract

Objective To investigate the effects of lymph from ischemic/reperfused intestine on the inflammatory factors and Toll-like receptor 4 (TLR4) ligand high mobility group box-1 (HMGB1) in TLR4 deficient (TLR4-/-) mice. Methods A total of 20 SD rats weighing (300±20)g were randomly assigned into two groups: lymph drainage group (group N, lymph drainage for 180 minutes without other treatment) and intestinal ischemia/reperfusion group (group I/R, draining the lymph for 180 minutes while clipping the superior-mesenteric artery for 60 minutes followed by 120-minute reperfusion). Thirty-two TLR4-/- mice and thirty-two C57BL/6 wild type (WT) mice were each divided into 4 sub-groups (n=8), injected with different fluids through the caudal vein: group N with normal lymph; group I/R with I/R lymph; group Edt with endotoxin; group HMGB1 with HMGB1 protein. The mice were sacrificed 180 minutes after the injection for sample collection. Results The levels of endotoxin and HMGB1 in the lymph drainage of the group I/R rats were significantly higher than that of the group N rats [(0.034±0.050)Eu/ml vs. (0.017±0.023)Eu/ml, P=0.033; (4.293±0.883)ng/ml vs. (0.509±0.128)ng/ml, P=0.006]. In the mice injected with HMGB1, the mucosa thickness and villus height in the ileum of the WT mice were significantly lower than that of the TLR4-/-mice [(335.8±43.2)μm vs. (602.1±37.5)μm, P=0.000; (273.0±31.7)μm vs. (404.5±18.6)μm, P=0.000]; in both WT and TLR4-/- mice injected with the I/R lymph drainage, the mucosa thickness and villus height were decreased, but the decrements were significantly lower in TLR4-/- mice; there were no statistically significant differences in the levels of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), endotoxin, and HMGB1 between the TLR4-/- and the WT mice injected with normal lymph or endotoxin. In the mice injected with I/R lymph drainage, the levels of inflammatory factors in the TLR4-/- mice were significantly lower than those in the WT mice [TNF-α: (28.637±5.166)pg/ml vs. (41.917±8.175)pg/ml, P=0.000; IL-6: (60.900±24.729)pg/ml vs. (110.265±28.545)pg/ml, P=0.000]. In the mice injected with HMGB1, the levels of inflammatory factors in the TLR4-/- mice were significantly decreased compared with those in the WT mice [TNF-α: (20.865±6.464)pg/ml vs. (31.059±6.204)pg/ml, P=0.004; IL-6: (36.268±8.977)pg/ml vs. (76.677±14.099)pg/ml, P=0.000]. Conclusions The concentrations of endotoxin and HMGB1 are significantly increased during intestinal I/R in rats. After injection of I/R lymph drainage, endotoxin, and HMGB1, the levels of inflammatory factors and HMGB1 in the mice injected with I/R lymph drainage are significantly higher than those in the mice injected with normal lymph; the levels of inflammatory factors and local damage of intestinal mucosa are significantly reduced in the TLR4-/- mice than in the WT mice. Thegut-lymphpathway may play a key role in the intestinal I/R injury. Key words: Intestinal ischemia/reperfusion; Toll-like receptor 4 deficient mice; Inflammatory factors; High mobility group box-1; Endotoxin