肝I/R损伤中髓系细胞与内皮细胞TLR4对PMN迁移的体外比较

Comparison of PMN migration between myeloid cells TLR4 and endothelial cell TLR4 in hepatic ischemia/reperfusion injury in vitro

目的探讨髓系细胞及肝窦内皮细胞Toll样受体4(TLR4)在小鼠肝缺血再灌注(I/R)损伤中对外周血多形核细胞(PMN)迁移的影响。方法以TLR4野生型小鼠(C3H/HeN)与TLR4变异型小鼠(C3H/HeJ)为对象,将供体小鼠骨髓细胞转入经射线照射的受体小鼠体内,制作4组嵌合体:A组为髓系细胞TLR4+/+和内皮细胞TLR4+/+;B组为髓系细胞TLR4+/+和内皮细胞TLR4-/-;C组为髓系细胞TLR4-/-和内皮细胞TLR4+/+;D组为髓系细胞TLR4-/-和内皮细胞TLR4-/-,然后建立4组嵌合体小鼠肝I/R损伤模型。将提取的肝组织炎性蛋白及外周血分离的PMN分置于Transwell小室上下2个空间内检测PMN迁移情况。以C3α为阳性对照,缓冲液为阴性对照。结果与C组(67.10±6.89)比较,A组PMN迁移数目(148.95±11.04)明显增多,差异有统计学意义(P<0.05);与D组(63.15±8.86)比较,B组PMN迁移数目(142.65±8.67)明显增多,差异有统计学意义(P<0.05)。而A组与B组PMN迁移数目差异无统计学意义(P>0.05),C组与D组PMN迁移数目差异亦无统计学意义(P>0.05)。结论在肝I/R损伤中的PMN迁移中髓系细胞TLR4而非内皮细胞TLR4起主导作用。

Objective To investigate the effect of PMN migration between myeloid cells TLR4 and LSEC TLR4 in hepatic ischemia/reperfusion（I/R） injury in mice.Methods TLR4 wild-type mice（C3H/HeN） and TLR4 mutant mice（C3H/HeJ） were the research subjects,bone marrow cells of the donor mice were injected into the irradiated recipient ones,and four chimeric models were established as follows：class A,myeloid cells TLR4＋/＋ [recipient（R） HeN]and LSECTLR4＋/＋[donor（D） HeN];class B,myeloid cells TLR4＋/＋（R,HeN） and LSECTLR4-/-（D,HeN）;class C,myeloid cellsTLR4-/-（R,HeN） and LSECTLR4＋/＋（D,HeN）;class D,myeloid cellsTLR4-/-（R,HeN） and LSECTLR4-/-（D,HeN）.Each mouse of the four chimeric models underwent hepatic I/R injury.Finally,inflammatory protein extracted from liver tissue and PMN isolated from peripheral blood were placed into separate wells of Transwell,and PMN migration was recorded by calculating the numbers of PMN.C3a was used as a positive control and buffer as a negative control.Results Compared with class C（67.10±6.89）,there was statistical increase of PMN migrating numbers in class A（148.95±11.04）（P0.05）.Compared with class D（63.15±8.86）,class B（142.65±8.67）had statistical increase of PMN migrating numbers（P0.05）.While there was no significant difference between class A and class B（P0.05）,and class C and class D had no significant difference（P0.05）as well.Conclusions Myeloid cells TLR4,not endothelial cell TLR4,play a leading role of PMN migration in hepatic I/R injury.