Tsai Wen-Hui, Shih Chung-Hung, Feng Shan-Yu, Chang Shao-Chi, Lin Yu-Chieh, Hsu Hui-Chi
Department of Respiratory Therapy, Taipei Medical University, Taipei, Taiwan, ROC.
Department of Physiology, National Yang-Ming University, Taipei, Taiwan, ROC.
J Chin Med Assoc. 2014 Jul;77(7):367-73. doi: 10.1016/j.jcma.2014.04.008. Epub 2014 Jun 5.
Phagocytic clearance of apoptotic neutrophils by tissue macrophages is a crucial component in the resolution phase of acute inflammation. However, the number of tissue macrophages is low and not likely to cope satisfactorily with the excess number of dying neutrophils. Although recent studies have reported that neutrophils are able to engulf apoptotic neutrophils, the mechanisms by which living neutrophils are attracted to apoptotic neutrophils are poorly defined. Increased amounts of CX3CL1 and microparticles (MPs) are rapidly released by apoptotic cells, and are involved in the chemoattraction of mononuclear phagocytes toward apoptotic cells. The current study investigated the role of CX3CL1 in the chemoattraction of all-trans retinoic acid (ATRA)-treated NB4 (ATRA-NB4) cells toward apoptotic cells.
Conditioning medium and MPs were harvested from apoptotic ATRA-NB4 cell cultures to determine their effects on living ATRA-NB4 cells by transmigration assay and adhesion assay. The cytokine levels in the conditioning medium were determined by enzyme-linked immunosorbent assay. Expression of CX3CR1 (a receptor of CX3CL1) on ATRA-NB4 cells was determined by flow cytometric analysis.
ATRA-NB4 cells transmigrated toward the apoptotic ATRA-NB4 cells, and this chemoattraction was partially inhibited when the CX3CR1 on ATRA-NB4 cells was blocked by its specific antibody. Both exogenous CX3CL1 and MPs released by apoptotic ATRA-NB4 cells were able to enhance the chemoattraction of ATRA-NB4 cells toward apoptotic cells or the adhesion of ATRA-NB4 cells to endothelial cells. CX3CL1 was expressed on the surface of MPs, and blocking this CX3CL1 with its specific antibody was able to partially inhibit the chemoattractive property of MPs.
CX3CL1, in either the free or MP form, is released rapidly by apoptotic ATRA-NB4 cells after induction of apoptosis to mediate the chemoattraction of living ATRA-NB4 cells toward apoptotic cells.
组织巨噬细胞对凋亡中性粒细胞的吞噬清除是急性炎症消退阶段的关键组成部分。然而，组织巨噬细胞数量较少，不太可能令人满意地应对过量死亡的中性粒细胞。尽管最近的研究报道中性粒细胞能够吞噬凋亡的中性粒细胞，但存活的中性粒细胞被凋亡中性粒细胞吸引的机制尚不清楚。凋亡细胞会迅速释放大量的CX3CL1和微粒（MPs），它们参与单核吞噬细胞向凋亡细胞的趋化作用。本研究调查了CX3CL1在全反式维甲酸（ATRA）处理的NB4（ATRA-NB）细胞向凋亡细胞趋化中的作用。
从凋亡的ATRA-NB4细胞培养物中收集条件培养基和MPs，通过迁移试验和黏附试验来确定它们对存活的ATRA-NB4细胞的影响。用酶联免疫吸附测定法测定条件培养基中的细胞因子水平。通过流式细胞术分析测定ATRA-NB4细胞上CX3CR1（CX3CL1的受体）的表达。
ATRA-NB4细胞向凋亡的ATRA-NB4细胞迁移，当ATRA-NB4细胞上的CX3CR1被其特异性抗体阻断时，这种趋化作用被部分抑制。凋亡的ATRA-NB4细胞释放的外源性CX3CL1和MPs都能够增强ATRA-NB4细胞向凋亡细胞的趋化作用或ATRA-NB4细胞与内皮细胞的黏附。CX3CL1表达于MPs表面，用其特异性抗体阻断该CX3CL1能够部分抑制MPs的趋化特性。
凋亡诱导后，凋亡的ATRA-NB4细胞迅速释放游离形式或MP形式的CX3CL1，以介导存活的ATRA-NB4细胞向凋亡细胞的趋化作用。
