Brooks Andrew C, Rickards Karen J, Cunningham Fiona M
Department of Veterinary Basic Sciences, Royal Veterinary College, Hawkshead Lane, North Mymms, Hertfordshire, AL9 7TA, UK.
Vet Immunol Immunopathol. 2011 Feb 15;139(2-4):141-7. doi: 10.1016/j.vetimm.2010.09.008. Epub 2010 Oct 7.
The chemokine, CXCL8, is a potent chemoattractant but it has also been shown to attenuate the migratory response of human neutrophils to the bacterial peptide, FMLP; this could lead to retention of cells in infected tissue and, potentially, to enhanced clearance of bacteria. This study has examined the effect of CXCL8 on equine neutrophil migration and adherence in response to PAF and LTB(4), chemoattractants that may play a role in non-infectious inflammatory conditions of the horse associated with neutrophil recruitment to the target tissue. The effects of CXCL8 on PAF- and LTB(4)-induced responses were determined using a ChemoTx plate migration assay and by measuring adhesion to protein-coated plastic. The CXCR1/2 antagonist, SB225002, was used to investigate whether the observed effects were receptor mediated and the role of cAMP was examined by measuring intracellular cAMP following exposure to agonists alone and in combination and by establishing the effect of dibutyryl cAMP on neutrophil migration. CXCL8, LTB(4) and PAF each induced migration and adhesion. Exposure of neutrophils to a combination of CXCL8 and PAF reduced the magnitude of the responses to that of unstimulated cells. In contrast, although the effect was less than additive, the response to co-stimulation with CXCL8 and LTB(4) were not nearly as pronounced. CXCL8 acted in a receptor mediated manner, the attenuation of PAF-induced responses being reversed by SB225002 at a concentration that blocks CXCR2. CXCL8, PAF and LTB(4) alone increased intracellular cAMP. In co-incubation studies, combination of CXCL8 with PAF led to an additive increase in cAMP whereas no increase above that obtained in response to LTB(4) alone was seen. Dibutyryl cAMP significantly reduced neutrophil migration in response to either CXCL8 or PAF alone. These results demonstrate that CXCL8, in addition to being a potent chemoattractant and pro-adhesive molecule for equine neutrophils, is able to attenuate responses to PAF and, to a much lesser extent, LTB(4). This effect, which appears to be CXCR2-mediated and cAMP dependent, could lead in vivo to trapping of cells at sites of inflammation resulting potentially in either enhanced clearance of injurious stimuli or increased local tissue damage by activated cells.
趋化因子CXCL8是一种有效的趋化剂，但也已证明它可减弱人类中性粒细胞对细菌肽FMLP的迁移反应；这可能导致细胞滞留在感染组织中，并有可能增强细菌的清除。本研究检测了CXCL8对马中性粒细胞迁移和黏附的影响，这些细胞是对血小板活化因子（PAF）和白三烯B4（LTB4）产生反应，这两种趋化剂可能在与中性粒细胞募集到靶组织相关的马的非感染性炎症中发挥作用。使用ChemoTx平板迁移试验并通过测量对蛋白包被塑料的黏附来确定CXCL8对PAF和LTB4诱导反应的影响。使用CXCR1/2拮抗剂SB225002来研究观察到的效应是否由受体介导，并通过单独和联合暴露于激动剂后测量细胞内cAMP以及确定二丁酰cAMP对中性粒细胞迁移的影响来研究cAMP的作用。CXCL8、LTB4和PAF均可诱导迁移和黏附。将中性粒细胞暴露于CXCL8和PAF的组合中会使反应幅度降低至未刺激细胞的水平。相比之下，尽管该效应小于相加效应，但与CXCL8和LTB4共同刺激的反应并不那么明显。CXCL8以受体介导的方式起作用，PAF诱导反应的减弱可被浓度足以阻断CXCR2的SB225002逆转。单独的CXCL8、PAF和LTB4均可增加细胞内cAMP。在共孵育研究中，CXCL8与PAF的组合导致cAMP的相加增加，而未观察到高于单独对LTB4反应所获得的增加。二丁酰cAMP可显著降低中性粒细胞对单独的CXCL8或PAF的反应性迁移。这些结果表明，CXCL8除了是马中性粒细胞的有效趋化剂和促黏附分子外，还能够减弱对PAF的反应，并在较小程度上减弱对LTB4的反应。这种效应似乎是由CXCR2介导且依赖于cAMP，在体内可能导致细胞在炎症部位滞留，从而可能增强对有害刺激的清除或增加活化细胞对局部组织的损伤。
