To better define the activity of soluble CXCL16 in the recruitment of polymorphonuclear neutrophils (PMNs) in vivo, utilizing a novel animal model of gout involving engraftment of SCID mice with normal human synovial tissue (ST) injected intragraft with gouty human synovial fluid (SF).
For in vitro studies, a modified Boyden chemotaxis system was used to identify CXCL16 as an active recruitment factor for PMNs in gouty SF. Migration of PMNs could be reduced by neutralization of CXCL16 activity in gouty SF. For in vivo analyses, fluorescent dye-tagged PMNs were injected intravenously into SCID mice while, simultaneously, diluted gouty SF containing CXCL16, or depleted of CXCL16 by antibody blocking, was administered intragraft. In addition, the receptor for CXCL16, CXCR6, was inhibited by incubating PMNs with a neutralizing anti-CXCR6 antibody prior to injection into the mouse chimeras. Recruitment of PMNs to the gouty SF-injected normal human ST was then examined in this SCID mouse chimera system.
CXCL16 concentrations were highly elevated in gouty SF, and PMNs were observed to migrate in response to CXCL16 in vitro. Normal human ST-SCID mouse chimeras injected intragraft with gouty SF that had been depleted of CXCL16 during PMN transfer showed a significant reduction of 50% in PMN recruitment to engrafted tissue as compared with that after administration of sham-depleted gouty SF. Similar findings were achieved when PMNs were incubated with a neutralizing anti-CXCR6 antibody before injection into chimeras.
Overall, the results of this study outline the effectiveness of the human-SCID mouse chimera system as a viable animal model of gout, serving to identify the primary function of CXCL16 as a significant mediator of in vivo recruitment of PMNs to gouty SF.