Matrix and cytokine regulation of leukocyte recruitment and activation at sites of inflammation

Cytokines and chemokines as well as the extracellular matrix (ECM) regulate leukocyte recruitment and local activation. Hyaluronan (HA) is a major glycosaminoglycan constituent of the ECM. Although HA typically exists as high molecular weight polymers (HMW-HA), lower molecular weight species (LMW-HA) accumulate under inflammatory conditions. To investigate the cellular requirements for activation of peripheral macrophages by HA, we compared the responses of murine resident and thioglycollate-eIicited peritoneal macrophages to HMW- and LMW-HA. LMW-HA selectively stimulated production of the chemokines RANTES, MIP-1α and MIP-1β by elicited macrophages. Further, LMW-HA induced production of biologically active interleukin-12 (IL-12), a pro-inflammatory cytokine not previously known to be regulated be cell-matrix interactions, which was markedly inhibited by an anti-CD44 mAb that blocks HA binding but was unaffected in CD44−/− macrophages. Freshly explanted resident peritoneal macrophages did not respond to LMW-HA. However, pre-culture in vitro for 16 hrs. prior to stimulation led to adhesion dependent priming for LMW-HA induced cytokine and chemokine production by resident macrophages. We examined regulation of LMW-HA induced chemokine and IL-12 production by IFN-γ and TNF, two cytokines with well defined pro-inflammatory activities. While IFN-γ primed macrophages to produce high levels of IL-12 in response to LPS or LMW-HA, it inhibited MIP-1α and MIP-1β production. Furthermore, pre-exposure to TNF inhibited IFN-γ priming of enhanced IL-12 production in response to LMW-HA or LPS mediated by IL-10 dependent and IL-10-independent mechanisms. To determine if TNF inhibition of IFN-γ induced IL-12 production contributed to resolution of an inflammatory response in vivo, the response of TNF+/+ and TNF−/− mice injected with heat-killed C. parvum were compared. TNF −/− mice developed a delayed, vigorous inflammatory response leading to death with increased serum levels of IL-12 and IFN-γ while TNF+/+ mice exhibited a prompt response that resolved. Treatment with neutralizing anti-IL-12 antibody led to resolution of the granulomatous response to C. parvum in the spleens and livers of TNF −/− mice. The kinetics and balance of the multiple cytokines produced and modification of the ECM during an inflammatory response may regulate its magnitude and duration