Recruitment and targeting of inflammatory cells during infection is a complex process involving multiple steps that are enabled, in part, by chemokine-dependent signals. In this study, we demonstrated that Ly6Chigh monocytes are rapidly recruited from bone marrow to the liver during L. monocytogenes infection. CCR2 deficiency impairs recruitment and leads to a greater bacterial burden in the liver. We provided evidence that CCR2-mediated signals principally enable monocyte egress from bone marrow and that once Ly6Chigh monocytes migrate into the circulation, they are targeted to hepatic foci of infection and provide immune defense, without a requirement for additional CCR2-mediated signals. Our study indicated that monocyte recruitment to hepatic foci involves the focal induction of adhesion molecules.
Our histologic data and previous studies (8
) showed that the distribution of bacterial infection in the liver is localized and restricted to inflammatory foci that consist of recruited leukocytes and infected hepatocytes. Intravital imaging demonstrated that blood flow to the central region of infected hepatic foci was markedly diminished. Bacterial infection seems to be restricted by the recruitment of inflammatory monocytes to the region surrounding bacterially infected foci. In contrast to previous studies, in which leukocyte recruitment to the liver was studied in the setting of widespread inflammation (29
), our study focused on monocyte recruitment to focal sites of infection and inflammation. We found that bacterial infection selectively increased expression of the adhesion molecule ICAM-1 in regions immediately adjacent to infected foci. Focal induction of ICAM-1 during L. monocytogenes
infection may result from localized production and diffusion of TLR ligands from areas of bacterial infection and hepatocyte necrosis.
Leukocyte recruitment in liver sinusoids occurs by distinct mechanisms that differ from the tether-and-roll paradigm governing the adhesion and emigration in other vascular compartments (41
). Hepatic sinusoidal endothelium does not express selectins, and selectins are not essential for recruitment of cells to sinusoids (42
). In fact, visualization of leukocyte movement in the liver revealed that adhesion in sinusoids often occurred independent of rolling (41
). Hepatic sinusoids are fenestrated and lack a basal lamina and tight junctions. Thus, the recruitment of monocytes from sinusoids to foci of infection likely follows a scheme that differs from chemokine-dependent endothelial transmigration. Although most previous studies showed that leukocyte chemotaxis induced by chemokines is inhibited by pertussis toxin, certain chemokine receptors were shown to couple to pertussis toxin-sensitive and -resistant G proteins (43
). Therefore, it is possible that pertussis toxin-insensitive signals mediated by chemokine receptors contribute to monocyte recruitment.
Previous studies showed that neutrophil sequestration in inflamed liver sinusoids following systemic LPS administration is mediated by the interaction of CD44 on neutrophils with extracellular hyaluronan (HA) (40
). Systemic administration of LPS, in addition to resulting in hepatic inflammation, induces the expression of serum-derived HA-associated protein, which enhances the association of HA with CD44. In our experiments, we found that Ab-mediated blockade of CD44 markedly reduced the number of Ly6Chigh
monocytes detected by FACS in L. monocytogenes
-infected livers (). Thus, our studies demonstrated a role for CD44 in inflammatory monocyte recruitment to the liver. However, it is unclear whether L. monocytogenes
infection induces serum-derived HA-associated protein expression and, thus, promotes monocyte recruitment by enhancing interactions between CD44 and HA or whether HA expression is altered in areas adjacent to foci of L. monocytogenes
infection. Further studies are necessary to determine how CD44 contributes to the targeting of monocytes to liver during L. monocytogenes
Studies with CX3CR1-GFP mice using intravital microscopy documented the patrolling behavior of resident monocytes in noninflamed dermis (44
). In those studies, specific Ab-mediated blockade of LFA-1 led to detachment of CX3CR1-expressing monocytes from the endothelial cell surface, suggesting that LFA-1 was essential for maintaining monocyte attachment to the endothelium under conditions of shear force. Although we observed similar patrolling of Ly6Chigh
monocytes in L. monocytogenes
-infected liver, LFA-1 blockade did not result in a reduction in monocyte recruitment to the liver or detachment of monocytes from the sinusoid wall or from foci of infection. It is possible that the role of LFA-1 in patrolling differs between the inflammatory CCR2+
monocyte subset and the CX3CR1+
subset. Alternatively, adhesion mechanisms during infection may differ from those in the absence of inflammation. In addition, monocyte patrolling in liver sinusoids may differ from that in dermal vasculature.
In summary, we demonstrated that Ly6Chigh
monocytes are rapidly recruited from bone marrow to foci of infection in the liver where they mediate clearance of L. monocytogenes
. Although monocyte egress from bone marrow is CCR2 dependent, entry into the liver parenchyma and localization to foci of bacterial infection are chemokine receptor independent. Focal induction of ICAM-1 facilitates the targeting of inflammatory monocytes to regions immediately adjacent to foci of bacterial infection. Ly6-Chigh
monocytes are increasingly recognized as essential mediators of defense against viral, bacterial, fungal, and protozoan pathogens (8
). Deciphering the mechanisms that enable monocyte trafficking to peripheral sites of bacterial infection may provide new approaches to enhance immune defense against these pathogens.