O22
SLIT2 IMPAIRS MONOCYTE CHEMOTAXIS AND RENAL EXPRESSION IS
DOWNREGULATED IN ACUTE ALLOGRAFT REJECTION
Mallik, M1, Rui-Crowm M 1, Yan, H1, Chaukos, D1, Coffman, T M2, Robinson, L A1.
1The Hospital for Sick Children, Toronto,Canada
2Duke University Hospital, Durham NC, USA
Migration of circulating leukocytes into the injured kidney is a key component of
allograft rejection and ischemia reperfusion injury. There is increasing evidence that
monocytes/macrophages play a critical role in both of these processes. Leukocyte
migration is orchestrated by chemokines, a family of small chemoattractant cytokines. As
multiple chemokines and receptors exist, there is much redundancy within the system.
Thus, strategies targeting inhibition of one pathway may result in its function being
assumed by another. Generalised chemokine blockade could therefore lead to more
effective inhibition of leukocyte chemotaxis.
The neuronal guidance cue, Slit, and its transmembrane receptor Roundabout (Robo)
prevent axonal migration during development. Recently Slit and Robo have also been
shown to impair migration of several types of leukocytes.
We studied expression of Slit2 in a murine model of acute renal allograft rejection. By
immunohistochemistry, Slit2 was expressed mainly in tubular epithelial cells. Slit2
expression was reduced in allografts compared to isografts at 2, 4 and 6 weeks post
transplantation.
Using flow cytometry, immunofluorescence microscopy and RT-PCR we demonstrated
that Robo1 is expressed by primary human peripheral blood monocytes, THP-1
monocytic cells and murine RAW macrophages. We used Transwell chambers to study
the effects of Slit2 on monocyte migration towards 3 different families of chemokines
(CXC-, CC-, CX3C-). Administration of Slit2 resulted in a 3-5 fold reduction in migration
towards these different chemokine gradients. To determine whether Slit also affects other
immune functions of monocytes/macrophages, phagocytosis was assessed by measuring
the ability of RAW cells to ingest latex beads opsonised with human IgG in the presence
or absence of Slit2. Slit2 had no effect on phagocytic index (no. ingested particles/no.
cells).
Our findings suggest that in acute allograft rejection, Slit2 expression is downregulated
thereby allowing circulating monocytes to infiltrate the graft. Our studies further
demonstrate that Slit/Robo prevent monocyte chemotaxis whilst preserving
monocytic/macrophage phagocytosis. Targeting this mechanism will allow the
development of novel strategies to prevent leukocyte influx associated with allograft
rejection, without compromising other key leukocyte immune functions.