Supplementary Material for: MIP-3α Expression in Macrophages Is NOD Dependent

<i>Background:</i> The first identified susceptibility gene for Crohn’s disease, <i>NOD2</i>, acts as a sensor for the bacterial-wall peptidoglycan fragment muramyl dipeptide (MDP) and activates the transcription factor nuclear factor-ĸB (NF-ĸB). Upon NF-ĸB activation, intestinal macrophages (IMACs) induce expression of macrophage inflammatory protein (MIP)-3α to attract memory T lymphocytes. We therefore investigated the influence of NOD2 ligation of IMAC differentiation and functional MIP-3α induction. <i>Methods:</i> Human embryonal kidney HEK293 cells were transfected with NOD2 wild-type (NOD2^WT) and the NOD2 SNP13 variant (NOD2^L1007fsinsC) and stimulated with MDP. Recruitment of CD45R0⁺ and Th17 cells was determined by immunohistochemistry. <i>Results:</i> Endogenous NOD2 stimulation was followed by a dose-dependent increase in MIP-3α secretion in MONO-MAC-6 (MM6) cells. MIP-3α mRNA was also significantly (* p &lt; 0.05) induced in HEK293 transfected with NOD2^WT via MDP ligation. In vivo cell-cell contacts between IMACs and CD45R0⁺ memory T cells as well as recruitment of Th17 cells in patients of NOD2 variants were unchanged as compared to wild-type patients. <i>Conclusion:</i> Our data demonstrate a dose-dependent increase in MIP-3α secretion in the human myeloid cell line MM6 upon MDP. However, MIP-3α-driven recruitment of Th17 cells or CD45R0⁺ memory T lymphocytes is not affected in patients carrying heterozygous NOD2 variants.