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Blood, Vol. 113, Issue 4, 887-892, January 22, 2009
The macrophage scavenger receptor CD163 functions as an innate immune sensor for bacteria
Blood Fabriek et al.
113: 887
Supplemental materials for: Fabriek et al
Files in this Data Supplement:
- Figure S1. Peptide CD163p2 from the second SCRC domain of CD163 induces bacterial aggregation (JPG, 39 KB)
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CD163p1–9 peptides (50 µg/ml) were added to S.mutans and aggregation was monitored. Representative photographs from triplicate incubations are shown. Apart from CD163p2 none of the other peptides agglutinated S.mutans. CD163p7 is shown as a representative control peptide.
- Figure S2. Characterization of novel blocking and non-blocking antibodies against human CD163 (JPG, 28.9 KB)
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THP EV and THP CD163 cells. Effect of mAb EDhu2, EDhu3 and EDhu4 directed against CD163 on: (A) Binding of DsRed-transfected E. coli to THP EV cells and THP CD163 cells, expressed relative to controls as means of the geometric mean fluorescence intensity values ± SD from triplicate incubations. (B) E. coli-induced TNFα production by THP EV and THP CD163 cells. Means ± SD from 3 independent experiments are shown. Statistics: two-tailed students T test. Note that mAb EDhu2 and EDhu3 essentially prevent bacterial binding and cytokine production, whereas EDhu4 shows little inhibition.
- Figure S3. Comparison of surface expression of CD163 introduced into CHO cells and THP-1 cells (JPG, 39.4 KB)
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CHO cells transfected with empty vector or full length CD163, and THP-1 cells retrovirally transduced with empty vector or full length CD163 were analyzed by flow cytometry using the anti-CD163 mAb EDhu1. Note that the CHO-CD163 cells have considerably higher CD163 expression than the THP-CD163 as indicated by a ~10 fold higher mean fluorescence intensity.
- Figure S4. CD163 does not contribute substantially to bacterial binding and/or phagocytosis by monocytes (JPG, 32.2 KB)
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Human monocytes were preincubated with blocking antibodies (EDhu2 and EDhu3) and non-blocking antibodies (EDhu1 and EDhu4) directed against CD163 allowed to phagocytose FITC-labelled Streptococus mutans bacteria for 2 hours. Binding and/or phagocytosis was assessed by flow cytometry. Histograms show the bacterial fluorescence of control cells (solid line) and monocytes that had been allowed to phagocytose bacteria after preincubation with the indicated antibodies (grey-filled histogram). Monocytes incubated in absence of bacteria are shown in the red histogram. Parallel confocal microscopic evaluation of the samples indicated that 80–90% of the bacteria had been internalized by the monocytes.
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