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Blood, Vol. 110, Issue 2, 587-595, July 15, 2007
Tumor-derived hyaluronan induces formation of immunosuppressive macrophages through transient early activation of monocytes
Blood Kuang et al.
110: 587
Supplemental materials for Kuang et al, Vol. 110, Issue 2, 587-595
Files in this Data Supplement:
- Table S1. Primers used in RT-PCR for hyaluronan synthase isoforms (PDF, 14.0 KB)
- Table S2. TSNs downregulated cell surface markers for Mφ activation (PDF, 60.9 KB) -
Monocytes were cultured in medium alone (Med) or with 15% of the indicated TSN for 6 days. Thereafter, M were left untreated (Med) or were stimulated with LPS or IFN- for 18 hours, and then stained with fluorochrome-conjugated anti-CD64, anti-CD86, or anti-HLA-DR or relevant control Ab. Values represent the mean (± SD) of the mean fluorescence intensity (MFI) from 10 separate experiments.
- Table S3. TSN or INT-HA caused transient early activation of monocytes (PDF, 17.9 KB) -
Monocytes were cultured in medium alone (Med) or with 15% TSN or 50 µg/mL INT-HA (HA60) for 18 hours or 48 hours and their expression of surface molecules (MFI) were determined. The data shown for 16 hour exposure are the mean (± SD) of four separate experiments, and the values for 48 h are the mean of two separate experiments. ND, not determined.
- Figure S1. Polymyxin B did not affect the cytokine production in TSN-stimulated monocytes (PDF, 44.4 KB) -
Monocytes were pre-incubated with (solid bars) or without (open bars) 10 µg/mL polymyxin B for 15 minutes and then stimulated with LPS or TSN for 18 hours. Cytokine production was determined by ELISA and values represent the mean (± SE) of 4 experiments; **P<0.01, compared with monocytes treated with polymyxin B.
- Figure S2. Confocal microscopic analysis of the IL-10 expression in Mφ from cancer nest (PDF, 55.9 KB) -
Frozen hepatocellular carcinoma (HCC) tissue sections were double stained with mouse antihuman CD68 and rabbit antihuman IL-10 Ab (Abcam, Cambridge, MA) followed by Alexa Fluor 488-conjugated goat anti-mouse IgM and Alexa Fluor 568-conjugated goat anti-rabbit IgG (Molecular Probes, Eugene, OR). These figures show that most macrophages (green) in the cancer nests were positive for IL-10 (red).
- Figure S3. Hyaluronan levels in TSN from different tumor cells (PDF, 58.9 KB) -
TSN were prepared as described in “Tumor cell lines and preparation of tumor culture supernatants, Materials and methods” and the concentration of hyaluronan in TSN was determined by an ELISA kit from Adlitteram Diagnostic Laboratories (San Diego, CA). (A) HA levels in the TSN from eight different solid tumors and two leukemia cell line cells. (B) HA levels in the TSN from parental, mock-transfected, and pSi-HAS2-transfected cells. The illustrated values are the mean of two separate experiments.
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