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Blood, Vol. 110, Issue 9, 3226-3233, November 1, 2007
RNA fingerprints provide direct evidence for the inhibitory role of TGFß and PD-1 on CD4+ T cells in Hodgkin lymphoma
Blood Chemnitz et al.
110: 3226
Supplemental materials for: Chemnitz et al
Files in this Data Supplement:
- Figure S1. Inhibition of T cell proliferation by TGFβ and PD-1 (PDF, 30.2 KB) -
Freshly isolated primary human CD4+ T cells were labeled with CFSE and left unstimulated or were stimulated with the indicated magnetic beads in the absence or presence of 30 ng/ml TGF . After 4 days CFSE dilution was analyzed by flow cytometry. The overall percentage of dividing cells is displayed inside the corresponding dot plot. Two different experiments are shown in A and B, reflecting the range of inhibition of T cell proliferation. The smallest concentration for TGF leading to maximum reduction of IFN- production (30 ng/ml TGF) was used to determine transcriptional changes as the “RNA-fingerprint” of TGF on human CD4+ T cells. Similarly, the minimum concentration of PD-1 inducing complete blockade of T cell activation was used for further analysis.
- Figure S2. Supervised classification (PDF, 34.4 KB) -
CD4+ T cells were isolated from lymph nodes of 4 patients with HL, 5 patients with RLN and 3 patients with FL. cRNA was hybridized to HG-U133A Affymetrix arrays. The RNA- fingerprints of (A and C) TGF and (B and D) PD-1 were used to separate transcriptional profiles of HL from RLN samples and HL from FL samples, respectively. Left panels: Supervised classification using PAM; for each sample the posterior probability, i.e. the percentage of certainty of a correct class prediction is plotted. Right panels: Supervised classification using SVMs. A fourfold table comparing the predicted class labels to the actual class labels is depicted.
- Figure S3. PCA and supervised classification on the Illumina platform comparing RLN with HL samples (PDF, 36 KB) -
CD4+ T cells were isolated from lymph nodes of 5 patients with HL and 4 patients with RLN. cRNA was hybridized to Sentrix® whole genome bead chips 6 × 2. The RNA- fingerprints of (A) TGF and (B) PD-1 were used to separate transcriptional profiles of HL from RLN samples. Left panels: Result of principle components analysis with the first three principal components plotted. Middle panels: Supervised classification using PAM; for each sample the posterior probability, i.e. the percentage of certainty of a prediction is plotted. Right panels: Supervised classification using SVMs. A fourfold table comparing the predicted class labels to the actual class labels is depicted.
- Figure S4. PCA and supervised classification on the Illumina platform comparing RLN with FL samples (PDF, 35.5 KB) -
CD4+ T cells were isolated from lymph nodes of 6 patients with FL and 4 patients with RLN. cRNA was hybridized to Sentrix® whole genome bead chips 6 × 2. The RNA- fingerprints of (A) TGF and (B) PD-1 were used to separate transcriptional profiles of HL from RLN samples. Left panels: Result of principle components analysis with the first three principal components plotted. Middle panels: Supervised classification using PAM; for each sample the posterior probability, i.e. the percentage of certainty of a prediction is plotted. Right panels: Supervised classification using SVMs. A fourfold table comparing the predicted class labels to the actual class labels is depicted.
- Table S1. Genes comprising the TGFβ fingerprint (after cross-annotation) (XLS, 43 KB)
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Fold changes of genes in vitro and in vivo are shown.
- Table S2. Genes comprising the PD-1 fingerprint (after cross-annotation) (XLS, 24 KB)
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Fold changes of genes in vitro and in vivo are shown.
- Table S3. Differentially expressed genes (FC>2, p-value<0.05, diff>100) between HL vs RLN samples and FL vs RLN samples (XLS, 36.0 KB)
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