Genes differentially regulated by VEGF-A and VEGF-C156S

List of genes which are differentially regulated by VEGF-A and VEGF-C156S stimulatio

Downregulated Genes after VEGFC156S stimulation

List of genes corresponding to the downregulated CAGE peaks (p < 0.05, log2FC<-0.6) for each time poin

Upregulated Genes after VEGFC156S stimulation

List of genes corresponding to the upregulated CAGE peaks (p < 0.05, log2FC>0.6) for each time poin

Regulatory relationship of monocyte's core TRN elements.

<p>(A) Relative expressions to monocyte are represented in colors. Higher relative expression is depicted as intensifying green color. The value of each relative expression is the average of biological replicates (n = 3). (B) Illustration of hierarchical network of monocyte core TRN elements. Each node (circle) indicates monocyte core TRN elements and green nodes represent the identified TRN inducers. When the TRN inducers or NR4A2 upregulate the gene expression to more than 5% of that of monocyte, an edge was drawn. An edge from an upper node to lower node indicates positive regulation.</p

Motif activity revealed reconstructed monocyte TRN.

<p>(A) The activity of 10 monocyte-associated motifs were shown as a line chart in FIB-mock, FIB-SPI1, and FIB-4Fs. Vertical axis represents the motif activity. (B) SPI1 motif activity was shown as bar plot. Each bar represent an average of biological replicates and error bar represent s.d. (n = 3).</p

The monocyte core TRN elements.

<p>Rfc = differential expression fold change rank.</p><p>Rco = co-occurrence rank.</p><p>IS = Importance Score (1/Rfc+1/Rco).</p

Reconstruction of Monocyte Transcriptional Regulatory Network Accompanies Monocytic Functions in Human Fibroblasts

<div><p>Transcriptional regulatory networks (TRN) control the underlying mechanisms behind cellular functions and they are defined by a set of core transcription factors regulating cascades of peripheral genes. Here we report SPI1, CEBPA, MNDA and IRF8 as core transcription factors of monocyte TRN and demonstrate functional inductions of phagocytosis, inflammatory response and chemotaxis activities in human dermal fibroblasts. The Gene Ontology and KEGG pathway analyses also revealed notable representation of genes involved in immune response and endocytosis in fibroblasts. Moreover, monocyte TRN-inducers triggered multiple monocyte-specific genes based on the transcription factor motif response analysis and suggest that complex cellular TRNs are uniquely amenable to elicit cell-specific functions in unrelated cell types.</p> </div

Cell feature assessments in reconstructed fibroblasts.

<p>(A) Morphological changes were visualized by microscopy. The cells were stained with Pallodin-Rhodamin (Yellow), Hoechst33342 (Blue), and Whole Cell Stains (Red). (B–D) Phagocytic latex beads were visualized (B) and the mean intensity of the ingested beads were confirmed by flow cytometry (C–D). (B) Beads-ingested cells are indicated by white arrows. Red color represents DID cell membrane staining, blue color represents nuclei, and green color represents the latex beads. (C) Beads ingested cells was quantified based on the beads fluorescence by flow cytometry in FIB-mock, FIB-SPI1 and DIB-4Fs 2 houes after beads addition.. The cells were cultured with 0.002 v/v%. Vertical and horizontal axes represent cell count and fluorescent intensity, respectively. Beads ingested cells were gated. The analysis was performed in triplicates, showing the similar results. (D) The flow cytometric analysis of the phagocytosis was summarized. Black, blue, and red lines represent FIB-mock, FIB-SPI1, and FIB-4Fs, respectively. The mean fluorescent intensity of ingested beads was measured by flow cytometry. The vertical axis and horizontal axis represent mean beads fluorescent intensity and incubation time, respectively. Error bars represent standard deviation (s.d.) (E) The expression change of <i>TNF</i>, <i>IL6</i>, <i>IL1A</i>, <i>IL1B</i>, <i>IL8</i>, <i>CCL2</i>, <i>CXCL10</i> and <i>IFNB1</i> induced by LPS treatment were measured by qRT-PCR in FIB-mock, FIB-SPI1, and FIB-4Fs. The cells were treated with LPS for 24 hours at the final concentration of 10 µg/µl. The bar represents the relative expression of LPS-treated cells as compared to untreated cells and error bar represents s.d. These experiments were repeated three times. * represents <i>P</i>-value≤0.05, ** represents <i>P</i>-value≤0.01 (t-test). The scale bar is 50 µm.</p

Core TRN elements isolation workflow.

<p>Key TFs were selected based on differential gene expression analysis and text mining. Left flow represents steps of the differential gene expression analysis. Gene expression profiling of primary human monocytes and that of human dermal fibroblasts were archived by using Human WG-6 v3.0 Expression beadschips (n = 3). Right flow represents the text mining. The text mining ranked the co-occurrence of “Monocyte” and “TF Name”. Finally, those two methods were integrated and isolated top 20 TFs as core monocyte TRN elements.</p

Expression of monocyte markers.

<p>Expression of monocyte markers.</p