## 3 Results
### Functional Classification of DEGs in Response to GP Extracts  
Based on the heatmap results, most DEGs identified in GP-all showed similar gene expression patterns. Therefore, we conducted a gene enrichment analysis using only the DEGs from GP-all. DEGs from GP-all were divided into upregulated and downregulated genes. Of the 125 upregulated genes, 121 were assigned GO term annotations. Genes assigned to biological regulation (50 genes) were most abundant, followed by those associated with metabolic process (48 genes) and response to stimulus (40 genes) when annotated by the biological process (Figure 6A). Genes associated with the membrane (37 genes) were most abundant, followed by those associated with the nucleus (28 genes) and endmembrane system (26 genes) when annotated by cellular components. Genes related to protein binding (48 genes) were most abundant, followed by ion binding (29 genes) and transferase activity (16 genes) when annotated by molecular function. Of the 51 downregulated genes, a high number were assigned to biological processes (26 genes), responses to stimuli (19 genes), and metabolic processes (19 genes) (Figure 6B). Many downregulated genes were associated with membranes (19 genes), protein-containing complexes (13 genes), and cell projections (nine genes). Two GO terms, endosome (four genes) and chromosome (one gene), were identified only among the upregulated genes. Compared to the upregulated genes assigned to 16 GO terms according to molecular functions, the downregulated genes were assigned to only 13 GO terms. Three GO terms, antioxidant activity (one gene), oxygen binding (one gene), and enzyme regulator activity (eight genes), were identified only in the upregulated genes. To identify the enriched functions for the 176 DEGs identified from GP-all, we conducted gene set analyses using the WebGestalt program, which contains several databases. For this, 125 upregulated and 51 downregulated genes were subjected to gene set analysis (Tables S5 and S6). GO enrichment analyses showed a similar number of GO terms for up- (52 terms) and downregulated genes (53 terms) according to the biological processes (Figure 7). However, based on cellular components and molecular functions, the number of GO terms for upregulated genes was higher than that for downregulated genes. For upregulated genes, two GO terms, response to growth factor (GO:0070848) and heart development (GO:0007507), were the most significant functional GO terms assigned to the biological processes (Figure 8A). The 11 genes assigned to growth factors were _LTBP1_, _MYCCD_, _VTN_, _SOX5_, _ERBB4_, _TH_, _CLDN5_, _FGF1_, _GRB10_, _TMEM204_, and _SOX2_ (Table S7). The nine genes related to heart development were _LTBP1_, _SCNA5_, _DNAHS_,_MYOCD, ERBB4, TH, CLDN5, CSRP3_, and _GLII_. Moreover, monooxygenase activity (GO:0004497)-associated GO term was the most significant molecular function, and four genes, _TPH1_, _CYP24A1_, _CYP2F1_, and _TH_, were involved (Table S7 and Figure 8B). Many upregulated genes were targeted to several cellular components, such as synapse (nine genes), supramolecular fiber (eight genes), neuron projection (10 genes), and integral components of the plasma membrane (12 genes), Golgi apparatus (11 genes), endoplasmic reticulum (15 genes), cytoplasmic region (six genes), and NLRP3 inflammasome complex (one gene) (Table S7). Figure 5: Comparison of the numbers of DEGs in four different conditions. Venn diagrams display the number of (**A**) upregulated genes and (**B**) downregulated genes among the four conditions. Yellow colored box indicates commonly identified transcripts in four conditions. (**C**) Heatmap shows the expression of the 176 DEGs identified in GP-all under the four different conditions. Red and green colors indicate up- and downregulated genes, respectively. DEG, differentially-expressed genes; GP, _Gynostemma pentaphylum_.