Bioinformatics Analysis and Experimental Validation of Epigallocatechin-3-gallate Against Iopromide-induced Injury in HEK-293 CellsviaAnti-oxidative and Anti-inflammation Pathways

Figures

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  Figure 1. Effect of cell viability on EGCG (50, 100 and 200 μM)-treated HEK293 cells. HEK-293 cells were treated with EGCG (50, 100 and 200 μM) for 24 h and cell viability was assessed using the MTT assay. The experiments were repeated thrice, and the data are presented as mean±standard deviation.

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  Figure 2. EGCG protected HEK-293 cells against the iopromide (50 mgI/ml)-induced cell viability decrease. (A) HEK-293 cells were treated with iopromide (50 mgI/ml) and/or EGCG (100 and 200 μM), and cell viability was measured with the MTT assay. Results were analyzed using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test; ***p<0.001 and ###p<0.001. The experiments were repeated thrice, and the data are presented as mean±standard deviation. (B) Changes in the morphology of EGCG (100 and 200 μM)-treated HEK-293 cells.

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  Figure 3. EGCG protected HEK-293 cells against iopromide-induced apoptotic cell death. HEK-293 cells were treated with iopromide (50 mgI/ml) and/or EGCG (100 and 200 μM). (A) Apoptotic cells (TUNEL positive cells) were measured by the TUNEL assay. (B) Caspase-3 enzyme activity was measured. Results were analyzed using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test; ***p<0.001 and ###p<0.001.

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  Figure 4. Volcano plot of whole-transcriptome sequencing analysis of EGCG (200 μM)-treated iopromide (50 mgI/ml)-injured HEK-293 cells.

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  Figure 5. The ingenuity canonical pathways of whole-transcriptome sequencing analysis of EGCG (200 μM)-treated iopromide (50 mgI/ml)-injured HEK-293 cells.

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  Figure 6. Network depicting associations among various genes of the anti-inflammatory pathway (A) and apoptosis of kidney pathway (B), by IPA analysis of whole-transcriptome sequencing analysis of EGCG (200 μM)-treated iopromide (50 mgI/ml)-injured HEK-293 cells.

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  Figure 7. Biological process (BP) of whole-transcriptome sequencing analysis of EGCG (200 μM)-treated iopromide (50 mgI/ml)-injured HEK-293 cells by Gene Ontology (GO) analysis. The top four relevant categories for EGCG’s effects on iopromide-induced nephrotoxicity was the nitric oxide mediated signal transduction, response to hydrogen peroxide, cellular response to oxidative stress, and response to oxidative stress. These categories are the only four with p-value <0.005.

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  Figure 8. Effect of intracellular ROS of EGCG-treated iopromide-injured HEK-293 cells. HEK-293 cells were treated with iopromide (50 mgI/ml) and/or EGCG (100 and 200 μM), and intracellular ROS was measured by the DCFH-DA fluorescence probe. Results were analyzed using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test; ***p<0.001 and ###p<0.001.

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  Figure 9. Effect of antioxidant activity of EGCG-treated iopromide-injured HEK-293 cells. HEK-293 cells were treated to iopromide (50 mgI/ml) and/or EGCG (100 and 200 μM), and antioxidant activity was measured by the DPPH radical scavenging activity. Results were analyzed using one-way analysis of variance (ANOVA) followed by Tukey’s post hoc test; ***p<0.001 and ###p<0.001.