TNF directly stimulates TNF activation and expression of the selected surrogate markers in human pluripotent stem cell derived-kidney organoids. (A) TNF activation scores were calculated from bulk RNAseq data obtained from kidney organoids treated with vehicle control (VC) or 1, 5, 25ng/ml TNF for 3 or 20h. Quantification of (B) CCL2 (left) and TIMP1 (right) transcript levels in kidney organoid cell lysates by qRT-PCR relative to control, and of (C) MCP-1 (left) and TIMP-1 (right) protein levels in kidney organoid culture supernatant by ELISA normalized to total protein, generated from the same samples following treatment with 5 ng/ml TNF or vehicle control for 24h. Each data point was generated from a unique sample and represents the average of analysis in triplicate. Long bar, mean; short bar, 1 S.D. ; *p-value < 0.05 by Student’s t-test. Representative experiment (1 of 4 independent) shown.