Fig 2: Histological analysis of TCF21 in human glomerular diseases. a, b The human normal glomeruli showed that TCF21 weakly expressed in the nuclei of podocytes linear marking along with synaptopodin (SYNPO, a) and nephrin (NPHS1, b). c In patients with nephrotic syndrome, MGN, NPHS1-stain somewhat decreased, but TCF21 highly expressed in the cellular body of podocytes along the lateral side of NPHS1-positivity. d In the sample of NGD defined score 1, TCF21 weakly expressed in the nuclei of segmental podocytes. e In IgAN sample defined score 2, TCF21 expressed in the nuclei of global podocytes. f In nephrotic syndrome, MGN sample defined score 4, TCF21 expression globally expanded in the cytoplasm of podocytes. g The comparison of TCF21 histological score among various glomerular diseases. Their groups were statistically significant among each glomerular disease (p < 0.01). h The relationship between TCF21 histological score and urinary protein level. The urinary protein level was also statistically significant among each histological TCF21 expression level (p < 0.01). Statistical analysis was performed with the Kruskal–Wallis test (g, h).

Fig 3: Urinary TCF21 levels in human glomerular diseases. a Urinary TCF21 levels showed a weak positive correlation with urinary protein levels (r 0.4639). b Urinary TCF21 concentration was statistically significant among each glomerular disease, and higher in nephrotic syndrome (p < 0.01). c, d Urinary TCF21 levels were positively associated with TCF21 histological score in total 50 cases (c) and 19 cases with MGN (d) (c: p < 0.01, d: p = 0.01, respectively). e The comparison of urinary TCF21 levels in MGN patients with or without anti-PLA2R positivity. It was not statistically significant (p = 0.28). f The change ratio of urinary TCF21 in treatment response of nephrotic patients. The change ratio was calculated from urinary TCF21 level at pre-/post-treatment. It was downward greater in remission group than in non-remission group (p < 0.01). g Continuous urinary TCF21 and protein levels during the remission-induction treatment in a case of steroid-sensitive MCD. Statistical analysis was performed with Spearman’s rank correlation coefficient (a), the Kruskal–Wallis test (b, c) and Mann Whitney U test (d–f).

Fig 4: In vitro experiments using Tcf21-expressing murine podocyte cell line. a Based on DEGs of Tcf21-MPs/Control-MPs using microarray, the KEGG pathway frequency analysis presented up-regulated and down-regulated gene groups. b–e: Both cellular morphology of Control-MPs and Tcf21-MPs showed similar shape in low-power (b, c) and mid-power magnification (d, e). f The area dimension of both MPs was not statistically different (p = 0.54). g In high-power magnification, Tcf21-MP possessed rich actin-stained dendrites, filopodia. h The number of filopodia was significantly higher in Tcf21-MP than in Control-MP (p < 0.01). i Cell proliferation assay. Cell proliferation of Tcf21-MP was significantly lower than that of Control-MP at 24, 48 and 72 h (p = 0.04, 0.02, < 0.01, respectively). j–m: Wound assay for cell migration. The images showed at the experimental beginning (j, k) and 24hrs after the beginning (l, m). n The migration ability of TCF21-MP was significantly lower than that of Control-MP (p < 0.01). Statistical analysis was performed with Mann Whitney U test (f, h, i, n).

Fig 5: Mechanism hypothesis of TCF21 expression in injured podocytes of glomerular disease. TCF21 expresses in nucleus of normal podocytes on glomerular capillary (left). By contrast, TCF21 highly expresses in both nucleus and cytoplasm of injured podocytes on glomerular disease including nephrotic syndrome (right). Therefore, TCF21 overexpression may have cellular functional effect in injured podocyte. This image was drawn by MEDICAL EDUCATION INC. (Tokyo, Japan), and allcopyright was assigned to corresponding author, J.U.