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. 2019 Dec 13;9:19037. doi: 10.1038/s41598-019-55442-x

Figure 2.

Figure 2

Urinary culture cells mainly consist of tubular epithelial cells (TEC) and present HLA-ABC and -DR-molecules upon IFNγ and TNFα treatment. (a) Representative flow cytometric characterization of urinary culture cells. Urine was collected from kidney-transplant patients after transplantation and the cell pellet was seeded in culture media after centrifugation. The colonies were then expanded in proliferation media for 1–3 weeks. After harvesting, the cells were stained for epithelial cell marker cytokeratin, proximal and distal renal tubular cell markers CD13 and EpCam, and for fibroblast marker CD90. One representative example of 22 individual donors is demonstrated. (b) Quantification of TEC (cytokeratin+ CD90-) and Fibroblasts (cytokeratin- CD90+ ) among living urinary culture cells (n = 22). Cells were analysed after 3–6 weeks in culture. Statistical comparison was done with Wilcoxon matched-pairs signed rank test for not normally distributed samples. (c) Representative phase-contrast microscopy of two samples of urinary cell cultures. The upper picture shows the characteristic dome formation of TEC (white circle). The cell morphology shown in the lower picture indicates epithelial cells. Magnification 20 × (left) and 10 × (right). A representative example for 22 individual donors is shown. (d–g) Expression levels indicated by median fluorescence intensity (MFI) of HLA-ABC (d,e) and HLA-DR ((f,g) on urine-derived TEC with and without addition of 20 ng/ml IFNγ and 10 ng/ml TNFα for 24 h (n = 18). Harvested cells were analysed by flow cytometry. Histograms are representative for n = 18. Statistical analysis was done with Wilcoxon matched-pairs signed rank test for not normally distributed samples.