Fig. 4

RFP-GFP-LC3 adenovirus which labeled LC3 was transfected into human podocytes (HPCs), and representative images of immunofluorescence were detected by laser scanning confocal microscope. Cells were counterstained with 4,6-diamidino-2-phenylindole (DAPI; blue). Autophagosomes (GFP+RFP+LC3 puncta) were labelled with yellow (merge from green and red). The zoom area enlarges representative images of human podocytes. Arrowheads indicate autophagosomes in human podocytes. Original magnification 400×. By detecting and analyzing the number of yellow dots, autophagic activity can be measured. Numbers of yellow dots in 30 random HPCs under transmission electron microscope were counted. All statistical analyses were performed by using SPSS version 21.0 (SPSS Inc., Chicago, IL, USA) and GraphPad Prism 6 (GraphPad Software, Inc., La Jolla, CA, USA) for macOS. A P value of less than 0.05 was considered statistically significant. Data were expressed as mean ± standard deviation and median with range or frequency, as appropriate. Intergroup comparisons were made by using Pearson chi-squared test for categorical variables and Student’s t test, Mann–Whitney U test, or Wilcoxon test for continuous variables, as appropriate. *P <0.05 versus control (CON) group; #P<0.05 versus immunoglobulin G (IgG) group (n = 30)