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. 2022 Nov 17;324(1):F106–F123. doi: 10.1152/ajprenal.00149.2022

Figure 1.

Figure 1.

Knockin of green fluorescent protein (GFP) into one allele of fetal liver kinase 1 (Flk1), a gene encoding vascular endothelial growth factor (VEGF) receptor type 2 (VEGFR2), caused VEGFR2 haploinsufficiency and peritubular capillary rarefaction in the kidney. A: schema for heterozygous knockin of GFP into the VEGFR2 locus. B: homozygous knockin of GFP into the VEGFR2 locus led to embryonic lethality. PCR genotyping was carried out at weaning. The kidneys were harvested from wild-type (WT) mice (without knockin of GFP into the flk1 locus, i.e., VEGFR2+/+) and mice with heterozygous knockin of GFP into the flk1 locus (VEGFR2+/−). Mice were fed with normal phosphate chow at 6 wk of age. C: immunofluorescent images for endothelial marker (PE; red), collagen type IV (Col IV; blue), and exogenous inserted gene (GFP; green) in kidney sections. C, left: representative immunofluorescent images. Scale bar = 50 µm. C, right: quantitative analysis of PE staining density on kidney sections with ImageJ software. D: immunoblot analysis for endothelial marker (CD31), VEGFR2, VEGFA, GFP, and β-actin in total kidney lysates. D, left: representative immunoblots. D, right: quantitative analysis of all experiments from two groups. E: quantitative analysis of mRNA of CD31, VEGFR2, and VEGFA in the kidneys with quantitative PCR. F: immunoblot analysis for phosphorylated (p-)Erk and p-Akt as well as total (T-)Erk and Akt in total kidney lysates. F, left: representative blots. F, right: quantitative analysis of all immunoblots from two groups. C–F: quantitative data are expressed with scatterplots of individual data points (open circles indicate male mice and pink circles indicate female mice) and means ± SD (bars and errors) of all mice from each group. Statistical significance was evaluated by an unpaired Student’s t test, and significance was accepted when **P < 0.01 between two groups. The sample number in each group is presented in parentheses underneath each corresponding bar. PE, a cocktail of primary antibodies against platelet-endothelial cell adhesion molecule-1 and endomucin; VR2, VEGFR2.