Figure 7. Cystic epithelia in human PKD... | Rockefeller University Press

Figure 7.

$Figure 7. Cystic epithelia in human PKD display CA and ciliogenesis defects. (A) H&E staining of normal and ADPKD kidney sections showing the typical architecture of renal tubules, compared with small and large cysts in PKD samples, respectively. (B) Immunofluorescence staining of normal and PKD kidney specimens with antibodies against γ-tubulin or centrin to identify centrosomes, and DAPI to mark nuclei. Normal kidney epithelia predominantly contained the normal complement of centrosomes (one to two foci), whereas a significant fraction of cystic epithelia contained greater than two foci per cell. Cystic tubules of PKD patients display both low (mild) and high (severe) levels of CA. White lines in normal kidneys outline a typical nephron tubule, and cysts of different sizes in PKD samples. Bars (insets), 2 µm. (C) Normal and PKD kidney sections were immunostained for centrosomes and cilia. Normal kidney cells contain one centrosome and assemble a single primary cilium (inset magnified twofold). Cystic cells containing excess centrosomes either fail to assembly a cilium (aciliated) or form more than one cilium (superciliated). Nuclei were stained with DAPI. Bars (insets), 5 µm. (D) Representative image of cystic cells in PKD tissue captured using superresolution SIM, highlighting the presence of more than one cilium in cells with CA. (E) Quantification of the percentage of cells with CA relative to cyst size. Centrosomes were marked with γ-tubulin, and centrosome number quantified per nucleus in normal nephron epithelia (control) and epithelial cells lining the cyst walls (PKD). n = 961 cells from 80 tubules analyzed from four normal kidneys, 501 cells from 40 small cysts (0–100 µm), and 1,423 cells from 37 large cysts (>100 µm) in seven PKD patient samples. Box plots represent the median, maximum, and minimum values for each dataset. One-way ANOVA was performed to determine statistically significant differences between samples (*, P < 0.05). (F) Quantification of ciliary defects in cystic epithelia of PKD patients. The percentage of cells with zero, one, or more than one cilium was determined from cyst-lining cells that contained either normal centrosome number, compared with cells with CA. n = 1,521 cells scored from eight PKD kidneys. Statistical significance was calculated using a two-tailed unpaired t test (*, P < 0.05).$

Cystic epithelia in human PKD display CA and ciliogenesis defects. (A) H&E staining of normal and ADPKD kidney sections showing the typical architecture of renal tubules, compared with small and large cysts in PKD samples, respectively. (B) Immunofluorescence staining of normal and PKD kidney specimens with antibodies against γ-tubulin or centrin to identify centrosomes, and DAPI to mark nuclei. Normal kidney epithelia predominantly contained the normal complement of centrosomes (one to two foci), whereas a significant fraction of cystic epithelia contained greater than two foci per cell. Cystic tubules of PKD patients display both low (mild) and high (severe) levels of CA. White lines in normal kidneys outline a typical nephron tubule, and cysts of different sizes in PKD samples. Bars (insets), 2 µm. (C) Normal and PKD kidney sections were immunostained for centrosomes and cilia. Normal kidney cells contain one centrosome and assemble a single primary cilium (inset magnified twofold). Cystic cells containing excess centrosomes either fail to assembly a cilium (aciliated) or form more than one cilium (superciliated). Nuclei were stained with DAPI. Bars (insets), 5 µm. (D) Representative image of cystic cells in PKD tissue captured using superresolution SIM, highlighting the presence of more than one cilium in cells with CA. (E) Quantification of the percentage of cells with CA relative to cyst size. Centrosomes were marked with γ-tubulin, and centrosome number quantified per nucleus in normal nephron epithelia (control) and epithelial cells lining the cyst walls (PKD). n = 961 cells from 80 tubules analyzed from four normal kidneys, 501 cells from 40 small cysts (0–100 µm), and 1,423 cells from 37 large cysts (>100 µm) in seven PKD patient samples. Box plots represent the median, maximum, and minimum values for each dataset. One-way ANOVA was performed to determine statistically significant differences between samples (*, P < 0.05). (F) Quantification of ciliary defects in cystic epithelia of PKD patients. The percentage of cells with zero, one, or more than one cilium was determined from cyst-lining cells that contained either normal centrosome number, compared with cells with CA. n = 1,521 cells scored from eight PKD kidneys. Statistical significance was calculated using a two-tailed unpaired t test (*, P < 0.05).