Preeclampsia impedes foetal kidney development by delivering placenta-derived exosomes to glomerular endothelial cells

Abstract

Background: Foetal renal dysplasia is still the main cause of adult renal disease. Placenta-derived exosomes are an important communication tool, and they may play an important role in placental (both foetal and maternal) function. We hypothesize that in women with preeclampsia, foetal renal dysplasia is impeded by delivering placenta-derived exosomes to glomerular endothelial cells.

Methods: In the present study, we established a PE trophoblast oxidative stress model to isolate exosomes from supernatants by ultracentrifugation (NO-exo and H/R-exo) and collected normal and PE umbilical cord blood plasma to isolate exosomes by ultracentrifugation combined with sucrose density gradient centrifugation (N-exo and PE-exo), then we investigated their effects on foetal kidney development by in vitro, ex vivo and in vivo models.

Results: The PE trophoblast oxidative stress model was established successfully. After that, in in vitro studies, we found that H/R-exo and PE-exo could adversely affect glomerular endothelial cell proliferation, tubular formation, migration, and barrier functions. In ex vivo studies, H/R-exo and PE-exo both inhibited the growth and branch formation of kidney explants, along with the decrease of VE-cadherin and Occludin. In in vivo studies, we also found that H/R-exo and PE-exo could result in renal dysplasia, reduced glomerular number, and reduced barrier function in foetal mice.

Conclusions: In conclusion, we demonstrated that PE placenta-derived exosomes could lead to foetal renal dysplasia by delivering placenta-derived exosomes to foetal glomerular endothelial cells, which provides a novel understanding of the pathogenesis of foetal renal dysplasia. Video Abstract.

Keywords: Foetal renal dysplasia; Human glomerular endothelial cell; Hypoxia; Placenta-derived exosomes; Preeclampsia; Reoxygenation.

Fig. 1

HIF1-α expression and exosomes identification. A and B Western blot of HIF1-α protein expression. C ELISA of HIF1-α concentration. D TEM image of exosomes, Scale: 100 nm. E NTA results showing the diameters of exosomes. F Western blot showing positive results for the specific placenta-derived exosome markers CD63, TSG101, and PLAP, and negative results of Golgi marker GM130. G Exosomes concentrations of NO-exo, H/R-exo, N-exo and PE-exo. Values are presented as means ± SD, ns, P > 0.05, ***P < 0.001, ****P < 0.0001

Fig. 2

Function of trophoblast-derived exosomes on HGECs. A Confocal microscopy showed that PKH67-labelled exosomes were internalized by HGECs, Scale: 100 μm. B and C EdU staining was used to examine proliferation after exosomes (100 μg/ml) treatment, Scale: 50 μm. D and F The reaction of cultured HGECs to exosomes (100 μg/mL) was examined in the angiogenesis assay, Scale: 200 μm. E and G Transwell experiments were used to examine HGECs migration after incubation with exosomes (100 μg/mL), Scale: 200 μm. H and I The endothelial monolayer barrier function of cells was tested in the model after treatment with exosomes (100 μg/ml). J Western blot analysis of the expression of VE-cadherin and Occludin in exosomes. K qRT-PCR of the expression of VE-cadherin and Occludin in exosomes. Values are presented as means ± SD, ***P < 0.001, ****P < 0.0001

Fig. 3

Function of trophoblast-derived exosomes on kidney explants. A Schematic diagram of the culture process. B Fluorescence microscopy showed that PKH67-labelled exosomes were internalized by kidney explants, Scale: 500 μm. C Microscopy showed kidney explants growth situation, Scale: 200 μm. D Fluorescence microscopy showed kidney explants, Scale: 500 μm. E Kidney explants growth surfaces. F Western blot analysis of the expression of VE-cadherin and Occludin; G, qRT-PCR of the expression of VE-cadherin and Occludin. Values are presented as the means ± SD, **P < 0.01, ***P < 0.001 ****P < 0.0001

Fig. 4

Function of trophoblast-derived exosomes on foetal mouse kidney. A Schematic diagram of culture process. B In vivo imaging system showed that DIR-labelled exosomes were internalized by Fetal mouse. C Appearance of fetal mice. D Foetal mice weight. E Glomerular numbers of foetal mice. F Staining of kidney segmens with PAS, Scale: 50 μm. G IHC of the expression of VE-cadherin, Scale: 20 μm. H IHC of the expression of Occludin, Scale: 20 μm. I Western blot of VE-cadherin and Occludin. J, qRT-PCR of the expression of VE-cadherin and Occludin. Values are presented as means ± SD, **P < 0.01, ***P < 0.001 ****P < 0.0001

Fig. 5

Schematic diagram of the experimental process