PMC

Effect of 8-Br cGMP and DEX on the F-actin architecture in mouse podocytes. The 24-h incubation with 8-Br cGMP resulted in a pronounced disassembly of the transcellular stress bundles, accompanied by the concentration of fibers in cortical regions. In the presence of DEX with or without 8-Br cGMP, F-actin structure remained unaffected and was similar to the structure in the control cells

Blocking of NPR-C receptors with C-ANP_4-23 did not influence the inhibitory effect of DEX on ANP-stimulated cGMP synthesis in mouse podocytes. The cells were preincubated for 24 h with DEX, and C-ANP_4-23 was added 1 min prior to the 15-min stimulation with ANP, as indicated in Materials and Methods. Results are presented as means from four independent experiments ±SEM. *P < 0.05 versus control

a Preincubation of mouse podocytes with 1 µM DEX upregulated the NPR-A mRNA already after 4 h, whereas increase in NPR-C mRNA could be observed after 24 h. Quantitative real-time PCR of NPR levels were normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). b Similarly to mRNA, expression of NPR-A and NPR-C protein increased upon DEX treatment. Flow cytometry results are expressed as the mean fluorescence intensity (MFI). All results are presented as means from three independent experiments ± SEM. *P < 0.05 versus control

Dexamethasone did not influence the inhibitory effect of Ang II on ANP-dependent production of cGMP. Mouse podocytes were preincubated for 24 h in the presence or absence of 1 µM DEX. 0.1 μM ANP and/or 1 μM Ang II were added for 15 min, as described in “.” Results are presented as means from 4 independent experiments ±SEM. **P < 0.01 versus control

The influence of 8-Br cGMP and DEX on mouse podocyte motility. Upper panel scrape-wound assay, DAPI staining. The serum-starved podocytes were incubated for 24 h with 100 µM 8-Br cGMP with or without 1 µM DEX, as described in “” Untreated cells served as the control. Lower panel the results showing that 8-Br cGMP significantly increased the number of migrating podocytes, while DEX inhibited this effect. The data represent the mean ±SEM from three independent experiments performed in duplicate

Ability of mouse (a and b) and rat (c and d) podocytes to produce cGMP varied depending on the time of exposure to 1 µM DEX. The cells cultured in 12-well plates were stimulated with 0.1 µM ANP or 1 µM SNAP for 15 min. a and c cGMP synthesis increased in ANP-stimulated cells after a 4-h preincubation with DEX. b and d Following a 24-h preincubation with DEX, the ANP- dependent cGMP production was suppressed, while SNAP-induced cGMP was elevated, as compared to the controls. Co-incubation of podocytes with ANP and SNAP did not reverse the inhibitory effect of DEX on ANP-induced synthesis of cGMP. The DEX-dependent effects were abolished in the presence of RU486, a specific GR antagonist. Results are presented as means from 5 to 6 independent experiments performed in duplicate ±SEM. *P < 0.05, **P < 0.01, ***P < 0.001