Influence of Angiotensin II on cell viability and apoptosis in rat renal proximal tubular epithelial cells in in vitro studies

Figure 1.. Angiotensin II receptors expression in NRK-52E cells. (a, b) Localization illustrated by immunofluorescence reaction: (a) AT1 receptor—cell membrane (white arrow) and cytoplasm (yellow arrows); (b) AT2 receptor—cell membrane (white arrow), cytoplasm (blue arrow) and nucleus (yellow arrow). Magnification 400×; (c) Representative Western Blot indicated expression of each receptor. Molecular weights of protein bands are denoted on the left side of the images. Lines: 1—control, 2—Ang II 10−9 M, 3—Ang II 10−9 M + LOS 10−6 M, 4—Ang II 10−9 M + PD 10−6 M, 5—Ang II 10−9 M + LOS 10−6 M + PD 10−6 M.

Figure 2.. Cell viability of NRK-52E cells incubated with Ang II 10−9 M at different times (6, 12, 24, 48 h) (a) SRB method, (b) MTT method. Untreated cells were considered as a control. *p < 0.05.

Figure 3.. Cell viability of NRK-52E cells preincubated for 1 h with losartan (LOS) 10−6 M and then exposed to Ang II 10−9 M at different times (6, 12, 24, 48 h) (a) SRB method, (b) MTT method. Cells incubated only with Ang II were considered as a control. *p < 0.05.

Figure 4.. Cell viability of NRK-52E cells preincubated for 1 h with PD123319 (PD) 10−6 M and then exposed to Ang II 10−9 M at different times (6, 12, 24, 48 h) (a) SRB method, (b) MTT method. Cells incubated only with Ang II were considered as a control. *p < 0.05.

Figure 5.. Cell viability of NRK-52E cells preincubated for 1 h with losartan (LOS) 10−6 M and PD123319 (PD) 10−6 M, simultaneously and then exposed to Ang II 10−9 M at different times (6, 12, 24, 48 h) (a) SRB method, (b) MTT method. Cells incubated only with Ang II were considered as a control. *p < 0.05.

Figure 6.. Detection of apoptosis in NRK-52E cells by TUNEL technique (a) Effect of Ang II 10−9 M on proportion of apoptotic cell nuclei (%) in NRK-52E cell line at different times (6, 12, 24, 48 h) (*p < 0.05); (b) Effect of blockade Ang II (AT1 and AT2) receptors in Ang II-treated NRK-52E cells on proportion of apoptotic nuclei at different times (6, 12, 24, 48 h). Cells incubated only with Ang II were considered as a control. *p < 0.05; (c) TUNEL technique detected apoptotic nuclei (brown) in NRK-52E cells after incubation with Ang II 10−9 M. Magnification 600×.

Figure 7.. Caspase-3 activity in NRK-52E cells: (a) after incubation with Ang II 10−9 M at different times (6, 12, 24, 48 h); (b) in control (Ang II 10−9 M) and experimental groups (Ang II 10−9 M + LOS 10−6 M, Ang II 10−9 M + PD 10−6 M, Ang II 10−9 M + LOS 10−6 M + PD 10−6 M) at different times (6, 12, 24, 48 h). *p < 0.05.

Figure 8.. Morphological alterations of NRK-52E cells showed in TEM after incubation with Ang II (10−9 M). (a) fragmentation of nucleus and nuclear chromatin condensation (arrow); (b) cell fragmentation and forming apoptotic bodies (arrow).