Priming prevent nephrotoxic acute renal failure through stimulation of antioxidant defense mechanism

Duarte, Fernanda; Pessoa, Edson Andrade; Reis, Luciana Aparecida; Schor, Nestor; Borges, Fernanda Teixeira

doi:10.5935/0101-2800.20160025

Fernanda Duarte

Universidade Federal de São Paulo.

Edson Andrade Pessoa

Universidade Federal de São Paulo.

Luciana Aparecida Reis

Universidade Federal de São Paulo.

Nestor Schor

Universidade Federal de São Paulo.

Fernanda Teixeira Borges Correspondence to: Fernanda Teixeira Borges. Universidade Federal de São Paulo. Rua Botucatu, nº 740, São Paulo, Brasil. CEP: 04023-900 E-mail: ft.borgesft@unifesp.br

Universidade Federal de São Paulo.

Universidade Cruzeiro do Sul.

Correspondence to: Fernanda Teixeira Borges. Universidade Federal de São Paulo. Rua Botucatu, nº 740, São Paulo, Brasil. CEP: 04023-900 E-mail: ft.borgesft@unifesp.br

Conflict of interest statement

We have no direct or indirect commercial financial incentive associated with publishing this article. Additionally, the authors have no conflicts of interest, and the source of extra-institutional funding is indicated in the manuscript. All authors participated in the design, interpretation of the study and analysis of the data and review of the manuscript; FD, EAP, LAP conducted the experiments, FD and FTB wrote the manuscript and NS critically read the manuscript.

Figures (6)
Tables (1)

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Figure 1
Viability in vitro. Necrotic (A) and apoptotic (B) cell death in HK-2 cells under control conditions (CTL), cells primed with gentamicin (Genta, 2 mM) for 24 h and again subjected to the antibiotic after 8 days (PI-8d) and 16 days (PI-16d). (*) p < 0.05 compared with CTL. (+) p < 0.05 compared with Genta.

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Figure 2
The ET-1 effect in vitro. ppET-1 expression measured by real-time PCR (A), ET-1 synthesis measured by western blot and graphic quantification (B), cellular proliferation measured by MTT assay (C), and necrotic and apoptotic cell death (D) in HK-2 cells under control conditions (CTL), stimulated with gentamicin (Genta, 2 mM), primed with gentamicin (PI), primed with gentamicin and concomitantly treated with the ET receptor antagonist bosentan (PI+BOS), or treated with bosentan only (BOS, 10 nM). (*) p < 0.05 compared with CTL. (#) p < 0.05 compared with PI and (+) p < 0.05 compared with Genta.

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Figure 3
The HO-1 effect in vitro. HO-1 expression measured by real-time PCR (A), HO-1 synthesis measured by western blot (B), cellular proliferation measured by MTT assay (C), and necrotic and apoptotic cell death (D) in HK-2 cells under control conditions (CTL), stimulated with gentamicin (Genta, 2 mM), primed with gentamicin (PI), primed with gentamicin and concomitantly treated with the HO-1 inducer hemin (PI+HEMIN), or primed with gentamicin and concomitantly treated with the HO-1 inhibitor ZNPP (PI+ZNPP). (*) p < 0.05 compared with CTL. (+)p < 0.05 compared with Genta; (#) p < 0.05 compared with PI.

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Figure 4
Histochemistry analysis. Light microscopy sections (4 µm) of kidney immunohistochemistry staining with Heme-oxygenase 1 (HO-1), catalase (CAT), superoxide dismutase 1 (SOD) and endothelin-1 (ET-1) antibodies in control rats (CTL), animals stimulated with gentamicin during 10 days (GENTA), animals primed with GENTA during 3 days and after one week, stimulated with the antibiotic during 10 days (PI). (*) p < 0.05 compared with the CTL group.

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Figure 5
Antioxidant activity in vivo. (A) Bilirubin concentration in units of µg/ml of protein, (B) SOD activity in units of U/mg of protein, (C) catalase activity in units of U/mg of protein in the kidney, and (D) quantification of the antioxidant fraction of the plasma in mM of control group animals (CTL), animals stimulated with gentamicin for 10 consecutive days (Genta, 40 mg/kg), and animals primed with gentamicin for 3 days and stimulated again with gentamicin for 10 days after one resting week (PI). Baseline (white bars): values of all groups before stimulation, Post-T (black bars): values obtained after stimulation with gentamicin in all groups. (*) p < 0.05 compared with control. (+) p < 0.05 compared with the Genta group.

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Figure 6
Oxidative stress in vivo. (A) Lipid peroxidation in urine indicated by malondialdehyde and expressed in units of nM/mg of protein, (B) hydroperoxides, and (C) carbonylated protein in urine expressed in units of M/creatinine in control group animals (CTL), animals stimulated with gentamicin for 10 consecutive days (Genta, 40 mg/kg), and animals primed with gentamicin for 3 days and stimulated again with gentamicin for 10 days after one resting week (PI). Baseline (white bars): values of all groups before stimulation, Post-T (black bars): values obtained after stimulation with gentamicin in all groups. * p < 0.05 compared with control. + p < 0.05 compared with the Genta group.

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Table 1
Physiological parameters

	CTL		Genta		PI		PI + HEMIN		PI⁺ + p <0,05 vs. PI; BOS
	Baseline	os-T	Baseline	os-T	Baseline	os-T	Baseline	os-T	Baseline	os-T
Cr (mg/dL)	0,39 ± 0,08	0,45 ± 0,07	0,34 ± 0,01	5,09 ± 0,34^* * p < 0.05 vs. CTL;	0,31 ± 0,04	1,55 ± 0,35^# # p <0.05 vs. Genta	0,31 ± 0,04	0,48 ± 0,03^# # p <0.05 vs. Genta ⁺ + p <0,05 vs. PI;	0,53 ± 0,08	0,51 ± 0,04^# # p <0.05 vs. Genta ⁺ + p <0,05 vs. PI;
U (mg/dL)	35,32 ± 2,43	34,28 ± 2,52	34,99 ± 3,22	144,25 ± 1,5^* * p < 0.05 vs. CTL;	35,45 ± 2,57	101,33 ± 2,7^# # p <0.05 vs. Genta	35,42 ± 2,53	45,81 ± 1,62^# # p <0.05 vs. Genta ⁺ + p <0,05 vs. PI;	42,03 ± 6,19	46,53 ± 3,46^# # p <0.05 vs. Genta ⁺ + p <0,05 vs. PI;
uP (mg/24hs)	7,42 ± 0,41	6,81 ± 0,61	9,11 ± 1,12	34,29 ± 3,44^* * p < 0.05 vs. CTL;	4,21 ± 2,32	10,20 ± 2,01^# # p <0.05 vs. Genta	7,2 ± 1,6	17,0 ± 0,9^# # p <0.05 vs. Genta	7,24 ± 1,64	17,01 ± 0,91^# # p <0.05 vs. Genta
FENa (%)	0,43 ± 0,18	0,50 ± 0,11	0,31 ± 0,05	2,97 ± 0,61^* * p < 0.05 vs. CTL;	0,22 ± 0,03	1,46 ± 0,33^# # p <0.05 vs. Genta	0,21 ± 0,06	0,71 ± 0,01^# # p <0.05 vs. Genta ⁺ + p <0,05 vs. PI;	0,30 ± 0,05	0,56 ± 0,08^# # p <0.05 vs. Genta ⁺ + p <0,05 vs. PI;
Uv (ml)	8,15 ± 0,61	8,55 ± 0,73	12,12 ± 1,54	21 ± 2,35^* * p < 0.05 vs. CTL;	5,13 ± 2,30	7,81 ± 1,11^# # p <0.05 vs. Genta	8,82 ± 0,71	13,30 ± 0,82^# # p <0.05 vs. Genta	7,82 ± 1,74	9,62 ± 2,42^# # p <0.05 vs. Genta
BW (g)	224 ± 0,51	245 ± 0,62	224 ± 0,87	241 ± 0,7	233 ± 0,54	251 ± 0,59	226,6 ± 0,57	248,3 ± 0,45	226,6 ± 0,41	246,3 ± 0,74

[1] Correspondence to: Fernanda Teixeira Borges. Universidade Federal de São Paulo. Rua Botucatu, nº 740, São Paulo, Brasil. CEP: 04023-900 E-mail: ft.borgesft@unifesp.br

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Priming prevent nephrotoxic acute renal failure through stimulation of antioxidant defense mechanism

ABSTRACT

Introduction:

Methods:

Results:

Conclusion: