Bupropion decreases plasma levels of asymmetric dimethylarginine and ameliorates renal injury by modulation of Ddah1, Oatp4c1, Oct2, and Mate1 in rats with adenine-induced chronic renal injury

Abstract

Objective: The objective of the study was to investigate whether bupropion (BUP) or its circulation metabolites could decrease plasma level of asymmetric dimethylarginine (ADMA) and ameliorate renal injury by modulation of Ddah1, Oatp4c1, Oct2, and Mate1 in rats with adenine-induced chronic renal injury.

Methods: The study initially determined the effect of BUP and its metabolites on cell viability and apoptosis in HK2 cells in the presence and absence of ADMA. Secondly, the study explored whether long-term administration of BUP could reduce the plasma level of ADMA and mitigate renal damage. Thirdly, the expression and activity of Oct2, Ddah1, Mate1 and Oatp4c1 was determined by Western blot and UPLC-MS/MS.

Results: With 0.5 μmol/L ADMA, hydroxybupropion (HBUP, 100 nmol/L), threo-hydrobupropion (TBUP, 10 nmol/L and 1 μmol/L) reduced N-Acetyl-β-D-glucosidase (NAG) level. At 5 μmol/L ADMA, BUP (1 nmol/L-1 μmol/L), HBUP (1-100 nmol/L), and BUP cocktail enhanced survival. At 50 μmol/L ADMA, HBUP (10 nmol/L and 1 μmol/L), TBUP/erythro-hydrobupropion (EBUP) (10-100 nmol/L), and BUP cocktail stimulated survival. EBUP (1 and 100 nmol/L) lowered LDH. BUP (100 nmol/L) and TBUP (1 μmol/L) decreased NAG. TBUP (10 nmol/L, 1 μmol/L) and EBUP (100 nmol/L) inhibited apoptosis. In adenine-induced chronic renal injury rats, long-term administration of BUP significantly decreased the serum concentration of ADMA and creatinine by 12.78% and 38.85%, respectively, ameliorated interstitial lesions and fibrosis and upregulated Ddah1, Oatp4c1, Oct2, Mate1. BUP increased metformin renal clearance without affecting digoxin disposition.

Conclusion: Bupropion moderately decreases plasma levels of ADMA and ameliorates renal injury by modulation of Ddah1, Oatp4c1, Oct2, and Mate1.

Keywords: asymmetrical dimethylarginine; bupropion and its metabolites; chronic renal injury; metabolism enzyme; renal transporters.

FIGURE 1

HK2 cell survival in the presence of ADMA (A), BUP (B), HBUP (C), TBUP (D), EBUP (E) or BUP cocktail (F). Ctrl means normal cell group. BUP cocktail consisted of 15 nmol/L BUP+125 nmol/L HBUP+50 nmol/L EBUP+50 nmol/L TBUP. Data are expressed as mean ± SD (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Data were analyzed by one-way ANOVA.

FIGURE 2

Cell apoptosis with AO/EB staining. (A,B) different concentrations of ADMA; (C–G) 50 μmol/L ADMA + BUP/HBUP/TBUP/EBUP. Ctrl means normal cell group. *: compared with the first column of each panel. Data are expressed as mean ± SD (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001. Data were analyzed by one-way ANOVA.

FIGURE 3

The effect of BUP and BUP metabolites on LDH (A) and NAG (B) in the culture medium and cell lysate. Ctrl means normal cell group. Data are expressed as mean ± SD (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Data were analyzed by one-way ANOVA.

FIGURE 4

The effect of BUP cocktail on LDH (A) and NAG (B) in the culture medium and cell lysate. Ctrl means normal cell group. BUP cocktail consisted of 15 nmol/L BUP+125 nmol/L HBUP+50 nmol/L EBUP+50 nmol/L TBUP. Data are expressed as mean ± SD (n = 3). Data were analyzed by one-way ANOVA.

FIGURE 5

The effects of BUP on Cre, Bun, Alb (A) and ADMA, Cys-c, Kim-1, β2-MG (B) of plasma in renal injury rats. Data are expressed as mean ± SD (n = 5). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Data were analyzed by one-way ANOVA.

FIGURE 6

Effects of BUP on urea cumulative excretion (A) and urea clearance (B) of ADMA, nitrogen and Cre in renal injury rats. Data are expressed as mean ± SD (n = 5). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Data were analyzed by one-way ANOVA.

FIGURE 7

Histopathological evaluation of rat kidney tissue of different groups. Blue circle: tubular atrophy; red circle: interstitial fibrosis.

FIGURE 8

The protein expression of Oatp4c1, Oct2, Ddah1and Mate1 in rat kidney. (A) Long-term administration of BUP and followed by single-dose administration of DIG; (B) Long-term administration of BUP and followed by single-dose administration of drug cocktail. Cocktail consisted of a single i.v. dose of MET at 5 mg/kg, a single i.v. dose of FUR at 4 mg/kg and a single p.o. dose of RSV at 25 mg/kg. Data are expressed as mean ± SD (n = 5). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Data were analyzed by one-way ANOVA.

FIGURE 9

Mean plasma concentration-time profiles of DIG (A) and MET (B). Cocktail consisted of a single i.v. dose of MET at 5 mg/kg, a single i.v. dose of FUR at 4 mg/kg and a single p.o. dose of RSV at 25 mg/kg. Data are expressed as mean ± SD (n = 5).

FIGURE 10

Mean plasma concentration-time profiles of BUP, HBUP, TBUP and EBUP. Data are expressed as mean ± SD (n = 5).

FIGURE 11

Proposed mechanism (By Figdraw 2.0). At 50 μmol/L ADMA, HBUP (10 nmol/L and 1 μmol/L), TBUP/EBUP (10–100 nmol/L), and BUP cocktail stimulated survival in HK2 cells. EBUP (1/100 nmol/L) lowered LDH. BUP (100 nmol/L) and TBUP (1 μmol/L) decreased NAG. Long term administration of BUP and its metabolites moderately decrease the serum level of ADMA by modulating the Oct2, Mate1, and Oatp4c1-mediated renal transport of ADMA as well as Ddah1-mediated metabolism, which alleviates the exacerbation of ADMA and retards the progression of renal interstitial lesions and fibrosis. ADMA: Asymmetrical Dimethylarginine. BUP: Bupropion. Ddah1: Dimethylarginine dimethylaminohydrolase1. LDH: Lactate dehydrogenase. Mate1: Mammalian multidrug and toxin extrusion protein 1. NAG: N-Acetyl-β-D-glucosidase. Oatp4c1: Organic anion-transporting polypeptide 4c1. Oct2: Organic cation transporter2.