Effect of prednisolone on glyoxalase 1 in an inbred mouse model of aristolochic acid nephropathy using a proteomics method with fluorogenic derivatization-liquid chromatography-tandem mass spectrometry

Abstract

Prednisolone is involved in glucose homeostasis and has been used for treatment for aristolochic acid (AA) nephropathy (AAN), but its effect on glycolysis in kidney has not yet been clarified. This study aims to investigate the effect in terms of altered proteins after prednisolone treatment in a mice model of AAN using a proteomics technique. The six-week C3H/He female mice were administrated AA (0.5 mg/kg/day) for 56 days. AA+P group mice were then given prednisolone (2 mg/kg/day) via oral gavage for the next 14 days, and AA group mice were fed water instead. The tubulointerstitial damage was improved after prednisolone treatment comparing to that of AA group. Kidney homogenates were harvested to perform the proteomics analysis with fluorogenic derivatization-liquid chromatography-tandem mass spectrometry method (FD-LC-MS/MS). On the other hand, urinary methylglyoxal and D-lactate levels were determined by high performance liquid chromatography with fluorescence detection. There were 47 altered peaks and 39 corresponding proteins on day 14 among the groups, and the glycolysis-related proteins, especially glyoxalase 1 (GLO1), fructosebisphosphate aldolase B (aldolase B), and triosephosphate isomerase (TPI), decreased in the AA+P group. Meanwhile, prednisolone decreased the urinary amount of methylglyoxal (AA+P: 2.004 ± 0.301 μg vs. AA: 2.741 ± 0.630 μg, p < 0.05), which was accompanied with decrease in urinary amount of D-lactate (AA+P: 54.07 ± 5.45 μmol vs. AA: 86.09 ± 8.44 μmol, p < 0.05). Prednisolone thus alleviated inflammation and interstitial renal fibrosis. The renal protective mechanism might be associated with down-regulation of GLO1 via reducing the contents of methylglyoxal derived from glycolysis. With the aid of proteomics analysis and the determination of methylglyoxal and its metabolite-D-lactate, we have demonstrated for the first time the biochemical efficacy of prednisolone, and urinary methylglyoxal and its metabolite-D-lactate might be potential biomarkers for AAN.

Evaluation of the nephrotoxicity and safety of low-dose aristolochic acid, extending to the use of Xixin (Asurum), by determination of methylglyoxal and D-lactate

“…All animal protocols were approved by the Animal Care and Use Committee of Taipei Medical University (LAC-2019-0482). Six-week-old female C3H/He mice were obtained from the National Laboratory Animal Breeding and Research Center (Taipei, Taiwan) and randomly allocated to either the normal group (N) or the AA group (n = 24/group) [20]. Six mice in the same group were placed in per cage.…”

“…The AA group were given ad libitum access to distilled water containing 3 µg/mL AA (Sigma-Aldrich, Inc., MO, USA; AAI:AAII = 63:31; 0.5 mg/kg/day) as drinking water for 56 days. After day 56, the AA group mice received normal drinking water [20,21]. The N group received only normal drinking water.…”

“…The mice were humanely sacrificed on day 28, 56, 70 or 84 (n = 6/group). Whole blood samples were collected and the kidneys were excised [20,21].…”

“…Serum creatinine (Scr) was determined using a modified version of the previous protocol [14,22]. Briefly, 20 µL blood samples were mixed with 10 µL of 1 mM [20,21]. Sections were observed at a magnification of 200× using a G-330 light microscope (Optima, New Taipei City, Taiwan) and images were captured using a Nikon Coolpix 4500 camera (Nikon, Tokyo, Japan) [20,21].…”

“…On the other hand, renal injury increases both D-lactate and L-lactate, while prednisolone [18] or metformin [10] treatment alleviate histological damage and decrease D-lactate -rather than L-lactate or D/L-lactate-in AAN models. Nevertheless, the changes in methylglyoxal and D-lactate contents after long-term administration of AA have not been explored, although elevation of urinary D-lactate was observed in a previous study [20].…”