Original article
Chronic impairment of mitochondrial bioenergetics and β-oxidation promotes experimental AKI-to-CKD transition induced by folic acid

https://doi.org/10.1016/j.freeradbiomed.2020.04.016Get rights and content

Highlights

  • AKI-to-CKD transaction induced by folic acid is related to mitochondrial dysfunction.

  • Mitochondrial β-oxidation is dysfunctional during the AKI-to-CKD transaction.

  • Prevention of mitochondrial dysfunction during AKI can prevent the transition to CKD.

  • NAC administration before AKI prevents the mitochondrial function deterioration and CKD development.

Abstract

Recent studies suggest that mitochondrial bioenergetics and oxidative stress alterations may be common mechanisms involved in the progression of renal damage. However, the evolution of the mitochondrial alterations over time and the possible effects that their prevention could have in the progression of renal damage are not clear. Folic acid (FA)-induced kidney damage is a widely used experimental model to induce acute kidney injury (AKI), which can evolve to chronic kidney disease (CKD). Therefore, it has been extensively applied to study the mechanisms involved in AKI-to-CKD transition. We previously demonstrated that one day after FA administration, N-acetyl-cysteine (NAC) pre-administration prevented the development of AKI induced by FA. Such therapeutic effect was related to mitochondrial preservation. In the present study, we characterized the temporal course of mitochondrial bioenergetics and redox state alterations along the progression of renal damage induced by FA. Mitochondrial function was studied at different time points and showed a sustained impairment in oxidative phosphorylation capacity and a decrease in β-oxidation, decoupling, mitochondrial membrane potential depolarization and a pro-oxidative state, attributed to the reduction in activity of complexes I and III and mitochondrial cristae effacement, thus favoring the transition from AKI to CKD. Furthermore, the mitochondrial protection by NAC administration before AKI prevented not only the long-term deterioration of mitochondrial function at the chronic stage, but also CKD development. Taken together, our results support the idea that the prevention of mitochondrial dysfunction during an AKI event can be a useful strategy to prevent the transition to CKD.

Introduction

The term chronic kidney disease (CKD) describes a group of pathologies characterized by the progressive loss of renal function, resulting in a progressive decrement in the glomerular filtration rate (GFR), with or without an increment in the conventional clinical renal damage markers, like creatinine and blood urea nitrogen (BUN) [1,2]. In recent years, the number of patients with CKD has increased dramatically [[3], [4], [5], [6], [7]], representing a growing worldwide public health problem [8]. As a result of the lack of understanding of the complex pathological mechanisms involved in CKD development [9], the current clinical strategies and treatments do not significantly improve kidney function or prevent illness progression [10], thus highlighting the urgent need to investigate and understand the mechanisms involved in the generation and progression of CKD [11,12].

Furthermore, dysfunction in mitochondrial bioenergetics and redox state have recently emerged as key components of the sudden deterioration of renal functions in a relatively short time-interval, known as acute kidney injury (AKI). In fact, mitochondrial dysfunction in AKI is involved in tubular dysfunction, cell death, oxidative stress and inflammation [[13], [14], [15], [16], [17], [18], [19]]. Likewise, in both animal models and patients, a severe episode of AKI, or a series of them, results in CKD development [2,[20], [21], [22]]. The growing evidence suggests that renal mitochondrial alterations [9,23,24] favor hemodynamic alterations, Ca2+ deregulation, cell death, inflammation, fibrotic processes and epithelial-to-mesenchymal transition (EMT), which participate in the progression of several types of CKD in both non-diabetic and diabetic contexts [9,[22], [23], [24], [25], [26], [27]]. However, currently there is no information about the mitochondrial bioenergetics and redox state alterations over a time-course. Also, the possible effects of their attenuation in the progression of renal damage are not clear.

In this context, folic acid (FA)-induced kidney damage is a widely recurred to experimental model for AKI induction [28,29]. This model also induces chronic processes such as the persistence of cell death, EMT, cytokines release, inflammation and fibrosis, which lead to CKD transition usually in less than 28 days [28,30,31]. Furthermore, it recreates the AKI pathology reported in the clinic [32] and is highly reproducible [28,29]. Together, these make the FA model good for the exploration of the mechanisms involved in AKI-to-CKD transition [28,30,31]. In the present work, we refer to “the acute stage” as the period that begins with the administration of FA and ends 72 h later, bringing about the AKI-to-CKD transition or “the chronic stage”, which encompasses the time period between day 4 until day 28. It is important to highlight that although high doses of FA have pathological effects on kidney function [28,30,31], nephroprotective effects and the reduction of cardiovascular risk have also been reported by lower FA doses in CKD patients with elevated homocysteine levels, but with baseline vitamin B12 levels [33,34]. In fact, vitamin B12 and the metabolite of FA, 5-methyltetrahydrofolate (5-THF), act as cofactors of the enzyme methionine synthetase, that catalyzes the transfer to the methyl group from 5-THF to homocysteine, producing methionine [33]. Therefore, the protection in this context may be related to reduced homocysteine levels mediated by FA and vitamin B12 [[35], [36], [37]]. Additionally, it must be taken into account that the FA dose used in these papers (<10 mg/day) is much lower compared to the dose used in the FA-induced renal damage models (>250 mg/day) and it is well documented that FA at higher levels triggers prooxidant and proinflammatory states in kidneys [38,39]. So, a protective or prooxidant effect of FA could depend on the dose, the administration route and the preexistent kidney damage conditions.

In the case of renal damage induced by high levels of FA, the redox imbalance generated by FA metabolism is one of the main mechanisms involved in the genesis of the pathology [13,38,40], because FA metabolism requires higher levels of NADPH to reduce folate to THF [41,42], decreasing the antioxidant defense [13,38]. This redox imbalance is particularly harmful for kidney's mitochondria, due to the high internalization rates of folate in kidney [38,43,44] and because mitochondria store approximately 40% of folates [43,45]. Therefore, mitochondria are especially vulnerable to high doses of FA [13,46]. Additionally, the mitochondrial redox imbalance can be directly related to mitochondrial bioenergetics alterations [47]. In fact, we recently demonstrated that at 24 h after its administration FA induced in kidney mitochondria the decrease in glutathione (GSH) levels, the increase in glutathione disulfide (GSSG) and the decrease in total glutathione content (GSH + GSSG), that together with higher S-glutathionylation removal activity of mitochondrial glutaredoxin (Grx), leads to complex I (CI) dysfunction [13]. Furthermore, CI dysfunction induced by FA triggers the decrease in mitochondrial oxidative phosphorylation (OXPHOS) capacity, mitochondrial decoupling, loss of mitochondrial membrane potential (ΔΨm), increased oxidative stress and mitochondrial hydrogen peroxide (H2O2) production. Such effects were associated with AKI development [13]. Additionally, we also demonstrated that the pretreatment with N-acetyl-cysteine (NAC), a donor of sulfhydryl groups and a precursor of GSH [48,49] prevented the mitochondrial bioenergetics and redox state impairment, which prevented the development of AKI 24 h after FA administration [13].

The progressive nature of the renal disease induced by FA was further evidenced by the reduction in mRNA levels of mitochondrial electron transport system (ETS) components in the days 2–14 after FA administration [39,46], the persistence of oxidative stress in kidneys after FA administration [38,50,51] and the inflammatory and fibrotic processes [[52], [53], [54], [55]]. Interestingly, inflammatory and fibrotic processes have been related to mitochondrial impairment [[56], [57], [58]], suggesting that mitochondrial bioenergetics and redox state impairment is persistent in AKI-to-CKD transition in this model. However, such hypothesis has not yet been evaluated.

Therefore, in the present study, we characterize for the first time, the time course of mitochondrial bioenergetics and redox state alterations and their possible association to changes in mitochondrial mass and biogenesis in the AKI-to-CKD transition induced by FA. We also assess the effects of NAC pre-administration in the temporal evolution of mitochondrial bioenergetics and redox state alterations. Lastly, we evaluate if the prevention of mitochondrial dysfunction by NAC during AKI prevents transition to CKD in the FA model.

Section snippets

Reagents

Adenosine 5′-diphosphate sodium salt (ADP), amplex red, antimycin A, l-arginine, fat free bovine serum albumin (BSA), bromophenol blue, β-mercaptoethanol, carbonyl cyanide m-chlorophenylhydrazone (CCCP), cytochrome c from equine heart, D-(+)-glucose, D-mannitol, decylubiquinone (DUB), 2,6-dichlorophenolindophenol sodium salt hydrate (DCPIP), 5,5′-dithio-bis-(2-nitrobenzoic acid) (DTNB), ethylene glycol-bis(2-aminoethylether)-N,N,N′,N'-tetraacetic acid (EGTA), glycerol, GSH, glucose-6-phosphate

FA induces renal damage progression

Our first goal was to characterize the kidney damage progression induced by FA. Therefore, we evaluated the levels of the classic renal damage markers BUN and creatinine in plasma. As shown in Fig. 1A–C, at day 2 after its administration, FA induced a remarkable increase in BUN and creatinine plasma levels, as well as in the kidney weight/rat weight ratio compared to the control group. These alterations were prevented at day 2 by the pre-administration of NAC (Fig. 1A–C). Furthermore,

Discussion

The administration of high levels of FA leads to an increase of its metabolized product (tetrahydrofolate) in blood [42,43], which is then transported inside the cell by the reduced folate carrier exchanger and by the high-affinity folate receptor [42,43,45,68]. In the case of the kidney, folate metabolites accumulate in higher concentrations than in other tissues due to the expression of the high-affinity folate receptor in this organ, especially in PT cells [43]. The folate metabolites are

Conclusion

FA induces an acute stage of renal damage alterations in mitochondrial β-oxidation and CI-linked respiration. Although in a chronic stage of renal damage there is partial recovery in CI-linked respiration parameters, the mitochondrial β-oxidation alterations persist over time, triggering a permanent reduction in ATP production and mitochondrial decoupling, favoring the progression to CKD.

Finally, the renal mitochondrial preservation that results from NAC pre-treatment in FA-induced AKI may be a

Declaration of competing interestCOI

The authors report no conflict of interests.

Acknowledgements

Research conducted for this publication was supported by grants from “Consejo Nacional de Ciencia y Tecnología” (CONACyT, A1-S-7495 and 281967), Programa de Apoyo a Proyectos de Investigación e Innovación Tecnológica (PAPIIT, #IN202219) of Universidad Nacional Autónoma de México (UNAM), Programa de Apoyo a la Investigación y el Posgrado (PAIP, 5000-9105), and Fondos del Gasto Directo autorizado a la Subdirección de Investigación Básica from Instituto Nacional de Cardiología Ignacio Chávez to ET

References (97)

  • P. Rojas-Morales et al.

    Fasting reduces oxidative stress, mitochondrial dysfunction and fibrosis induced by renal ischemia-reperfusion injury

    Free Radic. Biol. Med.

    (2019)
  • A.M. El Nahas et al.

    Chronic kidney disease: the global challenge

    Lancet

    (2005)
  • O.E. Aparicio-Trejo et al.

    Mitochondrial bioenergetics, redox state, dynamics and turnover alterations in renal mass reduction models of chronic kidney diseases and their possible implications in the progression of this illness

    Pharmacol. Res.

    (2018)
  • B. Benipal et al.

    Influence of renal compensatory hypertrophy on mitochondrial energetics and redox status

    Biochem. Pharmacol.

    (2011)
  • M. Tamaki et al.

    Chronic kidney disease reduces muscle mitochondria and exercise endurance and its exacerbation by dietary protein through inactivation of pyruvate dehydrogenase

    Kidney Int.

    (2014)
  • A. Ortiz et al.

    Translational value of animal models of kidney failure

    Eur. J. Pharmacol.

    (2015)
  • C.M. Wyatt et al.

    Folic acid supplementation and chronic kidney disease progression

    Kidney Int.

    (2016)
  • C.M. Wyatt et al.

    Folic acid supplementation and chronic kidney disease progression

    Kidney Int.

    (2016)
  • A. Gupta et al.

    Folic acid induces acute renal failure (ARF) by enhancing renal prooxidant state

    Exp. Toxicol. Pathol.

    (2012)
  • O. Ruiz-Andres et al.

    The inflammatory cytokine TWEAK decreases PGC-1α expression and mitochondrial function in acute kidney injury

    Kidney Int.

    (2016)
  • G.S. Ducker et al.

    One-carbon metabolism in health and disease

    Cell Metabol.

    (2017)
  • F.H. Nazki et al.

    Folate: metabolism, genes, polymorphisms and the associated diseases

    Gene

    (2014)
  • J.T. Fox et al.

    Folate-mediated one-carbon metabolism

    Vitam. Horm.

    (2008)
  • L.J. Stallons et al.

    Suppressed mitochondrial biogenesis in folic acid-induced acute kidney injury and early fibrosis

    Toxicol. Lett.

    (2014)
  • R.J. Mailloux et al.

    Redox regulation of mitochondrial function with emphasis on cysteine oxidation reactions

    Redox Biol.

    (2014)
  • E.Y. Plotnikov et al.

    The role of mitochondria in oxidative and nitrosative stress during ischemia/reperfusion in the rat kidney

    Kidney Int.

    (2007)
  • A. Ortiz et al.

    Expression of apoptosis regulatory proteins in tubular epithelium stressed in culture or following acute renal failure

    Kidney Int.

    (2000)
  • K. Doi et al.

    Attenuation of folic acid-induced renal inflammatory injury in platelet-activating factor receptor-deficient mice

    Am. J. Pathol.

    (2006)
  • H. Zhao et al.

    Role of mitochondrial dysfunction in renal fibrosis promoted by hypochlorite-modified albumin in a remnant kidney model and protective effects of antioxidant peptide SS-31

    Eur. J. Pharmacol.

    (2017)
  • Y. Yuan et al.

    Mitochondrial dysfunction accounts for aldosterone-induced epithelial-to-mesenchymal transition of renal proximal tubular epithelial cells

    Free Radic. Biol. Med.

    (2012)
  • T. Sekine et al.

    Solute Transport, Energy consumption, and production in the kidney

  • O.E. Aparicio-Trejo et al.

    Mitochondrial bioenergetics, redox state, dynamics and turnover alterations in renal mass reduction models of chronic kidney diseases and their possible implications in the progression of this illness

    Pharmacol. Res.

    (2018)
  • C.D. Chancy et al.

    Expression and differential polarization of the reduced-folate transporter-1 and the folate receptor α in mammalian retinal pigment epithelium

    J. Biol. Chem.

    (2000)
  • L.S. Chawla et al.

    The severity of acute kidney injury predicts progression to chronic kidney disease

    Kidney Int.

    (2011)
  • P.T. Kang et al.

    Protein thiyl radical mediates S-glutathionylation of complex i

    Free Radic. Biol. Med.

    (2012)
  • M. Forkink et al.

    Complex I and complex III inhibition specifically increase cytosolic hydrogen peroxide levels without inducing oxidative stress in HEK293 cells

    Redox Biol.

    (2015)
  • R.P. Brandes et al.

    Nox family NADPH oxidases: molecular mechanisms of activation

    Free Radic. Biol. Med.

    (2014)
  • A.M. Garrido et al.

    NADPH oxidases and angiotensin II receptor signaling

    Mol. Cell. Endocrinol.

    (2009)
  • F. Jiang et al.

    NADPH oxidase-dependent redox signaling in TGF-β-mediated fibrotic responses

    Redox Biol.

    (2014)
  • H.H. Szeto et al.

    Protection of mitochondria prevents high-fat diet induced glomerulopathy and proximal tubular injury

    Kidney Int.

    (2016)
  • S. Hallan et al.

    Metabolomics and gene expression analysis reveal down-regulation of the citric acid (TCA) cycle in non-diabetic CKD patients

    EBioMed.

    (2017)
  • A. Voulgaris et al.

    Chronic kidney disease in patients with obstructive sleep apnea. A narrative review

    Sleep Med. Rev.

    (2019)
  • C. Li et al.

    N-acetylcysteine ameliorates cisplatin-induced renal senescence and renal interstitial fibrosis through sirtuin1 activation and p53 deacetylation

    Free Radic. Biol. Med.

    (2019)
  • C.Y. Liao et al.

    Protective effect of N-acetylcysteine on progression to end-stage renal disease: necessity for prospective clinical trial

    Eur. J. Intern. Med.

    (2017)
  • L. Xiao et al.

    The mitochondria-targeted antioxidant MitoQ ameliorated tubular injury mediated by mitophagy in diabetic kidney disease via Nrf2/PINK1

    Redox Biol.

    (2017)
  • O. Akinlolu

    Addressing the global burden of chronic kidney disease through clinical and translational research

    Trans. Am. Clin. Climatol. Assoc.

    (2014)
  • J.M. Forbes et al.

    Mitochondrial dysfunction in diabetic kidney disease

    Nat. Rev. Nephrol.

    (2018)
  • B. Feng et al.

    Silymarin protects against renal injury through normalization of lipid metabolism and mitochondrial biogenesis in high fat-fed mice

    Free Radic. Biol. Med.

    (2017)
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