Mitochondrial quality control in kidney injury and repair

Mitochondria regulate a variety of cellular processes that are closely associated with tissue injury and repair, such as cell death, cell proliferation and differentiation, metabolic adaptation and inflammation ( FIG. 1 ). Cell death. Mitochondria regulate various forms of cell death, including apoptosis and necrosis. Apoptosis is a regulated cell death process that can occur via intrinsic or extrinsic pathways. Intrinsic apoptosis is initiated by mitochondrial outer membrane permeabilization (MOMP), which releases pro-apoptotic factors such as cytochrome c and second mitochondria-derived activator of caspase (SMAC; also known as Diablo homologue, mitochondrial) from the intermembrane space (IMS) into the cytosol, where they activate caspase 9 and subsequent executioner caspases 10 . Under physiological conditions, cytochrome c either exists in its free form in the IMS or is anchored in the inner mitochondrial membrane (IMM) through binding to cardiolipin and acts as an electron carrier between respiratory complexes III and IV in the electron transport chain (ETC) 11 , 12 . Upon mitochondrial damage, ROS accumulation results in conversion of cytochrome c into a peroxidase that oxidizes cardiolipin, ultimately leading to the release of cytochrome c from the IMS into the cytosol, caspase activation and apoptosis 11 , 12 . Electron transport chain (ETC). A series of four protein complexes (complex I–IV) embedded in the inner mitochondrial membrane that transfer electrons from electron donors to electron acceptors via redox reactions. This process drives the transfer of protons across the inner mitochondrial membrane to produce ATP. MOMP can result from the pore-forming activity of pro-apoptotic members of the BCL-2 protein family, such as the apoptosis regulators BAX (also known as BCL-2-like protein 4) and BAK1 (also known as BCL-2 homologous antagonist/killer) or from high amplitude swelling of the matrix owing to increased permeability of the IMM to small solutes, a phenomenon termed mitochondrial permeability transition 13 . Extrinsic apoptosis occurs in response to perturbations of the extracellular microenvironment that are detected and relayed by plasma membrane receptors (also known as death receptors), resulting in activation of caspase 8 and downstream executioner caspases 10 . The intrinsic pathway of apoptosis may be linked to the extrinsic pathway through caspase 8-mediated cleavage of BH3-interacting domain death agonist (BID) and consequent translocation of the truncated BID to mitochondria, which induces MOMP, providing an amplification loop for apoptosis 14 . Necrosis was traditionally classified as a passive, unregulated cell death process that is characterized morphologically by cell swelling and plasma membrane rupture. However, various forms of regulated necrosis have now been identified. One such form, necroptosis, is critically dependent on receptor-interacting serine-threonine kinase 1/3 (RIPK1/3)-mediated activation of mixed line-age kinase domain-like protein (MLKL) 10 . Mitochondrial ROS (mtROS) and mtDNA have been shown to induce necroptosis. In tumour necrosis factor (TNF)-induced necroptosis, mtROS-driven autophosphorylation of RIPK1 is essential for RIPK3 recruitment into the necrosome to induce necroptosis 15 . In addition, TNF-induced release of mtDNA into the cytosol can activate DNA sensors to enhance RIPK3–MLKL-dependent necroptosis 16 . Mitochondrial permeability transition is typically associated with necrosis 17 . Mitochondria also regulate pyroptosis, which depends on the formation of plasma membrane pores by members of the gasdermin protein family and often occurs as a consequence of inflammatory caspase activation 10 , 18 . In addition, mitochondria may have a role in ferroptosis, a regulated form of necrosis that is initiated by iron-dependent lipid peroxidation and controlled by phospholipid hydroperoxide glutathione peroxidase (GPX4) 10 , 19 . Animal studies have demonstrated an involvement of apoptosis, pyroptosis and ferroptosis in AKI 20 – 23 . However, the role of mitochondria in various forms of regulated necrosis during kidney injury and repair remains unclear.

Inflammation is a complex biological response that is essential for repairing tissue after injury. However, chronic inflammation is associated with a range of diseases, including CKD and diabetes mellitus 28 . Mitochondria act as a central hub of pro-inflammatory signalling. mtROS are activators of inflammation that promote pro-inflammatory gene expression, activating NF-κB signalling and the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome 29 , 30 . Damage-associated molecular patterns derived from mitochondria, including formyl peptides and mtDNA, can bind to Toll-like receptors or nucleotide-binding oligomerization domain-containing protein (NOD)-like receptors, leading to inflammation 31 . Mitochondria also have roles in controlling the development, activation, differentiation and survival of diverse immune cell types, including T lymphocytes 32 . Several studies have demonstrated an important role of mitochondria in the regulation of inflammation during kidney injury. In mouse models of cisplatin-induced AKI and in cultured tubular cells exposed to cisplatin, mtDNA from damaged mitochondria in kidney tubular cells leaked into the cytosol, probably through BAX-activated pores in the outer mitochondrial membrane (OMM), resulting in activation of the cyclic GMP–AMP synthase (cGAS)–stimulator of interferon genes protein (STING) cytosolic DNA sensing pathway and thereby triggering kidney inflammation and AKI progression 33 . Similarly, in mice with defective mitochondria in kidney tubule cells owing to tubule-specific knockout of transcription factor A, mitochondrial ( Tfam ), mtDNA was released into the cytosol and activated the cGAS–STING pathway, resulting in kidney inflammation and fibrosis 34 .

The ETC, particularly on complexes I and III, is the major site of ROS generation within mitochondria 43 . Electrons that leak from the ETC react with oxygen (O 2 ) to form superoxide anions, which can be dismutated to hydrogen peroxide (H 2 O 2 ) by superoxidase dismutases (SODs). H 2 O 2 can be further reduced to water by antioxidant catalase, glutathione peroxidases (GPXs) and peroxiredoxin. Under physiological conditions, the mitochondrial antioxidant defence system keeps ROS levels low within the organelles and enables emission of low levels of mitochondrial H 2 O 2 into the cytosol, where they act as cell survival signalling molecules 44 – 46 . Under conditions of stress when mtROS production overwhelms the capacity of mitochondrial antioxidant defence or the antioxidant defence system is impaired, ROS accumulate. Although O 2 •− is fairly unreactive, it can react rapidly with nitric oxide to form the potent oxidant and nitrating agent peroxynitrite and other reactive nitrogen species that have high reactivity with various biomolecules. H 2 O 2 can damage enzymes by oxidizing their thiol groups 47 and can produce highly reactive hydroxyl radicals in the presence of Fe 2+ cations via the Fenton reaction 48 . Increased mitochondrial H 2 O 2 emission to the cytosol as a result of mtROS accumulation expands oxidative damage outside of the mitochondria. As a result, a balance of ROS production and scavenging within mitochondria is essential for maintaining mitochondrial function and cell viability. Fenton reaction A catalytic process that converts hydrogen peroxide (H 2 O 2 ) into highly reactive hydroxyl free radicals in the presence of ferrous iron (Fe(II)). Role in AKI and kidney repair. Experimental studies have demonstrated an inadequate ROS-scavenging capacity within mitochondria in kidney tubules during AKI and subsequent kidney repair, as demonstrated by increases in mtROS in tubular cells and the beneficial effects of supplementation of exogenous mitochondria-targeted antioxidants during these processes. In a mouse model of kidney fibrosis induced by IR, a persistent increase in ROS and oxidative stress, accompanied by a sustained reduction in the activities of SOD1, SOD2 and catalase, was shown in kidney tissue 16 days after ischaemia 49 . Supplementation of a cell-permeable SOD mimetic from 48 h to 14 days after IR dramatically reduced kidney fibrosis in this model. Treatment of mice with cisplatin resulted in a persistent increase in mtROS in the kidney tubules for up to 1 month, whereas treatment with a mitochondrial-specific SOD mimetic (GC4419) ameliorated AKI induced by a single dose of cisplatin and kidney fibrosis induced by repeated cisplatin doses 50 . Together, these findings suggest that the mitochondrial antioxidant defence system is impaired or overwhelmed during kidney injury and repair and that this inadequacy contributes to pathological processes. Peroxisomes are intracellular organelles that contain antioxidant enzymes (particularly catalase) and a fatty acid β-oxidation machinery in which electrons from various metabolites reduce O 2 to H 2 O 2 . Studies have demonstrated that peroxisomes have a role in the regulation of redox homeostasis in kidney tubular cells and peroxisome damage might contribute to AKI 51 . Reductions in peroxisome number and activity, as well as downregulation of catalase, were shown in the kidneys of mice with AKI induced by cisplatin or IRI 52 . Proximal tubule-specific overexpression of NAD-dependent protein deacetylase sirtuin 1 (SIRT1) restored peroxisome number and function in kidney tubular cells, which led to upregulation of catalase expression and mitigation of ROS and kidney injury in these models. Notably, overexpression of SIRT1 failed to restore mitochondrial number but preserved mitochondrial proteins as a secondary effect of ROS reduction via catalase, suggesting that peroxisome damage-induced ROS contributes to mitochondrial damage and AKI. The potential relationship between peroxisomes and mitochondria in the pathogenesis of kidney diseases awaits further investigation. Excessive mtROS cause oxidative damage to mitochondrial components, which further increases mtROS production, forming a vicious circle, with an increased tendency towards cell death. Cardiolipin is a unique phospholipid that exclusively anchors in the IMM and has critical roles in the regulation of mitochondrial cristae formation and organization of the respiratory chain complex. Interaction with cardiolipin anchors cytochrome c in the IMM and thus prevents its release into the cytosol 11 , 53 . ROS induce cardiolipin peroxidation, which converts cytochrome c from an electron carrier into a peroxidase that enhances cardiolipin peroxidation, ultimately leading to mitochondrial dysfunction and cell death 11 , 53 . Experimental studies suggest that ROS-mediated peroxidation of cardiolipin contributes critically to the development of AKI and incomplete kidney repair after AKI.

Mitochondrial function is highly dependent on protein homeostasis within the organelles. However, mitochondrial protein homeostasis is challenging in both physiological and pathological conditions because of the complex mitochondrial proteome, which consists of proteins encoded by nuclear and mitochondrial genomes, and the continuous exposure of mitochondrial proteins to mtROS. Coordinated gene expression between nuclear and mitochondrial genomes, efficient importing of nuclear-encoded mitochondrial proteins from the cytosol into the mitochondria, accurate protein sorting to distinct mitochondrial subcompartments and protein maturation by folding, as well as correct assembly of relevant proteins into functional complexes, are essential for mitochondrial protein homeostasis during mitochondrial biogenesis and remodelling. Efficient refolding or removal of damaged proteins to prevent deleterious protein aggregation within the organelles is also critically important for normal mitochondrial function. The mitochondrial protein quality control system consists of chaperones that catalyse protein folding and ATP-dependent proteases that remove unwanted and unrepaired proteins 68 ( FIG. 2 ). When the capacity of this quality control system is overwhelmed by excessive amounts of unfolded or misfolded proteins, the mitochondrial unfolded protein response (UPR mt ) is induced. In the UPR mt , signals released from mitochondria trigger transcription of nuclear genes that encode mitochondrial chaperones to expand the mitochondrial protein folding capacity and thereby prevent deleterious protein aggregation within the organelles 69 . Impaired capacity of mitochondrial protein quality control systems may compromise mitochondrial function and jeopardize cell viability; such impairment has been associated with mitochondria-related and age-related degenerative diseases such as Parkinson disease and Alzheimer disease 70 . Role in AKI and kidney repair. Disturbances in mitochondrial protein homeostasis usually arise from either mutations that alter mitochondrial protein sequences or accumulation of mtROS that alter protein structure by direct oxidative modification. Recessive mutations in TRAP1 , which encodes heat shock protein 75 kDa, mitochondrial (TRAP1; also known as HSP75), a mitochondrial chaperone that is highly expressed in the proximal tubules and thick medullary ascending limbs of the loop of Henle, have been linked to congenital abnormalities of the kidney and urinary tract 71 , suggesting a critical role of TRAP1 in kidney tubular pathophysiology. Upregulation of TRAP1 and heat shock 70 kDa protein 9 (HSPA9) was detected in kidney tissue from animals with AKI 47 , 72 , 73 , suggesting a possible occurrence of UPR mt in this disorder. Notably, the endoplasmic reticulum unfolded protein response (UPR ER ) has been implicated in the pathogenesis of AKI and incomplete kidney repair after AKI 74 – 76 . Although the UPR mt and UPR ER operate within distinct organelles and involve organelle-specific chaperones and proteases, both pathways are rapidly activated and crosstalk with each other in response to extrinsic stimuli, such as hypoxia, oxidative stress and metabolic disorders 77 , 78 , which are present in kidney tubules during AKI. The induction of UPR ER during AKI and subsequent kidney repair might therefore suggest that activation of the UPR mt also occurs under these conditions. Moreover, mitochondrial biogenesis is critically important for kidney recovery after AKI 79 . Given the essential role of mitochondrial protein quality control in polypeptide sorting, folding and subsequent assembly into functional complexes during mitochondrial biogenesis, it is conceivable that efficient pro-protein folding activity within mitochondria is essential for kidney recovery. However, the functional association between the mitochondrial protein quality control system, AKI and kidney repair remains largely unknown.

In comparison with nuclear DNA, mtDNA is particularly vulnerable to oxidative stress because of its lack of protective histone proteins and close proximity to the site of ROS production. As mtDNA encodes 13 proteins of the ETC, mtDNA damage may impair OXPHOS. Moreover, once the fraction of damaged mtDNA in an individual cell exceeds a certain threshold, cell death ensues 82 . Efficient repair or removal of damaged mtDNA is therefore essential for preserving normal mitochondrial function and cell survival. In mammalian cells, mitochondria possess most of the DNA repair pathways that are available in the nucleus, including base excision repair, mismatch repair, homologous recombination and non-homologous end joining 83 . These mechanisms are responsible for repairing distinct mtDNA lesions and involve both nuclear gene- and mitochondrial gene-encoded enzymes. In the case of severe, irreparable damage, mtDNA can be degraded through a mitophagy- and autophagy-independent manner 84 . The molecular mechanism that underlies autophagy-independent mtDNA degradation has not been characterized. Defects in mtDNA repair impair mitochondrial function, increase vulnerability to cell death and have been implicated in human diseases such as cancer 82 . Role in AKI and kidney repair. Nuclear DNA damage and the subsequent DNA damage response have been demonstrated to have important roles in AKI and kidney repair 85 , 86 . mtDNA damage also occurs in the kidneys during AKI and kidney fibrosis. Increased levels of 8-hydroxy-2-deoxy-guanosine, a marker of oxidative DNA damage, in kidney mtDNA were identified shortly after reperfusion in rat models of kidney IRI 87 , 88 and in mice with Staphylococcus aureus sepsis-induced kidney injury 89 . In addition, mtDNA deletion was found in kidney tubular cells during adefovir-induced nephrotoxicity 90 and intraperitoneal administration of cisplatin was shown to decrease kidney mtDNA content in mice 91 . As mtDNA deletions are most likely to be the result of the repair of double-strand breaks by recombination-based processes 92 , mtDNA deletion during AKI and kidney fibrosis suggests activation of mtDNA repair. Consistent with this hypothesis, mitochondrial levels of the base excision repair proteins N-glycosylase/DNA lyase and uracil DNA glycosylase were increased in kidney tissues following septic AKI 89 . In rats with kidney IRI, levels of mtDNA oxidation (shown by 8-hydroxy-2-deoxyguanosine staining) decreased during the recovery of kidney function, suggesting efficient mtDNA damage repair 88 . Although further investigation is needed, together these findings suggest that mtDNA damage and activation of mtDNA repair processes occur during kidney injury and repair.

Mitochondria are highly dynamic organelles that constantly undergo fusion and fission ( FIG. 3 ). These processes are an important adaptive response to metabolic or environmental stresses and also a critical defence mechanism to preserve a mitochondrial population that is healthy overall, especially when quality control mechanisms fail to repair or remove damaged mitochondrial components. Mitochondrial fusion facilitates the exchange of metabolites and substrates between mitochondria to ensure optimal function of the mitochondrial network and is required for the complementation of oxidatively damaged mitochondrial components to mitigate organelle stress 97 , 98 . Mitochondrial fission is required for mitotic segregation of mitochondria into daughter cells and separation of damaged or dysfunctional parts of mitochondria for autophagic degradation 40 , 99 . Mitochondrial fission may also facilitate apoptosis in response to severe cellular stress. Mitochondrial fusion and fission are governed by members of a family of conserved GTPases. Fission is a multistep process in which DRP1 has an essential role. During fission, DRP1 is recruited from the cytosol to the OMM at mitochondria–endoplasmic reticulum contact sites by its receptors mitochondrial fission factor (MFF), mitochondrial dynamics protein MID49 and/or MID51. DRP1 then oligomerizes to form a ring-like structure around the mitochondria that utilizes the energy from GTP hydrolysis to constrict the organelle 100 , 101 . Dynamin 2 is then recruited to the mitochondrial constriction neck to sever the OMM 102 . DRP1 may also have severing ability sufficient for mitochondrial fission 103 . Our studies suggest an involvement of endophilin B1 (also known as BIF1) in IMM fission. We found that endophilin B1 translocates to mitochondria during cell stress and binds prohibitin 2 on the IMM, resulting in release of the metalloendopeptidase OMA1 and subsequent proteolysis of the IMM fusion protein dynamin-like 120 kDa protein, mitochondrial (OPA1) 104 . Another study suggested that IMM fission is regulated by calcium influx 105 . Mitochondrial fusion involves fusion of the OMM, which is mediated by mitofusin 1 (MFN1) and MFN2, and fusion of the IMM, which is mediated by OPA1. During fusion, MFNs in the OMM of two adjacent mitochondria interact to tether the organelles. GTP hydrolysis-induced conformational changes in the MFNs drive docking of the OMMs and increase the contact surface area. Subsequently, MFNs oligomerize to ensure OMM fusion. Following OMM fusion, interactions between OPA1 and cardiolipin tether the IMMs for fusion 106 . Disruption of the balance between fusion and fission alters mitochondrial morphology and impairs mitochondrial function and cell viability; such disruption has been linked to neurodegenerative diseases and cancer 107 . Role in AKI and kidney repair. Mitochondrial fragmentation resulting from excessive fission and/or suppression of fusion has been implicated as a key event in mitochondrial damage and kidney tubule injury during AKI 40 , 108 . In rodent models of AKI induced by kidney IR or cisplatin toxicity, mitochondrial fragmentation occurred prior to tubular cell apoptosis and inhibition of fission attenuated tubular cell death and kidney injury 40 . Consistent with this finding, proximal tubule-specific deletion of Drp1 protected mice against kidney IR-induced tubular cell death, inflammation and kidney injury, and accelerated kidney recovery 41 . Upstream regulation of mitochondrial fragmentation following AKI has also been investigated in kidney tubular cells. In mice, cisplatin-induced AKI was associated with downregulation of SIRT3, which could be prevented by the antioxidant acetyl- l -carnitine or the AMP-activated protein kinase (AMPK) agonist 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) 109 . AICAR and acetyl- l -carnitine attenuated cisplatin-induced kidney injury and mitochondrial fragmentation in wild-type mice but not in Sirt3 -knockout mice, suggesting that downregulation of SIRT3 contributes critically to mitochondrial fragmentation in tubular cells during kidney injury. In cultured kidney tubular cells, overexpression of SIRT3 prevented cisplatin-induced recruitment of DRP1 to the OMM 109 . Mitochondrial uncoupling protein 2 (UCP2) might also be a negative regulator of mitochondrial fragmentation. Following kidney IR, Ucp2 -deficient mice exhibited more severe mitochondrial fragmentation and kidney injury than did wild-type mice, whereas mice with increased kidney expression of UCP2 showed reduced mitochondrial fragmentation and less severe kidney injury 110 . Overexpression of UCP2 reduced the recruitment of DRP1 to the mitochondria of kidney tubular cells following kidney IR. Protein numb homologue (NUMB) might also negatively regulate DRP1. In mice, proximal tubule-specific knockout of Numb increased mitochondrial fragmentation in kidney tubule cells under normal conditions 111 . Moreover, following cis

Mitophagy is a mechanism of selective degradation of excessive and defective mitochondria via the autophagy pathway. In addition to eliminating unwanted mitochondria during development and adjusting mitochondrial number to changes in metabolic demand 127 , 128 , mitophagy acts as a critical component of mitochondrial quality control mechanisms that identifies and tags severely damaged mitochondria for prompt elimination. Defects in mitophagy have been implicated in a variety of human disorders 129 . Mitophagy requires efficient coordination of mitochondrial recognition with subsequent engulfment of targeted mitochondria within autophagosomes ( FIG. 4 ). Two major mechanisms for labelling mitochondria and delivering them to autophagosomes have been defined, one of which is regulated by the serine/threonine-protein kinase PINK1, mitochondrial (PINK1)–E3 ubiquitin-protein ligase parkin pathway and the other by mitophagy receptors including BCL-2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP3), BCL-2/adenovirus E1B 19 kDa protein-interacting protein 3-like (BNIP3L), FUN14 domain-containing 1 (FUNDC1) and E3 ubiquitin-protein ligase SMURF1 (SMURF1) 129 , 130 . PINK1 is a mitochondrial protein kinase that is constitutively imported into mitochondria, where it is cleaved by protease presenilins-associated rhomboid-like protein, mitochondrial (PARL). Parkin is a cytosolic ubiquitin E3 ligase. Upon mitochondrial damage or depolarization, PINK1 import into mitochondria is prevented and thus it accumulates in the OMM. PINK1 recruits parkin from the cytosol to the damaged mitochondria and activates its E3 ligase activity via phosphorylation of parkin and ubiquitin 131 , 132 . Activated parkin builds poly-ubiquitin chains on the OMM proteins, which in turn recruit receptor proteins, such as calcium-binding and coiled-coil domain-containing protein 2 (NDP52) and optineurin. These receptors simultaneously bind to poly-ubiquitin chains in mitochondria and microtubule-associated proteins 1A/1B light chain 3B (LC3B) on autophagosome membranes, resulting in engulfment of targeted mitochondria within autophagosomes, subsequent autophagosome–lysosome fusion and degradation of the polyubiquitinated mitochondria by lysosomal hydrolases 133 , 134 . The PINK1–parkin pathway is also involved in a mitophagy-independent quality control pathway in which damaged mitochondrial cargo is enclosed into mitochondria-derived vesicles for lysosomal degradation 135 – 137 . However, the involvement of this quality control mechanism in the kidney has not been determined. Mitophagy receptors are also recruited to the OMM of damaged mitochondria, where they can bridge the mitochondria to LC3B on autophagosome membranes via their N-terminal LC3-interacting regions, ultimately targeting mitochondria for autophagosome engulfment and degradation by lysosomal hydrolases. Experimental evidence suggests that crosstalk between the PINK1–parkin pathway and mitophagy receptors has a role in regulating mitophagy 138 , 139 . Role in AKI and kidney repair. Accumulating evidence suggests an important role of mitophagy in the pathogenesis of AKI. In mouse models that underwent bilateral kidney ischaemia for 30 min followed by reperfusion for up to 48 h, we showed an increase in autophagy flux accompanied with increases in autophagosomes with engulfed mitochondria and degradation of mitochondrial proteins in proximal tubular cells 140 , 141 . By contrast, another study demonstrated that both autophagy and mitophagy were suppressed in kidney tubules in a mouse model of AKI that was induced by 30 min of bilateral kidney ischaemia followed by 24 h of reperfusion 142 . The cause of the discrepancy remains unclear, but the differing durations of reperfusion might have a role. Induction of mitophagy in kidney tubular cells was also detected in models of nephrotoxic AKI induced by cisplatin or contrast medium 143 , 144 . An increase in mitophagy in kidney tubular cells in the early stages of septic AKI followed by mitophagy impairment in the later phases of AKI was reported in a mouse model of septic AKI induced by caecal ligation and puncture (CLP) 145 . Peroxisome proliferator-activated receptor-γ co-activator 1α (PGC1α) was identified as a positive regulator of mitophagy that induces transcription factor EB (TFEB)-mediated lysosome biogenesis in kidney tubular cells following cisplatin-induced AKI in mice 146 . Further studies demonstrated that kidney IR-, cisplatin- and contrast medium-induced mitophagy in kidney proximal tubules was partially abrogated in mice with knockout of Pink1 or Park2 (which encodes parkin) 141 , 143 , 144 . Bnip3 knockout also reduced tubular cell mitophagy in a mouse model of ischaemic AKI 140 . These findings suggest that both the PINK1–parkin pathway and mitophagy receptors are involved in the regulation of mitophagy in kidney tubules. Pink1 , Park2 or Bnip3 deficiency aggravated IR-induced kidney injury and enhanced mitochond

Mitochondrial biogenesis is the generation of new mitochondrial mass and replication of mtDNA through the proliferation of existing mitochondria 157 . The master regulator of mitochondrial biogenesis, PGC1α, directly regulates an array of transcription factors to modulate expression of nuclear genes that are required for this process, including nuclear respiratory factor 1 (NRF1), nuclear factor erythroid 2-related factor 2 (NRF2), peroxisome proliferator-activated receptor-α (PPARα), steroid hormone receptor ERR1 and transcriptional repressor protein YY1 (REF. 158 ) ( FIG. 5 ). Role in AKI and kidney repair. Accumulating evidence supports a beneficial role of mitochondrial biogenesis in kidney injury and repair after AKI. PGC1α is highly expressed in the proximal tubules 159 where mitochondria are abundant. In mouse models of septic AKI induced by lipopolysaccharide or CLP, the levels of PGC1α and downstream OXPHOS genes in the kidney were suppressed proportionally to the degree of kidney injury and were restored to normal levels during kidney recovery 79 , suggesting a negative correlation between PGC1α expression in kidney tubules and AKI severity. Reduced kidney expression of PGC1α was also observed in animal models of kidney IRI or cisplatin-induced AKI compared with controls 159 , 160 and in kidney biopsy samples from patients with AKI compared with normal human kidney tissue sections 160 . Further studies demonstrated that global or tubule-specific Pgc1a deficiency delayed kidney recovery following lipopolysaccharide-induced AKI in mice 79 . Global PGC1A deficiency also reduced kidney recovery following IRI, whereas transgenic expression of PGC1α in tubular cells facilitated kidney repair and recovery after IRI 160 . Similarly, pharmacological activation of PGC1α accelerated kidney function recovery after IRI in mice 161 . PGC1A deficiency worsened mitochondrial damage, whereas transgenic expression of PGC1A improved mitochondrial function and increased mitochondrial mass in experimental models of AKI 162 , suggesting a key role of PGC1α in maintaining mitochondrial function, at least in part via regulation of mitochondrial biogenesis 163 . Mitochondria that have been newly generated by biogenesis may replace damaged and degraded mitochondria that have undergone mitophagy during AKI and thus contribute to the repopulation of kidney tubular cells with adequate numbers of mitochondria to meet the increased metabolic and energy demands of tubular recovery after acute injury. A plethora of stimuli, including ATP depletion, ROS, hypoxia, nitric oxide, cyclic guanosine monophosphate and nutrient deprivation are potential inducers of PGC1α expression 164 . Although ATP depletion, ROS accumulation and hypoxia occur in the kidney during injury, PGC1α expression is suppressed during AKI and restored during kidney recovery 79 . This finding suggests that an evolving balance of suppressive and inductive factors might determine the temporal profile of PGC1α expression during AKI and kidney repair. Emerging evidence suggests that the pro-inflammatory cytokine tumour necrosis factor (TNF) 79 , 165 and tumour necrosis factor ligand superfamily member 12 (TNFSF12; also known as TWEAK) suppress PGC1A expression during AKI 166 . Several studies have also shown that kidney IR-induced activation of mitogen-activated protein kinase 3 (MAPK3; also known as ERK1) and MAPK1 (also known as ERK2) contribute to the downregulation of PGC1α pathways and ultimately kidney injury in mouse models 167 , 168 . By contrast, 5-hydroxytryptamine receptor 1F (5-HT1F; encoded by Htr1f ) and β2 adrenergic receptor (encoded by Adrb2 ) are positive regulators of Pgc1a expression in mouse kidney cells 169 , 170 . Kidney proximal tubule-specific Htr1f or Adrb2 knockout reduced mitochondrial numbers in these cells under physiological conditions and to a greater extent in a mouse model of kidney IRI, and also increased the severity of kidney injury in this model 169 , 170 . In addition to transcriptional regulation of PGC1A expression, post-translational modifications, including phosphorylation, ubiquitylation, methylation, acetylation and GlcNAcylation, have important roles in regulating PGC1α activity 171 .

Given the critical role of mitochondrial dysfunction in kidney injury and abnormal kidney repair, specific interventions that target mitochondrial quality control mechanisms to preserve and restore mitochondrial function have emerged as promising therapeutic strategies to prevent and treat kidney injury, as well as accelerate kidney repair. A variety of compounds that target mitochondrial quality control mechanisms have been shown to protect against kidney injury and/or to accelerate kidney repair in AKI and CKD ( TABLE 2 ; FIG. 6 ). Treatment with mitochondria-targeted antioxidants, such as quinone analogues (MitoQ 65 , 178 , SkQ1 and SkQR 179 ), SOD mimetics (Mito-CP) 180 and SS-31 (REFS 55 , 181 , 182 ), before kidney injury has been shown to protect against AKI and CKD in animal models. Notably, in rats with kidney IRI, treatment with SS-31 starting 1 month after kidney ischaemia for 6 weeks preserved mitochondrial integrity, restored glomerular capillaries and podocyte structure and arrested glomerulosclerosis and interstitial fibrosis 42 . Moreover, this SS-31-mediated protection was sustained for ≥6 months after treatment ended. Despite their efficiency in the prevention of kidney injury in experimental models, the potential of mitochondria-targeted antioxidants as treatments for AKI and/or CKD awaits further investigation. Despite the exciting preclinical data, challenges remain for the clinical translation of mitochondria-targeted antioxidants in AKI and CKD. As discussed above, low and moderate levels of mtROS may act as signalling molecules in cell stress response pathways 183 . By scavenging and removing physiological levels of ROS involved in signalling, antioxidant supplements could have deleterious effects. In line with this notion, ischaemic preconditioning using a single event of kidney ischaemia and reperfusion (SIRPC) induced ROS production and protected against subsequent kidney IRI in mice, whereas administration of antioxidants after SIRPC reduced endogenous antioxidant expression and abolished the protective effect 184 . This finding suggests that SIRPC induced the production of ROS, which acted as signalling molecules to induce a protective antioxidant response. Accurate measurement of ROS levels before or during the administration of mitochondria-targeted antioxidants will likely be essential to avoid adverse effects and enable the safe use of these therapies, but methods and equipment for such measurement are not currently available. Inhibition of mitochondrial fragmentation also seems to be beneficial during kidney injury and repair. mdivi-1 is a chemical inhibitor of mitochondrial fission that acts by inhibiting DRP1 activity 185 . Administration of mdivi-1 before injury has been shown to suppress mitochondrial fragmentation in kidney cells and protect against AKI and CKD in experimental models 40 , 121 . Of note, inhibition of mitochondrial fission 3 days after ischaemia using mdivi-1 accelerated kidney repair after AKI and protected against the development of post-AKI kidney fibrosis in mice 41 . It is important to note that fission is essential for increasing mitochondrial number to populate newly regenerated tubular cells with adequate numbers of mitochondria during kidney recovery after AKI. In addition, fission is required for removal of damaged or dysfunctional mitochondria by the autophagic pathway 186 . Inhibition of mitochondrial fission may therefore produce unexpected results. Moreover, the specificity of mdivi-1 has been questioned owing to experimental evidence that it might also act as a reversible mitochondrial complex I inhibitor 187 . Thus, there is a need to identify more specific chemical inhibitors of mitochondrial fission and to optimize the timing of administration of mitochondrial fission inhibitors to protect the kidney. The therapeutic potential of inhibitors of mitochondrial fragmentation in kidney disease awaits further investigation. Enhancement of the removal of damaged and dysfunctional mitochondria by autophagy and/or mitophagy in kidney tubular cells is another attractive strategy for preventing AKI and DKD 188 , 189 . mTOR inhibitors, such as rapamycin, have been shown to enhance autophagic removal of damaged mitochondria in the kidney 190 , 191 , but their application might be limited because they also suppress cell proliferation 192 . A mTOR-independent agent, Tat-beclin 1, which is a peptide that comprises the essential sequence of beclin 1, has been shown to induce autophagy in skeletal muscle, cardiac muscle, pancreas and kidney tubules in vivo 193 , 194 . Urolithin A, a gut microbial metabolite of ellagic acid and related compounds, has been demonstrated to induce mitophagy and to have beneficial effects on age-related decline of muscle function following oral consumption in mouse models 195 . Oral administration of urolithin A also attenuated kidney injury in mouse models of AKI induced by cisplatin or CLP 196 , 197 , but the involvement o