Cisplatin-induced Tubular Injury Research Articles

PARVB deficiency alleviates cisplatin-induced tubular injury by inhibiting TAK1 signaling

Cisplatin-induced renal tubular injury largely restricts the wide-spread usage of cisplatin in the treatment of malignancies. Identifying the key signaling pathways that regulate cisplatin-induced renal tubular injury is thus clinically important. PARVB, a focal adhesion protein, plays a crucial role in tumorigenesis. However, the function of PARVB in kidney disease is largely unknown. To investigate whether and how PARVB contributes to cisplatin-induced renal tubular injury, a mouse model (PARVB cKO) was generated in which PARVB gene was specifically deleted from proximal tubular epithelial cells using the Cre-LoxP system. In this study, we found depletion of PARVB in proximal tubular epithelial cells significantly attenuates cisplatin-induced renal tubular injury, including tubular cell death and inflammation. Mechanistically, PARVB associates with transforming growth factor-β-activated kinase 1 (TAK1), a central regulator of cell survival and inflammation that is critically involved in mediating cisplatin-induced renal tubular injury. Depletion of PARVB promotes cisplatin-induced TAK1 degradation, inhibits TAK1 downstream signaling, and ultimately alleviates cisplatin-induced tubular cell damage. Restoration of PARVB or TAK1 in PARVB-deficient cells aggravates cisplatin-induced tubular cell injury. Finally, we demonstrated that PARVB regulates TAK1 protein expression through an E3 ligase ITCH-dependent pathway. PARVB prevents ITCH association with TAK1 to block its ubiquitination. Our study reveals that PARVB deficiency protects against cisplatin-induced tubular injury through regulation of TAK1 signaling and indicates targeting this pathway may provide a novel therapeutic strategy to alleviate cisplatin-induced kidney damage.

Decreased IFT88 expression with primary cilia shortening causes mitochondrial dysfunction in cisplatin-induced tubular injury.

The relevance of primary cilia shortening in kidney disease and its pathomechanism are largely unknown. Tubular damage in acute kidney injury (AKI) is strongly associated with mitochondrial dysfunction. Thus, we investigated the interaction between primary cilia and mitochondria in cisplatin-induced AKI mouse models. We observed that the expression of intraflagellar transport 88 (IFT88), a ciliary maintenance protein, was decreased in the renal cortex following tubular damage due to cisplatin-induced AKI. This result was consistent with the decreased IFT88 expression in cisplatin-treated RPTEC/TERT1 cells (human primary proximal tubular cells) parallel to the shortening of primary cilia, suggesting a causative link between tubular damage and IFT88-mediated cilia regulation. To address the effect of impaired primary cilia with decreased IFT88 expression on tubular function, RPTEC/TERT1 cells treated with cisplatin and knocked down for IFT88 using siRNA (IFT88-KD) were assessed for phenotypic changes and mitochondrial metabolic function. Both cisplatin and IFT88-KD caused primary cilia shortening, downregulated mitochondrial oxidative phosphorylation capacity, and had defective fatty acid oxidation and decreased ATP production. Furthermore, IFT88 overexpression enhanced mitochondrial respiration, which partially counteracted cisplatin-induced defective fatty acid oxidation. These results are indicative of the contribution of IFT88 to mitochondrial homeostasis. Our findings suggest that tubular mitochondrial dysfunction in cisplatin-induced AKI is mediated, at least in part, by a decrease in IFT88 expression with primary cilia shortening. That is, tubular mitochondrial damage followed by tubular injury in AKI may occur through alteration of IFT88 expression and subsequent ciliary shortening in tubular cells.NEW & NOTEWORTHY Here, we demonstrated organelle cross-talk between primary cilia and mitochondria of proximal tubular cells in cisplatin-induced acute kidney injury. The primary cilia-mitochondria interaction may open new avenues for the development of novel therapeutic approaches in the treatment of acute kidney injury.

POS-075 Decreased IFT88 causes cilia shortening and mitochondrial dysfunction in cisplatin-induced tubular injury

Renal primary cilia are associated with the pathogenesis of various diseases, including acute kidney injury (AKI). The length of the primary cilia dynamically change during the progression of diseases: tubular primary cilia shortened in cisplatin-induced AKI mouse model. However, its relevance in kidney disease and the underlying mechanism are largely unknown. Tubular damage in AKI closely links to mitochondrial dysfunction. Thus, we investigated the interaction between primary cilia and mitochondria in cisplatin-induced tubular injury. C57BL6 mice with cisplatin-induced AKI were euthanized at 72 h after disease induction to collect blood and kidney samples. In in vitro experiments, we used RPTEC/TERT1 cells, which are human proximal tubular epithelial cells that maintain the cilia length at high cell densities, and knocked down IFT88 (IFT88-KD), a cilia maintenance protein, by siRNA. Cisplatin-treated or IFT88-KD cells were assessed for the cellular phenotypic changes or mitochondrial metabolic function using the Flux Analyzer. We found that the expression of protein IFT88 was decreased in damaged tubules of cisplatin-induced AKI mice. These data were consistent with that IFT88 expression decreased in the cilia of cisplatin-treated RPTEC/TERT1 at mRNA and protein levels in association with shortening of the primary cilia, suggesting the pathogenic link between tubular damage and IFT88-mediated cilia alteration. Interestingly, IFT88-KD cells significantly exhibited shorter cilia as compared to control siRNA-transfected cells and showed downregulation of mitochondrial oxidative phosphorylation capacity and decreased ATP production, indicating the contribution of IFT88 to mitochondrial homeostasis. In addition, the knockdown of IFT88 decreases fatty acid oxidation (FAO) in the same direction as cisplatin treatment. Our findings suggest that tubular mitochondrial dysfunction in cisplatin-induced AKI is mediated by a decreased IFT88 with shortening cilia, at least in part. Tubular mitochondrial damage followed by tubular injury in AKI may occur by the alteration of IFT88 expression, and subsequent cilia shortening in the tubular epithelial cells.

Decreased IFT88 Causes Cilia Shortening and Mitochondrial Dysfunction in Cisplatin-Induced Tubular Injury

Decreased IFT88 Causes Cilia Shortening and Mitochondrial Dysfunction in Cisplatin-Induced Tubular Injury

Pharmacological inhibition of ataxia-telangiectasia mutated exacerbates acute kidney injury by activating p53 signaling in mice

The DNA damage response after kidney injury induces cell cycle arrest in renal tubular epithelial cells, resulting in the secretion of pro-fibrotic cytokines, thereby promoting interstitial fibrosis in a paracrine manner. Phosphorylation of ataxia-telangiectasia mutated (ATM) is the initial step in the DNA damage response and subsequent cell cycle arrest; however, the effects of ATM inhibition on the injured kidney have not been explored. Pharmacological ATM inhibition by KU55933 in cisplatin-treated mice did not ameliorate, but instead exacerbated cisplatin-induced DNA damage and tubular injury, thereby increasing mortality. Analysis of isolated tubular epithelia by FACS from bigenic SLC34a1-CreERt2; R26tdTomato proximal tubular-specific reporter mice revealed that KU55933 upregulated p53 and subsequent pro-apoptotic signaling in tubular epithelia of cisplatin-treated mice, leading to marked mitochondrial injury and apoptosis. In addition, KU55933 attenuated several DNA repair processes after cisplatin treatment, including single-strand DNA repair and Fanconi anemia pathways, suggesting that DNA repair after dual treatment of cisplatin and KU55933 was not sufficient to prevent the cisplatin-induced tubular injury. Our study suggested that ATM inhibition does not increase DNA repair after cisplatin-induced DNA damage and exacerbates tubular injury through the upregulation of p53-dependent pro-apoptotic signaling. Acute kidney injury must be carefully monitored when ATM inhibitors become available in clinical practice in the future.

Serum d-serine accumulation after proximal renal tubular damage involves neutral amino acid transporter Asc-1

Chiral separation has revealed enantio-specific changes in blood and urinary levels of amino acids in kidney diseases. Blood d-/l-serine ratio has been identified to have a correlation with creatinine-based kidney function. However, the mechanism of distinctive behavior in serine enantiomers is not well understood. This study was performed to investigate the role of renal tubules in derangement of serine enantiomers using a mouse model of cisplatin-induced tubular injury. Cisplatin treatment resulted in tubular damage histologically restricted to the proximal tubules and showed a significant increase of serum d-/l-serine ratio with positive correlations to serum creatinine and blood urine nitrogen (BUN). The increased d-/l-serine ratio did not associate with activity of a d-serine degrading enzyme, d-amino acid oxidase, in the kidney. Screening transcriptions of neutral amino acid transporters revealed that Asc-1, found in renal tubules and collecting ducts, was significantly increased after cisplatin-treatment, which correlates with serum d-serine increase. In vitro study using a kidney cell line showed that Asc-1 is induced by cisplatin and mediated influx of d-serine preferably to l-serine. Collectively, these results suggest that cisplatin-induced damage of proximal tubules accompanies Asc-1 induction in tubules and collecting ducts and leads to serum d-serine accumulation.

SUN-155 Mitochondrial DNA leakage causes inflammation via the cGAS-STING axis in cisplatin-induced acute kidney injury

Cisplatin is an anti-neoplastic drug that induces tubular inflammation. However, its detailed mechanisms are not fully understood. Stimulator of interferon genes (STING) andits upstream or downstream molecules (cGAS-STING axis) play an important role inself/non-self DNA-triggered signal transduction, leading to inflammation. Here, weinvestigated the roles of STING-mediated inflammation in cisplatin-induced tubular injury. The human proximal tubular cell line, HK-2, treated with 20 μM cisplatin or the renal cortex of WT C57BL/6 and STING KO mice injected with 25 mg/kg cisplatin for 48 or 72 h, were analyzed. The changes in cGAS-STING axis activation, mitochondrial DNA (mtDNA)leakage, or BAX expression were evaluated using real-time PCR, western blotting, or immunostaining analysis. Cisplatin and/or STING siRNA-treated HK-2 culture supernatants were analyzed using cytokine arrays and migration assays. ethidium bromide (EtBr) were used for mtDNA depletion, respectively. In HK-2 or kidney cortex of animals treated with cisplatin, cGAS and STING expression was upregulated and translocated from the ER to the Golgi apparatus, indicating STING activation by cisplatin. Subsequently, the cGAS-STING axis(TBK-1 and P65) was activated due to cisplatin-mediated phosphorylation. Cisplatin also induced pro-inflammatory molecule(IL-6, IL-8, ICAM-1, CXCL10, and GM-CSF) production and neutrophil chemotaxis, which were ameliorated by STING knockdown (P < 0.05). Interestingly, cisplatin-mediated mtDNA leakage to the cytosol activated cGAS-STINGaxis-mediated inflammation. In fact, EtBr-mediated mtDNA depletion inhibited the inflammation by cisplatin and mtDNA transfection facilitated cGAS-STING axis and inflammation in HK-2. Cisplatin-inducedBAX expression,which interacted with the mitochondrial permeability transition pore, suggested that mtDNA leakage was caused by the increase in BAX expression. Moreover, STING KO ameliorated kidney dysfunction and tubular injury accompanied with inhibition of inflammation. mtDNA leakage to the cytosol induces tubular inflammationby activating the cGAS-STING axis in cisplatin nephrotoxicity.

Numb Depletion Promotes Drp1-Mediated Mitochondrial Fission and Exacerbates Mitochondrial Fragmentation and Dysfunction in Acute Kidney Injury.

Mitochondrial fragmentation is a crucial mechanism contributing to tubular cell apoptosis during acute kidney injury (AKI). However, the mechanism of modulating mitochondrial dynamics during AKI remains unclear. Numb is a multifunction adaptor protein that is expressed in renal tubules. The aim of the present study was to evaluate the role of Numb in mitochondrial dysfunction during AKI. The expression of Numb was upregulated in both ischemia-reperfusion- and cisplatin-induced AKI. Depletion of Numb from proximal tubules (PT-Nb-KO) exacerbated AKI shown as more severe renal tubular damage and higher serum creatinine than wild-type mice. Numb depletion alone significantly increased mitochondrial fragmentation without altering mitochondrial mass and function, including adenosine triphosphate production, mitochondrial membrane potential, oxygen consumption, and reactive oxygen species production. However, mitochondrial fragmentation and dysfunction were significantly aggravated after cisplatin exposure in PT-Nb-KO mice. Mechanistically, Numb depletion triggered dynamin-related protein 1 (Drp1) recruitment to mitochondria by increasing the phosphorylation of Drp1 at serine 656 residue (human Drp1 ser637). Inhibiting the activity of Rho-associated coiled-coil containing protein kinase (ROCK) by Y-27632 attenuated phosphorylation of Drp1 ser656 and mitochondrial fragmentation in Numb-deficient cells. Administration of mdivi-1, a pharmacological inhibitor of Drp1, restored mitochondrial morphology, attenuated cisplatin-induced tubular injury, and renal dysfunction in PT-Nb-KO mice. Innovation and Conclusion: Our data suggest that Numb depletion promotes mitochondrial fragmentation by promoting the phosphorylation of Drp1 Ser637 and thus exacerbates cisplatin-induced mitochondrial dysfunction and tubular cell apoptosis. These findings add a novel insight into modulating mechanism of mitochondrial dynamics during AKI.

Reply: Oxytocin ameliorates cisplatin-induced nephrotoxicity in Wistar rats

Reply: Oxytocin ameliorates cisplatin-induced nephrotoxicity in Wistar rats

Urinary chemokine (C-C motif) ligand 2 (monocyte chemotactic protein-1) as a tubular injury marker for early detection of cisplatin-induced nephrotoxicity

Because of the difficulty in detecting segment-specific response in the kidney, we investigated the molecular events underlying acute kidney injury in the proximal tubules of rats with cisplatin (cis-diamminedichloroplatinum II)-induced nephrotoxicity. Microarray analysis revealed that mRNA levels of several cytokines and chemokines, such as interleukin-1beta, chemokine (C-C motif) ligand (CCL) 2, CCL20, chemokine (C-X-C motif) ligand (CXCL) 1, and CXCL10 were significantly increased after cisplatin treatment in both isolated proximal tubules and whole kidney. Interestingly, tubular CCL2 mRNA levels increased soon after cisplatin administration, whereas CCL2 mRNA levels in whole kidney first decreased and then increased. Levels of both CCL2 and kidney injury molecule-1 (KIM-1) in the whole kidney increased after cisplatin administration. Immunofluorescence analysis revealed CCL2 changes in the proximal tubular cells initially and then in the medullary interstitium. Urinary CCL2 excretion significantly increased approximately 3-fold compared with controls the day after cisplatin administration (5mg/kg), when no changes were observed plasma creatinine and blood urea nitrogen levels. Urinary levels of KIM-1 also increased 3-fold after cisplatin administration. In addition, urinary CCL2 rather than KIM-1 increased in chronic renal failure rats after administration of low-dose cisplatin (2mg/kg), suggesting that urinary CCL2 was selective for cisplatin-induced nephrotoxicity in renal impairment. These results indicated that the increase in cytokine and chemokine expression in renal epithelial cells might be responsible for kidney deterioration in cisplatin-induced nephrotoxicity, and that urinary CCL2 is associated with tubular injury and serves as a sensitive and noninvasive marker for the early detection of cisplatin-induced tubular injury.

Expressions of Lipid Oxidation Markers, N.EPSILON.-Hexanoyl Lysine and Acrolein in Cisplatin-Induced Nephrotoxicity in Rats

The purpose of this study was to evaluate whether N(ε)-hexanoyl lysine (N(ε)-HEL) and acrolein reflect the severity of cisplatin-induced nephrotoxicity. Immunoexpression of N(ε)-HEL and acrolein in kidneys and their urinary concentration were examined up to day 4 post-cisplatin injection in rats. Cisplatin-induced tubular injury was observed histopathologically on days 2-4 after injection and became more severe time-dependently. On days 2-4, N(ε)-HEL and acrolein were immunostained in the cytoplasm of damaged tubular cells. Their immunostaining intensity and urinary levels increased as tubular injury became more severe. These results suggest that expressions of N(ε)-HEL and acrolein were associated with the pathogenesis of cisplatin-induced nephrotoxicity.