Impaired fatty acid metabolism perpetuates lipotoxicity along the transition to chronic kidney injury
Anna Rinaldi, Hélène Lazareth, Virginie Poindessous, Ivan Nemazanyy, Julio L. Sampaio, Daniele Malpetti, Yohan Bignon, Maarten Naesens, Marion Rabant, Dany Anglicheau, Pietro E. Cippà, Nicolas Pallet
Anna Rinaldi, Hélène Lazareth, Virginie Poindessous, Ivan Nemazanyy, Julio L. Sampaio, Daniele Malpetti, Yohan Bignon, Maarten Naesens, Marion Rabant, Dany Anglicheau, Pietro E. Cippà, Nicolas Pallet
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Research Article Nephrology Transplantation

Impaired fatty acid metabolism perpetuates lipotoxicity along the transition to chronic kidney injury

Abstract

Energy metabolism failure in proximal tubule cells (PTCs) is a hallmark of chronic kidney injury. We combined transcriptomic, metabolomic, and lipidomic approaches in experimental models and patient cohorts to investigate the molecular basis of the progression to chronic kidney allograft injury initiated by ischemia/reperfusion injury (IRI). The urinary metabolome of kidney transplant recipients with chronic allograft injury and who experienced severe IRI was substantially enriched with long chain fatty acids (FAs). We identified a renal FA-related gene signature with low levels of carnitine palmitoyltransferase 2 (Cpt2) and acyl-CoA synthetase medium chain family member 5 (Acsm5) and high levels of acyl-CoA synthetase long chain family member 4 and 5 (Acsl4 and Acsl5) associated with IRI, transition to chronic injury, and established chronic kidney disease in mouse models and kidney transplant recipients. The findings were consistent with the presence of Cpt2–Acsl4+Acsl5+Acsm5– PTCs failing to recover from IRI as identified by single-nucleus RNA-Seq. In vitro experiments indicated that ER stress contributed to CPT2 repression, which, in turn, promoted lipids’ accumulation, drove profibrogenic epithelial phenotypic changes, and activated the unfolded protein response. ER stress through CPT2 inhibition and lipid accumulation engaged an auto-amplification loop leading to lipotoxicity and self-sustained cellular stress. Thus, IRI imprints a persistent FA metabolism disturbance in the proximal tubule, sustaining the progression to chronic kidney allograft injury.

Authors

Anna Rinaldi, Hélène Lazareth, Virginie Poindessous, Ivan Nemazanyy, Julio L. Sampaio, Daniele Malpetti, Yohan Bignon, Maarten Naesens, Marion Rabant, Dany Anglicheau, Pietro E. Cippà, Nicolas Pallet

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Figure 6

ER stress reduces CPT2 expression.

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ER stress reduces CPT2 expression. (A) Correlation between CKD stages an...
(A) Correlation between CKD stages and ER stress genes’ expression from the tubulointerstitial compartment of 201 individuals with CKD. Colors are z score–normalized to depict relative values within rows. P values were calculated with a 1-way ANOVA followed by a Dunnett’s multiple-comparison test. (B) Expression of HSPA5 and CPT2 measured by RT-qPCR in HK2 cells incubated with 2.5 μg/mL tunicamycin (Tun), 5 μg/mL brefeldin A (BFA), 0.25 μM thapsigargin (Tg), 1 μM dithiothreitol (DTT), 100 μM etoposide (Eto), or DMSO for 24 hours (n = 7–8). Bars represent mean ± SD. P values were calculated with a 1-way ANOVA followed by a Dunnett’s multiple-comparison test. (C) Time course analysis of the relative expression of CPT2 and HSPA5 transcripts measured by RT-qPCR in HK2 cells incubated with either with vehicle or with 0.25 μM Tg, 5 μg/mL BFA, or 2.5 μg/mL Tun for up to 24 hours (4 replicates). Bars represent mean ± SD. P values were computed with 2-way ANOVA. (D) Expression of Cpt2 and Hspa5 transcripts by quantitative PCR in kidney cortex of mice 48 hours after intraperitoneal injection of 1 mg/kg Tun or DMSO (n = 5 in DMSO group and 14 in Tun group). Bars represent mean ± SD. P values were computed with a Student’s t test. (E) Immunoblot representing Cpt2, Hspa5, and tubulin protein expression in kidney cortex of mice 48 hours after intraperitoneal injection of 1 mg/kg Tun or DMSO (n = 4). Bars represent mean ± SD. P values were computed with a Student’s t test. (F) Representative photomicrograph of Cpt2 and Hspa5 expression in kidney cortex of mice 48 hours after intraperitoneal injection of 1 mg/kg Tun or DMSO (n = 5). Original magnification, ×40. (G) OCR measured by Seahorse bioanalyzer in HK2 cells 48 hours posttransduction with siRNA targeting CPT2 under basal conditions and in response to incubation with DMSO or 2.5 μg/mL Tun for 24 hours. P values were computed with a Student’s t test, n = 5–6.