Renal tubule Cpt1a overexpression protects from kidney fibrosis by restoring mitochondrial homeostasis
Verónica Miguel, … , Ricardo Ramos, Santiago Lamas
Verónica Miguel, … , Ricardo Ramos, Santiago Lamas
Published January 19, 2021
Citation Information: J Clin Invest. 2021;131(5):e140695. https://doi.org/10.1172/JCI140695.
View: Text | PDF
Research Article Nephrology

Renal tubule Cpt1a overexpression protects from kidney fibrosis by restoring mitochondrial homeostasis

Abstract

Chronic kidney disease (CKD) remains a major epidemiological, clinical, and biomedical challenge. During CKD, renal tubular epithelial cells (TECs) present a persistent inflammatory and profibrotic response. Fatty acid oxidation (FAO), the main source of energy for TECs, is reduced in kidney fibrosis and contributes to its pathogenesis. To determine whether gain of function in FAO (FAO-GOF) could protect from fibrosis, we generated a conditional transgenic mouse model with overexpression of the fatty acid shuttling enzyme carnitine palmitoyl-transferase 1A (CPT1A) in TECs. Cpt1a-knockin (CPT1A-KI) mice subjected to 3 models of renal fibrosis (unilateral ureteral obstruction, folic acid nephropathy [FAN], and adenine-induced nephrotoxicity) exhibited decreased expression of fibrotic markers, a blunted proinflammatory response, and reduced epithelial cell damage and macrophage influx. Protection from fibrosis was also observed when Cpt1a overexpression was induced after FAN. FAO-GOF restored oxidative metabolism and mitochondrial number and enhanced bioenergetics, increasing palmitate oxidation and ATP levels, changes that were also recapitulated in TECs exposed to profibrotic stimuli. Studies in patients showed decreased CPT1 levels and increased accumulation of short- and middle-chain acylcarnitines, reflecting impaired FAO in human CKD. We propose that strategies based on FAO-GOF may constitute powerful alternatives to combat fibrosis inherent to CKD.

Authors

Verónica Miguel, Jessica Tituaña, J. Ignacio Herrero, Laura Herrero, Dolors Serra, Paula Cuevas, Coral Barbas, Diego Rodríguez Puyol, Laura Márquez-Expósito, Marta Ruiz-Ortega, Carolina Castillo, Xin Sheng, Katalin Susztak, Miguel Ruiz-Canela, Jordi Salas-Salvadó, Miguel A. Martínez González, Sagrario Ortega, Ricardo Ramos, Santiago Lamas

×

Figure 1

Characterization of doxycycline-inducible Cpt1a gene overexpression in the transgenic mouse model for the inducible Cpt1a gene in renal epithelial cells.

Options: View larger image (or click on image) Download as PowerPoint
Characterization of doxycycline-inducible Cpt1a gene overexpression in t...
(A) Schematic depicting the strategy to generate mice with inducible renal tubular epithelial cell–specific overexpression of CPT1A. (B) PCR analysis of offspring genotypes from these crosses. Genomic DNA analysis by PCR for the Pax8-rtTA allele generates a 595-bp band (Supplemental Table 1). The GFP, exon, and FRT PCRs are described in Supplemental Figure 1A. (C) mRNA levels of the Cpt1a gene were determined by qRT-PCR in total kidney tissue of mice treated with doxycycline for 3 weeks. Data represent the mean ± SEM (n = 6 mice). **P < 0.05 compared with kidneys from WT mice. (D) Immunoblots depicting protein levels of CPT1A and GFP in kidneys and livers of 3 individual mice per group. β-actin was used for normalization purposes. (E) Bar graphs represent the mean ± SEM of fold changes corresponding to densitometric analyses (n = 6 mice). **P < 0.01 compared with kidneys from WT mice. (F) Representative images of double immunofluorescence staining with the proximal tubular marker lotus tetragonolobus lectin (LTL), GFP, DAPI (nuclei), and merge of all 3. (G) Staining with DAPI (nuclei), CPT1A, and the mitochondrial marker ATP synthase beta-subunit (βF1). All panels show immunofluorescence images of kidneys from WT and Pax8-CPT1A mice after doxycycline administration. Scale bar: 100 μm (F and G). (H) Radiolabeled palmitate-derived CO2 and acid-soluble products (ASP) were determined after incubation of 14C-palmitate with kidney tissue from WT or Pax8-CPT1A mice after doxycycline treatment. Bar graphs represent the mean ± SEM (n = 4 mice). *P < 0.05 compared with kidneys from WT mice. Statistical significance between 2 independent groups was determined using nonparametric 2-tailed Mann-Whitney U test; more than 2 groups were compared with Kruskal-Wallis test.