RESUMO
Acute kidney injury (AKI) is a worldwide public health problem characterized by the massive loss of tubular cells. However, the precise mechanism for initiating tubular cell death has not been fully elucidated. Here, we reported that phosphoglycerate mutase 5 (PGAM5) was upregulated in renal tubular epithelial cells during ischaemia/reperfusion or cisplatin-induced AKI in mice. PGAM5 knockout significantly alleviated the activation of the mitochondria-dependent apoptosis pathway and tubular apoptosis. Apoptosis inhibitors alleviated the activation of the mitochondria-dependent apoptosis pathway. Mechanistically, as a protein phosphatase, PGAM5 could dephosphorylate Bax and facilitate Bax translocation to the mitochondrial membrane. The translocation of Bax to mitochondria increased membrane permeability, decreased mitochondrial membrane potential and facilitated the release of mitochondrial cytochrome c (Cyt c) into the cytoplasm. Knockdown of Bax attenuated PGAM5 overexpression-induced Cyt c release and tubular cell apoptosis. Our results demonstrated that the increase in PGAM5-mediated Bax dephosphorylation and mitochondrial translocation was implicated in the development of AKI by initiating mitochondrial Cyt c release and activating the mitochondria-dependent apoptosis pathway. Targeting this axis might be beneficial for alleviating AKI.
Assuntos
Injúria Renal Aguda , Citocromos c , Camundongos , Animais , Citocromos c/metabolismo , Fosfoglicerato Mutase/metabolismo , Proteína X Associada a bcl-2/metabolismo , Apoptose/fisiologia , Mitocôndrias/metabolismo , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/metabolismo , Proteínas de Transporte/metabolismo , Fosfoproteínas Fosfatases/metabolismoRESUMO
The pyroptosis of renal tubular epithelial cells leads to tubular loss and inflammation and then promotes renal fibrosis. The transcription factor Krüppel-like factor 4 (KLF4) can bidirectionally regulate the transcription of target genes. Our previous study revealed that sustained elevation of KLF4 is responsible for the transition of acute kidney injury (AKI) into chronic kidney disease (CKD) and renal fibrosis. In this study, we explored the upstream mechanisms of renal tubular epithelial cell pyroptosis from the perspective of posttranslational regulation and focused on the transcription factor KLF4. Mice were subjected to unilateral ureteral obstruction (UUO) surgery and euthanized on D7 or D14 for renal tissue harvesting. We showed that the pyroptosis of renal tubular epithelial cells mediated by both the Caspase-1/GSDMD and Caspase-3/GSDME pathways was time-dependently increased in UUO mouse kidneys. Furthermore, we found that the expression of the transcription factor KLF4 was also upregulated in a time-dependent manner in UUO mouse kidneys. Tubular epithelial cell-specific Klf4 knockout alleviated UUO-induced pyroptosis and renal fibrosis. In Ang II-treated mouse renal proximal tubular epithelial cells (MTECs), we demonstrated that KLF4 bound to the promoter regions of Caspase-3 and Caspase-1 and directly increased their transcription. In addition, we found that ubiquitin-specific protease 11 (USP11) was increased in UUO mouse kidneys. USP11 deubiquitinated KLF4. Knockout of Usp11 or pretreatment with the USP11 inhibitor mitoxantrone (3 mg/kg, i.p., twice a week for two weeks before UUO surgery) significantly alleviated the increases in KLF4 expression, pyroptosis and renal fibrosis. These results demonstrated that the increased expression of USP11 in renal tubular cells prevents the ubiquitin degradation of KLF4 and that elevated KLF4 promotes inflammation and renal fibrosis by initiating tubular cell pyroptosis.
RESUMO
Transforming growth factor-ß1 (TGF-ß1) is regarded as a key factor in promoting renal fibrosis during chronic kidney disease (CKD). Signaling transduction of TGF-ß1 starts with binding to TGF-ß type II receptor (Tgfbr2), a constitutively activated kinase that phosphorylates TGF-ß type I receptor (Tgfbr1), and then activates downstream Smad2/3 or noncanonical pathways. Previous studies show that cellular senescence is associated with the progression of CKD, and accelerated tubular cell senescence is implicated in promoting renal fibrosis. In the present study we investigated the renal parenchymal cell senescence in fibrosis from the sight of posttranslational regulation and focused on Tgfbr2, the important gatekeeper for TGF-ß1 downstream signaling. In mice with unilateral ureteral obstruction (UUO) and folic acid (FA)-induced fibrotic kidneys, we found that Tgfbr2 was markedly elevated without obvious change in its mRNA levels. As an important member of deubiquitinating enzymes, ubiquitin-specific protease 11 (Usp11) was also significantly increased in fibrotic kidneys, and co-distributed with Tgfbr2 in tubular epithelial cells. Pretreatment with Usp11 inhibitor mitoxantrone (MTX, 30 mg · kg-1 · d-1, i.p.) twice a week, for 2 weeks significantly attenuated the elevation of Tgfbr2, activation in downstream senescence-related signaling pathway, as well as renal senescence and fibrosis. In cultured mouse tubular epithelial cells (MTECs), treatment with angiotensin II (Ang-II, 10-7, 10-6 M) dose-dependently elevated both Tgfbr2 and Usp11 levels. Inhibition or knockdown on Usp11 attenuated Ang-II-induced elevation in Tgfbr2 level, and attenuated the activation of downstream senescent-related signaling pathway and as well as cell senescence. We conducted Co-IP experiments, which revealed that Usp11 was able to interact with Tgfbr2, and inhibition of Usp11 increased the ubiquitination of Tgfbr2. Taken together, these results demonstrate that the elevation of Usp11 under pathological condition is implicated in promoting renal fibrosis. Usp11 promotes the development of renal fibrosis by deubiquitinating Tgfbr2, reducing Tgfbr2 ubiquitination degradation, and then facilitating the activation of downstream senescent signaling pathway.
Assuntos
Senescência Celular , Enzimas Desubiquitinantes , Insuficiência Renal Crônica , Animais , Camundongos , Senescência Celular/fisiologia , Enzimas Desubiquitinantes/metabolismo , Células Epiteliais/metabolismo , Fibrose/metabolismo , Rim/patologia , Receptor do Fator de Crescimento Transformador beta Tipo II/metabolismo , Insuficiência Renal Crônica/patologia , Fator de Crescimento Transformador beta1/metabolismo , Ubiquitina/metabolismo , Obstrução Ureteral/complicaçõesRESUMO
Endoplasmic reticulum (ER) stress, resulting from the accumulation of misfolded and/or unfolded proteins in ER membranes, is involved in the pathogenesis of diabetic nephropathy (DN). The aim of this study was to investigate the role of ER stress inhibitors ursodeoxycholic acid (UDCA) and 4-phenylbutyrate (4-PBA) in the treatment of DN in db/db mice. Findings have revealed that diabetic db/db mice were more hyperglycemic than their non-diabetic controls, and exhibited a marked increase in body weight, water intake, urine volume, fasting plasma glucose, systolic blood pressure, glucose and insulin tolerance. UDCA (40 mg/kg/day) or 4-PBA (100 mg/kg/day) treatment for 12 weeks resulted in an improvement in these biochemical and physical parameters. Moreover, UDCA or 4-PBA intervention markedly decreased urinary albuminuria and attenuated mesangial expansion in diabetic db/db mice, compared with db/db mice treated with vehicle. These beneficial effects of UDCA or 4-PBA on DN were associated with the inhibition of ER stress, as evidenced by the decreased expression of BiP, phospho-IRE1α, phospho-eIF2α, CHOP, ATF-6 and spliced X-box binding protein-1 in vitro and in vivo. UDCA or 4-PBA prevented hyperglycemia-induced or high glucose (HG)-induced apoptosis in podocytes in vivo and in vitro via the inhibition of caspase-3 and caspase-12 activation. Autophagy deficiency was also seen in glomeruli in diabetic mice and HG-incubated podocytes, exhibiting decreased expression of LC3B and Beclin-1, which could be restored by UDCA or 4-PBA treatment. Taken together, our results have revealed an important role of ER stress in the development of DN, and UDCA or 4-PBA treatment may be a potential novel therapeutic approach for the treatment of DN.