Puromycin aminonucleoside triggers apoptosis in podocytes by inducing endoplasmic reticulum stress.
10.23876/j.krcp.2018.37.3.210
- Author:
Seo Yoon MIN
1
;
Dong Soo HA
;
Tae Sun HA
Author Information
1. Department of Pediatrics, Chungbuk National University College of Medicine, Cheongju, Korea. tsha@chungbuk.ac.kr
- Publication Type:Original Article
- Keywords:
Apoptosis;
Endoplasmic reticulum stress;
Podocyte;
Proteinuria;
Puromycin aminonucleoside
- MeSH:
Animals;
Apoptosis*;
Blotting, Western;
Caspase 12;
DNA Nucleotidylexotransferase;
Endoplasmic Reticulum Stress*;
Endoplasmic Reticulum*;
Flow Cytometry;
Humans;
Immunoglobulins;
In Situ Nick-End Labeling;
In Vitro Techniques;
Mice;
Models, Animal;
Nephrosis;
Nephrotic Syndrome;
Oxidative Stress;
Phosphatidylinositol 3-Kinases;
Podocytes*;
Proteinuria;
Puromycin Aminonucleoside*;
Puromycin*;
RNA, Small Interfering;
Sodium;
Transcription Factors
- From:Kidney Research and Clinical Practice
2018;37(3):210-221
- CountryRepublic of Korea
- Language:English
-
Abstract:
BACKGROUND: Puromycin aminonucleoside (PAN) is a known podocytotoxin. PAN-induced nephrosis is a widely used animal model for studying human idiopathic nephrotic syndrome. Abnormal protein accumulation associated with podocyte-specific endoplasmic reticulum (ER) stress damages cells structurally and functionally, which in turn induces apoptosis and severe proteinuria. In the present study, we investigated the effect of PAN on ER stress and apoptosis in podocytes in vitro. METHODS: Mouse podocytes were cultured and treated with various concentrations of PAN. ER stress markers were then evaluated by western blotting, and apoptosis was evaluated by fluorescence-activated cell sorting (FACS) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays. RESULTS: PAN treatment increased ER stress markers such as activating transcription factor (ATF) 6α and caspase-12 in a dose-dependent manner at 12 and 24 hours, respectively. These markers were reduced by chemical chaperones, such as sodium 4-phenylbutyric acid and tauroursodeoxycholic acid. PAN treatment also increased 78 kD glucose-regulated protein (GRP78)/binding immunoglobulin protein (BiP) at the earlier stage of 12 hours. PAN significantly induced podocyte apoptosis in concentration- and time-dependent manners, as seen using FACS and TUNEL assays. This result was improved by Nox4 siRNA, ATF6 siRNA, and chemical chaperones. LY294002, a PI3-kinase inhibitor, significantly boosted ER stress and apoptosis. PAN-induced ER stress increased oxidative stress and subsequently induced apoptosis, and could be mitigated by inhibition of PI3-kinase signaling. CONCLUSION: Our findings suggest that PAN induces ER stress in podocytes mainly through the GRP78/BiP, ATF6α, and caspase-12 pathways, which trigger apoptosis via induction of oxidative stress. This stress is mitigated by inhibiting PI3-kinase signaling.
