Quantitative real-time PCR analysis
Total RNA was extracted from kidney tissue using total RNA extraction Kit (BioTek, Winooski, VT, United States). Reverse transcription was performed using PrimeScript RT Reagent Kit (Takara Bio, Inc., Otsu, Japan) according to instruction. Reactions of PCR amplification were quantified using the iTaq Universal SYBR Green Supermix (Bio-Rad Laboratories, Inc.) in a PCR system (CFX Connect; Bio-Rad, Hercules, CA, United States). Results were presented as relative mRNA levels to GAPDH.