Author + information
- Zhao Shihao,
- Fuyang Zhang,
- Yunlong Xia,
- Wenjun Yan,
- Wei Wang and
- Ling Tao
Diabetes mellitus is a world-wide health problem. Studies have demonstrated that diabetes increases myocardial susceptibility to ischemia/reperfusion (I/R) injury. Nucleostemin (NS) is a nucleoli located protein, which regulates proliferation, embryogenesis, cell cycle progression, and pre-rRNA processing. Studies have found that NS expression is induced by myocardial infarction (MI). Supression of NS expression promoted cardiomyocytes apoptosis after I/R injury. But there is still no relevant report about the role of NS in ischemic vulnerability of diabetic heart. The present study aims to determine the role of NS in diabetic heart I/R injury.
To establish diabetic models, adult male C57BL/6J mice were fed with a high fat diet (HFD). After 12 weeks, mice fed with HFD or normal diet (ND) were randomly subjected to sham or myocardial I/R operation. After 30min of ischemia and 3h or 24h reperfusion, Western blot was used to determine myocardial NS expression and real-time PCR was used to determine internal transcribed spacer (ITS, an indicator of pre-rRNA) levels. We then utilized adenovirus to overexpress NS (NS-Ad). The NS-Ad interference was implemented by injection into the apex and anterolateral wall of the heart. 3 days after injection, mice was subjected to I/R operation (ischemia for 30min, reperfusion for 24h) and divided into 8 groups: ND+GFP-Ad, ND+NS-Ad, ND+GFP-Ad+HTT (Homoharringtonine, a selective 60S ribosome inhibitor; 1.3mg/kg, intraperitoneally injected at 1, 3, 5 days before I/R operation), ND+NS-Ad+HTT, HFD+GFP-Ad, HFD+NS-Ad, HFD+GFP-Ad+HTT, HFD+NS-Ad+HTT. 24h after reperfusion, echocardiograph was performed to evaluate cardiac function, evans blue/TTC double staining was used to assess infarct size, TdT-mediated dUTP nick end labeling (TUNEL) was used to measure cardiomyocytes apoptosis.
High fat diet successfully established diabetic mice models. Myocardial NS and pre-rRNA were downregulated in HFD group at 3h after reperfusion, but their expressions were quickly restored to normal at 24h after reperfusion. While myocardial NS expression and pre-rRNA levels were unaltered after I/R in ND group, suggesting that myocardial NS expression and pre-rRNA synthesis were impaired specifically under diabetic conditions after I/R. NS-Ad administration increased myocardial pre-rRNA levels after I/R. Echocardiograph showed that NS-Ad administration significantly improved cardiac function of HFD group after I/R, and this improvement was abrogated by HTT. NS overexpression reduced TUNEL-positive cardiomyocytes and infarct sizes, which was also impaired by HTT administration. These data suggested that the protective effect of NS on diabetic myocardial I/R injury is through activation of pre-rRNA synthesis.
In the diabetic heart, reduced NS expression resulted in dysfunctional pre-rRNA synthesis and exacerbated myocardial I/R injury. Upregulation of NS in the diabetic heart may be a novel strategy to ameliorate myocardial ischemic insults.