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| Gene Synthesis | Total RNA of rat heart was extracted using Trizol reagent (15596018, Thermo Fisher, USA) and reverse translated with High- Capacity cDNA Reverse Transcription Kit (4368814, Thermo Fisher, USA) following the manufacturer’s protocol. Real-time PCR was performed using the 7500 Real- Time PCR Products (Applied Biosystems) with PowerUpTM SYBRR Green Master Mix (A25742, Thermo Fisher, USA). The reaction program was as follows: 50 ˚C for 2 min, 95 ˚C for 2 min, following by 40 cycles of 95 ˚C for 15 min, 57 ˚C for 15 min, 72 ˚C for 1 min. Copy numbers of the GAPDH housekeeping gene (B661204- 0001, BBI Life Sciences), TNF-α and IL-1β were determined. The following primers were designed and synthesized by GenScript Biotech Corp (Nan Jing, China): TNF-α, forward 5’-GCATGATCCGAGATGTGGAA-3’, reverse 5’- TGAGAAGAGGCTGAGGCACA-3’(98bp); IL-1β, forward 5’-AAATGCCTCGTGCTGTCTGA-3’, reverse 5’- TTGGGATCCACACTCTCCAG-3’ (218 bp). The relative expression levels of the genes were performed using the comparative quantification cycle (2-ΔΔCT) method (21). Each sample was analyzed in triplicate. | Get A Quote |
Objective(s): Myocarditis is characterized by inflammatory cell infiltration in myocardial stroma. Attenuation of tumor necrosis factor (TNF)-α and interleukin (IL)-1β is a reliable mark for improving the prognosis. Protein kinase B (Akt) plays an important role in the development and progression of myocarditis. The specific role of the natural inhibitor of Akt, Deguelin, on myocarditis has not been reported. In this study, we used deguelin to investigate the effects of natural Akt inhibitor on myocarditis in experimental autoimmune myocarditis (EAM) rats. Materials and Methods: EAM rat models were made by using Lewis rats and Deguelin was injected intraperitoneally on day 3, 6, 9, 12 and 15 after successf... More
