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Published on: September 17, 2009
Establishing a low-expression renalase gene model in cardiac tissue of Sprague-Dawley rats
1Department of Cardiology, The Third Xiangya Hospital of Central South University, 410013, Changsha, Hunan, China.
Background:
Renalase is a novel secretory amino oxidase expressed in the kidney and heart. To study the protective mechanism of renalase in local heart tissue, we established a low-expression renalase model with lentivirus (LV)-mediated RNA interference technology.
Materials And Methods:
Three renalase-targeting oligonucleotides were designed after analyzing the mRNA of renalase. LV particles were prepared with LV expression systems (using the Trono 3 plasmid component system), after which LV-RNLS-shRNAs and LV-NC-shRNA were transfected into H9C2 cells in different cell culture plates. The optimal oligonucleotide was screened by real-time PCR and Western blot. These techniques were also used to detect renalase gene expression in the heart tissue.
Results:
In the cell screening experiment, the efficacy of the inhibition of renalase mRNA expression was 93.7 % and that of renalase protein expression was 83.1 % in H9C2 cells. When the oligonucleotide was injected into the pericardial cavities of the SD rats on the 10th day, it inhibited 63.9 % of the expression of renalase protein in the heart tissue.
Conclusion:
LV-RNLS-RNAi (19813-1) can be used to establish an optimal renalase low-expression model for further research on the renalase system.
