Author + information
- He Chunhui,
- Bangdang Chen,
- Yining Yang,
- Fen Liu,
- Xiaocui Chen,
- Mintao Gai and
- Yitong Ma
This study was designed to investigate the effect of NF-κB signaling pathway during acute myocardial infarction and chronic ischemic heart failure in aged mice. We used cytomegalovirus (CMV) promoter and recombinant single strand AAV9 vector and double strand AAV9 vector to validate tissue-specific expression, and screened to obtain better AAV9 vectors which can be used in gene therapy of heart disease. We used IKBα (inhibitor of kappa Bα, IKBα) gene recombinant AAV9 vector for the targeting transduction in aging mouse heart, to investigate whether exogenously introduced IKBα can be used in gene therapy, and lead to targeted depression of NF-κB, reduction of myocardial infarction, inhibition of myocardial apoptosis, and improve survive of ischemia heart failure in aging mice.
Male C57BL/6J mice with age of 15 to 18-month were selected, to establish acute myocardial infarction model via left coronary artery liagation. The myocardial infarct size, myocardial apoptosis, IKBα, and the expression of p65 of p50 in cytoplasm and nuclear were observed on day 3 and day 28. ssAAV9-CMV-eGFP and dsAAV9-CMV-eGFP vector with the dose of 1 × 1011 vg were injected into C57BL/6J mice via the tail vein, respectively. At 0 week, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks and 8 weeks of viral vector transduction, the laser confocal and Western Blot were used to detect GFP protein expression in heart, liver, spleen, lung, kidney, brain and skeletal muscle. We evaluated the tissue targeting effects of ssAAV9 and dsAAV9 vectors with CMV promoter in vivo.
It is observed that p65 and p50 positive cells are much higher in MI mice than in SH mice by immunohistochemistry methods at 3 d and 28 d (p65 410 vs 328, P< 0.05; IKBα 350 vs 101, P< 0.05). Expression of p65 in MI 3d, 28d group in old mice is higher than in SH mice, which means higher activities of NF-κB in mice after myocardial infarction. There were statistics differences in expression of MMP-2 and MMP-9 between SH and MI. Expression of MMP-2 and MMP-9 were higher in MI than in SH in at 3 d and 28 d (MMP-2 420 vs 582,P< 0.05; MMP-9 568 vs 365, P< 0.05). Apoptosis rate compare with SH were 11.5 ± 0.35% vs 0.29 ± 0.03%, P<0.001.
NF-κB signaling pathway is involved in myocardial infarction and consequent chronic heart failure in aged mice. CMV promoter recombinant dsAAV9 vectors can be expressed in cardiac more efficiently and stably for a long-term comparing to ssAAV9 vector. While, the CMV promoter activity can easily be silent in the liver, dsAAV9-CMV vector is an ideal vector for gene therapy of heart disease.