| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2005: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2004: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
Calreticulin (CRT) is a Ca^<2+>-binding protein of the endoplasmic reticulum and essential for cardiac development. Cardiac specific overexpression of calreticulin by α-myosin heavy chain promoter results in complete heart block and sudden death in transgenic mice. For further investigating mechanism of calreticulin in cardiac development and its function, we generate transgenic mice that cause temporal and spatial overexpression of calreticulin using cre-loxP system. By microinjection of the transgene which contains a CAG promotor, a loxP-flanked chloramphenicol acetyltransferas (CAT) gene, and CRT cDNA into pronuclear cells of C57BL/6 mice, we made loxP-CRT transgenic mice. The mice were cross-bred with Nkx2.5-Cre mice which express cre recombinase under control of Nkx2.5 promoter, a transcription factor essential for early cardiac development. The breeding were resulted in transgenic mice with cardiac specific overexpression of calreticulin (Nkx2.5-CRT transgenic mice). Cardiac calreticulin protein levels of Nkx2.5-CRT transgenic mice were increased by 44-fold compared with control mice. Overexpression of calreticulin was associated with PR interval, QRS interval prolongation, bradycardia, and sudden death observed from 6- to 10-week-old. Furthermore, some NKx2.5-CRT transgenic mice revealed marked edema in 7-week-old. RT-PCR analysis revealed that the expression of hyperpolerization-activated cyclic nucleotide-gated channel1 (HCN1), essential component for cardiac pace maker activity, was receded in the heart of Nkx2.5-CRT transgenic mice. These suggested that carleticulin affect cardiac arrhythmia with disruption of cardiac signaling, such as HCN families. In conclusion, these transgenic mice are a novel model of fatal arrhythmia, and are useful model for analysis of endoplasmic reticulum protein such as calreticulin in cardiogenesis.
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