[Recombinant plasmid pIRES2-EGFP-HCN2 improved pacing function in canine model of sick sinus syndrome].
OBJECTIVE
To construct plasmid expressing pacemaker gene pIRES2-EGFP-HCN2 and study its effects in transfected atrial myocytes in vitro and in canine model of sick sinus syndrome (SSS).


METHODS
mHCN2 gene was isolated from PTR plasmids and cloned into eukaryotic expression plasmid pIRES2-EGFP. Recombinant plasmids pIRES2-EGFP-HCN2 was transfected with by electroporation into neonatal atrial cardiomyocytes or injected to the sinoatrial (SA) region of canines with SSS induced by catheter and chemical ablation. pIRES2-EGFP-HCN2 expression was detected under fluorescence microscope and confirmed by reverse transcription-polymerase chain reaction (RT-PCR). Spontaneous beating rate in atrial cardiomyocytes was detected with light microscope.


RESULTS
EGFP expression was seen in transfected atrial cardiomyocytes 24 to 48 hours after transfection and the spontaneous beating rate was significantly increased than that in non-transfected atrial cardiomyocytes [(180 +/- 11) bpm vs (140 +/- 14) bpm, P < 0.05]. Heart rate was significantly increased 24 hours post recombinant plasmids pIRES2-EGFP-HCN2 injection compared to saline injection in canines with SSS [(150 +/- 13) bpm vs (105 +/- 17) bpm, P < 0.05]. Green fluorescence was also detected in frozen SA tissue sections of canines injected with recombinant plasmids pIRES2-EGFP-HCN2 and the production amplified by RT-PCR was about 300 bp which is consistent with mHCN2 gene fragment.


CONCLUSIONS
The recombinant eukaryotic expression plasmid pIRES2-EGFP-HCN2 can improve pacing function in atrial myocytes and in canine model of SSS.
Niu
Ping
P
Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan 430060, China.
Huang
Cong-Xin
CX
Zhao
Yue-Qiang
YQ
Yang
Bo
B
Zhao
Qing-Yan
QY
Wang
Teng
T
Fan
Guo-Hua
GH
D000818:Animals; D004195:Disease Models, Animal; D004285:Dogs; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D066298:In Vitro Techniques; D007473:Ion Channels / Q000235:genetics*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D010957:Plasmids; D012804:Sick Sinus Syndrome / Q000628:therapy*
D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D007473:Ion Channels