Adolescent ; Cardiomyopathies ; complications ; genetics ; Female ; Heart Ventricles ; abnormalities ; Humans ; Pedigree ; Sick Sinus Syndrome ; complications ; genetics

Electrocardiography ; GTP-Binding Protein beta Subunits/genetics* ; Humans ; Mutation ; Pedigree ; Sick Sinus Syndrome/genetics*

The effective therapeutics for the sinoatrial node (SAN) pacemaker dysfunction induced by SCN5A gene mutation this is still being explored recently. In this study, a two-dimensional experimental model of rabbit SAN-atrial cell system which proposed by Zhang et al., was used as a prototype, the gene mutation was considered, and effects of both the acid concentration and temperature were also introduced. The effects of acid concentration and temperature on sick sinus syndrome (SSS) at the tissue level were investigated by simulation. The results showed that the SAN abnormal pacemaker could be caused by the reduction of I(Na), which is induced by the two mutations of T220I and delF1617. The results also showed that if we properly adjusted the acid concentration and temperature of the system, not only could we increase the relevant currents, but also could we increase I(Na) which reduced by gene mutations, so that the pacemaking behavior of SAN tissue could return to normal state from abnormalities. The above simulation results imply that the abnormal pacemaking of SAN system may closely relate to the gene mutation of ion channel mutations, and the acid concentration and temperature may play a modulatory role. Our study could be useful for clinical medical diagnosis and therapy of cardiac disease.
Acids ; Animals ; Computer Simulation ; Mutation ; NAV1.5 Voltage-Gated Sodium Channel ; genetics ; Rabbits ; Sick Sinus Syndrome ; etiology ; genetics ; Sinoatrial Node ; pathology ; Temperature

OBJECTIVETo 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).

METHODSmHCN2 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.

RESULTSEGFP 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.

CONCLUSIONThe recombinant eukaryotic expression plasmid pIRES2-EGFP-HCN2 can improve pacing function in atrial myocytes and in canine model of SSS.

Animals ; Disease Models, Animal ; Dogs ; Gene Expression ; Gene Transfer Techniques ; Genetic Therapy ; Genetic Vectors ; Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels ; In Vitro Techniques ; Ion Channels ; genetics ; Myocytes, Cardiac ; metabolism ; Plasmids ; Sick Sinus Syndrome ; therapy