Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers.
We tested the ability of human mesenchymal stem cells (hMSCs) to deliver a biological pacemaker to the heart. hMSCs transfected with a cardiac pacemaker gene, mHCN2, by electroporation expressed high levels of Cs+-sensitive current (31.1+/-3.8 pA/pF at -150 mV) activating in the diastolic potential range with reversal potential of -37.5+/-1.0 mV, confirming the expressed current as I(f)-like. The expressed current responded to isoproterenol with an 11-mV positive shift in activation. Acetylcholine had no direct effect, but in the presence of isoproterenol, shifted activation 15 mV negative. Transfected hMSCs influenced beating rate in vitro when plated onto a localized region of a coverslip and overlaid with neonatal rat ventricular myocytes. The coculture beating rate was 93+/-16 bpm when hMSCs were transfected with control plasmid (expressing only EGFP) and 161+/-4 bpm when hMSCs were expressing both EGFP+mHCN2 (P<0.05). We next injected 10(6) hMSCs transfected with either control plasmid or mHCN2 gene construct subepicardially in the canine left ventricular wall in situ. During sinus arrest, all control (EGFP) hearts had spontaneous rhythms (45+/-1 bpm, 2 of right-sided origin and 2 of left). In the EGFP+mHCN2 group, 5 of 6 animals developed spontaneous rhythms of left-sided origin (rate=61+/-5 bpm; P<0.05). Moreover, immunostaining of the injected regions demonstrated the presence of hMSCs forming gap junctions with adjacent myocytes. These findings demonstrate that genetically modified hMSCs can express functional HCN2 channels in vitro and in vivo, mimicking overexpression of HCN2 genes in cardiac myocytes, and represent a novel delivery system for pacemaker genes into the heart or other electrical syncytia.
Institute of Molecular Cardiology, Departments of Physiology and Biophysics, SUNY Stony Brook, Stony Brook, NY, USA.
Irina
I
Potapova
Alexei
A
Plotnikov
Zhongju
Z
Lu
Peter
P
Danilo
Virginijus
V
Valiunas
Jihong
J
Qu
Sergey
S
Doronin
Joan
J
Zuckerman
Iryna N
IN
Shlapakova
Junyuan
J
Gao
Zongming
Z
Pan
Alan J
AJ
Herron
Richard B
RB
Robinson
Peter R
PR
Brink
Michael R
MR
Rosen
Ira S
IS
Cohen
D000109:Acetylcholine / Q000494:pharmacology; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured / Q000187:drug effects / Q000502:physiology / Q000637:transplantation; D002586:Cesium / Q000494:pharmacology; D018678:Cholinergic Agents / Q000494:pharmacology; D018920:Coculture Techniques; D004285:Dogs; D018274:Electroporation; D017629:Gap Junctions / Q000502:physiology / Q000648:ultrastructure; D015316:Genetic Therapy; D006339:Heart Rate; D006352:Heart Ventricles / Q000166:cytology; D006801:Humans; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D007473:Ion Channels / Q000502:physiology*; D017136:Ion Transport / Q000187:drug effects; D007545:Isoproterenol / Q000494:pharmacology; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects; D051379:Mice; D009124:Muscle Proteins / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques; D015221:Potassium Channels; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000502:physiology; D014162:Transfection
D000318:Adrenergic beta-Agonists; D018678:Cholinergic Agents; C578710:HCN2 protein, human; C578711:Hcn2 protein, mouse; C578712:Hcn2 protein, rat; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D007473:Ion Channels; D009124:Muscle Proteins; D015221:Potassium Channels; D011993:Recombinant Fusion Proteins; D002586:Cesium; D007545:Isoproterenol; D000109:Acetylcholine