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Optogenetics-enabled assessment of viral gene and cell therapy for restoration of cardiac excitability.

Ambrosi CM, Boyle PM, Chen K, Trayanova NA, Entcheva E - Sci Rep (2015)

Bottom Line: Multiple cardiac pathologies are accompanied by loss of tissue excitability, which leads to a range of heart rhythm disorders (arrhythmias).Taken directly, these results can help guide optogenetic interventions for light-based control of cardiac excitation.More generally, our findings can help optimize gene therapy for restoration of cardiac excitability.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY.

ABSTRACT
Multiple cardiac pathologies are accompanied by loss of tissue excitability, which leads to a range of heart rhythm disorders (arrhythmias). In addition to electronic device therapy (i.e. implantable pacemakers and cardioverter/defibrillators), biological approaches have recently been explored to restore pacemaking ability and to correct conduction slowing in the heart by delivering excitatory ion channels or ion channel agonists. Using optogenetics as a tool to selectively interrogate only cells transduced to produce an exogenous excitatory ion current, we experimentally and computationally quantify the efficiency of such biological approaches in rescuing cardiac excitability as a function of the mode of application (viral gene delivery or cell delivery) and the geometry of the transduced region (focal or spatially-distributed). We demonstrate that for each configuration (delivery mode and spatial pattern), the optical energy needed to excite can be used to predict therapeutic efficiency of excitability restoration. Taken directly, these results can help guide optogenetic interventions for light-based control of cardiac excitation. More generally, our findings can help optimize gene therapy for restoration of cardiac excitability.

No MeSH data available.


Related in: MedlinePlus

Inscription of light sensitivity by viral gene or cell delivery.(a) Schematic for gene delivery of (GD) of ChR2. In cardiomyocytes (CMs) where ChR2 transduction was successful (left), channels that conduct light-sensitive current (IChR2; blue) are trafficked to the sarcolemma; these cells are connected via gap junctions (orange) to normal, non-transduced heart cells (right), as in unmodified tissue. (b) Superimposed immunofluorescence images of cardiac cell monolayers with light-sensitivity inscribed by GD showing co-localization of markers for CMs (α-actinin, red) and ChR2 (eYFP, green). (c) EYFP intensity profile of ChR2-expressing CMs along the cross section indicated by the dotted white line in (b). (d) Schematic for cell delivery (CD) of ChR2. ChR2-rich donor cells (left) co-cultured with normal CMs (right) form a light-sensitive heterocellular monolayer. Membrane behaviour in donor cells is considered passive and represented by a leakage current (IPASV) with properties based on experimental measurements from HEK cells. (e) Superimposed immunofluorescence images of monolayers with light-sensitivity inscribed by CD showing distinct, adjacent CMs (red) and ChR2-expressing donor cell (green) regions. (f) EYFP intensity profile of ChR2-expressing donor cells along the cross section indicated by the dotted white line in (e). Generic cardiac ion channels (yellow) are shown in CM schematic. Scale bars in (b,d) are 10 μm.
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f1: Inscription of light sensitivity by viral gene or cell delivery.(a) Schematic for gene delivery of (GD) of ChR2. In cardiomyocytes (CMs) where ChR2 transduction was successful (left), channels that conduct light-sensitive current (IChR2; blue) are trafficked to the sarcolemma; these cells are connected via gap junctions (orange) to normal, non-transduced heart cells (right), as in unmodified tissue. (b) Superimposed immunofluorescence images of cardiac cell monolayers with light-sensitivity inscribed by GD showing co-localization of markers for CMs (α-actinin, red) and ChR2 (eYFP, green). (c) EYFP intensity profile of ChR2-expressing CMs along the cross section indicated by the dotted white line in (b). (d) Schematic for cell delivery (CD) of ChR2. ChR2-rich donor cells (left) co-cultured with normal CMs (right) form a light-sensitive heterocellular monolayer. Membrane behaviour in donor cells is considered passive and represented by a leakage current (IPASV) with properties based on experimental measurements from HEK cells. (e) Superimposed immunofluorescence images of monolayers with light-sensitivity inscribed by CD showing distinct, adjacent CMs (red) and ChR2-expressing donor cell (green) regions. (f) EYFP intensity profile of ChR2-expressing donor cells along the cross section indicated by the dotted white line in (e). Generic cardiac ion channels (yellow) are shown in CM schematic. Scale bars in (b,d) are 10 μm.

Mentions: As illustrated in Fig. 1a,b, in the case of GD, cardiomyocytes were directly infected with a custom-made adenovirus containing the transgene for ChR2-eYFP. Opsin-expressing cardiomyocytes then formed gap junctions with other cardiomyocytes (both opsin-expressing and unmodified)18. In the case of CD (Fig. 1d,e), optical sensitivity was achieved by incorporating ChR2-rich somatic (HEK) cells within the syncytium. The donor (HEK) cells, which were light-sensitive and conductive, but non-excitable, formed gap junctions with native, unmodified cardiomyocytes14. Regardless of delivery method (GD or CD), ChR2 was expressed as a membrane-specific ion channel (Fig. 1b,e, green). eYFP intensity profiles showed that the expression of the reporter (as a surrogate for ChR2 expression) was similar between transduced myocytes (GD) and ChR2-expressing donor cells (CD) in syncytial distributions (Fig. 1c,f).


Optogenetics-enabled assessment of viral gene and cell therapy for restoration of cardiac excitability.

Ambrosi CM, Boyle PM, Chen K, Trayanova NA, Entcheva E - Sci Rep (2015)

Inscription of light sensitivity by viral gene or cell delivery.(a) Schematic for gene delivery of (GD) of ChR2. In cardiomyocytes (CMs) where ChR2 transduction was successful (left), channels that conduct light-sensitive current (IChR2; blue) are trafficked to the sarcolemma; these cells are connected via gap junctions (orange) to normal, non-transduced heart cells (right), as in unmodified tissue. (b) Superimposed immunofluorescence images of cardiac cell monolayers with light-sensitivity inscribed by GD showing co-localization of markers for CMs (α-actinin, red) and ChR2 (eYFP, green). (c) EYFP intensity profile of ChR2-expressing CMs along the cross section indicated by the dotted white line in (b). (d) Schematic for cell delivery (CD) of ChR2. ChR2-rich donor cells (left) co-cultured with normal CMs (right) form a light-sensitive heterocellular monolayer. Membrane behaviour in donor cells is considered passive and represented by a leakage current (IPASV) with properties based on experimental measurements from HEK cells. (e) Superimposed immunofluorescence images of monolayers with light-sensitivity inscribed by CD showing distinct, adjacent CMs (red) and ChR2-expressing donor cell (green) regions. (f) EYFP intensity profile of ChR2-expressing donor cells along the cross section indicated by the dotted white line in (e). Generic cardiac ion channels (yellow) are shown in CM schematic. Scale bars in (b,d) are 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4664892&req=5

f1: Inscription of light sensitivity by viral gene or cell delivery.(a) Schematic for gene delivery of (GD) of ChR2. In cardiomyocytes (CMs) where ChR2 transduction was successful (left), channels that conduct light-sensitive current (IChR2; blue) are trafficked to the sarcolemma; these cells are connected via gap junctions (orange) to normal, non-transduced heart cells (right), as in unmodified tissue. (b) Superimposed immunofluorescence images of cardiac cell monolayers with light-sensitivity inscribed by GD showing co-localization of markers for CMs (α-actinin, red) and ChR2 (eYFP, green). (c) EYFP intensity profile of ChR2-expressing CMs along the cross section indicated by the dotted white line in (b). (d) Schematic for cell delivery (CD) of ChR2. ChR2-rich donor cells (left) co-cultured with normal CMs (right) form a light-sensitive heterocellular monolayer. Membrane behaviour in donor cells is considered passive and represented by a leakage current (IPASV) with properties based on experimental measurements from HEK cells. (e) Superimposed immunofluorescence images of monolayers with light-sensitivity inscribed by CD showing distinct, adjacent CMs (red) and ChR2-expressing donor cell (green) regions. (f) EYFP intensity profile of ChR2-expressing donor cells along the cross section indicated by the dotted white line in (e). Generic cardiac ion channels (yellow) are shown in CM schematic. Scale bars in (b,d) are 10 μm.
Mentions: As illustrated in Fig. 1a,b, in the case of GD, cardiomyocytes were directly infected with a custom-made adenovirus containing the transgene for ChR2-eYFP. Opsin-expressing cardiomyocytes then formed gap junctions with other cardiomyocytes (both opsin-expressing and unmodified)18. In the case of CD (Fig. 1d,e), optical sensitivity was achieved by incorporating ChR2-rich somatic (HEK) cells within the syncytium. The donor (HEK) cells, which were light-sensitive and conductive, but non-excitable, formed gap junctions with native, unmodified cardiomyocytes14. Regardless of delivery method (GD or CD), ChR2 was expressed as a membrane-specific ion channel (Fig. 1b,e, green). eYFP intensity profiles showed that the expression of the reporter (as a surrogate for ChR2 expression) was similar between transduced myocytes (GD) and ChR2-expressing donor cells (CD) in syncytial distributions (Fig. 1c,f).

Bottom Line: Multiple cardiac pathologies are accompanied by loss of tissue excitability, which leads to a range of heart rhythm disorders (arrhythmias).Taken directly, these results can help guide optogenetic interventions for light-based control of cardiac excitation.More generally, our findings can help optimize gene therapy for restoration of cardiac excitability.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY.

ABSTRACT
Multiple cardiac pathologies are accompanied by loss of tissue excitability, which leads to a range of heart rhythm disorders (arrhythmias). In addition to electronic device therapy (i.e. implantable pacemakers and cardioverter/defibrillators), biological approaches have recently been explored to restore pacemaking ability and to correct conduction slowing in the heart by delivering excitatory ion channels or ion channel agonists. Using optogenetics as a tool to selectively interrogate only cells transduced to produce an exogenous excitatory ion current, we experimentally and computationally quantify the efficiency of such biological approaches in rescuing cardiac excitability as a function of the mode of application (viral gene delivery or cell delivery) and the geometry of the transduced region (focal or spatially-distributed). We demonstrate that for each configuration (delivery mode and spatial pattern), the optical energy needed to excite can be used to predict therapeutic efficiency of excitability restoration. Taken directly, these results can help guide optogenetic interventions for light-based control of cardiac excitation. More generally, our findings can help optimize gene therapy for restoration of cardiac excitability.

No MeSH data available.


Related in: MedlinePlus