Cytoarchitecture of the Heart
The Ehler group aims to look at the organisation of the heart cell (cardiomyocyte) at a subcellular level; mainly from a cytoskeletal and signalling point of view. We want to find out how its cytoarchitecture is assembled during development and how adaptations to increased work load or during cardiac disease are made.
Heart cells have an extremely regular cytoskeleton with their contractile elements assembled to paracrystalline myofibrils. They are also mechanically and electro-chemically connected to each other at specialised sites of cell-cell contact: the intercalated discs. Any compromise to this extremely regular arrangement has immediate consequences on heart function.
The major focus of our research is elucidating how myofibrils and intercalated disks are assembled in heart cells during development; how they are affected in the diseased heart and what the functional basis is for the adaptations of cardiomyocytes during development.
Only if we understand what goes wrong in cardiomyopathy at the level of the cell and know more about the signalling pathways that cause these changes, will we be able to do more than just treating symptoms.
Current PhD students:
Ms Amy Pearce
Ms Beth Ormrod
Mr Mihai Pruna
Ms Thayane Crestani
Projects

Heart tissue
Embryonic heart tissue, where the myofibrils are stained in green, the microtubules are stained in red and the nuclei are stained in blue. Two dividing cardiomyocytes can be seen in the top right-hand corner of the image, where the myofibrils have been disassembled.

Freshly isolated heart cells
Freshly isolated adult heart cell showing the typical rod-like shape and the extremely ordered cytoskeleton, nuclei are shown in blue and cross-striated myofibrils in green (MyBP-C) and red (alpha-actinin).

Heart cells in culture
Heart cells in culture retain their regular cytoskeleton to a certain extent and continue to beat. The actin cytoskeleton was stained in red, the myofibrils in green (MyBP-C) and blue (sarcomeric myosin heavy chain).

Diseased heart
The intercalated discs (a specialised type of cell contacts) are altered in their composition and ultrastructure in dilated cardiomyopathy (i.e. a ballooned heart), while the myofibrils appear unchanged (intercalated discs are shown in green, myofibrils shown in red).
Publications
Methods and protocols
Chick heart whole mount preparations (Ehler et al. 1999)
The heart was dissected in cold PBS and fixed in 3% PFA (paraformaldehyde) in PBS with 0.002% Triton X-100 (PBT) for 1 hour.
After several washes in PBT and permeabilisation with 0.1%Triton X-100 in PBS for 30 minutes, the hearts were treated with hyaluronidase (1 mg/ml; Sigma, St Louis, MO, USA) in PBT for 30 minutes at RT (room temperature) in order to remove the cardiac jelly and to ensure access for the antibodies to the inner myocardial wall.
After further washes in PBT and blocking with 5% NGS (normal pre-immune goat serum), 1% BSA (bovine serum albumin) in Gold buffer for 30 minutes, the hearts were incubated with the primary antibody mixtures, diluted in blocking solution, overnight at 4°C.
After 6x 20 min washing in PBT the secondary antibodies were applied for either 6 hours at RT or overnight at 4°C.
The hearts were washed in PBT (6x 20 min) and mounted in 0.1 M Tris-HCl (pH 9.5)-glycerol (3:7) including 50 mg/ml n-propyl gallate as anti-fading reagent.
Same procedure works for mouse embryos, just incubations times have to be increased - enjoy!
Projects

Heart tissue
Embryonic heart tissue, where the myofibrils are stained in green, the microtubules are stained in red and the nuclei are stained in blue. Two dividing cardiomyocytes can be seen in the top right-hand corner of the image, where the myofibrils have been disassembled.

Freshly isolated heart cells
Freshly isolated adult heart cell showing the typical rod-like shape and the extremely ordered cytoskeleton, nuclei are shown in blue and cross-striated myofibrils in green (MyBP-C) and red (alpha-actinin).

Heart cells in culture
Heart cells in culture retain their regular cytoskeleton to a certain extent and continue to beat. The actin cytoskeleton was stained in red, the myofibrils in green (MyBP-C) and blue (sarcomeric myosin heavy chain).

Diseased heart
The intercalated discs (a specialised type of cell contacts) are altered in their composition and ultrastructure in dilated cardiomyopathy (i.e. a ballooned heart), while the myofibrils appear unchanged (intercalated discs are shown in green, myofibrils shown in red).
Publications
Methods and protocols
Chick heart whole mount preparations (Ehler et al. 1999)
The heart was dissected in cold PBS and fixed in 3% PFA (paraformaldehyde) in PBS with 0.002% Triton X-100 (PBT) for 1 hour.
After several washes in PBT and permeabilisation with 0.1%Triton X-100 in PBS for 30 minutes, the hearts were treated with hyaluronidase (1 mg/ml; Sigma, St Louis, MO, USA) in PBT for 30 minutes at RT (room temperature) in order to remove the cardiac jelly and to ensure access for the antibodies to the inner myocardial wall.
After further washes in PBT and blocking with 5% NGS (normal pre-immune goat serum), 1% BSA (bovine serum albumin) in Gold buffer for 30 minutes, the hearts were incubated with the primary antibody mixtures, diluted in blocking solution, overnight at 4°C.
After 6x 20 min washing in PBT the secondary antibodies were applied for either 6 hours at RT or overnight at 4°C.
The hearts were washed in PBT (6x 20 min) and mounted in 0.1 M Tris-HCl (pH 9.5)-glycerol (3:7) including 50 mg/ml n-propyl gallate as anti-fading reagent.
Same procedure works for mouse embryos, just incubations times have to be increased - enjoy!