Supported by the Heart and Stroke Foundations of Manitoba,

New Brunswick and Canada, Canadian Institutes for Health

Research and St. Boniface Hospital and Research Foundation.

doi:10.1016/j.yjmcc.2006.03.283

A80. The functional role of endoglin in mammalian

cardiovascular development

Gregory Anderson a, Michelle Letarte b, Mira Puri a.a Sunnybrook and Women’s Research Institute. b The Hospital

for Sick Children

A multitude of molecular mechanisms control the differen-

tiation and assembly of the cell lineages involved in

cardiovascular development, and our understanding of this

process has grown significantly through the characterization of

signaling modulators such as those in the TGF-h superfamily.

Endoglin is an ancillary transmembrane receptor for the TGF-hsuperfamily that is expressed in endothelial cells (ECs) from

early gestation to the adult stage. In humans, the critical role of

endoglin in vascular development is shown in patients with

Hereditary Hemorrhagic Telangiectasia type 1 (HHT-1), a

disease characterized by arteriovenous malformations in the

pulmonary, cerebral, and hepatic circulations.

Mice bearing a homozygous null mutation in endoglin

(Eng�/�) die at mid-gestation due to circulatory failure and

hemorrhage. Their ECs lack integrity, angiogenesis in the yolk

sac is abrogated, and their hearts fail to undergo the normal

morphogenetic looping that generates the four-chambered heart.

However, the specific role of endoglin in both blood vessel

maturation and cardiac development still needs to be clarified.

Preliminary experimental results using chimeric mice have

shown that endoglin is necessary for endocardial cushion for-

mation during heart development. However, very few chimeric

mice survive to birth, and thus, several embryonic stem (ES)-cell

based assays have been performed to characterize the ability of

Eng�/� cells to contribute to cardiovascular development.

We will show how we are designing relevant experiments to

further characterize this receptor. The information obtained

from these experiments will have important consequences for

patients who suffer from HHT-1, and will help to clarify the

role of TGF-h signaling in the developing mammalian embryo

and the implications it has in cardiac development.

doi:10.1016/j.yjmcc.2006.03.284

A47. Decreased levels of non-nuclear muscle LIM protein

are associated with cardiac dysfunction and heart failure

S.Y. Boateng a, R.J. Belin a, D.L. Geenen a, K.B. Margulies b,

P.P. de Tombe a, B. Russell a. a Univ. of Illinois at Chicago,

Chicago, IL. b Univ. of Pennsylvania, Philadelphia, PA

Prolonged hemodynamic overload results in cardiac hypertro-

phy with detrimental changes in myocardial gene expression and

morphology that predict morbidity and mortality. Stretch sensors,

necessary for myocyte enlargement, are thought to become

impaired as a result of prolonged overload. However, the

mechanisms by which chronic overload leads to failure are

poorly understood. Cardiac muscle LIM protein (MLP) is thought

to be a mechanosensor, possibly relaying signals from the

cytoskeleton to the nucleus. Therefore the subcellular location

of myocytic MLP may have important functional implications in

health and disease.

Using subcellular fractionation and immuno-cytochemistry,

we have determined some of the factors that regulate the

subcellular location of MLP in cardiac myocytes. We found

MLP in the cytoskeletal, membrane and nuclear subcellular

compartments. Cytoskeletal MLP was most prominent in the M-

band. In cultured cardiac rat neonatal myocytes, 10 AMisoproterenol treatment for 24 h increased nuclear MLP, whilst

10 AMverapamil decreased it. This suggests that calcium cycling

plays a role in determining the subcellular distribution of MLP.

However, cyclic stretch alone at 20% maximum strain, 1 Hz for

24 or 48 h (conditions that induce myocyte hypertrophy), did not

translocate MLP to the nucleus. This suggests that MLP nuclear

translocation requires more than mechanical strain alone.

Transcriptional blockade by actinomycin D showed that the

translocation of MLP to the nucleus is regulated by the

nucleolus.In failing human hearts and ventricles from aortic

banded andmyocardial infarcted rat hearts, there was a significant

accumulation of nuclear MLP and a significant decrease in other

non-nuclear compartments. Thus, cardiac dysfunction and failure

may be associated with aberrant MLP subcellular localization

following prolonged overload.

Funded by AHA 0630307N and NIH HL 62426.

doi:10.1016/j.yjmcc.2006.03.285

A99. Protection by constitutively active PKC( expression is

blocked by CL� channel inhibition in cardiomyocytes

Azadeh Boloorchi, Roberto J. Diaz, He Ping Zeng,

Alina Hinek, Taneya Hossain, Gregory J. Wilson. The Hospital

for Sick Children, Toronto, Canada

Overexpression of constitutively active PKC( has been showntomimic protection induced by ischemic preconditioning (IPC) in

hearts. We have shown that chloride (Cl�) channel dependent

enhanced cell volume regulation is a key mechanism in IPC. In

the present study, we tested in cardiomyocytes (1) if over-

expression of constitutively active PKC( confers a protection

similar to pharmacological activation of A1/A3 adenosine

receptors using N6-2-(4-aminophenyl) ethyl adenosine (AP-

NEA), and (2) if this protection requires Cl� channel activity.

Rabbit ventricular myocytes(5-8 hearts), cultured for 48 h, were

exposed to 60 min simulated ischemia (SI) followed by 60-min

simulated reperfusion (SR) and the percentage of dead myocytes

was assessed before and after SI/SR using trypan blue staining.

Myocytes were preconditioned either by transfection with a

recombinant adenoviral vector encoding both EGFP and mutant

PKC( genes to overexpress a constitutively active form of PKC((AdEGFPAE5) 44 h before exposure to SI/SR, or by treatment

ABSTRACTS / Journal of Molecular and Cellular Cardiology 40 (2006) 862–918864