2%, of the left ventricle). IPC significantly (P < 0.001) reduced

infarct size (% of area at risk, 33 T 2%) vs Ctrl (51 T 2%). Chel

or (V1-2 alone in the absence of IPC did not alter infarct size

vs Ctrl (preisch, 47 T 3% or 51 T 2%; or at reperfusion, 48 T1% or 47 T 1%). Conversely, pre-treatment with a non-selective

PKC inhibitor or PKC( specific antagonist abolished the

infarct-sparing effects of IPC (47 T 3% or 47 T 2%).

Furthermore, when the same inhibitors were administered at

the onset of the 3 hr reperfusion period, the cardioprotective

effects of IPC were also reversed (48 T 3% or 48 T 2%). Thus,

these data suggest that IPC reduces infarct size by mechanisms

involving PKC( during reperfusion.

doi:10.1016/j.yjmcc.2006.03.419

A113. Ultrastructural and immuno-characterization of

cardiac progenitor cells in the embryonic myocardium

Feixiong Zhang, Kishore B.S. Pasumarthi

The mammalian heart is formed from the progenitor cells

present in the primary heart field (PHF) and anterior heart

field (AHF). Several rare populations of myogenic progen-

itor cells have been identified in the postnatal myocardium.

However, there is no information available on the existence

of cardiac progenitor cells in the embryonic heart post-

chamber specification. While markers for the AHF are

clearly defined (Isl1 and FGF10), markers exclusively

expressed in the PHF are unknown. The transcription factor,

Nkx2.5 is expressed in progenitor cells of both heart fields

as well as in the differentiated myocardial cells later in the

development. We examined subcellular characteristics of

E11.5 ventricular myocardial cells using Transmission

Electron Microscopy (TEM) and immunogold labeling

techniques. At the ultrastructural level, we observed three

different cell populations within the myocardial layer which

include undifferentiated cells (41 T 7%), moderately

differentiated cells (38 T 3%) and mature cardiomyocytes

(21 T 10%). Undifferentiated cells contained a large nucleus

and very sparse cytoplasm with no myofibrillar bundles.

Moderately differentiated cells contained randomly arranged

myofilaments in the cytoplasm. In contrast, mature cardio-

myocytes exhibited well developed sarcomere structures. All

three cell types formed cell to cell junctions with adjacent

cells via desmosomes. Intercalated discs were only observed

in many of the differentiated cells. We also confirmed the

presence of similar undifferentiated cells albeit at low levels

(5–7%) in the E18.5 myocardium. Furthermore, we tested

whether these distinct cell populations were also positive for

markers such as Nkx2.5, Isl1 and ANF. Preponderance of

anti-Nkx2.5 label was found in undifferentiated and moder-

ately differentiated cell types. Anti-ANF label was found

only in the cytoplasmic compartment of moderately differ-

entiated and mature myocardial cells. All the undifferentiated

cells were negative for anti-ANF labeling. We did not find

immunogold labeling with Isl1 in all these myocardial cell

types. Our results indicate that undifferentiated myocardial

cells expressing only Nkx2.5 may represent progenitor cell

population while cells expressing Nkx2.5 and or ANF

represent differentiating myocytes.

doi:10.1016/j.yjmcc.2006.03.420

A35. Prevention of pore-formation by voltage-dependent

anion channel protects against mitochondrial dysfunction

and cell death

Jun Zhang, Thomas M. Vondriska, David A. Liem,

Shushi Nagamori, Jeff Abramson, Guangwu Wang

Rachna Ujwal, Chenggong Zong, Michael J. Zhang,

James N. Weiss, Ronald H. Kaback, Peipei Ping. Depts. of

Physiology and Medicine/Division of Cardiology, UCLA,

Los Angeles, CA

Despite the increasingly recognized role of mitochondrial

permeability transition (MPT) in cell death following stress,

the underlying regulatory mechanisms of the MPT pore

remain elusive. Understanding the contributions of individ-

ual components of the MPT pore to modulate MPT is a

prominent challenge in this regard. In the outer membrane,

the voltage-dependent anion channel (VDAC) has been

implicated as a central component of the MPT pore,

however, the fundamental regulation of the 3 known

mammalian isoforms of VDAC is largely unknown due to

confounding technical difficulties with studying membrane

proteins. Using functionally viable recombinant VDAC

proteins and custom-made isoform-specific antibodies, we

conducted full characterization of VDAC isoforms with

respect to their expression pattern, contributions to the MPT

pore, and functional regulation. We found that VDAC1/2,

but not VDAC3, were associated with hexokinase II-

enriched membrane fractions. Functional pore formation

was determined using proteoliposomes; VDAC3 appeared to

be less prone to form a pore in response to ROS as

compared to VDAC1 and 2. Regulation of VDAC isoforms

and MPT pore was investigated using PKC( as a model

system. Enzymatic active PKC( was sufficient to attenuate

ROS-induced pore-formation in proteoliposomes containing

VDAC1/2, as well in isolated cardiac mitochondria. Lastly,

the physiological role of VDAC was definitively examined

via administration of the novel VDAC inhibitor (Ro68-

3400) to mice at the clinically relevant time point at which

the heart is reperfused. Inhibition of VDAC afforded a

powerful infarct-sparing effect in vivo, equivalent in

magnitude to that engendered by the most effective

protective strategies currently known. Collectively, these

data provide the first insight into functional regulation of

VDAC isoforms in the context of MPT in mammalian

tissues and indicate that kinase-dependent attenuation of

VDAC pore-formation prevents mitochondrial dysfunction

and cell death.

doi:10.1016/j.yjmcc.2006.03.423

ABSTRACTS / Journal of Molecular and Cellular Cardiology 40 (2006) 862–918 913