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ROBERT L. PRICE , Ph.D.
You,
S., Ohmori, M., Pena, M.M.O., Nassri, B., Quiton, J., Al-Assad,
Z.A., Liu, L., Wood, P.A., Berger, S.H., Liu, Z., Wyatt,
M.D., Price, R.L., Berger, F.G., Hrushesky, W.J. M. 2006.
Developmental abnormalities in multiple proliferative
tissued of Apc Min/+ mice. Int. J. Path. 87:227-236
Germ-line mutation of the Apc gene has
been linked to familial adenomatous polyposis (FAP) that
predisposes to colon cancer. Apc(Min/+) mice, heterozygous
for the Apc gene mutation, progressively develop small
intestinal tumours in a manner that is analogous to that
observed in the colon of patients with FAP (Su et al. 1992;
Fodde et al. 1994; Moser et al. 1995). We have studied the
effects of Apc gene mutation on murine intestinal and
extra-intestinal, proliferatively active tissues. We have
contrasted the histology to that of the age- and sex-matched
wild-type C57BL/6 mice. Histological assessment of the
normal appearing intestinal mucosa demonstrates minimal
change in size of crypts. In contrast, villi are longer in
the ileum of Apc(Min/+) mice relative to C57BL/6 mice at 12
and 15 weeks of age. Vigorous splenic haematopoiesis in
Apc(Min/+) mice was seen at 12 and 15 weeks of age, as
reflected by marked splenomegaly, increased splenic
haematopoietic cells and megakaryocytes. Peripheral blood
counts, however, did not differ between C57BL/6 and Apc(Min/+)
mice at 15 weeks of age. Lymphoid depletion in Apc(Min/+)
mice was characterized by diminished numbers of splenic
lymphoid follicles and small intestinal Peyer's patches. The
ovaries of 12- and 15-week-old Apc(Min/+) mice exhibited
increased numbers of atretic follicles, and estrous cycling
by serial vaginal smears showed tendency of elongation in
the mutant mice during these age ranges. The testicles of
10-week-old Apc(Min/+) mice showed increased numbers of
underdeveloped seminiferous tubules. Collectively, these
data suggest that, in addition to its obvious effects upon
intestinal adenoma formation, Apc gene mutation causes
impairment of developmental and apparent differentiation
blockade in proliferative tissues, including those of the
haematopoietic system, lymphoid and reproductive tract.
Slamani, M., Krol, A., Beaumont, J.,
Price, R.L., Coman, I.L. and Lipson, E.D. 2006. Application
of Phase Correlation to the Montage Synthesis and 3D
Reconstruction of Large Tissue Volumes from Confocal Laser
Scanning Microscopy. Microscopy and Microanl. 12:106-112.
Bullard, T.A., Borg, T.K. and Price,
R.L. 2005. The expression and role of protein kinase C in
neonatal cardiac myocyte attachment, cell volume and
myofibril formation is dependent upon the composition of the
extracellular matrix. Microscopy and Microanal.
11(3):224-234.
Center for Cellular and Molecular
Cardiology, University of Rochester Medical Center,
Rochester, NY 14642, USA.
The extracellular matrix (ECM) is a dynamic component of
tissues that influences cellular phenotype and behavior. We
sought to determine the role of specific ECM substrates in
the regulation of protein kinase C (PKC) isozyme expression
and function in cardiac myocyte attachment, cell volume, and
myofibril formation. PKC isozyme expression was ECM
substrate specific. Increasing concentrations of the PKC
delta inhibitor rottlerin attenuated myocyte attachment to
randomly organized collagen (1, 5, and 10 microM), laminin
(5 and 10 microM), aligned collagen (5 and 10 microM), and
fibronectin (10 microM). Rottlerin significantly decreased
cell volume on laminin and randomly organized collagen, and
inhibited myofibril formation on laminin. The PKC alpha
inhibitor Go 6976 inhibited attachment to randomly organized
collagen at 6 nM but did not affect cell volume. The general
PKC inhibitor Bisindolylmalemide I (10 and 30 microM) did
not affect myocyte attachment; however, it significantly
decreased cell volume on randomly organized collagen. Our
data indicate that PKC isozymes are expressed and utilized
by neonatal cardiac myocytes during attachment, cell growth,
and myofibril formation. Specifically, it appears that PKC
delta and/or its downstream effectors play an important role
in the interaction between cardiac myocytes and laminin,
providing further evidence that the ECM influences cardiac
myocyte behavior.
Nakayama, M., Yan, X., Price, R.L.,
Borg, T.K., Ito, K., Sanbe, A., Robbins, J. and Lorell, B.H.
2005. Chronic ventricular myocyte specific overexpression of
angiotensin II type 2 receptor results in intrinsic myocyte
contractile dysfunction. A. J. Physiol Heart Circ Phsiol
288(1): H317-327.
Department of Medicine, Cardiovascular
Division, Beth Israel Deaconess Medical Center and Harvard
Medical School, Boston, MA 02215, USA.
ANG II type 2 receptor (AT(2)) is upregulated in failing
hearts, but its effect on myocyte contractile function is
not known. We measured fractional cell shortening and
intracellular Ca(2+) concentration transients in left
ventricular myocytes derived from transgenic mice in which
ventricle-specific expression of AT(2) was driven by the
myosin light chain 2v promoter. Confocal microscopy studies
confirmed upregulation of AT(2) in the ventricular myocytes
and partial colocalization of AT(2) with AT(1). Three
components of contractile performance were studied. First,
baseline measurements (0.5 Hz, 1.5 mmol/l extracellular
Ca(2+) concentration, 25 degrees C) and study of contractile
reserve at faster pacing rates (1-5 Hz) revealed
Ca(2+)-dependent contractile dysfunction in myocytes from
AT(2) transgenic mice. Comparison of two transgenic lines
suggested a dose-dependent relationship between magnitude of
contractile dysfunction and level of AT(2) expression.
Second, activity of the Na(+)/H(+) exchanger, a dominant
transporter that regulates beat-to-beat intracellular pH,
was impaired in the transgenic myocytes. Third, the
inotropic response to beta-adrenergic versus ANG II
stimulation differed. Both lines showed impaired contractile
response to beta-adrenergic stimulation. ANG II elicited an
increase in contractility and intracellular Ca(2+) in
wild-type myocytes but caused a negative inotropic effect in
myocytes from AT(2) transgenic mice. In contrast with
beta-adrenergic response, the depressed response to ANG II
was related to level of AT(2) overexpression. The depressed
response to ANG II was also present in myocytes from young
transgenic mice before development of heart failure. Thus
chronic overexpression of AT(2) has the potential to cause
Ca(2+)- and pH-dependent contractile dysfunction in
ventricular myocytes, as well as loss of the inotropic
response to ANG II.
Goodwin RL, Nesbitt T, Price RL, Wells
JC, Yost MJ, Potts JD. Three-dimensional model system of
valvulogenesis. Dev Dyn. 2005 May;233(1):122-9
Valvular defects are among the most
common and deleterious of all cardiac malformations. The
early cardiac cushions are located in the atrioventricular
(AV) canal of the embryonic heart. These cushions contain
cells that are the primordia of the cardiac valves and
membranous septa. Significant progress has been made in
delineating the molecular mechanisms that regulate the early
steps of cushion formation; however, little is known about
how these cushions differentiate into valve leaflets. Here,
a new three-dimensional collagen tube culturing system was
tested for its ability to sustain the development and
maturation of the AV cushion anlagen. We report that AV
cushion tissues grown within the collagen tube scaffold
recapitulate aspects of AV valve development both at the
molecular and morphological levels. Furthermore, our results
indicate that valve leaflet formation in the tube model is
dependent on the presence of cardiac myocytes. Copyright
2005 Wiley-Liss, Inc.
Goldsmith EC, Hoffman A, Morales MO,
Potts JD, Price RL, McFadden A, Rice M, Borg TK.
Organization of fibroblasts in the heart. Dev Dyn. 2004
Aug;230(4):787-94
Cardiac fibroblasts are organized into a
three-dimensional network in the heart. This organization
follows the endomysial weave network that surrounds groups
of myocytes. Reverse transcriptase-polymerase chain
reaction, Western blots, and immunohistochemistry were used
to show that discoidin domain receptor 2 (DDR2) was specific
for cardiac fibroblasts and not expressed on endothelial
cells, smooth muscle cells, or cardiac myocytes. DDR2 is
expressed early in development and in the adult heart. High
voltage electron microscopy (HVEM), scanning electron
microscopy, and laser scanning confocal microscopy document
the three-dimensional organization of fibroblasts in the
heart. Antibodies against connexin 43 and 45 showed
different patterns but confirmed, along with HVEM, that
fibroblasts are connected to each other as well as cardiac
myocytes. The implications of this arrangement of
fibroblasts can be important to cardiac function. The
signaling of DDR2 and the expression of matrix
metalloproteinase 2 in relation to collagen turnover and
remodeling is discussed. Copyright 2004 Wiley-Liss, Inc.
Waller LN, Fox N, Fox KF, Fox A, Price
RL. Ruthenium red staining for ultrastructural visualization
of a glycoprotein layer surrounding the spore of Bacillus
anthracis and Bacillus subtilis. J Microbiol Methods. 2004
Jul;58(1):23-30
Ruthenium red is a polycationic stain
used to visualize acid polysaccharides on the outer surface
of cells. Ruthenium red staining followed by electron
microscopic analysis was used to demonstrate the presence of
an external glycoprotein layer surrounding the spore of both
Bacillus anthracis and Bacillus subtilis. This layer is less
apparent with traditional staining methods used for electron
microscopy. Renografin gradients were used to purify B.
subtilis spores. These purified spores displayed greatly
enhanced staining with ruthenium red, indicating nonspecific
binding of renografin, which has a major carbohydrate
constituent, methylglucamine. For B. anthracis, staining
with ruthenium red was sufficiently intense that it was not
significantly enhanced by renografin purification. In
addition to demonstrating a previously undiscovered layer
surrounding the spores of B. subtilis, the results help
explain a long-standing controversy as to ultrastructural
differences among these genetically closely related
organisms. Ruthenium red staining provides an important
addition to the identification of surface glycoproteins in
studies to define similarities and differences in the
exosporium layers of Bacillus species. Copyright 2004
Elsevier B.V.
Yost MJ, Baicu CF, Stonerock CE,
Goodwin RL, Price RL, Davis JM, Evans H, Watson PD, Gore CM,
Sweet J, Creech L, Zile MR, Terracio L. A novel tubular
scaffold for cardiovascular tissue engineering. Tissue Eng.
2004 Jan-Feb;10(1-2):273-84
We have developed a counter rotating cone
extrusion device to produce the next generation of
three-dimensional collagen scaffold for tissue engineering.
The device can produce a continuously varying fibril angle
from the lumen to the outside of a 5-mm-diameter collagen
tube, similar to the pattern of heart muscle cells in the
intact heart. Our scaffold is a novel, oriented, type I
collagen, tubular scaffold. We selected collagen because we
believe there are important signals from the collagen both
geometrically and biochemically that elicit the in vivo
-like phenotypic response from the cardiomyocytes. We have
shown that cardiomyocytes can be cultured in these tubes and
resemble an in vivo phenotype. This new model system will
provide important information leading to the design and
construction of a functional, biologically based assist
device.
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