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From the Departments of Physiology and Medicine and The
Cardiovascular Research Laboratories,*
UCLA School of
Medicine, Los Angeles, California; Mayo Clinic
Scottsdale,
Scottsdale, Arizona; and the
Department of Physiology and Biophysics,
College of Medicine, University of Illinois at Chicago,
Chicago, Illinois
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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myosin heavy chain promoter to
target expression of this chimera to the cardiac myocyte,
transgenic mice were generated that had varied levels of transgene
expression. Multiple transgenic founders that expressed the transgene
at high levels, died perinatally and exhibited replacement
fibrosis. Lines that survived showed 1) hypertrophic changes concordant
with reduction in endogenous ß1 integrin levels,
or 2) reduced basal contractility and relaxation as well as alterations
in components of integrin signaling pathways. These data support an
important role for ß1 integrin in normal cardiac
function.
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Integrins are heterodimeric cell-surface receptors composed of and
ß subunits, that function as adhesive and signaling molecules, as
well as mechanotransducers.1,2
In non-cardiac cells, it
has been demonstrated that integrins respond to abnormal strain in a
manner similar to that which would be found during pressure or volume
overload in the heart.3
ß1
integrin is a dominant integrin ß subunit expressed in heart. Two of
the four splice variants of ß1 integrin,
ß1A and ß1D are
expressed on cardiac myocytes. They are identical with the exception of
the last 24 amino acid residues of their respective cytoplasmic
domains. The expression of ß1A and
ß1D isoforms is developmentally regulated in
cardiac cells. ß1A is expressed during
embryogenesis while ß1D expression begins late
in development and eventually becomes the dominant
ß1 integrin isoform expressed on adult cardiac
myocytes. In previous work, we demonstrated that both isoforms could
participate in the hypertrophic response of cultured cardiac
ventricular myocytes.4,5
Increased expression of
ß1 integrin augmented both morphological and
biochemical characteristics of the hypertrophic response. In contrast,
expression of the chimeric protein Tacß1A, a
protein that disrupts integrin adhesion and signaling,6
suppressed the expression of atrial natriuretic factor (ANF), a marker
gene of hypertrophic induction. These data implicated integrin mediated
adhesion and signaling in the in vitro cardiac hypertrophic
response pathway.
Based on these results, we hypothesized that alteration of integrin function in the cardiac cell, through cardiac myocyte-specific expression of the Tacß1A chimeric molecule in the transgenic mouse could provide novel insights into the role of integrins in the heart. We produced lines of transgenic animals that expressed varied amounts of Tacß1A in the cardiac myocyte. Multiple transgenic founders which expressed the transgene at high levels, died perinatally with significant replacement fibrosis. Lines that survived showed 1) hypertrophic changes concordant with reduction in endogenous ß1 integrin levels, or 2) reduced basal contractility and relaxation, as well as alterations in components of integrin signaling pathways. These data suggest that ß1 integrin expression and signaling are required for normal murine cardiac form and function.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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All animals were housed in compliance with the NIH Guide to Care of Laboratory Animals in an AALAC approved facility.
Transgene Construction
A construct encoding a chimeric protein consisting of the
extracellular and transmembrane domain of the Tac subunit of the human
interleukin-2 (IL-2) receptor fused to the cytoplasmic domain of
ß1A integrin (Tac-ß1A)
in plasmid pcDNA was a kind gift of S. LaFlamme and K. Yamada (NIDR,
NIH, Bethesda, MD) and has been described previously.4
The
Tac-ß1A fragment was subcloned into the
SalI site of an myosin heavy chain (MHC) promoter
construct (clone 26, obtained from J. Robbins, University of
Cincinnati, Cincinnati, OH). To generate the
Tac-5 construct, the
ß1A cytoplasmic domain portion of the
Tacß1A construct was removed by digestion with
HindIII and XhoI and replaced with the
5 cytoplasmic domain. The cytoplasmic domain
of 5 was generated by reverse transcriptase
polymerase chain reaction (PCR) from RNA isolated from the human 293
cell line (ATCC). Oligonucleotides used for the amplification contained
a HindIII site on the 5' end of the forward primer and an
XhoI site on the 5' end of the reverse primer.
Southern and Northern Blots and Polymerase Chain Reaction
Southern blotting or PCR was used to identify potential founders that had integrated the transgene.7 For these procedures, genomic DNA was isolated from weanling mice by tail or toe clips. For Northern blot studies, total RNA was extracted from freshly isolated cardiac tissue with the TRIzol Reagent (Life Technologies, Rockville, MD). For Southern and Northern blotting, a 760 bp SstI-HindIII fragment composed of the human interleukin-2 receptor extracellular and transmembrane domain was excised from pBluescript-Tacß1A, and used as a probe. The ANF probe was as described previously.8 ß-myosin heavy chain (ßMHC) transcript was detected using an oligonucleotide probe(5'-CAAAGGCTCCAGGTCTGAGGGCTTCAC-3'). For Southern blotting, hybridizations were performed in 6x standard saline citrate (SSC), 5x Denhardt’s solution, 0.5% sodium dodecyl sulfate (SDS), 100 µg/ml of herring sperm DNA at 60°C overnight, followed by washing in 0.1x SSC, 0.1% SDS at 62.5°C. Similar conditions were used for Northern blotting, but the posthybridization washing temperature was increased to 65°C for the ANF and ßMHC probes. For PCR, two primers specific for the coding region of Tac subunit portion of the transgene were synthesized: forward primer: 5'-CATACCTGCTGATGTGGGGAC-3', and reverse primer: 5'-CCCTGCAGTGACCTGGAAGGC-3'. Tac-ß1A transgenic animals were identified by the presence of a 371-bp product.
Adenoviral Production and Culture of Neonatal Ventricular Myocytes
Adenoviral construction and culture of neonatal rat ventricular myocytes was performed as described previously.4,5
Histology and Immunofluorescent Microscopy
Sections (5 µm) of paraffin-embedded hearts were stained with hematoxylin and eosin or with Masson’s Trichrome. ß1D integrin protein was detected in 5-µm cryosectioned specimens of murine heart by immunostaining with a polyclonal isoform-specific antibody (no. 186) that has been previously characterized.5 Microscopic analysis was performed using a Nikon Diaphot microscope equipped with epifluorescent optics.
Lysates and Western Blot Analysis
Lysate preparation and Western blotting was performed as previously described.9 Hearts were dissected, rinsed in PBS, and immediately homogenized on ice in 1 ml ice-cold RIPA buffer (158 mmol/L NaCl, 10 mmol/L Tris-HCl pH 7.2, 1 mmol/L EGTA, 1 mmol/L orthovanadate, 1% Triton-X100, 1% Na deoxycholate, 0.1% SDS, 100 µmol/L leupeptin, 5 IU/ml aprotinin, 10 µg/ml soybean trypsin inhibitor, 10 mmol/L benzamidine, 1 mmol/L phenylmethylsulfonyl fluoride) and incubated for 10 minutes on ice. Lysates were clarified by centrifugation (16,000 x g, 15 minutes, at 4°C). Protein content of the lysate was determined using the BCA protein assay (Biorad Laboratories, Hercules, CA). For immunoblotting, equal amounts of protein (10–20 µg) were electrophoresed on an 8 to 16% gradient SDS-polyacrylamide gel electrophoresis gel (Novex, Carlsbad, CA) and transferred onto nitrocellulose. Immunoblotting was performed using the following antibodies: monoclonal 7G7/B6 (ATCC) to detect the Tac extracellular domain, polyclonal MC555, specific for the ß1A cytoplasmic domain, polyclonal no. 186 specific for the ß1D cytoplasmic domain,5 monoclonal anti-phosphotyrosine 4G10, monoclonal anti-Src GD11, polyclonal anti-FAK (all Upstate Biotech, Lake Placid, NY), polyclonal p44/42 MAP kinase, and monoclonal phospho-p44/42 MAP kinase E10 (New England Biolabs, Beverly, MA). Signals were quantitated by densitometry using Alphaease software (Alpha Innotech, San Leandro, CA).
Hemodynamics and Transverse Aortic Constriction
Eight- to 20-week-old male and female transgenic and negative littermate control mice were used for all studies, with ages matched for a particular analysis. Animals were anesthetized via intraperitoneal injection of 100 mg/kg ketamine and 5 mg/kg xylazine and placed on a warming pad. Heart rate and temperature were continuously monitored. Surgery was performed and hemodynamic measurements were obtained as previously described.10
Briefly, after anesthetization and intubation, the right carotid artery was exposed. A 1.4 French Millar catheter (Millar Instruments, Houston, TX) was inserted and advanced until a left ventricular pressure tracing was visualized. The catheter was adjusted so that no catheter trapping was evident. The animals were recovered from the initial procedure and baseline pressure measurements were obtained. Sequential injections of isoproterenol (0.01, 0.02, and 0.05 µg) were administered with at least a 5-minute recovery period between injections. Measurements were acquired with Hem Software (Notocord Systems, Croissy, France). Heart rate and left ventricular pressure were recorded 150 seconds after the injections and averaged over a 10-second period. Maximum and minimum dP/dT were calculated during this 10-second period. The animals were given a lethal dose of KCl to terminate the experiment.
Pressure overload (POL) hypertrophy was induced via transverse aortic constriction, using previously published techniques.8 Animals were anesthetized and monitored as above. A midline cervical incision was made and the trachea was exposed. Animals were intubated with a 20 gauge blunt-tipped needle and then connected to a mechanical ventilator. The chest was entered at the left second intercostal space. The thymus was deflected to expose the aorta. A constriction was made in the transverse aorta by tying a 7-0 silk suture over a 27-gauge needle. The pneumothorax was evacuated and the chest was closed. The animals were extubated approximately 20 minutes after the surgery and allowed to recover before being returned to their cages. Sham animals underwent an identical procedure without aortic constriction. On postoperative day 7, the animals were again anesthetized and intubated. The carotid arteries were exposed and cannulated with flame-stretched PE50 tubing. Carotid pressures were recorded to assess for adequate gradients between the left and right carotid pressures. The animals were then sacrificed, and the heart chambers were weighed at the conclusion of the procedure.
Determination of Calcium Sensitivity Using Skinned Fiber Bundles
Calcium sensitivity was determined as previously described.11 Briefly, hearts were isolated from transgenic or age-matched wild-type mice (10–13 days or 21–24 days old) and rinsed in cold high relaxing (HR) solution (10 mmol/L EGTA, 2 mmol/L MgCl2, 79.2 mmol/L KCl, 5.4 mmol/L Na2ATP, 12 mmol/L Creatine phosphate, 20 mmol/L MOPS, pH 7.0 [ionic strength 150 mmol/L]) plus protease inhibitors (2.5 µg/ml pepstatin A, 1 µg/ml leupeptin, and 50 µmol/L phenylmethylsulfonyl fluoride). Left ventricular papillary muscles were removed and dissected further into fiber bundles approximately 150 µm in diameter and 4 to 5 mm in length. Bundles were incubated in 1% Triton-X100 for 30 minutes to skin the fiber bundle and subsequently sarcomere length was set to 2.0 µm as determined by the laser diffraction pattern. All fibers used had a final maximal contraction that was at least 90% of the initial maximal contraction. The force-pCa (log of molar [Ca+2]) relation was fit to the Hill equation with nonlinear regression analysis (Prizm, GraphPad, San Diego, CA) to derive pCa50 and the Hill coefficient. One fiber bundle per animal was analyzed.
Statistics
Data are presented as mean ± SEM. Statistical differences were determined by t-test with P < 0.05 indicating significant differences.
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Results |
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Previous studies have shown that expression of free cytoplasmic
domains of ß1 integrin in various cultured cell
lines disrupts normal integrin function.6,12-15
To
further investigate the role of ß1 integrin in
cardiac myocytes, we assessed the effect of expression of
Tac-ß1A in primary cultured cardiac myocytes.
High-level expression of Tac-ß1A or
Tac-5 in cardiac myocytes was obtained by
infection of cultured myocytes with recombinant adenoviral expression
vectors (Figure 1A)
. Expression of the
control Tac-5 chimeric protein does not
inhibit integrin function in noncardiac cells.6
In
cultured neonatal ventricular myocytes, it similarly did not produce
any significant morphological alterations or affect myocyte adhesion to
the substrate as compared to uninfected cells (Figure 1, B and C)
. This
indicates that expression of the Tac extracellular transmembrane domain
does not affect cardiac myocytes. In contrast, high-level expression of
Tac-ß1A produced significant morphological
alterations in the infected cells by 16 hours post infection. At later
time points, Tac-ß1A expression resulted in
myocyte detachment (Figure 1, D and E)
. This is consistent with our
previous finding that expression of the Tac-ß1A
chimera in primary cultured cardiac myocytes disrupted adrenergically
induced hypertrophic signaling events.4
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Based on these and our previous results,4
we
hypothesized that transgenic expression of free
ß1A integrin cytoplasmic domains in the cardiac
myocyte would disrupt integrin function and signaling in the targeted
cells and provide important information about the role of integrins in
the intact heart. The MHC promoter construct used for generation of
these mice has been previously well characterized and is known to
become up-regulated in the murine ventricle perinatally and continue
expression in the adult.16,17
The MHC
-Tacß1A construct was used to generate 15
independent transgenic lines with varied level of transgene expression
as assessed via Western blot analysis. (Figure 2)
. Six independent founder animals,
which had the highest relative level of transgene expression, all died
perinatally and had diffuse fibrotic replacement of the myocardium
(Figure 3, C and D)
. Additional lines of
animals with high-level transgene expression (nos. 36 and 56 shown in
Figure 2
) survived and bred appropriately, but showed evidence of a
dilated and hypertrophic phenotype (Figure 3, E and F)
and expressed
molecular markers of hypertrophic induction (Figure 4)
, whereas other lines (eg, nos. 44 and
74 in Figure 2
) showed no basal histological or molecular abnormalities
(data not shown).
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Integrins are known to be important mechanotransducers in
non-cardiac cells.18
To further explore the role of
integrins in the cardiac myocyte, we analyzed transgenic animals from
lines that had high transgene expression (lines 44 and 74 of Figure 2
),
but showed no basal morphological or molecular abnormalities. First we
evaluated the hemodynamic properties of these mice, as compared to
littermate control animals using invasive techniques. Basal left
ventricular contractility and relaxation were significantly depressed
in the transgenic animals as compared to controls, yet following
infusion of the inotropic agent isoproterenol, this abnormality could
be overcome (Figure 5)
. No significant
difference was seen in left ventricular pressure or heart rate between
the transgenic or control groups. To assess whether this functional
abnormality of the myocardium was a direct consequence of intrinsic
myocyte dysfunction, we evaluated the Ca2+
sensitivity of isometric tension in skinned fiber bundles from
transgenic and control animals. No difference in the pCa-tension
relationship or Hill coefficient was detected in the transgenic fiber
bundles (data not shown).
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MHC-Tacß1A animals developed similar
degrees of left ventricular hypertrophy after 7 days of POL (22.4
± 2.9% increase in left ventricular weight/body weight over sham
control) as compared to wild-type littermate control animals (21.4
± 2.1% increase in left ventricular weight/body weight over
sham-operated control). Altered Phosphorylation of Focal Adhesion Kinase (FAK) and Extracellular Regulated Kinase (ERK), as Well as Reduction of Endogenous ß1 Protein, Were Found in Tac-ß1A Transgenic Animals
Integrins are devoid of intrinsic kinase activity and rely on the assembly of cytoskeletal and signaling proteins following integrin ligation for efficient signal transduction. To assess the effect of transgene expression on signaling events in the cardiac myocyte, we evaluated the phosphorylation profile of proteins that are known components of integrin signaling cascades. Basal (sham-operated) protein values were assessed and compared to the protein values after hemodynamic loading caused by aortic constriction. Densitometry of Western blot analyses was performed for each signaling molecule, where the activated form of each protein was normalized to the total amount of that particular protein.
We first examined the effect of transgene expression on the
phosphorylation of FAK, a cytoplasmic tyrosine kinase that is thought
to be a key intermediary of signaling through integrins.19
The level of phosphorylated FAK in sham-operated transgenic animals was
significantly reduced compared to sham-operated controls. Further, in
the wild-type control animals, FAK phosphorylation was reduced after 7
days of hemodynamic loading, whereas no change was detected in the POL
transgenic mice (Figure 6, A and B)
.
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, when
wild-type animals underwent aortic constriction, they significantly
increased phosphorylated p42 ERK relative to total p42 ERK protein
and trended toward similar increases in the p44 isoform
phosphorylation. In contrast, no significant change was noted in the
phosphorylation of either ERK isoform in the hemodynamically loaded
transgenic animals. Similarly, we evaluated phosphorylation of c-Src in
sham-operated or aortic constricted wild-type and transgenic animals
(Figure 6, E and F)
. No significant differences were noted in c-Src
normalized phosphorylation in the wild-type animals after aortic
constriction, and no differences were noted between the wild-type and
transgenic animals.
Finally, we evaluated the expression level of endogenous
ß1D integrin protein in the context of the
transgenic animals. As shown in Figure 7
,
the level of expression of the endogenous ß1
integrin was reduced in those animals with high-level expression of the
Tacß1A transgene. The reduction of the endogenous
ß1 integrin expression was apparent by western
blotting (Figure 7A)
and by immunostaining of cardiac tissue (Figure 7, B and C)
.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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In our previous work, we found that increased expression of
ß1 integrin augmented the hypertrophic response
of cultured neonatal ventricular myocytes, whereas expression of the
chimeric protein used in the current study,
Tac-ß1A, inhibited this response.4
The Tac-ß1A chimeric protein does not
heterodimerize with integrin subunits nor does it bind to
extracellular matrix ligands. Previous studies with various cultured
cell lines have demonstrated that expression of this or similar
chimeric proteins acts to dominantly inhibit endogenous
ß1 integrin function, perhaps by titrating
limiting amounts of cytoplasmic factors which bind to the
ß1 integrin cytoplasmic tail and are required
for normal ß1 integrin
function6,12,13,21-23
. Here we further demonstrated that
high-level expression of Tac-ß1A in cardiac
myocytes in vitro altered cell morphology and down-regulated
adhesion to extracellular matrix. In contrast, expression of the
control construct Tac-5 had no effect on
myocyte morphology or adhesion, indicating that these alterations are
due to the presence of free ß1A cytoplasmic
domains rather than expression of the Tac component of the chimeric
molecule. When the Tac-ß1A chimeric molecule is
expressed, it must compete with the endogenous
ß1 integrins that are present in the cells.
Therefore, increased cellular expression of the
Tac-ß1A chimera effectively titrates
progressive alteration of endogenous integrin function. Relative to
this, the phenotypes observed in Tac-ß1A
transgenic mice correlated with the level of expression.
The highest relative expression of Tac-ß1A resulted in reduction of endogenous ß1 integrin expression with fibrotic replacement of the myocardium and perinatal mortality. Normal cell-matrix adhesion is required for appropriate cardiac development, as has been illustrated by the embryonic lethal phenotypes of several integrin and extracellular matrix knockout animals.24-27 As overexpression of Tac-ß1A has been demonstrated to disrupt integrin receptor affinity for extracellular matrix, it is likely that the perinatal lethality in animals with the highest level of transgene expression was caused by abnormalities in myocyte adhesion to extracellular matrix. This work is consistent with the phenotype of transgenic mice that overexpress constitutively active ras-related c3 botulinum toxin substrate-1 (rac1) in the cardiac myocyte. These rac1 transgenic mice have been proposed to have alterations in cellular adhesion and in mechanotransductive properties of the heart.28
A second distinct phenotype was present in transgenic animals with substantial, but relatively less, transgene expression than those that died perinatally. Surviving lines with the highest level of transgene expression developed compensatory hypertrophy in the absence of any provoked hemodynamic stimulus. Other surviving lines with lesser relative chimera expression showed (i) no basal morphological abnormalities, (ii) depressed baseline positive and negative dP/dT, (iii) normal pCa-tension and Hill coefficient of fiber bundles isolated from hearts of transgenic animals, and (iv) disturbance in phosphorylation of several molecules implicated in integrin signaling.
Hypertrophy or remodeling of the myocardium to optimize the cardiac performance can result from mechanical stress on the heart from either pressure or volume loading, as well as cardiac myocyte death. Ventricular hypertrophy is an important adaptive mechanism that allows the heart to maintain its output. Recent data have shown modulation of integrin expression and extracellular matrix in the hypertrophied myocardium.29-32 This is particularly noteworthy given the demonstration that integrins can function as mechanotransducers, translating mechanical signals to biochemical signals.1,33 It is possible, therefore, that integrins may play a role in the translation of increased mechanical stress into the cardiac cellular response of hypertrophy.
We examined whether transgene expression would alter normal cardiac contractile function and prevent adequate compensatory hypertrophy from occurring in the transgenic animals. This concept was tested in the mice that had lower relative transgene expression. Abnormal contractility and relaxation were detected at rest, but no alterations in intrinsic function of the myofilaments were found. These functional abnormalities could be overcome by adrenergic stimulation of the transgenic myocardium. Additionally, after hemodynamic loading via aortic constriction, we found equal induction of cardiac hypertrophy in wild-type and transgenic hearts as measured by heart weight normalized to body weight or tibial length. This is in contrast to the transgenic animals with higher level transgene expression that developed a hypertrophic response without any provocation, indicative of a compensatory response for intrinsically impaired cardiac function. Therefore, it is likely that the animals tested for their hypertrophic response to aortic banding retained sufficient residual integrin function to effect relatively normal mechanotransduction.
In recent years, the role of the cardiac myocyte cytoskeleton in cardiac function has gained significant attention.34,35 ß1 integrin is a critical component of the linkage between the extracellular matrix and cytoskeleton. In striated muscle, it is also localized at the junctional structures and may serve a unique role in orchestrating appropriate organization of the support structure of the contracting myocyte.36 In this role, the cytoplasmic domain of ß1 has been shown to interact with several important structural components of the cytoskeleton such as vinculin and talin. Disturbance of vinculin function has been found to alter normal myofibrillar organization and, when fully ablated in the mouse, results in fetal death at embryonic day 9.5 to 10 with noncontracting hearts.37,38 With the onset of heart failure, vinculin has been found to alter its localization away from intercalated disks into the cell body of myocytes.39 Thus, it is possible that with disturbance of integrin function in our animals, this critical linkage was disrupted, leading to the unique phenotypes we saw both perinatally and in the adult animals. These results are in agreement with the data from chimeric animals composed of ß1 integrin null cells, where severe alterations in myofibril formation was noted in the few ß1 null cells found in the chimeric heart.24 It is interesting to note that preliminary work by our own group using a cardiac-specific knockout approach to ß1 integrin ablation has shown similar results with replacement fibrosis and progression to heart failure in adult mice.
Determining the signaling pathways involved in cardiac hypertrophy is essential to the understanding of normal cardiac development and response of the heart to increased hemodynamic load. Ligand binding of integrins leads to their clustering. To propagate integrin signaling, focal adhesion complexes enriched in adapter and signaling molecules are subsequently assembled. Integrin signaling activates pathways and effector molecules known to be components of signaling from other cardiac receptors implicated in the hypertrophic response.9,32,40 In addition, increased matrix deposition and altered integrin expression is a characteristic of in vivo models of hypertrophy.29,30,41-43 Thus, we assayed for alterations in various molecules that are components of the integrin signaling cascades. Previous experiments that tested the effects of autonomously expressed ß1 integrin cytoplasmic domains showed that at modest expression levels, the chimera could interfere with phosphorylation of FAK.13 At higher expression levels, the chimera could induce constitutive phosphorylation of FAK.6,13 Consistent with these data are the results from our transgenic animals with moderate levels of transgene expression; FAK phosphorylation was depressed basally and was not changed after aortic constriction, as was detected in the wild-type animals. Normal FAK function has been suggested to be a cell survival factor.44 Impaired induction of p42 ERK phosphorylation after hemodynamic loading was also observed in transgenic animals. These results suggest that expression of free ß1A integrin cytoplasmic domains disrupted several of the downstream events that are required for basal integrin signaling as well as events that may be crucial to modification of the myocyte stressed by a hemodynamic load, consistent with other investigators’ work in the pressure-overloaded cat ventricle.45
In conclusion, this study has shown that cardiac myocyte-specific expression of free ß1A integrin cytoplasmic domains that alters normal integrin function leads to disturbed cardiac function, histological abnormalities, modification of molecules instrumental in integrin signaling, and perinatal death in a manner that correlates with expression level of the transgene.
This work is in agreement with the concept proposed by others that cytoskeletal disturbances of the cardiac myocyte may ultimately lead to a cardiomyopathic phenotype.35 Further work is underway to alter ß1 integrin function conditionally in the intact myocardium to obtain further insights into the role of integrin-mediated adhesion and signaling in cardiac function.
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Acknowledgements |
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Footnotes |
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Supported by grants from the National Institutes of Health (HL10262–01S to R. S. K., HL42977 to J. C. L., HL 22231–22 to R. J. S., and HL57872 to R. S. R.), the American Heart Association (to C. J. B.), and the UCLA Laubisch Cardiovascular Research Fund.
R. S. K. and S.-Y. S. contributed equally to this study.
Accepted for publication December 12, 2000.
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F. G. Spinale Matrix Metalloproteinases: Regulation and Dysregulation in the Failing Heart Circ. Res., March 22, 2002; 90(5): 520 - 530. [Abstract] [Full Text] [PDF]
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S.-Y. Shai, A. E. Harpf, C. J. Babbitt, M. C. Jordan, M. C. Fishbein, J. Chen, M. Omura, T. A. Leil, K. D. Becker, M. Jiang, et al. Cardiac Myocyte-Specific Excision of the {beta}1 Integrin Gene Results in Myocardial Fibrosis and Cardiac Failure Circ. Res., March 8, 2002; 90(4): 458 - 464. [Abstract] [Full Text] [PDF]
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A. Braun, B. L. Trigatti, M. J. Post, K. Sato, M. Simons, J. M. Edelberg, R. D. Rosenberg, M. Schrenzel, and M. Krieger Loss of SR-BI Expression Leads to the Early Onset of Occlusive Atherosclerotic Coronary Artery Disease, Spontaneous Myocardial Infarctions, Severe Cardiac Dysfunction, and Premature Death in Apolipoprotein E-Deficient Mice Circ. Res., February 22, 2002; 90(3): 270 - 276. [Abstract] [Full Text] [PDF]
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 P. J. Schmeissner, H. Xie, L. B. Smilenov, F. Shu, and E. E. Marcantonio Integrin Functions Play a Key Role in the Differentiation of Thymocytes In Vivo J. Immunol., October 1, 2001; 167(7): 3715 - 3724. [Abstract] [Full Text] [PDF]
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D. Bouvard, C. Brakebusch, E. Gustafsson, A. Aszodi, T. Bengtsson, A. Berna, and R. Fassler Functional Consequences of Integrin Gene Mutations in Mice Circ. Res., July 30, 2001; 89(3): 211 - 223. [Abstract] [Full Text] [PDF]
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R. S. Ross and T. K. Borg Integrins and the Myocardium Circ. Res., June 8, 2001; 88(11): 1112 - 1119. [Abstract] [Full Text] [PDF]
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A. Braun, B. L. Trigatti, M. J. Post, K. Sato, M. Simons, J. M. Edelberg, R. D. Rosenberg, M. Schrenzel, and M. Krieger Loss of SR-BI Expression Leads to the Early Onset of Occlusive Atherosclerotic Coronary Artery Disease, Spontaneous Myocardial Infarctions, Severe Cardiac Dysfunction, and Premature Death in Apolipoprotein E-Deficient Mice Circ. Res., February 22, 2002; 90(3): 270 - 276. [Abstract] [Full Text] [PDF]
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S.-Y. Shai, A. E. Harpf, C. J. Babbitt, M. C. Jordan, M. C. Fishbein, J. Chen, M. Omura, T. A. Leil, K. D. Becker, M. Jiang, et al. Cardiac Myocyte-Specific Excision of the {beta}1 Integrin Gene Results in Myocardial Fibrosis and Cardiac Failure Circ. Res., March 8, 2002; 90(4): 458 - 464. [Abstract] [Full Text] [PDF]
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