© 1996 American Heart Association, Inc.
Articles |
Endothelial Cell Tyrosine Kinase Receptor and G Protein–Coupled Receptor Activation Involves Distinct Protein Kinase C Isoforms
From the Franz Volhard Clinic, Virchow Klinikum at the Max Delbrück Center for Molecular Medicine, Humboldt University of Berlin, Berlin, Germany.
Correspondence to Hermann Haller, MD, Franz-Volhard-Klinik, Wiltberg Strasse 50, 13122 Berlin, Germany.
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Abstract |
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Abstract Protein kinase C (PKC) is a family of serine/threonine protein kinase isoforms that is important to intracellular enzymes for both tyrosine kinase receptors and G protein–coupled receptors. However, which isoforms are linked to which class of receptors in endothelial cell signaling is not known. Moreover, the PKC isoforms in endothelial cells have not been thoroughly characterized. We tested the hypothesis that specific PKC isoforms are involved in different signaling pathways. PKC isoform expression was assessed by using reverse transcription polymerase chain reaction and Western blotting. The spatial distribution of PKC after stimulation of the cells with basic fibroblast growth factor (bFGF) and thrombin was examined by using confocal microscopy. Expression of PKC
, 
, 
, 
, and 
was
detectable on both the mRNA and protein levels. In resting cells, PKC
and were mostly distributed in the cytosol, while PKC and
were also present in the nucleus. Nuclear immunoreactivity of
PKC and increased significantly between passages 1 and 3. The
phorbol ester TPA induced a rearrangement of PKC and a
translocation of PKC and to the nucleus. Treatment of
endothelial cells with TPA for 24 hours caused PKC ,
, and to disappear, while PKC was not influenced by TPA.
bFGF induced a rapid assembly of PKC along cytosolic structures,
followed by a translocation of the isoform toward the perinuclear
region and into the nucleus. bFGF had a similar effect on PKC . In
contrast, thrombin had a smaller effect on nuclear translocation of PKC
, did not influence PKC , and induced a rapid nuclear
translocation of PKC . Thus, tyrosine kinase receptor activation via
bFGF induced a rapid association of PKC and with nuclear
structures, while activation of the G protein–coupled thrombin
receptor increased mostly nuclear PKC . The translocation of PKC
isoforms into the nucleus by growth-promoting factors may be
important for the induction of endothelial cell growth.
Key Words: tyrosine kinase receptors • protein kinase C isoforms • endothelial cells • thrombin • basic fibroblast growth factor
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Introduction |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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The calcium- and phospholipid-dependent, serine/threonine protein kinase PKC is an important second messenger in ECs. Endothelial PKC has been implicated in the expression and regulation of adhesion molecules,1 in the expression of endothelin-1,2 and in the proliferative response of ECs to hormones and growth factors.3 4 Furthermore, endothelial PKC appears to mediate the intracellular effects of shear stress5 and may also be important to angiogenesis.6 Investigating PKC is difficult because PKC is not a single entity but instead consists of several distinct isoforms with different regulatory and biochemical properties.7 8 These isoforms are expressed on separate genes and may play different roles in cell signaling and cell function.8 An analysis of isoform expression and distribution is necessary to investigate the role of PKC in signaling. The early notion that resting PKC is located in the cytosol and is translocated to the cell membrane on stimulation has been expanded by the finding that PKC is associated with cytoskeletal proteins, nuclear proteins, and intracellular membranes.9 10 We studied the expression and distribution of PKC isoforms in cultured human umbilical ECs by using RT-PCR, Western blotting, and confocal microscopy. We specifically tested the hypothesis that thrombin and bFGF, which exert their effects via separate intracellular pathways, have different effects on PKC isoform distribution.
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Methods |
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Materials and Culture Procedures
All materials, if not stated otherwise, were purchased from Sigma and Merck. M-MLV reverse transcriptase and Taq polymerase were obtained from Perkin Elmer. Human umbilical vein ECs were isolated from umbilical cords by chymotrypsin treatment and prepared.11 Subcultures 1 through 3 were used for the experiments.
RT-PCR Analysis
RNA Isolation
Total RNA was extracted from unstimulated, confluent primary
cultures between passages 1 and 2 by using a guanidinium salt method
derived from MacDonald et al.12 Cells were lysed in 5 mL
guanidinium thiocyanate buffer (buffer 1) containing 4 mol/L
guanidinium thiocyanate, 25 mmol/L sodium citrate (pH 6.5), 0.5%
(wt/vol) N-lauroylsarcosine, 0.1 mol/L
ß-mercaptoethanol, and 0.1% (wt/vol) antifoam A. The RNA was
precipitated by the addition of 0.5 vol ethanol, collected by
centrifugation (5 minutes at 10 000g) and
redissolved in buffer 1. The insoluble debris was removed by a brief
centrifugation. After the addition of 0.15 vol/percent
acetic acid and 65 vol/percent ethanol, the RNA was precipitated
overnight at -20°C. The precipitate was collected by
centrifugation for 10 minutes at 10 000g
and dissolved in 4 mL guanidinium hydrochloride buffer (buffer 2) with
7.5 mol/L guanidinium hydrochloride, 25 mmol/L EDTA, and 11 mmol/L
ß-mercaptoethanol. After the addition of 0.3 vol/percent acetic
acid and 50 vol/percent ethanol, the RNA was precipitated for 20
minutes at -20°C. The last step was repeated twice with gradual
reduction of buffer volumes. The final precipitation was prolonged to 1
hour, and the pellet was dissolved in 0.5 mL buffer 3 with 0.3 mol/L
NaCl, 10 mmol/L Tris-HCl (pH 7.5), 1 mmol/L EDTA, and 0.1% (wt/vol)
sodium dodecyl sulfate. The RNA was precipitated overnight at
-20°C by the addition of 2 vol ethanol. The precipitate was
collected by centrifugation for 30 minutes at
10 000g, washed with 70% ethanol, and dissolved in 200
µL TE. The A 260/A 280 ratio was 1.8.
RT-PCR
The RNA was analyzed by using an RT-PCR assay based on
the RNA PCR kit protocol supplied by Perkin Elmer. The RT was done in a
20-µL volume, applying 1 µg total RNA and 10 or 20 pmol specific
downstream primers, at 52°C for 20 minutes. The subsequent PCR step
was performed in a volume of 100 µL under optimized MgCl2
concentrations and temperature conditions with 10 or 20 pmol specific
primer for 35 cycles. The typical cycle profile used on the Trio
(Biometra) was 20 seconds at 96°C, 60 seconds TA (50°C to 62°C),
and 90 seconds at 73°C. The experiments were accompanied by water and
RNA controls in which the RNA or reverse transcriptase, respectively,
was omitted. Functional control of the primers was achieved by
additions of human fetal brain cDNA (Clonetech) to the PCR step.
Primer Development
The PKC isoform-specific primer pairs (TIBMolbiol) were
derived from EMBL Data Library sequence information based on the
sequence entries for PKC (HSPKCA113 ), PKC ß
(HSPKB14 ), PKC 
(HSPKG14 and
HSPKCG15 ), PKC (HSPKCD13Y16 ), PKC
(HSPKCE17 ), PKC (HSPKCZ15 ), and PKC
(HSPKC18 ). The genetic analysis was done by using
PC/GENE (IntelliGenetics). The primers hybridize to the regulatory
domain of the PKC isoforms, producing PCR products, which include
the V3 region of the corresponding isoform. The primers,
which were designed by using Oligo 4.0 (Med Probe), were used for
specific RT-PCRs (Table
). The specificity of the primers
was tested on a human fetal brain cDNA bank (Clonetech) and verified by
restriction analysis and partial sequencing of the
products.
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Immunoblotting and
Immunocytochemistry
Immunocytochemistry and confocal microscopy were performed by
using a Bio-Rad MRC 500 confocal imaging system (Bio-Rad Laboratories)
with an argon laser.19 We used highly specific,
affinity-purified polyclonal antibodies directed against peptide
sequences of PKC that reacted specifically with the ß, , , ,
and subspecies of PKC (antibodies were from GIBCO; the antibody
against PKC was a monoclonal from UBI). Specificity was
demonstrated by using specific oligopeptides that prevent binding of
the antibodies to the isoforms.20 The secondary
fluorescein isothiocyanate–conjugated anti-mouse
IgG antibody was obtained from Pierce Chemicals. At least 10 to 18
cells from each of at least five experiments were examined under each
experimental condition. The results were reproduced by two separate
investigators and multiple experiments were performed. Quantification
of the signal intensity in the nuclear region was done by using AREA
functions in the MRC software. The nuclear region was outlined
manually, and the calculated mean fluorescent intensity was
obtained.
Statistical analysis was performed on a Macintosh II computer (Apple Inc) by using a commercial program (StatView, Cricket Software Inc); results are expressed as mean±SEM. The Wilcoxon test was used, and a probability of less than.05 was considered significant.
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Results |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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We first investigated PKC expression in resting cells at passages 1 and 2. Four independent total RNA preparations were made that led to the semiquantitative representation of PKC isoform expression shown in Fig 1
. A strong signal for PKC was
present (Fig 1A). No signals for PKC ß and were observed.
Signals for PKC , , and were identified (Fig 1B), and a
positive signal was present for PKC (Fig 1C). Thus, we found
expression of PKC , , , , and in all preparations
(n=13). A strong signal for PKC , , and was observed in each
preparation, while PKC and mRNA were more difficult to detect.
This finding could indicate that these two PKC isoforms are more weakly
expressed in ECs. No mRNA for PKC and ß was observed in any of
the preparations investigated.
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We next investigated whether these PKC isoforms were demonstrable at
the protein level. Fig 2 shows the Western blot
analysis of PKC isoforms. PKC and showed a strong
expression, with single bands at 84 and 87 kD, respectively. No
immunoreactivity for PKC ß was observed. PKC displayed two
protein bands, a strong, upper band with a molecular weight of 82 kD
and a lower band at 81 kD. PKC showed a single protein band at 84
kD. No bands were observed with antibodies against PKC (not
shown).
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We then addressed the question of the intracellular isoform
distribution. Fig 3 shows the immunocytochemistry of PKC
isoforms , , , and by fluorescence confocal
microscopy. PKC showed a dense, punctuated pattern and was
distributed in the cytoplasmic, perinuclear, and nuclear areas. PKC
showed a coarse fibrillar pattern and was exclusively located in the
cytosol. PKC showed a strong, patchy pattern and was localized
throughout the cytosol and in the nucleus. PKC was present in
the cytosol, with a faint uniform staining of the nuclear area. Lane 2
shows the effect of a 10-minute treatment with the phorbol ester TPA
(100 nmol/L) on the cellular distribution of the PKC isoforms. TPA
induced an increase in the immunoreactivity of PKC , , and ,
while PKC was mostly unaffected. TPA led to an increase in
fluorescence intensity in the nuclear area for PKC and .
PKC showed an increased density in the cytosol and a fibrillar
pattern. Lane 3 shows the effect of a 24-hour treatment with TPA (100
nmol/L) on PKC isoforms. TPA induced a marked decrease in the
immunoreactivity of PKC , , and , while PKC was not
affected. Lane 4 gives the negative control for the immunocytochemistry
experiments. Shown are staining without the specific antibody (PKC )
or staining in the presence of immunogenic peptide (other PKC
isoforms). For experiments with TPA we performed controls by using
corresponding concentrations of the vehicle dimethyl sulfoxide (data
not shown). In these control experiments we did not observe any
differences in the untreated cells.
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During the course of these experiments we noted that the cellular
distribution of some of the PKC isoforms was affected by the number of
passages. Fig 4 shows the differences in cellular
distribution of PKC and between passages 1 and 3. PKC in
cells from passage 1 was mostly located in the cytosolic area, with
only sparse patches in the cell nucleus. By passage 3 more nuclear
staining was present, and immunoreactivity had decreased in the
cytosol. PKC at passage 1 showed only punctated areas of
immunoreactivity in the perinuclear region and no nuclear staining, but
by passage 3 a dramatic increase in PKC immunoreactivity in the
cytosol was observed, and nuclear immunoreactivity was present. For
the other PKC isoforms, and , no difference in cellular
distribution was observed in the different passages.
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We next investigated the effects of bFGF (100 nmol/L) and thrombin (1 U/mL) on the cellular distribution of the PKC isoforms in ECs from passage 3.
bFGF and thrombin induced an increase in nuclear immunoreactivity
of PKC (Fig 5). However, there was a marked difference between the different agonists. bFGF had
a strong effect, with an increase of PKC in the perinuclear and
nuclear regions that was already present at 5 minutes
and did not decrease after 10 minutes. In contrast, thrombin induced a
slower and smaller response. bFGF and thrombin had similar
effects on the spatial distribution of PKC
(Fig 6). Both agonists induced an increase in the
cytosolic and perinuclear immunoreactivity of PKC . The cytosolic
staining of PKC changed to a more fibrillar pattern. This effect on
PKC was similar to that observed after treatment with TPA. However,
the effect of thrombin and bFGF on PKC was transient, and
immunoreactivity was decreased by 10 minutes. bFGF greatly increased
the nuclear immunoreactivity of PKC , but thrombin led to a slight
decrease in cytosolic and nuclear PKC immunoreactivity at 5 and 10
minutes (Fig 7). The effect of bFGF on nuclear PKC
was persistent and unaltered by 10 minutes. bFGF led to a small effect
on nuclear PKC at 5 minutes, followed by an increase in cytosolic
immunofluorescence at 10 minutes (Fig 8). The fine meshed PKC pattern under resting
conditions was changed to a more coarse, granular pattern. In contrast,
thrombin induced a rapid and marked nuclear translocation of PKC at
5 minutes that decreased markedly at 10 minutes.
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Fig 9 shows the time course of the relative changes in
PKC isoform immunoreactivity after stimulation with bFGF (100 nmol/L)
and thrombin (1 U/mL). Significant differences (P<.05)
between bFGF and thrombin were observed for PKC and .
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Discussion |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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The important findings in this study are that bFGF induced nuclear translocation of the PKC isoforms
and and led to a cytosolic
rearrangement of PKC ; in contrast, thrombin had no effect on PKC
but instead induced a rapid nuclear translocation of PKC . These
findings could in part serve to explain how different classes of
agonists result in contrasting effects in ECs. To our knowledge, PKC
isoforms have not previously been thoroughly characterized in ECs. We
showed that cultured human ECs express PKC isoforms , , , ,
and . We then analyzed the different intracellular locations
of the PKC isoforms. We showed that under resting conditions only PKC
and were present in the nucleus, while PKC and
showed mostly cytosolic staining. The expression level and
intracellular distribution of PKC and were associated with
phenotypic alterations. Both changed between passages 1 and 3, with a
marked increase in PKC immunoreactivity that was associated with
enhanced nuclear staining. PKC showed a similar but somewhat less
prominent pattern. TPA increased PKC and nuclear staining and
altered the cytosolic pattern of PKC .
We detected the expression of PKC , , , , and in our
cells with RT-PCR but could not find expression of PKC ß and .
Differences in tissue expression and distribution of PKC have been
reported.7 21 PKC , , and are present in
almost all tissues, while PKC , ß, and are expressed only in
certain cell types. Moreover, discrepancies exist in the literature
regarding the expression of PKC ß in ECs.22 23 24 25 26 27 A
possible explanation of this discrepancy is the state of
differentiation of the ECs investigated. We used primary EC cultures
only at passages 1 through 3. Expression of von Willebrand
factor was still present at higher passages. However, as shown in
our study, expression levels of some PKC isoforms changed at higher
passages and were already altered by passage 3. This finding is
supported by the findings of Mattila et al,27 who observed
significant expression of PKC ß only in
a human umbilical vein–derived EC line. A second possible
explanation is that PKC ß expression is species dependent and differs
in the cells investigated.23 24 Marked differences may
also exist between ECs from different vascular beds. For instance, PKC
ß may be expressed in ECs from arteries but not in ECs from umbilical
veins.28
We observed an increase of PKC immunoreactivity from passages 1
through 3 even though the ECs of passage 3 displayed unaltered
expression of von Willebrand factor and showed a typical
cobblestone pattern. PKC may play a role in the dedifferentiation
process of ECs in culture; others have observed increased expression of
PKC in dedifferentiating vascular smooth muscle
cells.29 The increased PKC expression may be of
functional relevance, as overexpression of this isoform leads to
increased proliferation in NIH 3T3 cells.30
The spatial distribution of PKC in resting ECs showed distinctive patterns for the specific isoforms. We mostly observed a translocation of PKC from the cytosol to the nucleus; a significant translocation to the plasma membrane was not visible in our experiments. We and others have demonstrated that the translocation of PKC from the cytosolic to the plasma membrane is quite variable and depends on the cell type used.21 In ECs a significant translocation to the plasma membrane was not visible in our experiments using confocal microscopy. The rather thin optical sections obtained by confocal microscopy do not allow the detection of small amounts of PKC attached to the EC plasma membrane. The variable intracellular distribution suggests that PKC exerts specific functions and acts on different substrates. Several groups have identified intracellular PKC receptors, which contribute to the subcellular distribution of PKC isoforms. These binding proteins include receptors for activated C kinase (so-called RACKs), which are localized in the cytoskeleton and nucleus.31 32 PKC is associated with nuclear membranes,9 10 32 33 34 and nuclear substrates of PKC have been identified.35 36 These substrates include proteins implicated in maintaining chromatin structure and in the replication or repair of DNA, such as topoisomerase II.37 These observations support the notion that PKC may perform important tasks within the cell nucleus.38
The isoform distribution pattern varied depending on the state of
activation of the cell or the agonists used. The phorbol ester TPA
induced an increase in the nuclear immunoreactivity of PKC and .
This shift of PKC isoforms to the nucleus was also observed after
exposure to bFGF or thrombin. Which signal directs PKC isoforms to the
nucleus is presently unclear. The presence of a nuclear
localization sequence in the regulatory domain of the enzyme has been
suggested.39 However, this motif is absent in PKC as
well as in the nonconventional PKC isoforms. In addition, a nuclear
localization sequence should be localized in the hinge region or the
catalytic domain of the molecule.40 Alternatively, PKC
itself may be directed to the nucleus by the action of PKC binding
proteins and may not necessarily rely on a nuclear localization
sequence.
Thrombin and bFGF exerted different effects on the subcellular
translocation of PKC isoforms. bFGF induced a marked increase in
nuclear immunoreactivity of PKC , while thrombin showed no effect.
Agonists that activate receptors with intrinsic tyrosine kinase
activity have different effects on PKC isoform distribution in
vascular smooth muscle cells than in G
protein–coupled receptors.41 Others have also
demonstrated differential translocation of PKC isoforms by thrombin and
platelet-derived growth factor.42 The lack of
nuclear PKC translocation by thrombin in our experiments suggests
that bFGF uses specific intracellular signals. bFGF leads to EC PKC
translocation in the absence of inositol-lipid
hydrolysis,43 suggesting that the effect of bFGF on PKC is
mediated by tyrosine phosphorylation and is calcium
dependent. However, the exact mechanisms of the bFGF-induced effect on
PKC localization remain unclear. Our results suggest that the bFGF
signaling pathway has a direct effect on PKC . Our findings could in
part explain how different classes of agonists result in contrasting
effects in ECs. They underscore an important role for PKC isoforms in
nuclear signaling and transcriptional control.
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Selected Abbreviations and Acronyms |
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Acknowledgments |
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This work was supported by a grant-in-aid from the Deutsche Forschungsgemeinschaft to Hermann Haller.
Received September 14, 1995; accepted January 5, 1996.
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 J.-K. Min, Y.-M. Kim, S. W. Kim, M.-C. Kwon, Y.-Y. Kong, I. K. Hwang, M. H. Won, J. Rho, and Y.-G. Kwon TNF-Related Activation-Induced Cytokine Enhances Leukocyte Adhesiveness: Induction of ICAM-1 and VCAM-1 via TNF Receptor-Associated Factor and Protein Kinase C-Dependent NF-{kappa}B Activation in Endothelial Cells J. Immunol., July 1, 2005; 175(1): 531 - 540. [Abstract] [Full Text] [PDF]
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 H. Tang, B. Low, S. A. Rutherford, and Q. Hao Thrombin induces endocytosis of endoglin and type-II TGF-{beta} receptor and down-regulation of TGF-{beta} signaling in endothelial cells Blood, March 1, 2005; 105(5): 1977 - 1985. [Abstract] [Full Text] [PDF]
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X. Li, C. N. Hahn, M. Parsons, J. Drew, M. A. Vadas, and J. R. Gamble Role of protein kinase C{zeta} in thrombin-induced endothelial permeability changes: inhibition by angiopoietin-1 Blood, September 15, 2004; 104(6): 1716 - 1724. [Abstract] [Full Text] [PDF]
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 C. Lindschau, P. Quass, J. Menne, F. Guler, A. Fiebeler, M. Leitges, F. C. Luft, and H. Haller Glucose-Induced TGF-{beta}1 and TGF-{beta} Receptor-1 Expression in Vascular Smooth Muscle Cells Is Mediated by Protein Kinase C-{alpha} Hypertension, September 1, 2003; 42(3): 335 - 341. [Abstract] [Full Text] [PDF]
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U. Tigges, B. Koch, J. Wissing, B. M. Jockusch, and W. H. Ziegler The F-actin Cross-linking and Focal Adhesion Protein Filamin A Is a Ligand and in Vivo Substrate for Protein Kinase C{alpha} J. Biol. Chem., June 20, 2003; 278(26): 23561 - 23569. [Abstract] [Full Text] [PDF]
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T. Minami, Md. R. Abid, J. Zhang, G. King, T. Kodama, and W. C. Aird Thrombin Stimulation of Vascular Adhesion Molecule-1 in Endothelial Cells Is Mediated by Protein Kinase C (PKC)-delta -NF-kappa B and PKC-zeta -GATA Signaling Pathways J. Biol. Chem., February 21, 2003; 278(9): 6976 - 6984. [Abstract] [Full Text] [PDF]
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 H. Lum and K. A. Roebuck Oxidant stress and endothelial cell dysfunction Am J Physiol Cell Physiol, April 1, 2001; 280(4): C719 - C741. [Abstract] [Full Text] [PDF]
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S. Hoshi, M. Goto, N. Koyama, K.-i. Nomoto, and H. Tanaka Regulation of Vascular Smooth Muscle Cell Proliferation by Nuclear Factor-kappa B and Its Inhibitor, I-kappa B J. Biol. Chem., January 14, 2000; 275(2): 883 - 889. [Abstract] [Full Text] [PDF]
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 E Genersch, K Hayess, Y Neuenfeld, and H Haller Sustained ERK phosphorylation is necessary but not sufficient for MMP-9 regulation in endothelial cells: involvement of Ras-dependent and -independent pathways J. Cell Sci., January 12, 2000; 113(23): 4319 - 4330. [Abstract] [PDF]
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 A. Skaletz-Rorowski, J. Waltenberger, J. G. Muller, E. Pawlus, K. Pinkernell, and G. Breithardt Protein Kinase C Mediates Basic Fibroblast Growth Factor–Induced Proliferation Through Mitogen-Activated Protein Kinase in Coronary Smooth Muscle Cells Arterioscler. Thromb. Vasc. Biol., July 1, 1999; 19(7): 1608 - 1614. [Abstract] [Full Text] [PDF]
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 C. M. Terry, J. A. Clikeman, J. R. Hoidal, and K. S. Callahan TNF-alpha and IL-1alpha induce heme oxygenase-1 via protein kinase C, Ca2+, and phospholipase A2 in endothelial cells Am J Physiol Heart Circ Physiol, May 1, 1999; 276(5): H1493 - H1501. [Abstract] [Full Text] [PDF]
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 R. Bychkov, K. Pieper, C. Ried, M. Milosheva, E. Bychkov, F. C. Luft, and H. Haller Hydrogen Peroxide, Potassium Currents, and Membrane Potential in Human Endothelial Cells Circulation, April 6, 1999; 99(13): 1719 - 1725. [Abstract] [Full Text] [PDF]
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M. Wellner, C. Maasch, C. Kupprion, C. Lindschau, F. C. Luft, and H. Haller The Proliferative Effect of Vascular Endothelial Growth Factor Requires Protein Kinase C-{alpha} and Protein Kinase C-{zeta} Arterioscler. Thromb. Vasc. Biol., January 1, 1999; 19(1): 178 - 185. [Abstract] [Full Text] [PDF]
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 S. M. Haffner The Importance of Hyperglycemia in the Nonfasting State to the Development of Cardiovascular Disease Endocr. Rev., October 1, 1998; 19(5): 583 - 592. [Abstract] [Full Text]
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 M. Gollasch, H. Haase, C. Ried, C. Lindschau, I. Morano, F. C. Luft, and H. Haller L-type calcium channel expression depends on the differentiated state of vascular smooth muscle cells FASEB J, May 1, 1998; 12(7): 593 - 601. [Abstract] [Full Text]
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 H. Haller, C. Lindschau, C. Maasch, H. Olthoff, D. Kurscheid, and F. C. Luft Integrin-Induced Protein Kinase C{alpha} and C{epsilon} Translocation to Focal Adhesions Mediates Vascular Smooth Muscle Cell Spreading Circ. Res., February 9, 1998; 82(2): 157 - 165. [Abstract] [Full Text] [PDF]
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W. S. Garver, M. A. Deeg, R. F. Bowen, M. M. Culala, E. L. Bierman, and J. F. Oram Phosphoproteins Regulated by the Interaction of High-Density Lipoprotein With Human Skin Fibroblasts Arterioscler. Thromb. Vasc. Biol., November 1, 1997; 17(11): 2698 - 2706. [Abstract] [Full Text]
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A. Hempel, C. Maasch, U. Heintze, C. Lindschau, R. Dietz, F. C. Luft, and H. Haller High Glucose Concentrations Increase Endothelial Cell Permeability via Activation of Protein Kinase C{alpha} Circ. Res., September 19, 1997; 81(3): 363 - 371. [Abstract] [Full Text]
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M. A. Deeg, R. F. Bowen, J. F. Oram, and E. L. Bierman High Density Lipoproteins Stimulate Mitogen-Activated Protein Kinases in Human Skin Fibroblasts Arterioscler. Thromb. Vasc. Biol., September 1, 1997; 17(9): 1667 - 1674. [Abstract] [Full Text]
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J.-J. Cheng, B.-S. Wung, Y.-J. Chao, and D. L. Wang Sequential Activation of Protein Kinase C (PKC)-alpha and PKC-epsilon Contributes to Sustained Raf/ERK1/2 Activation in Endothelial Cells under Mechanical Strain J. Biol. Chem., August 10, 2001; 276(33): 31368 - 31375. [Abstract] [Full Text] [PDF]
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