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*
Department of Dermatology and Emory Skin Diseases Research Core Center, Emory University School of Medicine, Atlanta, GA 30322;
Department of Dermatology, University of Washington, Seattle, WA 98195;
Departments of Physiology and Surgery, University of California, San Francisco, CA 94143; and
§
Veterans Administration Medical Center, Atlanta, GA 30322
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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A number of distinct neuropeptides are released by sensory C-fibers in the skin (10) where they exert a variety of modulatory actions on dermal and epidermal target cells. Substance P (SP) has been studied as the prototypic cutaneous neuropeptide because it has been shown to induce a wide range of inflammatory activities including vasodilation, immune cell activation, leukocyte infiltration, and cytokine secretion (reviewed in Refs. 11 and 12). SP mediates its effects on target cells by binding to cell surface G protein-coupled neurokinin receptors (NK-R) (13, 14, 15). In addition to SP, cutaneous C-fibers have also been shown to release several other neuropeptides including neurokinin A (NKA), neurokinin B (NKB), calcitonin gene-related peptide (CGRP), neurotensin (NT), and somatostatin (Sms) that may be important in certain neuroinflammatory processes in the skin (reviewed in 16 .
There is recent evidence that cutaneous neuroinflammatory interactions include the ability of neuropeptides to induce adhesion molecules on HDMEC (11, 17). We have detected NK-1, NK-2, and NK-3 receptor mRNA in cultured HDMEC and positive immunostaining of the NK-1R on dermal microvascular endothelium in human skin (17). SP engagement of the NK-1R resulted in a rapid increase in HDMEC intracellular Ca2+ levels, which could be specifically blocked by a selective NK-1R antagonist. SP-induced HDMEC activation was followed by increased mRNA and cell surface expression of ICAM-1. In the current study, we examine the effect of cutaneous nerve-released neuropeptides on microvascular endothelial cell VCAM-1 expression and function. These studies demonstrate the direct and specific ability of SP to induce VCAM-1 expression and function on these cells, thus further supporting the role of efferent neurological signals in mediating specific cutaneous inflammatory responses.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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HDMEC isolated from foreskins were obtained from the cell culture facility of the Emory Skin Disease Research Core Center (Atlanta, GA) (18). Experiments were conducted with cells in passages 3–5. HDMEC were cultured on a gelatinized (0.1%) surface in MCDB 131 (Life Technologies, Gaithersburg, MD) supplemented with 10% normal human serum (Irvine Scientific, Santa Ana, CA), 5 ng/ml epidermal growth factor (Clonetics, San Diego, CA), 1 mg/ml hydrocortisone acetate (Sigma, St. Louis, MO), 100 U/ml penicillin, 250 µg/ml amphotericin B, and 10 µg/ml streptomycin (Life Technologies). SV40 large T Ag-transformed HDMEC (HMEC-1) were generated by the Emory Skin Disease Research Core Center (19) and were cultured identically to primary isolated HDMEC. MOLT-4 T cells (derived from a patient with acute T cell leukemia; American Type Culture Collection, Manassas, VA) were cultured in RPMI 1640 medium supplemented with 10% FBS, 100 U/ml penicillin, 250 µg/ml amphotericin B, and 10 µg/ml streptomycin (Life Technologies).
Lyophilized SP, NKA, NKB, NT, CGRP, and Sms (Peninsula Laboratories,
Belmont, CA) were diluted in the appropriate volume of HDMEC assay
medium immediately before use. In selected studies, 1 µM NK-1
receptor antagonist GR82334 (Peninsula Laboratories) was added to
cultured endothelial cells 20 min before the addition of SP or
TNF-
. Polyclonal rat anti-human SP (American Research Products,
Belmont, MA) was diluted to 15 µg/ml in HDMEC medium. The
SP-degrading enzyme neutral endopeptidase (NEP) was a gift from Nigel
Bunnett (University of California, San Francisco, CA). Lyophilized
human recombinant TNF- and anti-TNF- Ab were obtained from
R&D Systems (Minneapolis, MN). Murine mAb P3C4 directed against human
VCAM-1 was a generous gift from Dr. Elizabeth Wayner (University of
Minnesota, Minneapolis, MN). Murine mAb 84H10 directed against human
ICAM-1, which functions as a blocking Ab was provided by Dr. Stephen
Shaw (National Institutes of Health, Bethesda, MD) (20).
In vivo expression of VCAM-1 in human skin microvascular endothelial cells by immunohistochemistry
Capsaicin-high potency (Zostrix, 0.075%, GenDerm, Lincolnshire, IL) was applied topically to the skin of four volunteers to stimulate the release of cutaneous neuropeptides and 4-mm punch biopsies were taken from treated sites at 6, 24, and 48 h as well as from an untreated site on the opposite limb taken immediately before capsaicin application. Tissue was embedded in O.C.T. (Tissue Tek; Miles, Elkhart, IN) and frozen at -70°C. Immunohistochemistry was performed on 8-µm sections using mouse anti-human VCAM-1 mAb P3C4 as the primary Ab (diluted 1:400), a biotinylated horse anti-mouse secondary Ab (Zymed, San Francisco, CA), and strept-avidin-biotin complex (Zymed) as a tertiary reagent to detect specific binding of the primary and secondary reagents. Samples were examined using a Nikon (New York, NY) Microshot SA microscope, and photographed at x120 magnification using a Nikon NFX-35 microscope camera. Control staining was performed on tissue processed without the primary Abs. No staining was obtained with either of the controls.
ELISA measurement of HDMEC cell surface VCAM-1 expression
HDMEC were plated onto 96-well plates and upon reaching 80%
confluence were either left untreated or stimulated for 16 h with 1,
10, 100, or 1000 nM of neuropeptides; SP, NKA, NKB, NT, CGRP, or Sms.
Cell viability was determined at the end of the stimulation
period by performing cell counts in separate plates. The addition of
300 U/ml TNF- for 16 h to cultured HDMEC was used as a positive
control for VCAM-1 cell surface induction. HDMEC VCAM-1 expression was
assessed using the anti-VCAM-1 mAb P3C4 or isotype-matched control
IgG by ELISA as described previously (21). Results represent the mean
of three values ± SD for each variable tested, and results are
representative of three independent assays.
HDMEC cell supernatant effects on VCAM-1 cell surface expression
To determine whether SP-treated HDMEC-conditioned medium
contained neuropeptide-induced factors that could be responsible for
the increased expression of VCAM-1, a series of supernatant transfer
experiments were performed. HDMEC were plated onto 96-well plates and
upon reaching 80% confluence were left untreated or treated with 10 nM
SP for 1–24 h. The addition of 300 U/ml TNF- for 16 h to
cultured HDMEC was used as a positive control for VCAM-1 cell surface
induction. After the stimulation period (1–24 h), these
cell-conditioned HDMEC supernatants (200 µl per sample) were removed
and, with or without the presence of 15 µg/ml polyclonal rat
anti-human SP Ab, 15 µg/ml polyclonal rat anti-human TNF-
Ab, or 1 µM NEP, transferred to fresh HDMEC 96-well plates from which
the medium had been removed. The HDMEC supernatants were then allowed
to incubate for 16 h. In selected experiments, supernatants
were "spiked" with 300 U/ml recombinant TNF-. HDMEC were then
assessed for VCAM-1 cell surface expression using mAb P3C4 or isotype
control IgG by ELISA as described previously (21).
VCAM-1 mRNA expression determination by Northern blot analysis
Cultured HDMEC (1 x 106 cells) were left
untreated or treated with 10 nM SP for 1, 3, 6, or 9 h or with 300
U/ml TNF- for 6 h. Total cellular RNA was isolated and Northern
blot analyses were done as described previously (21). A 0.6-kb
BglII fragment of the human VCAM-1 cDNA (provided by Dr. T.
Venkat Gopal, Otsuka America Pharmaceuticals, Rockville, MD) was used
as a VCAM-1 probe. Hybridization with radiolabeled ß-actin cDNA was
used for determination of lane loading consistency and for
normalization of ICAM-1 signal in the various conditions tested. The
autoradiograph was scanned on a flat bed scanner (La Cie, Beaverton,
OR) utilizing Photoshop software (Adobe Systems, Mountain View, CA).
The digitized image was then imported into a Power Point (Microsoft,
Redmond, WA) file where it was labeled and printed on a high-resolution
laser printer.
Determination of surface VCAM-1 expression on HMEC-1 by FACS
For FACS studies, we used the HDMEC cell line HMEC-1. We have
shown previously that VCAM-1 expression in these cells is similar to
that of HDMEC. Cultured HMEC-1 were left untreated or stimulated with
10 nM SP for 16 h. In selected experiments, cells were pretreated
with 1 µM of the selective NK-1R antagonist GR82334 for 20 min before
the addition of 10 nM SP or 300 U/ml TNF-. HMEC-1 were detached from
tissue culture plates by Versene 1:5000 (Life Technologies) and washed
with PBS (minus). Aliquots were transferred into tubes for Ab staining.
HMEC-1 were incubated with mouse anti-human VCAM-1 mAb P3C4, or
isotype control mouse anti-human IgG (H+L) (Jackson ImmunoResearch,
West Grove, PA) at a final concentration of 10 µg/ml for 1 h on
ice. Cells were washed twice and incubated with FITC-conjugated
affinity purified goat F(ab')2 anti-mouse IgG (H+L)
(Jackson ImmunoResearch) at a final concentration of 10 µg/ml for
1 h on ice. Subsequently, cells were washed twice and analyzed by
a FACscan flow cytometer (Becton Dickinson, Raleigh, NC) equipped with
CellQuest software for data acquisition and analysis. The forward
scatter threshold was set to permit analysis of viable endothelial
cells.
Adherence of MOLT-4 T cells to HDMEC
Binding assays measuring cellular adherence to HDMEC were
performed with human MOLT-4 T cells, which preferentially express the
VCAM-1 ligand (22), the integrin 1ß4, also
referred to as very late Ag-4 (VLA-4), and consequently adhere to
VCAM-1 on target cells (23). HDMEC were plated in 24-well plates
overnight, then either left untreated or treated for 16 h with
0.1–100 nM SP or 300 U/ml TNF- diluted in HDMEC assay medium in
triplicate wells, and then plates were washed three times with
HBSS+. MOLT-4 cells were incubated in the presence of 500
µCi of 51Cr for 1 h at 37°C at 5%
CO2 and then washed twice with HBSS+. Blocking
studies were performed on untreated or HDMEC treated for 16 h with
1 or 10 nM SP, by adding either anti-VCAM-1 mAb P3C4 or
isotype-matched anti-ICAM-1 mAb 84H10 at a final concentration of
10 µg/ml and allowing HDMEC to incubate for 20 min at 37°C
at 5% CO2 before the addition of MOLT-4 cells.
MOLT-4 cells were then added to untreated and variously treated HDMEC
wells at a concentration of 70,000 MOLT-4 cells per well and incubated
for 20 min at 37°C. The supernatant was removed and the
MOLT-4-overlayed HDMEC were washed gently three times with
HBSS+ to remove nonadherent MOLT-4 cells. One percent SDS
was added and allowed to incubate for 15 min at room temperature. Each
well was then swabbed with two cotton-tipped swabs that were counted in
a gamma counter. Adherence was calculated as % MOLT-4 binding =
[(cpm per well - background cpm)/(cpm added counts -
background cpm)] x 100, as described previously (24).
Photographs of unlabeled MOLT-4 T cells adhering to untreated or
SP-treated HDMEC were taken using Kodak (Rochester, NY) Ektachrome 66
Tungsten film on an OM2 camera (Olympus, New Hyde Park, NY) mounted on
an Olympus CK2 microscope.
Statistical analysis
For statistical analysis, Student’s t tests were performed. Values of p > 0.005 were not considered significant.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The in vivo effect of C-fiber-released neuropeptides on dermal
endothelial cell VCAM-1 expression was examined. A capsaicin-containing
cream (Zostrix) was applied topically to human volunteers to stimulate
the release of neuropeptides including SP from cutaneous sensory nerves
(25). Biopsies were obtained at 0, 6, 24, and 48 h, and evaluated
by immunohistochemistry for VCAM-1 expression associated with
microvascular endothelial cells in the dermis. As shown in
representative photomicrographs from one individual, microvascular
VCAM-1 staining was present in very low levels in untreated skin (Fig. 1
a). VCAM-1 immunoreactivity
increased slightly 6 h after capsaicin application (Fig. 1b). A significant increase in dermal endothelial cell
VCAM-1 staining was detected 24 h after application of topical
capsaicin (Fig. 1c), which was decreased in intensity by
48 h after capsaicin application (Fig. 1d). In other
subjects, VCAM-1 expression persisted at 48 h after application of
capsaicin (data not shown). Biopsies obtained from symmetrical
untreated sites showed no increased VCAM-1 staining (data not shown).
Thus, the release of neuropeptides by cutaneous sensory nerves results
in increased HDMEC VCAM-1 expression in vivo.
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A number of neuropeptides are detected in normal and inflamed skin
(10). The effect of several of these agents known to be released by
cutaneous sensory nerves on HDMEC VCAM-1 expression was investigated by
ELISA (Fig. 2). Untreated HDMEC (-) have
no constitutive VCAM-1 present on their surface. Similarly, treatment
with an isotype control IgG in place of the anti-VCAM mAb P3C4 had
no effect on HDMEC VCAM-1 surface expression. As indicated, incubation
with 1, 10, 100, or 1000 nM SP for 16 h up-regulated cell surface
VCAM-1 on HDMEC. The optimal 7-fold induction was observed with the 10
nM SP dose (Fig. 2). In contrast, incubation for 16 h with 1, 10,
100, or 1000 nM of NKA, NKB, NT, CGRP, or Sms had no effect on HDMEC
VCAM-1 induction (Fig. 2). Cell viability as determined by cell counts
after stimulation was not affected by any of the neuropeptide
treatments (data not shown). TNF- served as a positive control for
HDMEC VCAM-1 cell surface induction (Fig. 2).
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To test whether SP directly up-regulates the expression of HDMEC
VCAM-1 or whether this was in part due to the induction by SP of
HDMEC-secreted soluble factors that in turn could be responsible for
increased VCAM-1 expression, we performed a series of supernatant
transfer experiments (Fig. 3). Initially,
we determined that HDMEC treated between 1 and 24 h with 10 nM SP
resulted in a maximal 7-fold up-regulation of cell surface VCAM-1
expression at 16 h (Fig. 3A) as measured by ELISA.
TNF- significantly up-regulated VCAM-1 expression compared with
untreated cells and served as a positive control (Fig. 3A).
Isotype control IgG in place of the anti-VCAM-1 Ab showed no change
in VCAM-1 expression as compared with untreated HDMEC (data
not shown). The effect of supernatants obtained from SP-treated HDMEC
on VCAM-1 expression was measured (Fig. 3, B–D). As
indicated in Fig. 3B, conditioned medium from SP-treated
HDMEC added to freshly cultured HDMEC for 16 h was capable of
augmenting VCAM-1. This was most apparent with supernatants derived
from the short SP-HDMEC incubation time points (1–6 h). The results in
Fig. 3B are consistent with the high degree of protein
lability of the SP peptide in cell culture studies. After 3 h,
most of the SP bioactivity is lost in the HDMEC supernatants as
measured by SP receptor activation studies (data not shown).
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C). This complete abrogation of VCAM-1 induction with the
Ab to SP strongly suggests that no factor that could induce VCAM-1
(other than SP) is present in the SP-treated supernatants. No change in
HDMEC VCAM-1 expression was observed following treatment with
anti-TNF- Ab (data not shown). In addition, recent
transcriptional studies in our laboratory indicate that the addition of
neutralizing Abs to both TNFR-I and TNFR-II had no effect on the
ability of SP to transcriptionally activate either the ICAM-1 or VCAM-1
gene (our unpublished observations).
TNF- added to the supernatants in the studies conducted in Fig. 3C induced high levels of HDMEC VCAM-1 expression, thus
ruling out a nonspecific toxic effect of the SP Ab on HDMEC (Fig. 3D). When untreated and SP-treated HDMEC supernatants were
incubated with NEP, which selectively degrades SP, and transferred to
fresh HDMEC, there was no up-regulation of VCAM-1, although
TNF--treated/NEP-treated supernatants were still capable of inducing
VCAM-1 on HDMEC (data not shown). These data support the direct role of
SP in the induction of HDMEC VCAM-1 expression.
SP induces HDMEC VCAM-1 mRNA
To determine whether SP is capable of modulating VCAM-1 mRNA in
HDMEC, Northern blot analysis was performed (Fig. 4A). HDMEC were treated with
10 nM SP for 1, 3, 6, or 9 h, or for 6 h with 300 U/ml
TNF-. HDMEC VCAM-1 mRNA expression was normalized to ß-actin mRNA
expression by densitometric analysis for each experimental condition
(Fig. 4B). HDMEC do not constitutively express basal levels
of VCAM-1 mRNA. HDMEC VCAM-1 mRNA was significantly induced
1–3 h after treatment with SP, which was further augmented after
6 h and 9 h of exposure to SP (Fig. 4). TNF- induction of
HDMEC VCAM-1 mRNA expression served as a positive control for this
study. Thus, SP is capable of inducing HDMEC VCAM-1 mRNA.
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The effect of SP on immortalized HMEC-1 cell surface VCAM-1
expression was examined by FACS analysis and mean log10
fluorescence values were compared. Untreated HMEC-1 were calculated as
having a mean channel fluorescence of 1.90 for surface VCAM-1 staining
(Fig. 5, dotted lined
histograms). HMEC-1 treated with 10 nM SP for 16 h resulted
in a >6.5-fold increase in mean fluorescence, with a mean channel
fluorescence of 12.73 for surface VCAM-1 (Fig. 5B) compared
with control untreated cells. Pretreatment of HMEC-1 with an NK-1R
antagonist before the addition of SP resulted in a mean channel
fluorescence of 2.26, demonstrating the important role of the NK-1R in
SP-induced VCAM-1 up-regulation (Fig. 5C). The NK-1R
antagonist alone had no effect on VCAM-1 cell surface expression (data
not shown). TNF--treated HMEC-1 displayed high levels of cell
surface VCAM-1 expression, with a mean fluorescence of 45.2 and served
as a positive control (Fig. 5D). TNF--mediated HMEC-1
VCAM-1 up-regulation was not blocked by pretreatment with an NK-1R
antagonist (data not shown). In addition, no increase in fluorescence
occurred when an isotype matched control Ab was used on SP-treated
HMEC-1 (Fig. 5A). Therefore, these data demonstrate SP
up-regulates the expression of cell surface VCAM-1 on HMEC-1, primarily
through interaction with the NK-1R.
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The biological consequence of SP-induced HDMEC VCAM-1 cell surface
expression was determined using a quantitative cellular adhesion assay
with MOLT-4 T cells (22). MOLT-4 T cells bear the counter-receptor
ligand for VCAM-1, VLA-4 (4ß1) (3). Although
51Cr-labeled MOLT-4 T cells showed minimal binding to
untreated cultured HDMEC (Fig. 6), the
adhesion of MOLT-4 T cells to SP-treated HDMEC increased following the
addition of 0.1 nM SP for 16 h. The enhanced binding of MOLT-4
cells to 1 nM SP-treated HDMEC was significant as well. As indicated,
10 nM SP induced a 4-fold increase in the adhesion of MOLT-4 cells to
HDMEC. There was a small inductive increase in MOLT-4 cell adhesion to
HDMEC treated with 100 nM SP. The specificity of VCAM-1 in the
adherence of MOLT-4 T cells to HDMEC after SP treatment was examined.
Leukocyte binding to SP-treated HDMEC could be specifically
blocked by pretreatment of cells with an anti-VCAM-1 blocking Ab
before the addition of labeled MOLT-4 T cells, whereas HDMEC pretreated
with a blocking anti-ICAM-1 Ab (20) showed no decrease in
VCAM-1/VLA-4 binding to MOLT-4 cells (Fig. 6). TNF- treatment
of HDMEC served as a positive control for enhanced MOLT-4 binding (Fig. 6).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and IL-1, respectively
(34, 35). SP has been shown to preferentially enhance the migration of
eosinophils in asthmatic patients (36). Understanding the role of SP in
leukocyte trafficking will aid in elucidating its contribution to the
pathogenesis of certain inflammatory diseases.
By the expression of specific cell adhesion molecules, as well as
release of cytokines and chemokines, the microvascular endothelium is a
key cell in regulating leukocyte trafficking into inflamed tissue. The
nature of the inflammatory stimulus determines whether lymphocytes,
monocytes, neutrophils, or eosinophils predominate, exercising
specificity in molecular signals that control traffic of leukocyte
subsets. The main classes of adhesion molecules that play a part in
trafficking are the selectins, the integrins, members of the Ig
superfamily, and sialoadhesins (37). In skin, P- and E-selectins
mediate initial leukocyte adhesion, whereas ICAM-1 and VCAM-1 mediate
subsequent, firm adhesion and transendothelial migration. ICAM-1 binds
leukocytes via ß2 integrins and can therefore recruit
both granulocytes and monocytes from the bloodstream to sites of tissue
injury, whereas VCAM-1 binds monocytes, eosinophils, and lymphocytes
expressing the integrins 4ß1 and
4ß7 specifically supporting the
recruitment of chronic inflammatory cells (38, 39). SP has been
suggested as playing a role in the pathogenesis of psoriasis, where
possibly VCAM-1/VLA-4-mediated chronic inflammation exists (40).
Our present evaluation of the effect of SP on cutaneous microvascular endothelial cell VCAM-1 expression further supports the role of neuropeptides in the regulation of leukocyte trafficking in the skin. The ability of neuropeptides to influence adhesion molecule expression on microvascular endothelial cells was shown in our recent studies of SP-induction of dermal endothelial ICAM-1 expression (17). Additionally, SP was found to up-regulate adhesins P-selectin and E-selectin in normal human skin (36) when incubated for a limited time period (0–8 h). We demonstrate induction of dermal microvascular endothelial cell VCAM-1 expression in vivo after topical application of the neuropeptide-releasing agent capsaicin. Cutaneous sensory nerve-released neuropeptides NKA, NKB, NT, CGRP, and Sms had no effect on HDMEC VCAM-1 cell surface up-regulation across a broad concentration range, whereas SP significantly increased VCAM-1 cell surface expression. Additionally, SP was capable of directly up-regulating HDMEC surface VCAM-1, without the participation of other HDMEC-derived factors. This direct SP induction resulted in increased levels of VCAM-1 mRNA and NK-1R-mediated VCAM-1 cell surface expression. SP treatment resulted in functional VCAM-1 induction as demonstrated by the ability of VCAM-1 on HDMEC to bind its ligand VLA-4 on MOLT-4 T cells.
SP is known to induce the secretion of cytokines from various immune cells (32, 35, 41). We determined whether SP exposure resulted in HDMEC secretion of soluble factors capable of inducing VCAM-1 expression. Our study indicates that SP was capable of directly inducing increased HDMEC VCAM-1 expression without the participation of HDMEC-derived factors. In vivo, cytokine release may play a role in endothelial adhesion molecule up-regulation, but cytokine release is neither necessary nor required for SP-mediated induction of VCAM-1 on cultured microvascular endothelial cells.
In summary, our studies demonstrate that SP is capable of directly regulating dermal microvascular endothelial VCAM-1 expression, primarily through activation of the NK-1 receptor. These findings further support the role of neuropeptide modulation of leukocyte recruitment in the skin during cutaneous inflammatory reactions. The coordination of signals such as neuropeptide release and adhesion molecule up-regulation may play a role in the development of a wide range of inflammatory responses and wound healing. Further investigation into the interactions of various neuropeptides, adhesion molecules, and other inflammatory mediators should provide a rational basis for controlling inflammatory disorders that may have a significant neurogenic component such as rheumatoid arthritis, asthma, psoriasis, and atopic eczema.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. John C. Ansel, Department of Dermatology, Emory University School of Medicine, 5001 Woodruff Memorial Research Building, Atlanta, GA 30322. E-mail address:
3 Abbreviations used in this paper: HDMEC, human dermal microvascular endothelial cell(s); SP, substance P; NK-R, neurokinin receptor; NKA, neurokinin A; NKB, neurokinin B; CGRP, calcitonin gene-related peptide; NT, neurotensin; Sms, somatostatin; NEP, neutral endopeptidase; VLA-4, very late Ag-4.
Received for publication July 8, 1998. Accepted for publication October 15, 1998.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and interleukin-1 in dermal microvascular endothelial cells. Blood. 87:211.
-mediated induction of ICAM-1 expression via activation of STAT proteins. J. Invest. Dermatol. 108:295.[Medline]
gene expression in murine mast cells. J. Immunol. 150:1.[Abstract]
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 M. T. Tran, R. N. Lausch, and J. E. Oakes Substance P Differentially Stimulates IL-8 Synthesis in Human Corneal Epithelial Cells Invest. Ophthalmol. Vis. Sci., November 1, 2000; 41(12): 3871 - 3877. [Abstract] [Full Text]
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 P. G. McLean, A. Ahluwalia, and M. Perretti Association between Kinin B1 Receptor Expression and Leukocyte Trafficking across Mouse Mesenteric Postcapillary Venules J. Exp. Med., July 31, 2000; 192(3): 367 - 380. [Abstract] [Full Text] [PDF]
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 P. D. Drummond The Effect of Peripheral Opioid Block and Body Cooling on Sensitivity to Heat in Capsaicin-Treated Skin Anesth. Analg., April 1, 2000; 90(4): 923 - 927. [Abstract] [Full Text] [PDF]
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K. L. Quinlan, S. M. Naik, G. Cannon, C. A. Armstrong, N. W. Bunnett, J. C. Ansel, and S. W. Caughman Substance P Activates Coincident NF-AT- and NF-{kappa}B-Dependent Adhesion Molecule Gene Expression in Microvascular Endothelial Cells Through Intracellular Calcium Mobilization J. Immunol., November 15, 1999; 163(10): 5656 - 5665. [Abstract] [Full Text] [PDF]
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 K. A. DeFea, Z. D. Vaughn, E. M. O'Bryan, D. Nishijima, O. Dery, and N. W. Bunnett The proliferative and antiapoptotic effects of substance P are facilitated by formation of a beta -arrestin-dependent scaffolding complex PNAS, September 26, 2000; 97(20): 11086 - 11091. [Abstract] [Full Text] [PDF]
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