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Departments of
*
Pediatrics,
Medicine, and
Pathology, Duke University Medical Center, Durham, NC 27710;
Department of Biology, University of North Carolina, Chapel Hill, NC 27599; and
¶ Department of Immunology, The Scripps Research Institute, La Jolla, CA 92037
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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-activated murine
endothelial cells under low-shear conditions. Treatment with either
anti-FKN or anti-CX3CR1-blocking Ab significantly
inhibited PBMC binding, indicating that a large proportion of leukocyte
binding to murine endothelium occurs via the FKN and CX3CR1
adhesion receptors. To determine the functional significance of FKN in
rejection, we treated cardiac allograft recipients with daily
injections of anti-CX3CR1 Ab. Treatment with the
anti-CX3CR1 Ab significantly prolonged allograft
survival from 7 ± 1 to 49 ± 30 days (p
< 0.0008). These studies identify a critical role for FKN in the
pathogenesis of acute rejection and suggest that FKN may be a useful
therapeutic target in rejection. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Recently, the human chemokine fractalkine
(FKN)3 and its murine
homologue neurotactin have been identified as novel chemokines with a
unique transmembrane chemokine/mucin hybrid structure (9, 10). FKN not only promotes leukocyte activation, but, unlike
other chemokines, can mediate each individual step of the leukocyte
adhesion cascade (11). Specifically, FKN interacts with
its unique receptor, CX3CR1, to affect firm
adhesion of resting monocytes, resting and activated
CD8+ T lymphocytes, and activated NK cells. Like
all the other known chemokine receptors, CX3CR1
belongs to the large family of G protein-coupled receptors
(12). Although little is known about
CX3CR1 signaling mechanisms, calcium flux that is
triggered by CX3CR1 is pertussis toxin sensitive,
suggesting that it couples to Gi
(12).
Human CX3CR1 mRNA is expressed in T lymphocytes, monocytes, and NK cells (12), and the rat homologue RBS11 is found in leukocytes, microglia, and astrocytes (13, 14, 15). The equivalent murine receptor, mCX3CR1, is found in PBL and multiple organs, with particular abundance in brain and lung (16). Given that CX3CR1-bearing leukocyte subsets are precisely the ones found within acutely rejecting allografts, we postulated that FKN might play a functional role in rejection. We tested this hypothesis using mouse models. Our studies demonstrate that 1) FKN expression is enhanced on rejecting mouse cardiac allografts; 2) enhanced FKN expression on activated mouse endothelial cells promotes increased leukocyte adhesion; and 3) inhibition of FKN-CX3CR1 signaling with an anti-CX3CR1 Ab significantly prolongs survival of mouse cardiac allografts in a vigorous model of acute rejection. Thus, the FKN-CX3CR1 pathway for leukocyte trafficking and activation plays an integral role in the pathogenesis of allograft rejection.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Mouse PBMC were isolated from heparinized whole blood by centrifugation over lymphocyte separation medium (Organon Teknika, Durham, NC) as previously described (17). Contaminating RBC were lysed, and PBMC were washed three times with PBS. After the final wash step, PBMC were resuspended in warm FCS and incubated at 37°C with 10 µCi of 51Cr/million PBMC for 1 h. Unincorporated 51Cr was removed by washing, and the labeled cells were resuspended in medium. Py-4-1 endothelial cells, derived from hemangiomas of polyomavirus transgenic mice (18), were grown in RPMI 1640 medium (Life Technologies, Gaithersburg, MD) supplemented with 10% FCS.
Antibodies
Abs used in these studies included purified polyclonal rabbit anti-mouse FKN directed against the chemokine domain of mouse FKN (Torrey Pines Biolabs, San Diego, CA) and purified polyclonal rabbit anti-rat CX3CR1 Ab (Torrey Pines Biolabs) (13, 14), each used at concentration of 20 µg/ml. The secondary Ab used in these studies was biotin-conjugated goat anti-rabbit IgG adsorbed for human, mouse, and rat serum proteins (Jackson ImmunoResearch Laboratories, West Grove, PA), used at a final concentration of 10 µg/ml. The tertiary reagent was streptavidin-FITC (PharMingen, San Diego, CA), used at 5 µg/ml. For flow cytometry studies, described below, the Abs used were FITC-conjugated CD3e (145-2C11), CD4 (L3T4), CD8a (53-6.7), IgM (R6-60.2), Pan-NK (DX5), NK1.1 (PK136), and CD11b (Mac-1, M1/70), all from PharMingen, and F4/80 from Serotec (Raleigh, NC).
Flow cytometry
Py-4-1 cells were stimulated for 4 h with recombinant
murine TNF- 100 U/ml (Genentech, San Francisco, CA) and then lifted
from the cell culture flask using enzyme-free cell dissociation buffer
(Life Technologies). Cells were washed in PBS containing 2% bovine
calf serum and then incubated with polyclonal anti-FKN Ab at 4°C
for 45 min. Cells were washed and then incubated with biotinylated goat
anti-rabbit IgG for 45 min at 4°C. After another wash, the cells
were incubated with streptavidin-FITC for 45 min at 4°C. Following a
final wash, the cells were fixed in 2% formaldehyde in PBS and
analyzed by flow cytometry using a FACScan flow cytometer (Becton
Dickinson, San Jose, CA).
In vitro adhesion assays
Leukocyte adhesion assays were performed as described previously
with minor modifications (17). Briefly, Py-4-1 cells were
grown to confluence in 24-well culture plates, activated with TNF-,
washed, and incubated at 37°C for 20 min with medium or anti-FKN
Ab. 51Cr-labeled PBMC (0.5 x
106/well) were resuspended in 400 µl of medium
alone or medium containing anti-CX3CR1 Ab at
20 µg/ml. The leukocyte suspension was added to the endothelial
monolayer under gentle rocking conditions (10 cycles/min). After 30 min
the medium was decanted, and the wells were gently washed. Adherent
cells were lysed by treating with 1% Triton in PBS. Total binding was
determined by measuring individual well-associated cpm using a gamma
counter. To determine whether treatment with
anti-CX3CR1 Ab altered the phenotype of PBMC
bound to the activated endothelium, a parallel set of experiments was
performed in which endothelial monolayers were overlaid with
nonradiolabeled PBMC under the conditions described above. At the
conclusion of the assay, cells were gently lifted from culture wells
using enzyme-free cell dissociation buffer (Life Technologies), and
FACS analysis performed, gating on the leukocyte rather than
endothelial cell population. Cell surface markers evaluated included
CD3, CD4, CD8, IgM, NK1.1, pan-NK, and F4/80.
Mouse cardiac transplantation
Heterotopic murine cardiac transplants were performed as
described previously (19). Recipient C57BL/6
(H-2b) mice were anesthetized using isoflurane
and prepared by separating the vena cava and aorta between the renal
vasculature and the iliac bifurcation. The donor heart was dissected
from an MHC-mismatched (DBA/2 x BALB/c)F1
(H-2d) mouse, and an end-to-side anastomosis was
created between the recipient aorta and the donor heart ascending
aorta. A similar vascular anastomosis was created between the donor
superior vena cava and the inferior vena cava of the recipient.
Surgical mortality of the recipients was 
10%. Allograft survival
was determined by directly palpating the cardiac impulse through the
abdominal wall, with graft failure defined as the cessation of a
palpable heartbeat. For immunostaining experiments, cardiac allografts
and isografts from untreated recipients were harvested at 3 and 7 days
posttransplantation.
Anti-CX3CR1 Ab treatment
Heterotopic mouse heart transplants were performed as described above. On the day before transplantation, 75 µl of undiluted CX3CR1 antiserum or control rabbit serum was injected i.p. into prospective recipient C57BL/6 (H-2b) mice. Thereafter, recipient mice received similar injections each day for a period of up to 4 wk following transplantation, after which time the therapy was discontinued. Cardiac allograft survival was determined as described above. To further examine the effect of anti-CX3CR1 on the nature of rejection, we evaluated the histopathology of cardiac allografts in the experimental groups. On day 3 or 7 after transplantation, the allografts were removed and placed in 10% buffered formalin. The hearts were then sectioned and stained with hematoxylin and eosin, and the slides were reviewed by a pathologist (D.N.H.) who was masked to the treatment groups.
Immunohistochemistry
The ventricles were cut in cross-section and fixed in formalin.
Formalin-fixed hearts were paraffin-embedded, sectioned, and mounted on
slides. Slides were subsequently deparaffinized by immersing in xylene
and graded ethanols. Immunohistochemical staining for FKN was performed
using the pressure cooker method for Ag retrieval. Briefly, slides were
incubated in an epitope decloaking chamber (Biocare Medical, Walnut
Creek, CA) at 120°C and 
15–20 psi for 25 min according to the
manufacturer’s specifications. Slides were demarcated using a
lipophilic pen and were incubated for 30 min at room temperature with
normal goat serum. Slides were washed four times with PBS containing
0.5% BSA, then incubated with either anti-FKN Ab or normal rabbit
serum at room temperature for 45 min. Slides were washed four times
with PBS containing 0.5% BSA, then incubated with biotin-conjugated
goat anti-rabbit IgG at room temperature for 30 min. Slides were
washed again, then incubated with ABC Elite developing reagent (Vector
Laboratories, Burlingame, CA) at room temperature for 30 min. This was
followed by another wash and incubation with Tris-HCl containing
diaminobenzidine and hydrogen peroxide (0.6%) for 3 min. Slides were
then placed under running water for 2 min, stained with hematoxylin for
15 s, washed under running water for 2 min, and then placed in
water containing NH4OH (1/500) for 15 s.
Slides were sequentially immersed in graded ethanols and xylene, then
mounted using Acrytol (Surgipath, Richmond, IL).
Statistical analysis
Results are expressed as the mean ± SEM unless otherwise indicated. Student’s t test was used to determine the level of significance of differences in mean values between treatment groups. For nonparametric analysis of the allograft survival study, a Mann-Whitney U test was used.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Since the endothelium of the allograft represents the initial site
of interactions with leukocytes from the recipient, we examined
expression of FKN in the mouse endothelial cell line Py-4-1. As shown
in Fig. 1
, low levels of FKN expression
could be detected in quiescent Py-4-1 cells by flow cytometry using
anti-FKN Ab. Exposure of Py-4-1 cells to TNF-, a proinflammatory
cytokine present at high levels in rejecting allografts, caused a
marked up-regulation of FKN expression. These data suggest that in the
mouse, similar to previous reports in human cells, endothelial
activation by proinflammatory cytokines such as TNF- is associated
with enhanced cell surface expression of FKN.
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To determine the functional consequences of enhanced FKN
expression in activated Py-4-1 cells, we examined binding of freshly
isolated mouse PBMC to confluent Py-4-1 monolayers under low-shear
conditions. Quiescent endothelial monolayers bound PBMC at low levels,
but binding increased by 4-fold in response to activation with TNF-
(data not shown). To define the contribution of the FKN-
CX3CR1 pathway to leukocyte adhesion to the
endothelial cell line, PBMC were incubated with
anti-CX3CR1 Ab before the assay. The
anti-CX3CR1 Ab caused a modest (12%), but
significant, reduction in leukocyte binding (p
< 0.01). Preincubation of PBMC with control rabbit IgG had no effect
on binding to endothelial cells (Fig. 2).
Pretreatment of Py-4-1 cells with an anti-FKN Ab caused a more
marked (33%) reduction in PBMC binding (p <
0.01). Combined treatment with the two Abs further reduced PBMC binding
(62%; p < 0.01). Thus, a significant portion of PBMC
binding to activated murine endothelial cells is mediated by the
FKN-CX3CR1 pathway.
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FKN expression in rejecting cardiac allografts
To determine whether FKN expression might be regulated by an
inflammatory response in vivo, we examined expression of FKN protein in
a mouse model of cardiac transplant rejection. We examined FKN
expression in mouse cardiac allografts at early and late time points
during rejection. FKN expression in allografts was compared with that
in nonrejecting isografts in tissue sections using polyclonal rabbit Ab
against mouse FKN. Negligible FKN staining was detected in nonrejecting
isografts at any time (Fig. 3, C and F). In contrast, on day 3 after transplant
when leukocyte infiltrates were first detectable, FKN was prominently
expressed diffusely on endothelium of cardiac allografts (Fig. 3A). FKN was also detected on epicardium and endocardial
surfaces. By day 7, further enhancement of FKN expression was detected,
including apparent expression on myocardium (Fig. 3D). No
staining was observed with nonimmune rabbit serum (Fig. 3, B
and E). Thus, enhanced FKN expression accompanies cardiac
allograft rejection.
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To determine the functional importance of the up-regulation of FKN
expression in allograft rejection, we treated recipient animals with a
neutralizing anti-CX3CR1 Ab. Beginning on the
day before transplantation, recipient animals received daily i.p.
injections of 75 µl of anti-CX3CR1 Ab or
nonimmune rabbit serum for up to 4 wk. In the animals that received
rabbit serum (n = 9), the mean survival of cardiac
allografts was 7 ± 1 days (Fig. 4).
Treatment with anti-CX3CR1 Ab significantly
prolonged graft survival to a mean of 49 ± 30 days (Fig. 4;
p < 0.0008). Although all the control allografts
underwent rejection on day 7 or 8, the survival of
anti-CX3CR1-treated grafts ranged from 12–91
days (Table I). In long-surviving
anti-CX3CR1-treated animals, Ab
administration was discontinued after 4 wk, and these grafts continued
to survive for a mean of 33 ± 23 days, resulting in total mean
survival of 61 ± 29 days (Fig. 4). Therefore, treatment with
anti-CX3CR1 Ab with no additional
immunosuppression induces a significant prolongation of cardiac
allograft survival. These findings indicate a unique role for the
FKN-CX3CR1 pathway in promoting allograft
injury.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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A general understanding of the process of leukocyte recruitment into sites of inflammation has been developed using simple in vitro and in vivo models (1). This process consists of systematic, stepwise interactions between adhesion molecules expressed on endothelium and the circulating leukocyte. During its later phases, chemokine signaling through G protein-coupled receptors facilitates firm adhesion and migration of the leukocyte across the endothelium (2, 20, 21). Signals from chemokine receptors promote and facilitate leukocyte activation and thus may also contribute to the intensity of the inflammatory response. Although it is generally assumed that this paradigm explains the recruitment and accumulation of immune cells within rejecting tissues, the relative contributions of each component of this recruitment process to allograft rejection have not been explicitly characterized.
We have studied the potential role of a recently identified member of
the chemokine family, FKN, in rejection. FKN is unusual among
chemokines in that it is a membrane-bound molecule with a chemokine
domain coupled to a mucin stalk (22). In a mouse
endothelial cell line, we found that expression of FKN is markedly
up-regulated when the cells are activated by TNF-. These findings
are consistent with previous studies of Bazan and associates using
HUVEC, where TNF- induced expression of FKN mRNA and protein
(9). Since TNF- is highly expressed in rejecting
allografts, this regulatory pathway for FKN expression may have direct
relevance in rejection.
Human FKN mRNA expression is present in normal heart, brain, kidney, lung, pancreas, and colon (9). Likewise, in normal mice, neurotactin mRNA is especially prominent in brain and, to a lesser extent, in kidney, lung, and heart (10). The studies by Harrison et al. (14) further pinpoint the site of FKN expression in the CNS of the rat to neurons. Similar to our studies in cultured endothelial cells, studies by Pan et al. (10) have suggested that inflammatory stimuli may also enhance FKN expression in vivo. These authors found increased levels of neurotactin staining on vascular endothelium in brain following LPS treatment (10). Enhanced neurotactin expression in activated microglia from mice with severe experimental autoimmune encephalomyelitis has also been documented. We found only negligible levels of FKN expression in nonrejecting cardiac isografts. In contrast, expression of FKN was markedly up-regulated in allografts as early as 3 days after transplantation. FKN expression became more prominent as the rejection process progressed. The most intense staining for FKN was present on endothelial surfaces, where it might be involved in the early events associated with trafficking of circulating leukocytes into the graft. However, definite staining was also found on epicardial surfaces and cardiac myocytes in ventricular tissue. Although FKN expression in myocytes has recently been described (23), its functional significance is not clear.
FKN has dual functions in leukocyte migration: as a chemokine and also
as a mediator of leukocyte capture and firm adhesion (11, 12). For example, under physiologic conditions of flow, FKN and
CX3CR1 interact to mediate sequential steps of
the leukocyte adhesion cascade, including rapid capture, stable arrest,
and activation (11). Firm leukocyte adhesion mediated by
FKN-CX3CR1 is not sensitive to pertussis toxin,
suggesting that it does not require integrin activation. Although most
previous studies of FKN-dependent leukocyte adhesion have been
performed using human systems, we find that neurotactin has a similar
role in the mouse. In our studies a substantial portion (>60%) of
PBMC binding to TNF--activated murine endothelial cells is mediated
by an FKN-CX3CR1-dependent pathway. Although
anti-FKN Ab blockade inhibited binding by 33%,
anti-CX3CR1 treatment only blocked 12% of
adhesion, suggesting incomplete blockade of this pathway. In contrast,
blocking both arms of the FKN-CX3CR1pathway using
Ab therapy dramatically decreased leukocyte-endothelial binding by
62%. Taken together, these data suggest that up-regulation of FKN on
endothelial surfaces of a rejecting allograft should have potent
effects to promote leukocyte adhesion.
The propensity for a given cell population to be influenced by FKN is determined by expression of CX3CR1, the receptor for FKN. For example, in microchemotaxis assays, soluble FKN is a potent chemoattractant for CX3CR1-bearing leukocytes (12). CX3CR1 expression has been detected in a range of leukocyte populations, including CD14+ monocytes, CD8+ lymphocytes, and CD16+ NK cells (12). These cells are all typically present among the infiltrating cell population observed in rejecting allografts. Similar to other chemokine receptors, such as CCR1 and CCR2 (24), CX3CR1 expression in T lymphocytes is up-regulated by cytokines such as IL-2 (12). Since the microenvironment of the rejecting allograft is rich in proinflammatory cytokines, the capacity for FKN responses may be maximized during rejection.
To determine whether enhanced expression of FKN plays a role in
allograft rejection, we treated cardiac transplant recipients with a
neutralizing Ab against CX3CR1. In a vigorous
model of allograft rejection, blockade of the
FKN-CX3CR1 pathway significantly increased
allograft survival. In comparing the individual animals, the impact of
anti-CX3CR1 Ab treatment was variable. The
reasons for this variability are not clear, but they may relate to
variations in the degree of CX3CR1 inhibition
that was achieved or to interactions with other factors, such as
ishemic injury. Nonetheless, as depicted in Table I
, a definite
beneficial effect of therapy was seen in each animal that received
anti-CX3CR1 Ab. These data indicate that the
FKN-CX3CR1 pathway has a unique function to
promote allograft rejection.
Our findings add to the accumulating body of evidence suggesting an important role for chemokines in the pathogenesis of allograft rejection. For example, enhanced expression of a number of C-C chemokines has been demonstrated in murine allograft models as well as in rejecting transplants in humans (5). Because of the range of chemokines that are expressed in rejection and the substantial overlap in functions between chemokine family members, some authors suggested that inhibition of individual chemokines or chemokine receptors was unlikely to have a substantive impact on the course of rejection. Nonetheless, several reports have identified specific chemokines and chemokine receptors that appear to have unique and nonoverlapping roles in the pathogenesis of allograft rejection. For example, the absence of CCR1 significantly prolonged allograft survival in a murine model of acute cardiac allograft rejection (25). However, the effects of CCR1 deficiency were more marked across isolated class I or class II MHC differences. Our data suggest that the FKN-CX3CR1 pathway also has a nonredundant role in allograft rejection. The efficacy of anti-CX3CR1 Ab in prolonging the survival of cardiac allografts with complete MHC disparity between donor and host suggests that the FKN-CX3CR1 pathway may be a useful therapeutic target for antirejection therapies.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Lisa A. Robinson, Box 3959, Duke University Medical Center, Durham, NC 27710.
3 Abbreviation used in this paper: FKN, fractalkine.
Received for publication June 6, 2000. Accepted for publication September 1, 2000.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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R. Donadelli, C. Zanchi, M. Morigi, S. Buelli, C. Batani, S. Tomasoni, D. Corna, D. Rottoli, A. Benigni, M. Abbate, et al. Protein Overload Induces Fractalkine Upregulation in Proximal Tubular Cells through Nuclear Factor {kappa}B- and p38 Mitogen-Activated Protein Kinase-Dependent Pathways J. Am. Soc. Nephrol., October 1, 2003; 14(10): 2436 - 2446. [Abstract] [Full Text] [PDF]
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C. Hundhausen, D. Misztela, T. A. Berkhout, N. Broadway, P. Saftig, K. Reiss, D. Hartmann, F. Fahrenholz, R. Postina, V. Matthews, et al. The disintegrin-like metalloproteinase ADAM10 is involved in constitutive cleavage of CX3CL1 (fractalkine) and regulates CX3CL1-mediated cell-cell adhesion Blood, August 15, 2003; 102(4): 1186 - 1195. [Abstract] [Full Text] [PDF]
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R. A. Tripp, A. Dakhama, L. P. Jones, A. Barskey, E. W. Gelfand, and L. J. Anderson The G Glycoprotein of Respiratory Syncytial Virus Depresses Respiratory Rates through the CX3C Motif and Substance P J. Virol., June 1, 2003; 77(11): 6580 - 6584. [Abstract] [Full Text] [PDF]
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M. D. Silverman, D. O. Zamora, Y. Pan, P. V. Texeira, S.-H. Baek, S. R. Planck, and J. T. Rosenbaum Constitutive and Inflammatory Mediator-Regulated Fractalkine Expression in Human Ocular Tissues and Cultured Cells Invest. Ophthalmol. Vis. Sci., April 1, 2003; 44(4): 1608 - 1615. [Abstract] [Full Text] [PDF]
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D. N. Streblow, C. Kreklywich, Q. Yin, V. T. De La Melena, C. L. Corless, P. A. Smith, C. Brakebill, J. W. Cook, C. Vink, C. A. Bruggeman, et al. Cytomegalovirus-Mediated Upregulation of Chemokine Expression Correlates with the Acceleration of Chronic Rejection in Rat Heart Transplants J. Virol., February 1, 2003; 77(3): 2182 - 2194. [Abstract] [Full Text] [PDF]
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 N. G. Inston and P. Cockwell The evolving role of chemokines and their receptors in acute allograft rejection Nephrol. Dial. Transplant., August 1, 2002; 17(8): 1374 - 1379. [Full Text] [PDF]
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A. M. Fong, S. M. Alam, T. Imai, B. Haribabu, and D. D. Patel CX3CR1 Tyrosine Sulfation Enhances Fractalkine-induced Cell Adhesion J. Biol. Chem., May 24, 2002; 277(22): 19418 - 19423. [Abstract] [Full Text] [PDF]
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 K. Balabanian, A. Foussat, P. Dorfmuller, I. Durand-Gasselin, F. Capel, L. Bouchet-Delbos, A. Portier, A. Marfaing-Koka, R. Krzysiek, A.-C. Rimaniol, et al. CX3C Chemokine Fractalkine in Pulmonary Arterial Hypertension Am. J. Respir. Crit. Care Med., May 15, 2002; 165(10): 1419 - 1425. [Abstract] [Full Text] [PDF]
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R. Abdi, T. Thi Bich Huong, A. Sahagun-Ruiz, P. M. Murphy, B. M. Brenner, E. L. Milford, and D. H. McDermott Chemokine Receptor Polymorphism and Risk of Acute Rejection in Human Renal Transplantation J. Am. Soc. Nephrol., March 1, 2002; 13(3): 754 - 758. [Abstract] [Full Text] [PDF]
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A. Ludwig, T. Berkhout, K. Moores, P. Groot, and G. Chapman Fractalkine Is Expressed by Smooth Muscle Cells in Response to IFN-{gamma} and TNF-{alpha} and Is Modulated by Metalloproteinase Activity J. Immunol., January 15, 2002; 168(2): 604 - 612. [Abstract] [Full Text] [PDF]
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K. Red-Horse, P. M. Drake, M. D. Gunn, and S. J. Fisher Chemokine Ligand and Receptor Expression in the Pregnant Uterus : Reciprocal Patterns in Complementary Cell Subsets Suggest Functional Roles Am. J. Pathol., December 1, 2001; 159(6): 2199 - 2213. [Abstract] [Full Text] [PDF]
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K. J. Garton, P. J. Gough, C. P. Blobel, G. Murphy, D. R. Greaves, P. J. Dempsey, and E. W. Raines Tumor Necrosis Factor-alpha -converting Enzyme (ADAM17) Mediates the Cleavage and Shedding of Fractalkine (CX3CL1) J. Biol. Chem., October 5, 2001; 276(41): 37993 - 38001. [Abstract] [Full Text] [PDF]
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C.-L. Tsou, C. A. Haskell, and I. F. Charo Tumor Necrosis Factor-alpha -converting Enzyme Mediates the Inducible Cleavage of Fractalkine J. Biol. Chem., November 21, 2001; 276(48): 44622 - 44626. [Abstract] [Full Text] [PDF]
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), Ab therapy was discontinued. Cardiac allografts
were monitored daily, and the length of survival of each transplant
noted. Results are expressed as the daily percent survival of
allografts.