| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Regular Articles |
From the Center for Experimental Therapeutics and Reperfusion
Injury,*
Department of Anesthesiology, Perioperative and
Pain Medicine, Brigham and Women’s Hospital, Harvard Medical School,
Boston, Massachusetts; the Department of Bacteriology and
Immunology,
University of Helsinki and the
Helsinki University Central Hospital, Helsinki, Finland; Alexion
Pharmaceuticals,
New Haven, Connecticut; the
Department of Clinical Pathology,§
Boston
University School of Medicine, Boston, Massachusetts; and the
Department of Emergency Medicine,¶
Brigham and
Women’s Hospital, Harvard Medical School, Boston, Massachusetts
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The complement system is comprised of
three different cascades: the classical, the alternative, and the
recently described lectin complement pathway (LCP).16,17
The LCP is an antibody-independent cascade initiated by binding of
mannose-binding lectin (MBL) to cell surface
carbohydrates.18
MBL, also known as mannose-binding
protein (MBP), is structurally similar to C1q and may predate C1q
evolutionarily.19
Associated with MBL are two novel
C1r2C1s2-like serine
proteases, MBL-associated serine protease (MASP)-1 and
MASP-2,16
that cleave C2 and C4 to form the classical
pathway C3 convertase.20
Thus, unlike the classical
complement pathway, activation of the LCP does not require antibody or
C1 (Figure 1)
. Although there is
experimental evidence linking the classical and alternative complement
pathways to human disease, the role of the LCP is just beginning to be
evaluated.21
Further, potent, selective inhibitors of the
LCP have not yet been described.
|
, we investigated
whether the LCP could mediate complement activation following
endothelial oxidative stress. Further, we designed specific,
functionally inhibitory monoclonal antibodies (mAbs) against human MBL
to inhibit MBL-dependent complement activation. |
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
HUVECs were isolated and cultured as previously described.2,3 Cells were used during passages 1 through 3.
Isolation and Purification of MBL
MBL and MASPs were purified from fresh-frozen plasma as previously described.23 Western blot analysis confirmed the absence of IgG and/or IgM contamination.
MBL-Deficient Human Serum
MBL-deficient human serum (HS) was produced by treatment with 2 mmol/L phenylmethlysulfonyl fluoride and affinity chromatography using mannan cross-linked to agarose (Sigma Chemical Co., St. Louis, MO). The resultant MBL-deficient HS was dialyzed against Hanks’ balanced salt solution (HBSS) containing Mg2+ and Ca2+.
Immunization and mAb Production
Immunization, fusion, and production of anti-MBL mAbs were performed as previously described with a modification.24 Female Balb/C mice were immunized initially with human or rat MBL (50 µg, i.p.) in TiterMax (Sigma) and then with MBL (25 µg, i.p.) in PBS once a week for 4 weeks. Four days after the last immunization, the splenocytes were aseptically removed and fused with P301.24 The hybridoma supernatant was first screened against human or rat MBL in a solid phase antibody capture enzyme-linked immunosorbent assay (ELISA) consisting of human or rat MBL plated onto 96-well microtiter plates. Positive clones were subcloned to monoclonal status by limiting dilution.
Hybridomas were grown in Dulbecco’s modified Eagle’s medium (DMEM, Irvine Scientific, Santa Ana, CA) containing 10% Hyclone fetal bovine serum, 4 mmol/L L-glutamine, 1% penicillin/streptomycin, and 1% nonessential amino acids. Monoclonal antibodies were isotyped with a commercially available kit (Gibco BRL, Grand Island, NY). Antibodies were purified by protein G affinity chromatography, eluted with 100 mmol/L glycine, pH 3.0, and immediately neutralized in 1 mol/L Tris (1:10; v:v). All antibodies were dialyzed against PBS, concentrated, and sterile filtered.
Western Blot Analysis of MBL
To demonstrate that the anti-MBL mAbs were specific for human or rat MBL, and not MASP-1 or MASP-2, Western blot analysis was performed. Reduced human or rat MBL was resolved by 9% sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The gel was transferred to nitrocellulose and blocked with 10% nonfat dry milk overnight. Each anti-MBL mAb (10 µg/ml) was then incubated with the nitrocellulose in 3% nonfat dry milk for 1 hour at 4°C. The nitrocellulose was then washed and incubated with horseradish peroxidase-conjugated goat anti-mouse polyclonal antibody (1:3000, ICN, Aurora, OH) for 1 hour at 4°C. The nitrocellulose was then washed and developed with the ECL system (Amersham) and X-ray films (Kodak).
Complement Hemolytic Assays
HS was incubated with 0 or 50 µg/ml of the anti-human MBL mAbs for 30 minutes at 37°C. The HS was then diluted serially 1:2 (v:v) in gelatin veronal buffer. Classical complement pathway hemolytic assays (CH50) using sensitized chicken red blood cells were performed as described.7 Hemolytic assay of MBL-deficient HS was also performed to determine the effect of MBL removal on classical complement pathway activity.
MBL-Dependent C3 Deposition on Mannan-Coated Plates
A MBL-dependent C3 deposition ELISA was used as previously described.25 Briefly, mannan (Sigma, 500 µg/ml in 15 mmol/L sodium carbonate, pH 9.6) was added to 96 well microtiter plates for 12 to 16 hours at 4°C and then washed. Human or rat serum (2% final concentration) was incubated with (i) 0.1 to 100 mmol/L GlcNAc; (ii) various doses of mAb; or (iii) vehicle (veronal buffered saline containing Ca2+/Mg2+) for 30 minutes at room temperature. The plates were inoculated with 100 µl of treated or untreated human or rat serum and then incubated for 30 minutes at 37°C. The plates were then washed and 50 µl of horseradish peroxidase-conjugated goat anti-human or rat C3 polyclonal antibody (1:2000; ICN) was used to determine the relative amounts of C3 deposited on the wells. MBL-dependent C3 deposition was determined to be the amount of C3 deposited that could be inhibited by 100 mmol/L N-acetyl-D-glucosamine (GlcNAc). C3 deposition in each well was calculated as a percentage of C3 deposition compared to untreated wells (100%) and wells not receiving HS (0%). IC50 concentrations were calculated for each individual experiment. C3 deposition was evaluated using six wells per experimental group, and each experiment was repeated 4 to 5 times.
C3 Deposition ELISA
C3 deposition was measured on hypoxic/reoxygenated HUVECs as previously described.2,26 We have previously shown that the predominant C3 species present on HUVECs in this model is iC3b.2 Therefore, the data are presented as changes in iC3b deposition. HUVECs were grown to confluence and then subjected to 0 (normoxia) or 24 hours of hypoxia (1% O2). The cell media were aspirated and 100 µl of one of the following was added to each well: 1) 30% HS, 2) HBSS, 3) 30% HS + 3, 30, or 300 mmol/L GlcNAc, 4) 30% HS + 3, 30, or 300 mmol/L D-mannose, 5) 30% HS + 3, 30, 300 mmol/L L-mannose, 6) 30% MBL-deficient HS, or 7) 30% MBL-deficient HS + 0.6 µg/ml MBL. The cells were then reoxygenated for 3 hours, washed, and then fixed with 1% paraformaldehyde (Sigma) for 30 minutes. The cells were then washed and incubated with peroxidase-conjugated polyclonal goat anti-human C3 antibody (Cappel, West Chester, PA) for 1 hour at 4°C. The plates were washed and developed with ABTS, and the absorbances (405 nm) were measured using a microtiter plate reader (Molecular Devices, Sunnyvale, CA). Background optical density for the C3 deposition ELISA was obtained from cells to which only the anti-human C3 antibody was added or cells incubated with 30% heat-inactivated HS, and was subtracted from all groups. All ELISA experiments were performed 3 times using 6 wells per experimental group (n = 3).
Confocal Microscopy
HUVECs were grown on LabTech tissue culture microscope slides (NUNC) and exposed to hypoxia for 24 hours as described above. The slides were removed from the hypoxia chamber and reoxygenated for 3 hours in 30% HS treated with PBS (ie, vehicle), 3F8 (5 µg/ml), 2A9 (5 µg/ml), or 1C10 (50 µg/ml). The slides were then washed in PBS containing calcium and magnesium and fixed in 4% paraformaldehyde for 15 minutes, washed again, and blocked with 10% goat serum to prevent nonspecific secondary antibody staining. Fixation is necessary to avoid the loss of MBL from the cell surface from the multiple washing steps because, unlike covalently attached iC3b, MBL attachment is calcium-dependent and not covalent. Human MBL deposition was identified using biotinylated 1C10 and streptavidin-conjugated CY5 (blue; Jackson Immunoresearch, West Grove, PA). Human C3 deposition (green) was evaluated with a FITC-conjugated goat anti-human C3 F(ab')2 antibody (ICN). Following incubation with the appropriate antibodies, the slides were washed 3x 10 minutes, coated with anti-fade mounting media (Molecular Probes, Eugene, OR), covered and analyzed with a Zeiss confocal microscope as described.26 Controls with streptavidin-conjugated CY5 only were processed as above, omitting the primary antibody to determine nonspecific binding. All analyses were conducted at 40x magnification at the same pinhole, voltage, and laser settings. This experiment was performed three times (n = 3).
Animal Preparation and Protocols
The study protocol was approved by the ethical committee of the Meilahti Theoretical Institutes (Helsinki, Finland) and by the local health authority. Adult male Lewis rats (220–260 g) were anesthetized with sodium pentobarbital (50 mg/kg i.p.) and xylazine (6 mg/kg, i.m.). Rats were tracheostomized and ventilated with a SAR-380 small animal respirator (CWE Inc., PA). Expired CO2 was monitored continuously with a microcapnometer (CWE Inc.) and maintained at 4 to 5% by adjusting the respiratory rate and/or tidal volume. Myocardial ischemia and reperfusion was produced as described previously.27 Briefly, a left thoracotomy was performed and the left anterior descending coronary artery (LAD) was ligated 3 to 4 mm from its point of origin with 6–0 silk. Ischemia was produced by tightening the previously placed reversible ligature to occlude completely the vessel. Sham-operated animals underwent the same surgical procedures, but without ligation of the LAD suture. Animals were randomly divided into the following three groups: (i) 30 minutes ischemia followed by 30 minutes reperfusion (n = 4); (ii) 60 minutes ischemia, but no reperfusion (n = 4); and (iii) sham controls (n = 4). The rats were sacrificed by carbon dioxide inhalation and decapitation. Ischemia was confirmed in all rats by the presence of ventricular ectopy, discoloration of the ischemic area, and left ventricular (LV) dyskinesia. Ventricular ectopy and the return of color to the ischemic area confirmed reperfusion. The great vessels, atria, and right ventricle were removed at the end of the experimental protocol. The LV cavity was embedded in Tissue-Tek embedding medium (Ames Corp., Elkhart, IN), frozen with dry ice, and stored at -80°C.
Rat Myocardial Immunohistochemistry
Frozen sections (5 µm) were fixed for 5 minutes in cold (-20°C) acetone and then stained with a primary fluorescein isothiocyanate (FITC)-conjugated rabbit anti-rat C3 polyclonal antibody (pAb; Cappel) or a primary mouse anti-rat MBL mAb followed by a secondary FITC-conjugated goat anti-mouse pAb (Jackson Immunoresearch). Control incubations were done either by omitting the primary antibody or by using FITC-conjugated rabbit anti-horse IgG. The slides were mounted with Mowiol28 and examined with an Olympus Standard microscope equipped with a filter specific for FITC fluorescence.
Statistical Analysis
All data presented represent the mean and SE for n
determinations. Data analyses were performed using Sigma Stat (Jandel
Scientific, San Rafael, CA) and a P value of <0.05 was
considered significant. Endothelial C3 deposition on normoxic
versus hypoxic HUVECs was analyzed by two-way analysis of
variance. All pairwise multiple comparisons were made using the
Student-Newman-Keuls test. C3 deposition on treated cells (ELISA; Figures 2 and 3
) after hypoxia/reoxygenation was
normalized to hypoxic HUVECs reoxygenated in untreated 30% HS.
Means ± SE of the raw data used for normalization are presented
in Results and/or figure legends.
|
|
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Consistent with previous studies,2,3
a significant
increase in iC3b deposition was observed after oxidative stress
compared to normoxic HUVECs (OD405 = 0.14 ±
0.01 vs. 0.05 ± 0.01, respectively; P
< 0.05). The MBL inhibitory sugar, D-mannose, attenuated iC3b
deposition in a dose-dependent manner (Figure 2)
. Similarly, GlcNAc (30
mmol/L) inhibited iC3b deposition by 81 ± 2%. L-mannose (300
mmol/L) did not significantly inhibit iC3b deposition. These data
demonstrate that GlcNAc or D-mannose significantly attenuates
complement activation and iC3b deposition in a dose-dependent,
stereospecific manner, suggesting a possible role for MBL.
Deposition of iC3b after MBL Depletion and Reconstitution
MBL-deficient serum was produced to further evaluate the role of
MBL in this model. iC3b deposition (Figure 3)
on hypoxic HUVECs
reoxygenated in MBL-deficient HS was approximately 25% that in vehicle
control. Reconstitution of MBL-deficient HS with purified MBL restored
iC3b deposition. A complement hemolytic assay
(CH50) demonstrated that the classical complement
pathway activity of the MBL-deficient HS did not significantly differ
from that of complete HS (data not shown). Thus, these data suggest
that the C2-dependent LCP is primarily responsible for complement
activation and deposition after endothelial oxidative stress.
Anti-MBL mAb Production and Characterization
To demonstrate that MBL was responsible for complement activation
in this study, we developed novel, functionally inhibitory mAbs against
human MBL. Eleven parent hybridoma clones that recognized human MBL
were identified in a solid phase antibody-capture ELISA. After limiting
dilution and isotyping, eight mAbs that recognized human MBL in the
antibody-capture ELISA were identified. Clones 3F8, 2A9, and hMBL1.2
were isotyped as mouse IgG1
, whereas clone
1C10 was a mouse IgG2b. The other hybridomas
produced IgM antibodies and were not included in this study.
To demonstrate that the anti-human MBL mAbs recognized human MBL,
Western blot analysis of reduced and nonreduced MBL was performed.
Monoclonal antibodies 2A9, hMBL1.2, 1C10, and 3F8 recognized nonreduced
human MBL (MW ~600 kd; data not shown). As shown in Figure 4
, mAbs 2A9 (Lane 1), hMBL1.2 (Lane 2),
1C10 (Lane 3), and 3F8 (Lane 4) recognized reduced human MBL (MW ~32
kd). In particular, mAbs 2A9 and hMBL1.2 easily recognized reduced and
denatured human MBL. Although mAbs 1C10 and 3F8 also recognized reduced
and denatured human MBL, longer exposure times were necessary,
resulting in higher backgrounds. Addition of purified human MBL to each
of the mAbs before Western blot analysis inhibited detection of
immobilized MBL on the membranes. Further, immunoprecipitation of human
MBL with each of these mAbs yielded a 32-kd protein under reducing
conditions that was recognized by anti-human MBL pAbs (data not shown).
Thus, these antibodies are specific for human MBL and do
not recognize MASP-1 or MASP-2. In particular, clones 2A9 and hMBL1.2
are good mAbs for Western blot analysis of reduced human MBL. Clones
hMBL1.2, 2A9, and 3F8 have been deposited at the International
Depository Authority with American Type Culture Collection
designations of HB-12619, HB-12620, and HB-12621, respectively.
|
Selective activation of the LCP using mannan-coated plates and
human serum was performed as described25
to characterize
whether the anti-human MBL mAbs functionally inhibited MBL-dependent
complement activation. Monoclonal antibodies 3F8, 2A9, or hMBL1.2
inhibited C3 deposition on the mannose-coated plates in a
dose-dependent manner (Figure 5)
. Clone
1C10 failed to inhibit MBL-dependent C3 deposition in this assay. The
IC50 for 3F8, 2A9, hMBL1.2 and GlcNAc were
0.9 ± 0.3 nmol/L, 13.3 ± 3.3 nmol/L, 18.9 ± 7.8
nmol/L, and 15 ± 6 mmol/L, respectively. Thus, the functionally
inhibitory anti-human MBL mAbs (eg, 3F8, 2A9, and hMBL1.2) were about 6
orders of magnitude more effective than GlcNAc in this assay.
|
Complement Hemolysis Assays
Hemolysis assays were performed to demonstrate that the anti-human
MBL mAbs did not inhibit or activate (ie, immune complex formation) the
classical complement pathway. As demonstrated in Figure 6
, mAb 3F8 (eg, the best MBL inhibitory
antibody in the mannan assay in concentrations up to 50 µg/ml) did
not attenuate classical complement pathway activation (ie, hemolytic
activity of human serum). Similar findings were observed for mAbs
hMBL1.2, 1C10, and 2A9 (data not shown). Thus, the mechanism of LCP
inhibition is neither through activation (ie, immune complex formation)
or inhibition of the classical complement pathway.
|
Dual labeling for MBL (blue) and C3 (green) deposition on HUVECs
was performed to demonstrate MBL and C3 colocalization after oxidative
stress (Figure 7)
. Normoxic and hypoxic
HUVECs were reoxygenated in 30% HS treated with and without the
anti-human MBL mAbs 3F8 (5 µg/ml), 2A9 (5 µg/ml), or 1C10 (50
µg/ml). Small amounts of C3 (Figure 7A
1) and MBL (Figure 7A
2)
staining were observed under normoxic conditions, confirming our
previous finding of low level C3 deposition under normoxic
conditions.2,3
C3 (Figure 7B
1) and MBL (Figure 7B
2)
staining on hypoxic/reoxygenated HUVECs was increased compared to
normoxic cells. Clone 1C10 failed to inhibit C3 (Figure 7C
1) or MBL
(Figure 7C
2) deposition after oxidative stress, confirming its
inability to inhibit MBL. C3 (Figure 7
, D1 and E1) and MBL (Figure 7
,
D2 and E2) staining was attenuated on hypoxic/reoxygenated HUVECs
treated with mAbs 3F8 or 2A9, respectively (similar results were
observed with mAb hMBL1.2). Row 3 demonstrates that MBL and C3
colocalize on human endothelial cells under the conditions outlined
above. These data demonstrate that inhibition of MBL deposition with a
functionally inhibitory mAb decreases C3 deposition after endothelial
oxidative stress.
|
A previous study has demonstrated that complement activation and
deposition during the early phase of myocardial ischemia/reperfusion is
limited to the coronary vascular endothelium.1
We
investigated MBL (Figure 8
, C and D) and
C3 (Figure 8
, A and B) deposition in the ischemic/reperfused rat
myocardium by indirect immunofluorescence microscopy. Strong C3 and MBL
(clone 14C3.74) staining were observed throughout the ischemic area at
risk (Figure 8
, B and D, upper part) in rat hearts
(n = 4) subjected to 30 minutes ischemia
followed by 30 minutes reperfusion (Figure 8
, B and D, respectively).
In contrast, minimal C3 or MBL deposition was observed in rat
hearts (n = 4) subjected to 60 minutes of
ischemia without reperfusion (Figure 8
, A and C, respectively). No
staining for C3 or MBL was observed in the sham-operated rat hearts
(data not shown). These data are the first to demonstrate MBL
deposition during the early phase of myocardial ischemia/reperfusion.
Further, reperfusion appears to augment MBL deposition and complement
activation. Thus, these in vivo data are consistent with our
in vitro findings using isolated human endothelial cells.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Anoxic human endothelial cells were recently shown to activate
complement in the absence of immunoglobulin-binding, suggesting
antibody-independent complement activation.22
Additionally, we have shown that complement activation after
endothelial oxidative stress is C2-dependent.2
Because the
lectin or classical complement pathway is C2-dependent (Figure 1)
, we
investigated whether the antibody-independent LCP mediates complement
activation after oxidative stress. In the present study, inhibition of
MBL deposition with D-mannose, GlcNAc, or MBL-deficient serum
significantly attenuated iC3b deposition after endothelial oxidative
stress. Further, immunofluorescent confocal microscopy demonstrated
increased deposition and colocalization of MBL and C3 on
hypoxic/reoxygenated human endothelial cells. Inhibition of MBL
deposition with the anti-human MBL mAbs significantly attenuated C3
deposition. Together, these in vitro data suggest that MBL
initiates complement activation following endothelial oxidative stress.
Further, these data suggest that oxidative stress leads to the
formation of a complement-activating MBL ligand on human endothelial
cells. We are currently identifying and characterizing this
complement-activating MBL ligand.
Experimental in vivo data strongly support the role of the reperfused endothelium in activating complement after ischemia/reperfusion.1,11 It has been hypothesized that novel endothelial epitopes are up-regulated and that natural antibodies then activate the classical complement pathway.11 Indeed, complement activation after reperfusion of ischemic myocardium is initially limited to the coronary endothelium.1 In the present study, strong C3 and MBL staining were observed throughout the ischemic area at risk during the early phase of rat myocardial reperfusion. Furthermore, myocardial reperfusion augmented MBL deposition and complement activation in vivo compared to sham-operated or ischemia-only animals. This study is the first to demonstrate MBL deposition during the early phase of myocardial ischemia/reperfusion. This is an important observation, insofar as several previous studies using C4-deficient or C1 esterase inhibitor-treated animals have suggested that the classical complement pathway mediates complement activation after ischemia/reperfusion.6,11,29 However, the possible involvement of the LCP in these studies cannot be ruled out, as C1 esterase inhibitor also attenuates MASP-1 and MASP-2 activity, and LCP activation is inhibited in C4-deficient animals.30 Thus, the specific roles of the lectin and/or classical complement pathways in ischemia/reperfusion injury have not been fully elucidated. Generation of species-specific MBL inhibitors/antibodies or MBL/MASP knockout mice will aid in the elucidation of the role of the lectin versus classical complement pathways in animal models. To date, we have not been able to generate a functionally inhibitory antibody to rat MBL. This difficulty may be due to the presence of two rat MBL isoforms.31
Inhibition of MBL and the LCP can be accomplished by using specific carbohydrates including mannose, N-acetylglucosamine (GlcNAc), or oligomers of these sugars. Although these sugars are somewhat specific in attenuating MBL deposition/binding,32,33 their efficacy and half-life are relatively poor.34 Further, intravenous treatment with carbohydrate oligomers (ie, mannan) may directly activate complement in vivo, creating a complement-deprived animal. We thus designed specific, functionally inhibitory mAbs against human MBL to increase the efficacy and specificity of MBL inhibition. Using an MBL-dependent C3 deposition ELISA,25 these mAbs (eg, 3F8, 2A9, and hMBL1.2) were about 6 orders of magnitude more effective than GlcNAc in this assay. Furthermore, these antibodies were very potent inhibitors of MBL deposition after endothelial oxidative stress. Thus, anti-human MBL mAb treatment may represent a novel, specific therapeutic strategy for LCP-mediated tissue injury.
In summary, MBL and iC3b deposition are increased following human endothelial oxidative stress in vitro, rat MBL and C3 deposition is increased following myocardial reperfusion in vivo, and inhibition of human MBL deposition with GlcNAc, D-mannose, MBL-deficient serum, or anti-MBL mAbs attenuates complement activation after endothelial oxidative stress in vitro. These data suggest that MBL deposition on the vascular endothelium after oxidative stress may lead to LCP activation. Future studies with specific inhibitors of MBL in animal models will help define the role of MBL and the LCP in cardiovascular disease.
|
Footnotes |
|---|
Supported by HL-03854 (to C. D. C.), the Foundation for Anesthesia Education and Research (to C. D. C.), the Academy of Finland, the Sigrid Juselius Foundation, the State Subsidy to the Helsinki University Central Hospital, the Finnish Foundation for Cardiovascular Research (to A.V and S.P.), HL-52886 (to G. L. S.), and an American Heart Association Established Investigator Award (to G. L. S.).
Accepted for publication January 24, 2000.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
B and new protein synthesis. Immunopharmacology 1998, 39:39-50[Medline]
B translocation and vascular cell adhesion molecule-1 induction by complement: Inhibition with anti-C5 therapy or cGMP analogues. Arterioscler Thromb Vasc Biol 1999, 19:2623-2629This article has been cited by other articles:
 |
|
 |
|
  A. R. Kinderlerer, F. Ali, M. Johns, E. A. Lidington, V. Leung, J. J. Boyle, S. S. Hamdulay, P. C. Evans, D. O. Haskard, and J. C. Mason KLF2-dependent, Shear Stress-induced Expression of CD59: A NOVEL CYTOPROTECTIVE MECHANISM AGAINST COMPLEMENT-MEDIATED INJURY IN THE VASCULATURE J. Biol. Chem., May 23, 2008; 283(21): 14636 - 14644. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. R. La Bonte, G. Davis-Gorman, G. L. Stahl, and P. F. McDonagh Complement inhibition reduces injury in the type 2 diabetic heart following ischemia and reperfusion Am J Physiol Heart Circ Physiol, March 1, 2008; 294(3): H1282 - H1290. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Roos, M. R. Daha, J. van Pelt, and S. P. Berger Mannose-binding lectin and the kidney Nephrol. Dial. Transplant., December 1, 2007; 22(12): 3370 - 3377. [Full Text] [PDF]
|
|
|
|
 |
|
 G. Szeplaki, L. Varga, J. Laki, E. Dosa, S. Rugonfalvi-Kiss, H. O. Madsen, Z. Prohaszka, A. Kocsis, P. Gal, A. Szabo, et al. Low C1-Inhibitor Levels Predict Early Restenosis After Eversion Carotid Endarterectomy Arterioscler. Thromb. Vasc. Biol., December 1, 2007; 27(12): 2756 - 2762. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Banz, O. M. Hess, S. C. Robson, E. Csizmadia, D. Mettler, P. Meier, A. Haeberli, S. Shaw, R. A. Smith, and R. Rieben Attenuation of myocardial reperfusion injury in pigs by Mirococept, a membrane-targeted complement inhibitor derived from human CR1 Cardiovasc Res, December 1, 2007; 76(3): 482 - 493. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. D. Collard, S. K. Shernan, A. A. Fox, T. Bernig, S. J. Chanock, W. K. Vaughn, K. Takahashi, A. B. Ezekowitz, P. Jarolim, and S. C. Body The MBL2 'LYQA Secretor' Haplotype Is an Independent Predictor of Postoperative Myocardial Infarction in Whites Undergoing Coronary Artery Bypass Graft Surgery Circulation, September 11, 2007; 116(11_suppl): I-106 - I-112. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Szabo, J. Laki, H. O. Madsen, E. Dosa, Z. Prohaszka, S. Rugonfalvi-Kiss, M. Kokai, G. Acsadi, I. Karadi, L. Entz, et al. Early Rise in Serum VEGF and PDGF Levels Predisposes Patients With a Normal MBL2 Genotype to Restenosis After Eversion Endarterectomy Stroke, August 1, 2007; 38(8): 2247 - 2253. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. K. Hansen, M.-A. Gall, L. Tarnow, S. Thiel, C. D. Stehouwer, C. G. Schalkwijk, H.-H. Parving, and A. Flyvbjerg Mannose-binding lectin and mortality in type 2 diabetes. Arch Intern Med, October 9, 2006; 166(18): 2007 - 2013. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. T.V.M. de Jong Age-Related Macular Degeneration N. Engl. J. Med., October 5, 2006; 355(14): 1474 - 1485. [Full Text] [PDF]
|
|
|
|
 |
|
 M. Zhang, K. Takahashi, E. M. Alicot, T. Vorup-Jensen, B. Kessler, S. Thiel, J. C. Jensenius, R. A. B. Ezekowitz, F. D. Moore, and M. C. Carroll Activation of the Lectin Pathway by Natural IgM in a Model of Ischemia/Reperfusion Injury J. Immunol., October 1, 2006; 177(7): 4727 - 4734. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. T. Keller, S. I. van Leuven, M. C. Meuwese, N. J. Wareham, R. Luben, E. S. Stroes, C. E. Hack, M. Levi, K.-T. Khaw, and S. M. Boekholdt Serum Levels of Mannose-Binding Lectin and the Risk of Future Coronary Artery Disease in Apparently Healthy Men and Women Arterioscler. Thromb. Vasc. Biol., October 1, 2006; 26(10): 2345 - 2350. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Geleijns, A. Roos, J. J. Houwing-Duistermaat, W. van Rijs, A. P. Tio-Gillen, J. D. Laman, P. A. van Doorn, and B. C. Jacobs Mannose-Binding Lectin Contributes to the Severity of Guillain-Barre Syndrome J. Immunol., September 15, 2006; 177(6): 4211 - 4217. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Roos, M. P. Rastaldi, N. Calvaresi, B. D. Oortwijn, N. Schlagwein, D. J. van Gijlswijk-Janssen, G. L. Stahl, M. Matsushita, T. Fujita, C. van Kooten, et al. Glomerular Activation of the Lectin Pathway of Complement in IgA Nephropathy Is Associated with More Severe Renal Disease J. Am. Soc. Nephrol., June 1, 2006; 17(6): 1724 - 1734. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. M. Quaniers, J. Leruth, A. Albert, R. R. Limet, and J.-O. Defraigne Comparison of inflammatory responses after off-pump and on-pump coronary surgery using surface modifying additives circuit. Ann. Thorac. Surg., May 1, 2006; 81(5): 1683 - 1690. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. L. D. Riis, T. K. Hansen, S. Thiel, C. H. Gravholt, S. Gjedde, L. C. Gormsen, J. O. L. Jorgensen, J. Weeke, and N. Moller Thyroid hormone increases mannan-binding lectin levels Eur. J. Endocrinol., November 1, 2005; 153(5): 643 - 649. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. C. Walsh, T. Bourcier, K. Takahashi, L. Shi, M. N. Busche, R. P. Rother, S. D. Solomon, R. A. B. Ezekowitz, and G. L. Stahl Mannose-Binding Lectin Is a Regulator of Inflammation That Accompanies Myocardial Ischemia and Reperfusion Injury J. Immunol., July 1, 2005; 175(1): 541 - 546. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. L. Hart, K. A. Ceonzo, L. A. Shaffer, K. Takahashi, R. P. Rother, W. R. Reenstra, J. A. Buras, and G. L. Stahl Gastrointestinal Ischemia-Reperfusion Injury Is Lectin Complement Pathway Dependent without Involving C1q J. Immunol., May 15, 2005; 174(10): 6373 - 6380. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Hovind, T. K. Hansen, L. Tarnow, S. Thiel, R. Steffensen, A. Flyvbjerg, and H.-H. Parving Mannose-Binding Lectin as a Predictor of Microalbuminuria in Type 1 Diabetes: An Inception Cohort Study Diabetes, May 1, 2005; 54(5): 1523 - 1527. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Rugonfalvi-Kiss, E. Dosa, H. O. Madsen, V. Endresz, Z. Prohaszka, J. Laki, I. Karadi, E. Gonczol, L. Selmeci, L. Romics, et al. High Rate of Early Restenosis After Carotid Eversion Endarterectomy in Homozygous Carriers of the Normal Mannose-Binding Lectin Genotype Stroke, May 1, 2005; 36(5): 944 - 948. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. de Vries, S. J. Walter, C. J. Peutz-Kootstra, T. G.A.M. Wolfs, L.W. E. van Heurn, and W. A. Buurman The Mannose-Binding Lectin-Pathway Is Involved in Complement Activation in the Course of Renal Ischemia-Reperfusion Injury Am. J. Pathol., November 1, 2004; 165(5): 1677 - 1688. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Windbichler, B. Echtenacher, T. Hehlgans, J. C. Jensenius, W. Schwaeble, and D. N. Mannel Involvement of the Lectin Pathway of Complement Activation in Antimicrobial Immune Defense during Experimental Septic Peritonitis Infect. Immun., September 1, 2004; 72(9): 5247 - 5252. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Ohlenschlaeger, P. Garred, H. O. Madsen, and S. Jacobsen Mannose-Binding Lectin Variant Alleles an |
