Advertisement
JBC

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Originally published In Press as doi:10.1074/jbc.M407057200 on August 24, 2004

J. Biol. Chem., Vol. 279, Issue 44, 46129-46134, October 29, 2004
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
279/44/46129    most recent
M407057200v1
Right arrow Submit a Letter to Editor
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowRequest Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Beller, T. C.
Right arrow Articles by Kanaoka, Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Beller, T. C.
Right arrow Articles by Kanaoka, Y.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

Targeted Gene Disruption Reveals the Role of the Cysteinyl Leukotriene 2 Receptor in Increased Vascular Permeability and in Bleomycin-induced Pulmonary Fibrosis in Mice*

Thomas C. Beller{ddagger}§, Akiko Maekawa{ddagger}§, Daniel S. Friend||**, K. Frank Austen{ddagger}§, and Yoshihide Kanaoka{ddagger}§{ddagger}{ddagger}

From the Departments of {ddagger}Medicine and ||Pathology, Harvard Medical School, and the §Division of Rheumatology, Immunology, and Allergy and the **Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts 02115

Received for publication, June 23, 2004 , and in revised form, July 26, 2004.


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 EXPERIMENTAL PROCEDURES
 RESULTS
 DISCUSSION
 REFERENCES
 
The cysteinyl leukotrienes (cys-LTs) mediate both acute and chronic inflammatory responses in mice, as demonstrated by the attenuation of the IgE/antigen-mediated increase in microvascular permeability and of bleomycin-induced pulmonary fibrosis, respectively, in a strain with targeted disruption of leukotriene C4 synthase to prevent cys-LT synthesis. Our earlier finding that the acute, but not the chronic, injury was attenuated in a strain with targeted disruption of the cysteinyl leukotriene 1 (CysLT1) receptor suggested that the chronic injury might be mediated through the CysLT2 receptor. Thus, we generated CysLT2 receptor-deficient mice by targeted gene disruption. These mice developed normally and were fertile. The increased vascular permeability associated with IgE-dependent passive cutaneous anaphylaxis was significantly reduced in CysLT2 receptor-null mice as compared with wild-type mice, whereas plasma protein extravasation in response to zymosan A-induced peritoneal inflammation was not altered. Alveolar septal thickening after intratracheal injection of bleomycin, characterized by interstitial infiltration with macrophages and fibroblasts and the accumulation of collagen fibers, was significantly reduced in CysLT2 receptor-null mice as compared with the wild-type mice. The amounts of cys-LTs in bronchoalveolar lavage fluid after bleomycin injection were similar in the CysLT2 receptor-null mice and the wild-type mice. Thus, in response to a particular pathobiologic event the CysLT2 receptor can mediate an increase in vascular permeability in some tissues or promote chronic pulmonary inflammation with fibrosis.


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
EXPERIMENTAL PROCEDURES
 RESULTS
 DISCUSSION
 REFERENCES
 
The cysteinyl leukotrienes (cys-LTs),1 leukotriene (LT) C4, LTD4, and LTE4, are lipid mediators (1, 2) that have been implicated in the pathobiology of allergic and asthmatic inflammatory responses on the basis of the efficacy of specific intervention with a biosynthetic inhibitor (3) or antagonists to a receptor subsequently shown to be the cysteinyl leukotriene 1 (CysLT1) receptor (4). Two types of human receptors for cys-LTs, designated CysLT1 receptor and CysLT2 receptor, belonging to the seven-transmembrane, G protein-coupled receptor family, were cloned and shown to be 38% homologous in their amino acid sequences (5–9). The rank order of affinities of the cys-LTs for the CysLT1 and CysLT2 receptors is LTD4 > LTC4 > LTE4 >> LTB4 and LTC4 = LTD4 >> LTE4 >> LTB4, respectively. The CysLT1 receptor is expressed on airway smooth muscle, monocytes/macrophages, eosinophils (5, 10, 11), a subset of neutrophils (11), mast cells (12, 13), and endothelial cells (14). The CysLT2 receptor is expressed on monocyte/macrophages, airway smooth muscle, cardiac Purkinje cells, adrenal medulla cells, peripheral blood leukocytes, brain cells (7), eosinophils (11, 15), mast cells (16), and endothelial cells (17, 18). We and others have reported that the mouse CysLT1 receptor can function as a receptor for LTD4 in transfected cells with a ligand preference similar to that of the human CysLT1 receptor (19–22). The mouse CysLT2 receptor exhibits a ligand profile of LTC4 ≥ LTD4 >> LTE4 (22, 23). In contrast to the extensive studies on the functions of the CysLT1 receptor (24, 25), the functional characterization of the CysLT2 receptor has been limited because of a lack of specific competitive antagonists. In particular, any in vivo role for the CysLT2 receptor remains unknown.

LTC4 synthase is the pivotal enzyme for the generation of LTC4, the parent compound of the cys-LTs (26–28). LTC4 synthase conjugates reduced glutathione to an epoxide intermediate, LTA4, generated from arachidonic acid by 5-lipoxygenase in the presence of the 5-lipoxygenase-activating protein (29). After carrier-mediated export (30), glutamic acid and glycine are sequentially cleaved from the glutathione moiety of LTC4 by {gamma}-glutamyl transpeptidase (31) or {gamma}-glutamyl leukotrienase (32) and dipeptidase (33) to form LTD4 and LTE4, respectively. We have shown that the increased vascular permeability in zymosan A-induced, monocyte/macrophage-mediated peritoneal inflammation and in IgE-dependent, mast cell-mediated passive cutaneous anaphylaxis (PCA) is reduced by ~50% in LTC4 synthase-null mice as compared with their wild-type littermates (34). We then demonstrated by targeted disruption of the CysLT1 receptor in mice that this alteration of vascular permeability is dominantly mediated through the CysLT1 receptor (35). Recently, we found that bleomycin-induced chronic pulmonary inflammation and fibrosis are diminished in LTC4 synthase-null mice, thereby implicating the cys-LTs in this pathobiology (36). However, CysLT1 receptor-null mice were not protected, and this finding suggested a proinflammatory role for the CysLT2 receptor in the chronic response. To define the role of the CysLT2 receptor in vivo, we generated CysLT2 receptor-null mice by targeted gene disruption. We then examined plasma protein extravasation and neutrophil infiltration in zymosan A-induced peritoneal inflammation, plasma protein extravasation in IgE-dependent PCA, and chronic pulmonary inflammation and fibrosis induced by intratracheal injection of bleomycin.


   EXPERIMENTAL PROCEDURES
TOP
 ABSTRACT
 INTRODUCTION
 EXPERIMENTAL PROCEDURES
RESULTS
 DISCUSSION
 REFERENCES
 
Generation of CysLT2 Receptor Gene-disrupted Mice—Based on the nucleotide sequences of the mouse CysLT2 receptor cDNA and genomic DNA (22, 23), a 5-kb mouse genomic DNA fragment containing exon III to 35 base pairs (bp) of exon VI was amplified by PCR with C57BL/6 mouse genomic DNA using a sense primer of 5'-GACACTCAGCAACGGAGGAATTTT-3' and an antisense primer of 5'-GACCTTGGTAGACATCAGGCATAC-3'. The amplified fragment was subcloned into a pBluescriptII vector (Stratagene). A 1139-bp fragment containing part of exon VI and the 3'-flanking region was amplified by PCR with a sense primer of 5'-GGGGATGTGCTACATAAGGCC-3' and an antisense primer of 5'-GGTAGTATCTGTAGTCAGTTTCTG-3'. A neo gene from a pMC1Neo plasmid (Stratagene) and then the 1139-bp fragment were inserted tandemly into the 3'-side of the 5-kb fragment so that the neo gene would disrupt the region encoding the N-terminal 264 amino acids of the mouse CysLT2 receptor. Finally, the herpes simplex virus thymidine kinase (TK) gene was inserted into the 5'-side of the 5-kb fragment (Fig. 1A). The resultant targeting vector was linearized and electroporated into a C57BL/6 mouse embryonic stem cell line (Specialty Media, Phillipsburg, NJ). The embryonic stem cells were selected with G418 (200 µg/ml; Invitrogen) and ganciclovir (2 µM; Sigma), and homologous recombination was confirmed by Southern blot analysis of EcoRI-digested genomic DNA from each embryonic stem cell clone with a 1250-bp 3'-fragment as a probe located outside the targeting vector (Fig. 1A). The embryonic stem cell clones, verified by Southern blot analysis, were microinjected into blastocysts from BALB/c mice; and chimeric males were obtained. Chimeras were bred to C57BL/6 females, and offspring were genotyped by Southern blot analysis of the tail DNA. Heterozygotes were intercrossed to obtain CysLT2 receptor(-/-) mice. All animal studies were approved by the Animal Care and Use Committee of the Dana-Farber Cancer Institute.



View larger version (37K):
[in this window]
[in a new window]
  		FIG. 1.
Generation of CysLT2 receptor gene-disrupted mice. A, genomic organization of the mouse CysLT2 receptor gene (upper), structure of the targeting vector (middle), and organization of the putative recombinant CysLT2 receptor allele (lower). Exons III–VI are shown as boxes with the coding regions in black. The location of the 1250-bp fragment used for Southern blot analysis is shown as a thick line. TK, herpes simplex virus thymidine kinase. Restriction site E, EcoRI. B, Southern blot analysis of EcoRI-digested tail DNAs from the pups of a CysLT2 receptor (+/-) male and a CysLT2 receptor (+/-) female mouse. C, Northern blot analysis of poly(A)+ RNA from the lung, spleen, and small intestine of CysLT2 receptor-null mice (-/-) and their wild-type littermates (+/+). Hybridizations were performed with a 32P-labeled mouse CysLT2 receptor cDNA probe (upper), with a 32P-labeled mouse CysLT1 receptor cDNA probe (middle), and with a 32P-labeled mouse glyceraldehyde-3-phosphate dehydrogenase (GAPDH) cDNA probe (lower).

 
Northern Blot Analysis—Total RNA from the lung and spleen tissue of CysLT2 receptor-null and wild-type mice was isolated with Tri-Reagent (Sigma) according to the manufacturer's protocol. Poly(A)+ RNA was then purified with Oligotex(dT) chromatography (Qiagen). A 4-µg sample of the poly(A)+ RNA was resolved by electrophoresis on a formaldehyde-denatured gel and transferred to a nylon membrane (Biodyne B, 0.45 µm; Pall Corp., Ann Arbor, MI) with 20x SSC for 24 h. The membrane was hybridized with 32P-labeled mouse CysLT2 receptor, CysLT1 receptor, or glyceraldehyde-3-phosphate dehydrogenase coding region at 42 °C for 18 h; washed with 2x SSC, 0.1% SDS at room temperature, and then with 0.2x SSC, 0.1% SDS at 60 °C; and exposed at –80 °C to a Kodak X-AR film with an intensifying screen for 7 days for the CysLT2 receptor and CysLT1 receptor, and for 1 day for glyceraldehyde-3-phosphate dehydrogenase.

Zymosan A-induced Peritoneal Inflammation—Each mouse received an intravenous injection of 0.5% Evans blue dye (10 ml of dye solution/kg of body weight) in phosphate-buffered saline (PBS) immediately before the intraperitoneal injection of 1 ml of zymosan A suspension (1 mg/ml in PBS; Sigma). Mice were killed by CO2 30, 60, 120, and 240 min after the injection of zymosan A, and the peritoneal cavity of each mouse was lavaged with 4 ml of cold PBS. Cells were sedimented from the lavage fluid by centrifugation at 500 x g for 5 min, and Evans blue dye extravasation was assessed by light spectrophotometry of the supernatants at 610 nm. The cell pellets were suspended in 100 µl of PBS, and the cells were counted. Cells (2 x 104) were cytospun onto a glass slide, stained with Diff-Quik (Dade Behring, Deerfield, IL), and analyzed for cell types. The total neutrophils in the lavage fluid were calculated from the percentage of the cell type and the total cell count.

PCA—Mice received intradermal injections of 25 ng of mouse monoclonal anti-dinitrophenyl IgE (Sigma) in 25 µl of saline in the right ear and 25 µl of saline only in the left ear. After 20 h, mice were injected intravenously with 100 µg of dinitrophenyl-human serum albumin (Sigma) and 1% Evans blue dye in 100 µl of PBS. The mice were killed by CO2 30 min after the intravenous injection, and a 6-mm diameter disk of tissue was obtained from the center of each ear. Each disk was incubated in 200 µl of formamide at 55 °C for 48 h. Extravasation of Evans blue dye was quantitated by spectrophotometric analysis of the formamide extracts at 610 nm, and vascular permeability enhancement was calculated as the net difference between the sensitized and control ears.

Intratracheal Injection of Bleomycin into Mice—Bleomycin sulfate (2.5 units/kg, Sigma) dissolved in saline (2.5 µl/g) or saline alone was administered intratracheally to mice that had been anesthetized with an intraperitoneal injection of pentobarbital (75 mg/kg).

Histologic Assessment of Lung Tissue—Twelve days after bleomycin injection, the lower lobes of the lung were isolated, fixed for 4 h at room temperature and overnight at 4 °C in 4% paraformaldehyde in 0.1 M sodium phosphate (pH 7.6), and washed twice with PBS. A portion of each lobe was embedded in paraffin in a routine manner and stained with hematoxylin and eosin. The majority of each lobe was dehydrated and embedded in accordance with the JB-4 kit from Polysciences (Warrington, PA). Sections were cut on a Reichert-Jung Supracut microtome (Leica, Deerfield, IL) with glass knives and picked up on glass slides. Some of the sections were stained using the chloroacetate esterase reaction with counterstaining by hematoxylin to depict neutrophils. To assess extracellular matrix deposition, some sections were stained with the Jones' periodic acid-methenamine silver method (37) modified with hematoxylin counterstain in the JB4-embedded lungs.

The degree of alveolar septal thickening was quantitated without knowledge of the particular strain as described previously (36). Briefly, 5 pictures of randomly selected areas of the lower lobes of each lung at low power (x10 magnifications) stained with chloroacetate esterase and hematoxylin were obtained, and the areas of septal thickening, defined as macrophage proliferation in the alveolar septa accompanied by fibroblasts, giant cells, and increased extracellular matrix, were outlined. After being converted to digital images with a flatbed computer scanner (UMAX PowerLook III) and software (Adobe Photoshop 7.0), the number of pixels contained in the areas of the digital images that had septal thickening and the number of pixels contained in the image of the entire lung field were determined with the histogram function. The total number of pixels outlined as septal thickening was divided by the total number of pixels in the entire lung field and multiplied by 100 to generate a percentage of area with septal thickening for each animal.

Bronchoalveolar Lavage Fluid Analysis in Mice with Bleomycin-induced Pulmonary Fibrosis—Twelve days after treatment with bleomycin or saline, mice in each group were killed by intraperitoneal injection of an overdose of pentobarbital. Bronchoalveolar lavage (BAL) fluid was obtained by lavage with 2.25 ml of PBS with 1 mM EDTA (0.75 ml x 3) through a 22-gauge intravenous canula. The fluid was centrifuged at 500 x g for 5 min, and the supernatants were retained for measurement of eicosanoids. The cell pellets were resuspended in 100 µl of PBS, and total and differential cell counts were obtained as described above.

Assays for eicosanoids were carried out on 2 ml of each BAL fluid supernatant to which 10,000 cpm of [3H]LTC4 (PerkinElmer Life Sciences) was added to calculate percentage recovery. Proteins were precipitated from the supernatant by the addition of a 4x volume of methanol and incubation on ice for 10 min. After centrifugation at 3,000 x g for 10 min, samples were dried in a vacuum centrifuge, resuspended in 50 mM sodium acetate (pH 4.0) with 10% methanol, and subjected to C18 Sep-Pak cartridge purification (Waters Associates, Milford, MA). The eluted samples obtained from each cartridge were dried by vacuum centrifugation and resuspended in enzyme immunoassay buffer. The amounts of cys-LTs, LTB4, and prostaglandin E2 were measured with enzyme immunoassay kits (Amersham Biosciences) according to the manufacturer's instructions.

Statistical Analysis—Results were expressed as mean ± S.E. Stu dent's t test was used for the statistical analysis in cases in which the variance was homogeneous, and Welch's test was used when the variance was heterogeneous. A value of p < 0.05 was considered significant.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 EXPERIMENTAL PROCEDURES
 RESULTS
DISCUSSION
 REFERENCES
 
Generation of CysLT2 Receptor Gene-disrupted Mice—A targeting vector was designed so that the neo gene insertion interrupts 85% of the coding region of the mouse CysLT2 receptor gene (Fig. 1A). Of 150 embryonic stem cell clones that survived in the presence of G418 and ganciclovir, two were identified as targeted clones by Southern blot analysis for the 2.6-kb band. The embryonic stem cell clones were microinjected into BALB/c mouse-derived blastocysts. Eight male chimeric mice (>50% chimerism by coat color) were obtained from each clone and bred to C57BL/6 females. The heterozygotes, verified by Southern blot analysis, were intercrossed to obtain mice homozygous for the CysLT2 receptor-null mutation. Southern blot analysis of EcoRI-digested DNA from the progeny demonstrated a 2.6-kb band for the disrupted allele and an ~10-kb band for the wild-type allele, respectively, as illustrated for one litter (Fig. 1B). Litter size from these intercrosses were normal (mean = 6.6 offspring per litter; n = 39 litters). Of 258 progeny geno-typed from these matings, 62 were homozygotes (24.0%), 130 were heterozygotes (50.4%), and 66 were wild-type (25.6%), consistent with the expected Mendelian frequency. In addition, a similar number of male and female (29 and 33) homozygotes were produced. The CysLT2 receptor-null mice that developed normally (mean body weight = 25.7 g at 8 weeks), were fertile, and exhibited no apparent clinical abnormalities up to 10 months of age. There were no microscopic abnormalities in the CysLT2 receptor-null mice (data not shown).

Because mouse CysLT2 receptor mRNA is most abundantly expressed in the lung, spleen, and small intestine by Northern blot analysis (22) and reverse transcription-PCR (23), we examined whether homologous recombination by the targeting vector disrupted expression of the CysLT2 receptor mRNA in these tissues. By Northern blot analysis with poly(A)+ RNAs, a 5.5-kb band was dominantly detected as previously reported (22) and a 1.8-kb band that is considered to be the mature transcript based on its size (23) was also faintly detected in these tissues of the wild-type mice. In contrast, those bands were not detected in the samples from CysLT2 receptor-null mice, although a 9-kb band was detected in the spleen of CysLT2 receptor-null mice (Fig. 1C). Because we used a CysLT2 receptor coding region probe, the 9-kb band was considered to be an unstable transcript that contained part of exon VI. Expression of an aberrant and unstable transcript was also the case for the LTC4 synthase-null mice (34). Importantly, there was no change in the CysLT1 receptor mRNA expression in these tissues by the disruption of the CysLT2 receptor gene (Fig. 1C).

Role of the CysLT2 Receptor in Zymosan A-induced Peritoneal Inflammation—Although we previously demonstrated that the early plasma protein extravasation in zymosan A-induced peritoneal inflammation is reduced by ~50% in LTC4 synthase-null mice (34) and is similarly reduced in CysLT1 receptor-null mice (35), we performed the same study to determine whether there exists any role for the CysLT2 receptor. Plasma protein extravasation assessed 30 to 240 min after zymosan A injection in CysLT2 receptor-null mice was similar at all time points compared with that of the wild-type mice (Fig. 2A).



View larger version (15K):
[in this window]
[in a new window]
  		FIG. 2.
Zymosan A-induced peritoneal inflammation in CysLT2 receptor-null mice. A, plasma protein extravasation. CysLT2 receptor-null mice ({circ}) and their wild-type littermates (•) were injected intraperitoneally with 1 ml of zymosan A in PBS (1 mg/ml) immediately after an intravenous injection of 0.5% Evans blue dye. Peritoneal lavage was performed 30, 60, 120, and 240 min after zymosan A injection, and the absorbance of the lavage fluid supernatant was measured at 610 nm to quantitate Evans blue dye extravasation. Values at each time point are the mean ± S.E. (n = 3–5 mice). Results are combined from two independent experiments. B, neutrophil accumulation. Cell pellets 4 h after zymosan A injection were assayed for total cell counts (black bars) and neutrophil counts (gray bars) in each peritoneal lavage fluid from CysLT2 receptor-null (right) and wild-type (left) mice. Values are the mean ± S.E. Results are combined from two independent experiments (n = 3–5 mice per group).

 
We also examined neutrophil infiltration, which generally peaks 240 min after the injection of zymosan A (32, 38, 39). As assessed with neutrophil counts 240 min after the injection, there was no significant difference in neutrophil infiltration between CysLT2 receptor-null and wild-type mice (Fig. 2B).

Role of the CysLT2 Receptor in Mast Cell-dependent PCA—We previously demonstrated that the local edema induced by IgE-dependent PCA is reduced by 50% in the LTC4 synthase-null mice as compared with their wild-type littermates (34) and that this cys-LT-mediated response is dominantly through the CysLT1 receptor (35). Nonetheless, to determine whether the CysLT2 receptor is involved, we examined plasma protein extravasation induced by IgE-dependent PCA in CysLT2 receptor-null mice. Unexpectedly, Evans blue dye extravasation in the ear of CysLT2 receptor-null mice was reduced in a preliminary experiment. Thus, we included the CysLT1 receptor-null mice in three more studies. Plasma protein extravasation was reduced by ~65% 30 min after antigen challenge as compared with the level in the wild-type littermates, and the reduction was similar to that of CysLT1 receptor-null mice (Fig. 3).



View larger version (15K):
[in this window]
[in a new window]
  		FIG. 3.
PCA in CysLT2 receptor-null mice. CysLT2 receptor-null mice (white bar), their wild-type littermates (black bar), and CysLT1 receptor-null mice (gray bar) received intradermal injections of 25 ng of mouse anti-dinitrophenyl monoclonal IgE in 25 µl of saline in the right ear and 25 µl of saline only in the left ear. After 20 h, mice were injected intravenously with 100 µg of dinitrophenyl-human serum albumin and 1% Evans blue dye in 100 µl of PBS. A 6-mm diameter ear disk was isolated 30 min after challenge, and Evans blue dye was extracted in 200 µl of formamide at 55 °C for 48 h. Extravasation of Evans blue dye was quantitated by spectrophotometric analysis at 610 nm. The difference between the absorbance of the right and left ear extracts representing the net extravasation in the sensitized ear was expressed as the mean ± S.E. Results are combined from three independent analyses (n = 10 mice per group). *, p < 0.001.

 
Role of the CysLT2 Receptor in Bleomycin-induced Pulmonary Fibrosis—We previously demonstrated that bleomycin-induced pulmonary chronic inflammation and fibrosis are attenuated in LTC4 synthase-null mice but exacerbated in CysLT1 receptor-null mice (36). To define a role for the CysLT2 receptor in the pathobiology of chronic inflammation with fibrosis, we injected bleomycin into the tracheas of CysLT2 receptor-null mice and their wild-type littermates and examined lung histology and cys-LT levels in the BAL fluid 12 days after the injection. The lung tissue of bleomycin-treated CysLT2 receptor-null mice had less septal thickening, with less accumulation of monocyte/macrophages, giant cells, fibroblasts, and eosinophils and less extracellular matrix deposition than that of their bleomycin-treated wild-type littermates (Fig. 4, a–f). We used the Jones' periodic acid-methenamine silver method with modifications to assess the deposition of extracellular matrix proteins beneath the pleura, around the bronchi, and in the alveolar septa. After bleomycin treatment, the amount of black fibrillar material was less in the lung tissue of the CysLT2 receptor-null mice as compared with lung tissue from the wild-type mice (Fig. 4, g–l).



View larger version (109K):
[in this window]
[in a new window]
  		FIG. 4.
Histologic assessment of bleomycin-induced pulmonary fibrosis in CysLT2 receptor-null mice. The lower lobes of the lungs were examined histologically 12 days after animals received an intratracheal injection of saline or bleomycin. Sections of lung from saline-treated wild-type mice (a, low power: x10 objective; d, high power: x50 objective) and bleomycin-treated wild-type (b, low power; e, high power) and CysLT2 receptor-null (c, low power; f, high power) mice were stained with hematoxylin and eosin (a–c), or by the chloroacetate esterase reaction with counterstaining with hematoxylin (d–f). Sections of lung from saline-treated wild-type mice (g, low power; j, high power) and bleomycin-treated wild-type (h, low power; k, high power) and CysLT2 receptor-null (i, low power; l, high power) mice were stained with the Jones' periodic acid-methenamine silver method with hematoxylin counterstain. Reticular fibers (type III collagen) and basement membranes were stained dark brown to black.

 
To quantitate the degree of bleomycin-induced inflammation and fibrosis, we analyzed the sections stained with chloroacetate esterase and hematoxylin by comparing digital images of the areas of alveolar septal thickening and the total area of the lower lung lobes of the mice 12 days after bleomycin injection. The area of the lower lobes with alveolar septal thickening in CysLT2 receptor-null mice was significantly reduced to only 39% that of wild-type mice (Table I). Bleomycin treatment significantly increased the amounts of cys-LTs in the BAL fluid in both wild-type mice and CysLT2 receptor-null mice. Despite the attenuation of septal thickening, bleomycin-treated CysLT2 receptor-null mice had levels of cys-LTs comparable with those of the bleomycin-treated wild-type mice (Table I). Bleomycin treatment did not increase the amounts of LTB4 in either wild-type mice or CysLT2 receptor-null mice. Although bleomycin treatment did increase the amount of prostaglandin E2 in wild-type mice, but not in CysLT2 receptor-null mice (Table I), the amounts of prostaglandin E2 recovered from both strains with bleomycin injury were not different.


View this table:
[in this window]
[in a new window]
  		TABLE I
Alveolar septal thickening and the amounts of cys-LTs, LTB4, and PGE2 in the BAL fluid in CysLT2 receptor-null mice and wild-type mice 12 days after treatment with bleomycin or saline Alveolar septal thickening was quantitated by digital imaging as described under "Experimental Procedures" and the amounts of cys-LTs, LTB4, and PGE2 in BAL fluid were measured by enzyme immunoassays. Data are combined from three independent experiments.

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 EXPERIMENTAL PROCEDURES
 RESULTS
 DISCUSSION
REFERENCES
 
We generated CysLT2 receptor-null mice by targeted gene disruption to seek in vivo evidence for the role of this receptor in cys-LT-mediated pathobiology. Despite the extensive characterization of the CysLT2 receptor with transfected cells (7–9, 22, 23) and human umbilical vein endothelial cells (17, 18, 40), little is known about the role of the second receptor in in vivo inflammatory responses, because of the lack of specific antagonists for it. We have found that the CysLT2 receptor mediates increased vascular permeability in IgE-dependent PCA and macrophage and fibroblast accumulation and extracellular matrix protein deposition in response to bleomycin.

LTC4 and LTD4 directly elicit transient vasoconstriction of small arterioles and then increased vascular permeability in postcapillary venules (41, 42), where contraction of endothelial cells is considered to be required for the plasma protein leakage (43). Because increased vascular permeability induced by either intraperitoneal injection of zymosan A or IgE-dependent PCA is attenuated in the CysLT1 receptor-null mice (35) to the same extent as in the LTC4 synthase-null mice (34), we anticipated that the effect of CysLT2 receptor deficiency on increased vascular permeability in these acute inflammatory responses would be negligible. Surprisingly, increased vascular permeability by IgE-dependent PCA, but not in zymosan-induced peritoneal inflammation, was attenuated in the CysLT2 receptor-null mice. Furthermore, this protection was equal to that observed in the CysLT1 receptor-null mice in the same set of experiments (Fig. 3). The lack of attenuation of the vascular leak in the peritoneal cavity in the CysLT2 receptor-null mice would imply heterogeneity of receptor distribution in the microvasculature across tissues (Fig. 2).

There was no impairment of mast cell degranulation by morphologic analysis (data not shown), indicating that the attenuation involved the microvasculature. The CysLT2 receptor mRNA is detected in endothelial cells of the mouse heart by in situ hybridization (23) and on human umbilical vein endothelial cells in a 1000-fold excess to the CysLT1 receptor by quantitative reverse transcription-PCR (17, 18). Moreover, the calcium response of the human umbilical vein endothelial cells to the cys-LTs was attributed to the CysLT2 receptor because there was no inhibition by a selective CysLT1 receptor antagonist and some inhibition by a partial CysLT2 receptor agonist. That the magnitude of the attenuation of the PCA was approximately equal in each of three independent experiments for the CysLT2 receptor-null and CysLT1 receptor-null mice and for the overall mean implies that the inhibition is linked, possibly through CysLT2 and CysLT1 receptor heterodimers. Heterodimer formation has been reported for other G protein-coupled receptors, such as opioid (44) and chemokine (45) receptors.

In this study, we have shown that CysLT2 is the receptor responsible for mediating the contribution of the cys-LTs to bleomycin-induced chronic pulmonary inflammation and fibrosis. The levels of cys-LTs in the BAL fluid of bleomycin-treated CysLT2 receptor-null mice and wild-type mice remained comparable (Table I). Thus, either the lack of cys-LT production as in the LTC4 synthase-null mice (36) or the lack of the CysLT2 receptor significantly attenuates the chronic injury (Fig. 4). In contrast, the CysLT1 receptor-null mice were found to have increased bleomycin-induced pulmonary inflammation and increased levels of cys-LTs in BAL fluid relative to their controls (36). The source of cys-LTs in this response is most likely alveolar and interstitial macrophages as cells of this lineage in several species generate cys-LTs in response to a range of ligands (46–49).

Mouse skin fibroblasts express both CysLT1 and CysLT2 receptors by in situ hybridization (22). Before the CysLT1 and CysLT2 receptors were defined by their cloning, it was shown that LTC4 and LTD4 could stimulate [3H]thymidine incorporation by in vitro cultured human skin fibroblasts in the presence of indomethacin and that the effect of LTC4 was slightly superior to that of LTD4 (50). In addition, LTC4 and, to a lesser extent, LTD4 could stimulate collagen synthesis of rat lung fibroblasts in vitro and LTC4 could directly bind to the cells (51). These earlier studies revealing a functional receptor for cys-LTs on fibroblasts analyzed in vitro are compatible with a CysLT2 receptor-mediated response based on the efficacy of LTC4. Our findings that CysLT2 receptor-null mice have a significant reduction in bleomycin-induced pulmonary fibrosis could reflect a dominant role for this receptor in the fibroblast component of the response.

We generated CysLT2 receptor-null mice and then demonstrated a role for the CysLT2 receptor in inflammatory responses in vivo. Our studies highlight the CysLT2 receptor as a potential therapeutic target for the treatment of idiopathic pulmonary fibrosis, for which no efficient interventions are available (52).


   FOOTNOTES
 
* This work was supported by National Institutes of Health Grants AI-07306, AI-31599, and HL-36110. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Back

Both authors contributed equally to this work. Back

{ddagger}{ddagger} To whom correspondence should be addressed: Brigham and Women's Hospital, Smith Bldg., Rm. 626C, One Jimmy Fund Way, Boston, MA 02115. Tel.: 617-525-1263; Fax: 617-525-1310; E-mail: ykanaoka{at}rics.bwh.harvard.edu.

1 The abbreviations used are: cys-LT, cysteinyl leukotriene; LT, leukotriene; PCA, passive cutaneous anaphylaxis; PBS, phosphate-buffered saline; BAL, bronchoalveolar lavage; CysLT1, cysteinyl leukotriene 1 receptor. Back


   ACKNOWLEDGMENTS
 
We thank Mi Xiao Donovan for technical assistance and Dr. Arlene Sharpe and Lina Du (Transgenic Core Facility,Brigham and Women's Hospital) for blastocyst injection of the embryonic stem cell clones.



   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 EXPERIMENTAL PROCEDURES
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Murphy, R. C., Hammarstrom, S., and Samuelsson, B. (1979) Proc. Natl. Acad. Sci. U. S. A. 76, 4275-4279[Abstract/Free Full Text]
  2. Lewis, R. A., Austen, K. F., Drazen, J. M., Clark, D. A., Marfat, A., and Corey, E. J. (1980) Proc. Natl. Acad. Sci. U. S. A. 77, 3710-3714[Abstract/Free Full Text]
  3. Israel, E., Dermarkarian, R., Rosenberg, M., Sperling, R., Taylor, G., Rubin, P., and Drazen, J. M. (1990) N. Engl. J. Med. 323, 1740-1744[Abstract]
  4. Manning, P. J., Watson, R. M., Margolskee, D. J., Williams, V. C., Schwartz, J. I., and O'Byrne, P. M. (1990) N. Engl. J. Med. 323, 1736-1739[Abstract]
  5. Lynch, K. R., O'Neill, G. P., Liu, Q., Im, D. S., Sawyer, N., Metters, K. M., Coulombe, N., Abramovitz, M., Figueroa, D. J., Zeng, Z., Connolly, B. M., Bai, C., Austin, C. P., Chateauneuf, A., Stocco, R., Greig, G. M., Kargman, S., Hooks, S. B., Hosfield, E., Williams, D. L., Jr., Ford-Hutchinson, A. W., Caskey, C. T., and Evans, J. F. (1999) Nature 399, 789-793[CrossRef][Medline] [Order article via Infotrieve]
  6. Sarau, H. M., Ames, R. S., Chambers, J., Ellis, C., Elshourbagy, N., Foley, J. J., Schmidt, D. B., Muccitelli, R. M., Jenkins, O., Murdock, P. R., Herrity, N. C., Halsey, W., Sathe, G., Muir, A. I., Nuthulaganti, P., Dytko, G. M., Buckley, P. T., Wilson, S., Bergsma, D. J., and Hay, D. W. (1999) Mol. Pharmacol. 56, 657-663[Abstract/Free Full Text]
  7. Heise, C. E., O'Dowd, B. F., Figueroa, D. J., Sawyer, N., Nguyen, T., Im, D. S., Stocco, R., Bellefeuille, J. N., Abramovitz, M., Cheng, R., Williams, D. L., Jr., Zeng, Z., Liu, Q., Ma, L., Clements, M. K., Coulombe, N., Liu, Y., Austin, C. P., George, S. R., O'Neill, G. P., Metters, K. M., Lynch, K. R., and Evans, J. F. (2000) J. Biol. Chem. 275, 30531-30536[Abstract/Free Full Text]
  8. Nothacker, H. P., Wang, Z., Zhu, Y., Reinscheid, R. K., Lin, S. H., and Civelli, O. (2000) Mol. Pharmacol. 58, 1601-1608[Medline] [Order article via Infotrieve]
  9. Takasaki, J., Kamohara, M., Matsumoto, M., Saito, T., Sugimoto, T., Ohishi, T., Ishii, H., Ota, T., Nishikawa, T., Kawai, Y., Masuho, Y., Isogai, T., Suzuki, Y., Sugano, S., and Furuichi, K. (2000) Biochem. Biophys. Res. Commun. 274, 316-322[CrossRef][Medline] [Order article via Infotrieve]
  10. Figueroa, D. J., Breyer, R. M., Defoe, S. K., Kargman, S., Daugherty, B. L., Waldburger, K., Liu, Q., Clements, M., Zeng, Z., O'Neill, G. P., Jones, T. R., Lynch, K. R., Austin, C. P., and Evans, J. F. (2001) Am. J. Respir. Crit. Care Med. 163, 226-233[Abstract/Free Full Text]
  11. Figueroa, D. J., Borish, L., Baramki, D., Philip, G., Austin, C. P., and Evans, J. F. (2003) Clin. Exp. Allergy 33, 1380-1388[CrossRef][Medline] [Order article via Infotrieve]
  12. Mellor, E. A., Maekawa, A., Austen, K. F., and Boyce, J. A. (2001) Proc. Natl. Acad. Sci. U. S. A. 98, 7964-7969[Abstract/Free Full Text]
  13. Sjostrom, M., Jakobsson, P. J., Juremalm, M., Ahmed, A., Nilsson, G., Macchia, L., and Haeggstrom, J. Z. (2002) Biochim. Biophys. Acta 1583, 53-62[Medline] [Order article via Infotrieve]
  14. Sjostrom, M., Jakobsson, P. J., Heimburger, M., Palmblad, J., and Haeggstrom, J. Z. (2001) Eur. J. Biochem. 268, 2578-2586[Medline] [Order article via Infotrieve]
  15. Mita, H., Hasegawa, M., Saito, H., and Akiyama, K. (2001) Clin. Exp. Allergy 31, 1714-1723[CrossRef][Medline] [Order article via Infotrieve]
  16. Mellor, E. A., Frank, N., Soler, D., Hodge, M. R., Lora, J. M., Austen, K. F., and Boyce, J. A. (2003) Proc. Natl. Acad. Sci. U. S. A. 100, 11589-11593[Abstract/Free Full Text]
  17. Lotzer, K., Spanbroek, R., Hildner, M., Urbach, A., Heller, R., Bretschneider, E., Galczenski, H., Evans, J. F., and Habenicht, A. J. (2003) Arterioscler. Thromb. Vasc. Biol. 23, e32-36[Abstract/Free Full Text]
  18. Sjostrom, M., Johansson, A. S., Schroder, O., Qiu, H., Palmblad, J., and Haeggstrom, J. Z. (2003) Arterioscler. Thromb. Vasc. Biol. 23, e37-41[Abstract/Free Full Text]
  19. Maekawa, A., Kanaoka, Y., Lam, B. K., and Austen, K. F. (2001) Proc. Natl. Acad. Sci. U. S. A. 98, 2256-2261[Abstract/Free Full Text]
  20. Mollerup, J., Jorgensen, S. T., Hougaard, C., and Hoffmann, E. K. (2001) Biochim. Biophys. Acta 1517, 455-459[Medline] [Order article via Infotrieve]
  21. Martin, V., Sawyer, N., Stocco, R., Unett, D., Lerner, M. R., Abramovitz, M., and Funk, C. D. (2001) Biochem. Pharmacol. 62, 1193-1200[CrossRef][Medline] [Order article via Infotrieve]
  22. Ogasawara, H., Ishii, S., Yokomizo, T., Kakinuma, T., Komine, M., Tamaki, K., Shimizu, T., and Izumi, T. (2002) J. Biol. Chem. 277, 18763-18768[Abstract/Free Full Text]
  23. Hui, Y., Yang, G., Galczenski, H., Figueroa, D. J., Austin, C. P., Copeland, N. G., Gilbert, D. J., Jenkins, N. A., and Funk, C. D. (2001) J. Biol. Chem. 276, 47489-47495[Abstract/Free Full Text]
  24. Brink, C., Dahlen, S. E., Drazen, J., Evans, J. F., Hay, D. W., Nicosia, S., Serhan, C. N., Shimizu, T., and Yokomizo, T. (2003) Pharmacol. Rev. 55, 195-227[Abstract/Free Full Text]
  25. Evans, J. F. (2003) Prostaglandins Leukotriene Essent. Fatty Acids 69, 117-122[CrossRef][Medline] [Order article via Infotrieve]
  26. Yoshimoto, T., Soberman, R. J., Spur, B., and Austen, K. F. (1988) J. Clin. Investig. 81, 866-871[Medline] [Order article via Infotrieve]
  27. Lam, B. K., Penrose, J. F., Freeman, G. J., and Austen, K. F. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 7663-7667[Abstract/Free Full Text]
  28. Welsch, D. J., Creely, D. P., Hauser, S. D., Mathis, K. J., Krivi, G. G., and Isakson, P. C. (1994) Proc. Natl. Acad. Sci. U. S. A. 91, 9745-9749[Abstract/Free Full Text]
  29. Samuelsson, B., Dahlen, S. E., Lindgren, J. A., Rouzer, C. A., and Serhan, C. N. (1987) Science 237, 1171-1176[Abstract/Free Full Text]
  30. Lam, B. K., Owen, W. F., Jr., Austen, K. F., and Soberman, R. J. (1989) J. Biol. Chem. 264, 12885-12889[Abstract/Free Full Text]
  31. Anderson, M. E., Allison, R. D., and Meister, A. (1982) Proc. Natl. Acad. Sci. U. S. A. 79, 1088-1091[Abstract/Free Full Text]
  32. Shi, Z. Z., Han, B., Habib, G. M., Matzuk, M. M., and Lieberman, M. W. (2001) Mol. Cell. Biol. 21, 5389-5395[Abstract/Free Full Text]
  33. Lee, C. W., Lewis, R. A., Corey, E. J., and Austen, K. F. (1983) Immunology 48, 27-35[Medline] [Order article via Infotrieve]
  34. Kanaoka, Y., Maekawa, A., Penrose, J. F., Austen, K. F., and Lam, B. K. (2001) J. Biol. Chem. 276, 22608-22613[Abstract/Free Full Text]
  35. Maekawa, A., Austen, K. F., and Kanaoka, Y. (2002) J. Biol. Chem. 277, 20820-20824[Abstract/Free Full Text]
  36. Beller, T. C., Friend, D. S., Maekawa, A., Lam, B. K., Austen, K. F., and Kanaoka, Y. (2004) Proc. Natl. Acad. Sci. U. S. A. 101, 3047-3052[Abstract/Free Full Text]
  37. Sheehan, D. C., and Hrapchak, B. B. (1980) in Theory and Practice of Histotechnology, 2nd Ed., Battelle Press, Detroit, MI
  38. Doherty, N. S., Poubelle, P., Borgeat, P., Beaver, T. H., Westrich, G. L., and Schrader, N. L. (1985) Prostaglandins 30, 769-789[CrossRef][Medline] [Order article via Infotrieve]
  39. Rao, T. S., Currie, J. L., Shaffer, A. F., and Isakson, P. C. (1994) J. Pharmacol. Exp. Ther. 269, 917-925[Abstract/Free Full Text]
  40. Di Gennaro, A., Carnini, C., Buccellati, C., Ballerio, R., Zarini, S., Fumagalli, F., Viappiani, S., Librizzi, L., Hernandez, A., Murphy, R. C., Constantin, G., De Curtis, M., Folco, G., and Sala, A. (2004) FASEB J. 18, 842-844[Abstract/Free Full Text]
  41. Dahlen, S. E., Bjork, J., Hedqvist, P., Arfors, K. E., Hammarstrom, S., Lindgren, J. A., and Samuelsson, B. (1981) Proc. Natl. Acad. Sci. U. S. A. 78, 3887-3891[Abstract/Free Full Text]
  42. Soter, N. A., Lewis, R. A., Corey, E. J., and Austen, K. F. (1983) J. Investig. Dermatol. 80, 115-119[CrossRef][Medline] [Order article via Infotrieve]
  43. Majno, G., Shea, S. M., and Leventhal, M. (1969) J. Cell Biol. 42, 647-672[Abstract/Free Full Text]
  44. Jordan, B. A., and Devi, L. A. (1999) Nature 399, 697-700[CrossRef][Medline] [Order article via Infotrieve]
  45. Mellado, M., Rodriguez-Frade, J. M., Vila-Coro, A. J., Fernandez, S., Martin de Ana, A., Jones, D. R., Toran, J. L., and Martinez, A. C. (2001) EMBO J. 20, 2497-2507[CrossRef][Medline] [Order article via Infotrieve]
  46. Rouzer, C. A., Scott, W. A., Cohn, Z. A., Blackburn, P., and Manning, J. M. (1980) Proc. Natl. Acad. Sci. U. S. A. 77, 4928-4932[Abstract/Free Full Text]
  47. Rouzer, C. A., Scott, W. A., Hamill, A. L., and Cohn, Z. A. (1982) J. Exp. Med. 155, 720-733[Abstract/Free Full Text]
  48. Orange, R. P., Moore, E. G., and Gelfand, E. W. (1980) J. Immunol. 124, 2264-2267[Abstract]
  49. Damon, M., Chavis, C., Godard, P., Michel, F. B., and Crastes de Paulet, A. (1983) Biochem. Biophys. Res. Commun. 111, 518-524[CrossRef][Medline] [Order article via Infotrieve]
  50. Baud, L., Perez, J., Denis, M., and Ardaillou, R. (1987) J. Immunol. 138, 1190-1195[Abstract/Free Full Text]
  51. Phan, S. H., McGarry, B. M., Loeffler, K. M., and Kunkel, S. L. (1988) Biochemistry 27, 2846-2853[CrossRef][Medline] [Order article via Infotrieve]
  52. Raghu, G., Brown, K. K., Bradford, W. Z., Starko, K., Noble, P. W., Schwartz, D. A., and King, T. E., Jr. (2004) N. Engl. J. Med. 350, 125-133[Abstract/Free Full Text]

Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?


This article has been cited by other articles:


Home page
 CirculationHome page
T. A. Doser, S. Turdi, D. P. Thomas, P. N. Epstein, S.-Y. Li, and J. Ren
Transgenic Overexpression of Aldehyde Dehydrogenase-2 Rescues Chronic Alcohol Intake-Induced Myocardial Hypertrophy and Contractile Dysfunction
Circulation, April 14, 2009; 119(14): 1941 - 1949.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
Y. Jiang, L. Borrelli, B. J. Bacskai, Y. Kanaoka, and J. A. Boyce
P2Y6 Receptors Require an Intact Cysteinyl Leukotriene Synthetic and Signaling System to Induce Survival and Activation of Mast Cells
J. Immunol., January 15, 2009; 182(2): 1129 - 1137.
[Abstract] [Full Text] [PDF]

Home page
 FASEB J.Home page
M. P. W. Moos, J. D. Mewburn, F. W. K. Kan, S. Ishii, M. Abe, K. Sakimura, K. Noguchi, T. Shimizu, and C. D. Funk
Cysteinyl leukotriene 2 receptor-mediated vascular permeability via transendothelial vesicle transport
FASEB J, December 1, 2008; 22(12): 4352 - 4362.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
A. Maekawa, Y. Kanaoka, W. Xing, and K. F. Austen
Functional recognition of a distinct receptor preferential for leukotriene E4 in mice lacking the cysteinyl leukotriene 1 and 2 receptors
PNAS, October 28, 2008; 105(43): 16695 - 16700.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
N. Uozumi, Y. Kita, and T. Shimizu
Modulation of Lipid and Protein Mediators of Inflammation by Cytosolic Phospholipase A2{alpha} during Experimental Sepsis
J. Immunol., September 1, 2008; 181(5): 3558 - 3566.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
S. Paruchuri, Y. Jiang, C. Feng, S. A. Francis, J. Plutzky, and J. A. Boyce
Leukotriene E4 Activates Peroxisome Proliferator-activated Receptor {gamma} and Induces Prostaglandin D2 Generation by Human Mast Cells
J. Biol. Chem., June 13, 2008; 283(24): 16477 - 16487.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
C. Thompson, A. Cloutier, Y. Bosse, C. Poisson, P. Larivee, P. P. McDonald, J. Stankova, and M. Rola-Pleszczynski
Signaling by the Cysteinyl-Leukotriene Receptor 2: INVOLVEMENT IN CHEMOKINE GENE TRANSCRIPTION
J. Biol. Chem., January 25, 2008; 283(4): 1974 - 1984.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
K. M. Vannella, T. R. McMillan, R. P. Charbeneau, C. A. Wilke, P. E. Thomas, G. B. Toews, M. Peters-Golden, and B. B. Moore
Cysteinyl Leukotrienes Are Autocrine and Paracrine Regulators of Fibrocyte Function
J. Immunol., December 1, 2007; 179(11): 7883 - 7890.
[Abstract] [Full Text] [PDF]

Home page
 NEJMHome page
M. Peters-Golden and W. R. Henderson Jr.
Leukotrienes
N. Engl. J. Med., November 1, 2007; 357(18): 1841 - 1854.
[Full Text] [PDF]

Home page
 BloodHome page
Y. Jiang, L. A. Borrelli, Y. Kanaoka, B. J. Bacskai, and J. A. Boyce
CysLT2 receptors interact with CysLT1 receptors and down-modulate cysteinyl leukotriene dependent mitogenic responses of mast cells
Blood, November 1, 2007; 110(9): 3263 - 3270.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Respir. Cell Mol. Bio.Home page
S. B. Early, E. Barekzi, J. Negri, K. Hise, L. Borish, and J. W. Steinke
Concordant Modulation of Cysteinyl Leukotriene Receptor Expression by IL-4 and IFN-{gamma} on Peripheral Immune Cells
Am. J. Respir. Cell Mol. Biol., June 1, 2007; 36(6): 715 - 720.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
Y. Jiang, Y. Kanaoka, C. Feng, K. Nocka, S. Rao, and J. A. Boyce
Cutting edge: interleukin 4-dependent mast cell proliferation requires autocrine/intracrine cysteinyl leukotriene-induced signaling.
J. Immunol., September 1, 2006; 177(5): 2755 - 2759.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
B. Uzonyi, K. Lotzer, S. Jahn, C. Kramer, M. Hildner, E. Bretschneider, D. Radke, M. Beer, R. Vollandt, J. F. Evans, et al.
Cysteinyl leukotriene 2 receptor and protease-activated receptor 1 activate strongly correlated early genes in human endothelial cells
PNAS, April 18, 2006; 103(16): 6326 - 6331.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
D. C. Kim, F. I. Hsu, N. A. Barrett, D. S. Friend, R. Grenningloh, I-C. Ho, A. Al-Garawi, J. M. Lora, B. K. Lam, K. F. Austen, et al.
Cysteinyl Leukotrienes Regulate Th2 Cell-Dependent Pulmonary Inflammation
J. Immunol., April 1, 2006; 176(7): 4440 - 4448.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
279/44/46129    most recent
M407057200v1
Right arrow Submit a Letter to Editor
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowRequest Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Beller, T. C.
Right arrow Articles by Kanaoka, Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Beller, T. C.
Right arrow Articles by Kanaoka, Y.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
 All ASBMB Journals   Molecular and Cellular Proteomics 
 Journal of Lipid Research   ASBMB Today 
Copyright © 2004 by the American Society for Biochemistry and Molecular Biology.
Advertisement
spacer
Advertisement
Advertisement