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At least 19 recordsLinked to original sources

PubMed1998

Cloning of the gene and cDNA for hamster chymase 2, and expression of chymase 1, chymase 2 and angiotensin-converting enzyme in the terminal stage of cardiomyopathic hearts.

Chymase is responsible for the formation of angiotensin II, which plays crucial roles in the pathogenesis of cardiovascular diseases. In the present study we determined the gene organization of a novel hamster chymase (hamster chymase 2) and analysed the expression of chymase 1, chymase 2 and angiotensin-converting enzyme (ACE) in hamster hearts at the terminal stage of cardiomyopathy. The gene encoding hamster chymase 2 is 3.2 kb in length and has five exons and four intervening sequences. The overall organization of this gene is similar to that of several other serine proteases. The deduced amino acid sequence revealed the existence of a preproenzyme composed of a signal peptide with 19 amino acids, a propeptide with two amino acids and a catalytic domain with 226 amino acids. The predicted full sequence of the catalytic domain was revealed to be very similar to the sequences of mouse mast-cell protease 5 (86%), rat mast-cell protease III (85%) and human chymase (70%) and less similar to hamster chymase 1 (56%). The expression of chymase 1 in heart was higher than that of chymase 2. The cardiac chymase-like activity, as well as the mRNA levels of chymase 1 and 2 of BIO 14.6 cardiomyopathic hamsters at the age of 60 weeks were increased 3.4-, 2.8- and 5.1-fold respectively compared with age-matched BIO F1B control hamsters. The cardiac ACE activity and the ACE mRNA level of cardiomyopathic hamsters were also increased 4.1- and 2.4-fold compared with those of age-matched controls. These results suggest that up-regulation of both ACE and chymases participates in the pathophysiology of the terminal stage of cardiomyopathy.

Amino Acid Sequence↗

PubMed2005

Mast cell tryptase and chymase in chronic leg ulcers: chymase is potentially destructive to epithelium and is controlled by proteinase inhibitors.

BACKGROUND: Numerous mast cells are present in chronic leg ulcers. Tryptase and chymase are the major mediators of mast cells, but their significance is mostly dependent on their activity. In addition, the proteinases may affect ulcer epithelialization. OBJECTIVES: To study levels and activity of tryptase and chymase in wash samples and biopsies from chronic leg ulcers and the possible effect of these proteinases on keratinocyte growth and adherence. METHODS: Wash samples were taken from 16 patients and a superficial shave biopsy was taken in eight of these patients; a second biopsy series was obtained from the edge of chronic venous leg ulcers (n = 6). RESULTS: Significant levels of soluble tryptase activity and histamine, but low levels of chymase activity, were measured in wash samples from chronic ulcers. No tryptase-inhibiting activity, but clear chymase-inhibiting activity, was detected in the wash samples. In superficial wound bed biopsies, relatively marked levels of chymase activity together with histamine and tryptase activity were detected. In the second biopsy series, about 80% of the mast cells belonged to the MC(TC) type (tryptase- and chymase-immunopositive). However, about 55-61% of the chymase-immunopositive cells displayed chymase activity and 64 +/- 17% of the tryptase-positive cells revealed immunoreactivity of alpha(1)-antichymotrypsin. As the activity of chymase and tryptase was detected in the ulcer base in a ratio of 1:8, a preparation containing both chymase and tryptase was partially purified from human skin yielding a similar activity ratio of 1:11-13. Treatment of fibronectin-coated plastic surfaces with this preparation decreased the adherence of cultured human keratinocytes, this effect being attributable mainly to chymase. In 2-day cultures using growth factor/serum-deficient low- or high-calcium medium, the tryptase-chymase preparation inhibited the slow growth and at higher concentrations it even induced detachment of keratinocytes. This effect was attributed to chymase, and it was partially regulated by heparin and histamine. CONCLUSIONS: Even though chymase is partially inactivated in chronic leg ulcers, accumulated mast cells in the close proximity of the epithelium edge and their chymase may impair keratinocyte adherence and migration.

Adult↗

PubMed2002

Recombinant human chymase produced by silkworm-baculovirus expression system: its application for a chymase detection kit.

Human chymase is a mast cell-derived serine proteinase, which is a non-angiotensin converting enzyme angiotensin II-generating enzyme. It appears to participate in various diseases, but it is unclear whether chymase plays major roles in physiological and pathophysiological functions in vivo. To obtain information on the physiological and pathophysiological functions of chymase and to search for diseases in which chymase participates, in the present study, we aimed at producing recombinant human chymase in large quantities and at developing an ELISA system using anti-human chymase antibodies. A recombinant human chymase was produced by a silkworm-baculovirus expression system. The recombinant chymase in active form was efficiently purified from larval hemolymph using cation-exchange and heparin column chromatography. This recombinant enzyme was enzymatically identical with native human chymase. On the other hand, the stability of the recombinant enzyme in cultured medium for mammalian cells at 37 degrees C was very high as compared with the stability of the native enzyme; 20% of the activity was maintained 120 h after addition of medium. These results indicated that the recombinant enzyme could also utilize in vitro and in vivo assay systems. We obtained several anti-chymase monoclonal antibodies by using the recombinant human chymase as antigen. These antibodies were used to construct an ELISA system for measuring the chymase concentration in blood. As a result of preliminary examination using this ELISA system, it was shown that the chymase concentration in each serum from hypertensive patients is significantly higher than in normal serum. The ELISA system will be applicable for clinical diagnosis and in vivo evaluation systems for chymase-targeting drugs.

Animals↗

PubMed2006

Pathological roles of angiotensin II produced by mast cell chymase and the effects of chymase inhibition in animal models.

The discovery of a new angiotensin II (Ang II) pathway generated by mast cell chymase has highlighted new biological functions for Ang II that is not related to the classic renin-angiotensin system (RAS). The conversion of Ang I to II occurs not only via the plasma angiotensin converting enzyme (ACE) or tissue ACE but also via chymase produced in the mast cells of humans, monkeys, dogs, and hamsters. The conversion by chymase has been especially found in morbid tissues following the migration of mast cells. The newly discovered functions of chymase are discussed in this review. During the vascular narrowing that occurs after vein grafting or balloon injury in dogs, chymase activity and Ang II concentrations along with intimal proliferation are significantly increased and chymase inhibitors completely suppressed these increase, though ACE inhibitors are ineffective. Similar results have also been confirmed in the dog arteriovenous fistula stenosis model. In both human and animal aneurysmal aortas, chymase activity is significantly increased, and chymase inhibitor has been shown to prevent the development of aneurysms in dogs. Chymase is activated in diseased hearts, and chymase inhibitors reduce both the mortality rates after acute myocardial infarction and the cardiac fibrosis that leads to the development of cardiomyopathy in hamsters. Chymase is also a pro-angiogenic factor, since the injection of chymase strongly facilitates angiogenesis in hamsters. We propose that chymase inhibitors are effective in the prevention of multiple cardiovascular disorders, especially at the local event level without any effect on the systemic blood pressure.

Angiogenesis Inhibitors↗

PubMed1997

Distinct multisite synergistic interactions determine substrate specificities of human chymase and rat chymase-1 for angiotensin II formation and degradation.

Human chymase and rat chymase-1 are mast cell serine proteases involved in angiotensin II (Ang II) formation and degradation, respectively. Previous studies indicate that both these enzymes have similar P1 and P2 preferences, which are the major determinants of specificity. Surprisingly, despite the occurrence of optimal P2 and P1 residues at the Phe8 downward arrow and Tyr4 downward arrow bonds (where downward arrow, indicates the scissile bond in peptide substrates) in Ang I (DRVYIHPFHL), human chymase cleaves the Phe8 downward arrow bond with an approximately 750-fold higher catalytic efficiency (kcat/Km) than the Tyr4 downward arrow bond in Ang II (DRVYIHPF), whereas rat chymase-1 cleaves the Tyr4 downward arrow bond with an approximately 20-fold higher catalytic efficiency than the Phe8 downward arrow bond. Differences in the acyl groups IHPF and DRVY at the Phe8 downward arrow and Tyr4 downward arrow bonds, respectively, are chiefly responsible for the preference of human chymase for the Phe8 downward arrow bond. We show that the IHPF sequence forms an optimal acyl group, primarily through synergistic interactions between neighboring acyl group residues. In contrast to human chymase, rat chymase-1 shows a preference for the Tyr4 downward arrow bond, mainly because of a catalytically productive interaction between the enzyme and the P'1 Ile5. The overall effect of this P'1 Ile interaction on catalytic efficiency, however, is influenced by the structure of the acyl group and that of the other leaving group residues. For human chymase, the P'1 Ile interaction is not productive. Thus, specificity for Ang II formation versus Ang II degradation by these chymases is produced through synergistic interactions between acyl or leaving group residues as well as between the acyl and leaving groups. These observations indicate that nonadditive interactions between the extended substrate binding site of human chymase or rat chymase-1 and the substrate are best explained if the entire binding site is taken as an entity rather than as a collection of distinct subsites.

Amino Acid Sequence↗

PubMed2000

Chymase as a proangiogenic factor. A possible involvement of chymase-angiotensin-dependent pathway in the hamster sponge angiogenesis model.

We investigated the profound involvement of chymase, an alternative angiotensin II-generating enzyme, in angiogenesis using a hamster sponge implant model. In vivo transfection of human pro-chymase cDNA or a direct injection of purified chymase into the sponges implanted resulted in marked increment of hemoglobin contents in the sponge granuloma tissues, demonstrating that chymase has an ability to elicit angiogenesis and is a potent angiogenic factor. Daily injection of basic fibroblast growth factor into the sponges implanted also induced angiogenesis, which was suppressed by the treatment with chymostatin, an inhibitor of chymase, or TCV-116, an antagonist of angiotensin II (Ang II) type 1 receptor. Expression of chymase mRNA and production of Ang II in the granuloma tissues were enhanced by the stimulation with basic fibroblast growth factor. Chymase activity in the sponge granulomas increased in parallel with the rise in hemoglobin contents, and mast cells observed in the granuloma tissues were positively stained with anti-chymase antibody. Exogenous administration not only of Ang II but of angiotensin I (Ang I) directly into the sponges could enhance angiogenesis. Chymostatin inhibited the angiogenesis induced by Ang I but not Ang II, suggesting the presence of a chymase-like Ang II-generating activity in the sponge granulomas. Our results may suggest a potential ability of chymase to promote angiogenesis through the local chymase-dependent and angiotensin-converting enzyme-dependent Ang II generating system in pathophysiological angiogenesis.

Angiotensin I↗

PubMed2005

Cardiac chymase: pathophysiological role and therapeutic potential of chymase inhibitors.

On release from cardiac mast cells, alpha-chymase converts angiotensin I (Ang I) to Ang II. In addition to Ang II formation, alpha-chymase is capable of activating TGF-beta1 and IL-1beta, forming endothelins consisting of 31 amino acids, degrading endothelin-1, altering lipid metabolism, and degrading the extracellular matrix. Under physiological conditions the role of chymase in the mast cells of the heart is uncertain. In pathological situations, chymase may be secreted and have important effects on the heart. Thus, in animal models of cardiomyopathy, pressure overload, and myocardial infarction, there are increases in both chymase mRNA levels and chymase activity in the heart. In human diseased heart homogenates, alterations in chymase activity have also been reported. These findings have raised the possibility that inhibition of chymase may have a role in the therapy of cardiac disease. The selective chymase inhibitors developed to date include TY-51076, SUN-C8257, BCEAB, NK320, and TEI-E548. These have yet to be tested in humans, but promising results have been obtained in animal models of myocardial infarction, cardiomyopathy, and tachycardia-induced heart failure. It seems likely that orally active inhibitors of chymase could have a place in the treatment of cardiac diseases where injury-induced mast cell degranulation contributes to the pathology.

Animals↗

PubMed1999

[Pathophysiological roles of chymase and effects of chymase inhibitor].

Human chymase forms angiotenin (ANG) I to ANG II, whereas the roles of ANG II generated by chymase and the effects of chymase inhibitors have been unclear. On the other hand, rat chymase could not convert ANG I to ANG II. In isolated rat arteries, the ANG I-induced vascular contraction was completely suppressed by angiotensin-converting enzyme (ACE) inhibitor only. However, 30% of ANG I-induced vascular contraction in isolated human arteries was suppressed by an ACE inhibitor, but the remainder was blocked by chymostatin. In hamster hypertensive models, ANG II formation by ACE, but not by chymase, in vascular tissues plays an important role in maintaining hypertension. ANG II formation also induces vascular remodeling such as neointima formation. After balloon injury of vessels in dog, chymase and ACE activities were significantly increased in the injured vessels. In this model, an ANG II receptor antagonist was effective in preventing neointimal formation after balloon injury of vessels in dog, but an ACE inhibitor was ineffective. In dog grafted veins, the activities of chymase and ACEmin the grafted vein were significantly increased 15- and 2-fold, respectively, compared with those in the symmetrical veins. The intimal area of the grafted vein was reduced by a chymase inhibitor. Therefore, chymase-dependent ANG II formation plays an important role in the proliferative response, and chymase inhibitors may appear useful for preventing vascular proliferation.

Angiotensin I↗

PubMed2003

[Role of chymase in vascular diseases and the efficacy of chymase inhibitor].

In vascular tissues, angiotensin II is cleaved from angiotensin I by chymase and angiotensin converting enzyme (ACE). In the normal state, chymase is stored in mast cells and has no angiotensin II-forming activity, while chymase is activated immediately where mast cells have been activated by local stimuli. A clinical trial of an angiotensin receptor blocker (ARB) for preventing restenosis after percutaneous transluminal coronary angioplasty was successful, but that of an ACE inhibitor was not. After balloon injury in dog vessels, chymase activity was significantly increased in the injured artery, and a chymase inhibitor and an ARB were effective in preventing the vascular proliferation, but an ACE inhibitor was ineffective. In dog grafted veins, intimal area, chymase activity, and angiotensin II concentration were significantly increased after the operation, while they were significantly suppressed by a chymase inhibitor. However, the chymase inhibitor, unlike ACE inhibitor and ARB, did not affect blood pressure. These reports indicate that local angiotensin II production by chymase is involved only in the injured vessels. Therefore, a chymase inhibitor may be useful for preventing vascular disorders without affecting blood pressure.

Angioplasty, Balloon, Coronary↗

PubMed2002

Development of a chymase inhibitor: pharmacological characterization of a chymase inhibitor in inflamed tissue remodeling and fibrosis.

Chymase, a chymotrypsin-like serine protease, has not only alternative angiotensin II-generating activity but also various activities involving inflammatory responses. However, little is known of its contribution to physiological functions. Therefore, chymase inhibitors are thought to be potentially useful as tools for elucidating the physiological functions of chymase and therapeutic agents. Within the last five years, many patents on non-peptide chymase inhibitors have been published. We developed a potent non-peptide chymase inhibitor BCEAB (4-[1-[[bis-(4-methyl-phenyl)-methy]-carbamoyl]-3-(2-ethoxy-benzyl)-4-oxo-azetidine-2-yloxy]-benzoic acid) and examined its effect on inflamed tissue remodeling and fibrosis using a hamster sponge implant model. BCEAB has high inhibitory activity against human chymase but not against angiotensin-converting enzyme, elastase and tryptase. In the hamster sponge implant model, oral administration of BCEAB for 15 days dose-dependently suppressed both the dry weight of granuloma tissues in the sponge discs and the amounts of hydroxyproline in the tissues gradually increased during the experimental period. These results suggest that chymase, at least in part, participates in the growth of granuloma tissues of inflammatory regions by stimulating fibroblast growth and extracellular matrix collagen deposition. Chymase inhibitors for oral administration, such as BCEAB, might be useful for clarifying the pathophysiological roles of chymase in vivo.

Administration, Oral↗

PubMed2003

Hepatic chymase level in chronic hepatitis: co-localization of chymase with fibrosis.

Chymase, secreted by mast cells, is associated with angiotensin II production and fibrosis of myocardium and renal interstitium. Assuming that chymase also is involved in liver fibrosis, we previously established an enzyme-linked immunosorbent assay for chymase in human liver tissue. In the present study, we explored the localization of mast cells in the liver using antibodies against human chymase and also studied relationships between hepatic chymase level and histologic findings in 49 patients with chronic hepatitis. By the international classification, fibrosis was staged from F0 to F4 and activity was graded from A1 to A3. Cells immunoreactive for chymase were seen throughout portal areas and intralobular sinusoidal walls, largely colocalizing with inflammation and fibrosis. Hepatic chymase levels in F3 and F4 cases were greater than in F1 and F2 (F3+F4, 30.8+/-41.2 ng/mg vs. F1+F2, 5.7+/-6.6 ng/mg; P<0.01). Chymase in A3 casese (39.4+/-50.8 ng/mg) was more abundant than in A1 (3.7+/-4.3 ng/mg) or A2 (12.8+/-19.4 ng/mg); P<0.05 for each. Our findings suggest that hepatic chymase level is implicated in fibrosis and activity in human liver disease.

Journal Article↗

PubMed2006

Effects of chymase on the macular region in monkeys and porcine muller cells: probable involvement of chymase in the onset of idiopathic macular holes.

OBJECTIVES: To investigate chymase involvement in idiopathic macular hole onset, the effects of chymase on monkey eyes and cultured Muller cells were investigated. METHODS: Immunohistochemistry using antinestin and antiglial fibrillary acidic protein antibodies was performed in a normal monkey eye. After chymase was injected into the monkey vitreous, histological changes in the retina were evaluated using the TdT-mediated dUTP nick-end labeling (TUNEL) assay. Expression of c-kit, a stem cell factor receptor, and nestin was examined in porcine Muller cells cultured with basic fibroblast growth factor. The effects of chymase on proliferation and TUNEL staining in Muller cells were also examined. RESULTS: The number of nestin and glial fibrillary acidic protein-positive cells was higher in the macula than in other regions. Thickening of the posterior hyaloid membrane and some apoptotic cells were found in the macula of chymase-treated eyes. The expression of c-kit and nestin in Muller cells was shown and enhanced when cultured with basic fibroblast growth factor. Exposure to chymase inhibited Muller cell proliferation and produced TUNEL-positive cells. CONCLUSIONS: There might be Muller cells possessing atypical properties near the macular region and chymase might cause fibrosis and apoptosis through these cells. These findings suggest that increased chymase activity may result in idiopathic macular hole onset.

Animals↗

PubMed1999

Inhibitors of chymase as mast cell-stabilizing agents: contribution of chymase in the activation of human mast cells.

There has long been evidence that inhibitors of chymotryptic proteinases can inhibit the degranulation of rodent mast cells, but their actions on human mast cells and the contribution of mast cell chymase itself have received little attention. We investigated the ability of the selective chymase inhibitor Z-Ile-Glu-Pro-Phe-CO(2)Me and other proteinase inhibitors to inhibit chymase and cathepsin G activity, and we examined their potential to modulate the responsiveness of mast cells dispersed from human skin, lung, and tonsil tissues. IgE-dependent histamine release from skin mast cells was inhibited by up to about 80% after preincubation with Z-Ile-Glu-Pro-Phe- CO(2)Me (up to 0.1 microM), 70% with chymostatin (17 microM), and 60% with soybean trypsin inhibitor (0.5 microM). The mast cell-stabilizing properties of chymase inhibitors appeared to be greater for skin mast cells than for those from lung, whereas tonsil mast cells were relatively unresponsive. There were marked differences in the time course of responses to inhibitors, and the effect was dependent on the stimulus, with calcium ionophore-induced histamine release being unaffected. Incubation of dispersed skin, lung, or tonsil cells for up to 45 min with purified chymase failed to induce histamine release, although preincubation of cells with chymase was able to suppress IgE-dependent activation. Chymase could thus contribute to mast cell degranulation and after secretion could provide a feedback mechanism to limit this process. Nevertheless, inhibitors of chymase can be potent mast cell stabilizers, particularly in the skin.

Cathepsin G↗

PubMed2002

Suppression of basic fibroblast growth factor-induced angiogenesis by a specific chymase inhibitor, BCEAB, through the chymase-angiotensin-dependent pathway in hamster sponge granulomas.

1. We investigated the profound involvement of mast cell chymase, an alternative angiotensin II-generating enzyme, in angiogenesis using a specific chymase inhibitor. We also studied the functional profiles of this novel inhibitor in basic fibroblast growth factor (bFGF)-induced angiogenesis. 2. In this study, angiogenesis was induced by daily injections of bFGF (0.3 micro g site(-1) day(-1)), angiotensin I (2 nmol site(-1) day(-1)) or angiotensin II (2 nmol site(-1) day(-1)) into sponges implanted to male hamsters subcutaneously for 7 days. Angiogenesis in the granulation tissue surrounding sponges was evaluated by measuring the haemoglobin (Hb) content and local blood flow as the parameters for angiogenesis. 3. A chymase inhibitor, BCEAB (4-[1-[[bis-(4-methyl-phenyl)-methyl]-carbamoyl]-3-(2-ethoxy-benzyl)-4-oxo-azetidine-2-yloxy]-benzoic acid), was simultaneously administered into the implanted sponges (2 or 5 nmol site(-1) day(-1), for 7 days) treated with bFGF and strongly suppressed the haemoglobin contents in sponge granulomas. In the studies using a laser doppler perfusion imager, BCEAB (5 nmol site(-1) day(-1)) also attenuated the bFGF-induced increase of local blood flow around the implanted sponge granuloma. 4. In bFGF-induced angiogenesis, chymase activity in sponge granulomas was substantially increased. It was also confirmed that the chymase activity increased by bFGF was significantly and dose-dependently inhibited by BCEAB (2, 5 nmol site(-1) day(-1)). 5. BCEAB inhibited the Hb contents and the expression of vascular endothelial growth factor (VEGF) mRNA induced by angiotensin I but not by angiotensin II. 6. These results suggest that the significance of chymase in bFGF-induced angiogenesis was confirmed, and a novel inhibitor, BCEAB, strongly suppresses the bFGF-induced angiogenesis through the chymase-angiotensin II-VEGF dependent pathway.

Angiotensins↗

PubMedDate not supplied

Measurement of activities in two different angiotensin II generating systems, chymase and angiotensin-converting enzyme, in the vitreous fluid of vitreoretinal diseases: a possible involvement of chymase in the pathogenesis of macular hole patients.

PURPOSE: To investigate possible involvement of chymase and angiotensin-converting enzyme (ACE) in the pathogenesis of vitreoretinal diseases, both of which are related to the production of angiotensin II. METHODS: We measured chymase and ACE activities in the vitreous in the 54 affected eyes of 54 patients who had undergone vitreous surgery for idiopathic macular holes (MH, n = 14), proliferative diabetic retinopathy (PDR, n = 14), idiopathic epiretinal membranes (ERM, n = 13), and rhegmatogenous retinal detachment (RRD, n = 13). RESULTS: Chymase activities in the vitreous from patients with MH, PDR, ERM, and RRD were 1.87 +/- 0.53, 0.06 +/- 0.04, 0.40 +/- 0.12, and 0.08 +/- 0.03 (mean +/- SE) mU/mg protein, respectively, and ACE activities in the vitreous humor were 0.18 +/- 0.09, 0.30 +/- 0.07, 0.01 +/- 0.01, and 0.03 +/- 0.02 (mean +/- SE) mU/mg protein, respectively. Chymase activity was significantly elevated in MH among these diseases (p < 0.01, Scheffe), and ACE was significantly activated in PDR compared to ERM and RRD (p < 0.05, Scheffe). CONCLUSIONS: Our results suggest that two different angiotensin II generating systems are activated in human vitreous humor; an increased activity of chymase may play a possible role in the formation of macular holes.

Aged↗

PubMed2001

An orally active chymase inhibitor, BCEAB, suppresses heart chymase activity in the hamster.

We investigated the effects of a novel chymase inhibitor, BCEAB (4-[1-[[bis-(4-methyl-phenyl)-methyl]-carbamoyl]-3-(2-ethoxy-benzyl)-4-oxo-azetidine-2-yloxy]-benzoic acid). The IC50 value of BCEAB for purified human chymase was 5.4 nM, whereas BCEAB did not inhibit the angiotensin-converting enzyme, elastase and tryptase. In isolated dog arteries, the IC50 value of BCEAB for the angiotensin I-induced contraction in the presence of 1 microM lisinopril was 2.8 microM. In the hamster, the heart chymase activities were significantly suppressed to 42.0% and 26.9% 3 h after oral administration of 100 and 300 mg of BCEAB/kg of body weight, respectively. In conclusion, BCEAB is a useful chymase inhibitor for studying the role of chymase in vivo.

Angiotensin II↗

PubMed1996

Fixation with Carnoy's fluid reduces the number of chymase-positive mast cells: not all chymase-positive mast cells are also positive for tryptase.

Mast cells in the nasal mucosa can be studied by means of monoclonal antibodies (mAb) against tryptase (T+MC) and chymase (C+MC). Fixation with acetone gives more positive cells than does fixation with Carnoy's fluid. In frozen biopsy specimens of allergic nasal mucosa fixed with acetone, the number of T+MC equals that of C+MC. When fixed with Carnoy's fluid, however, the number of T+MC is larger than the number of C+MC. The decrease in both T+MC and C+MC resulting from fixation with Carnoy's fluid is time-related and depends on the type of mAb used. Carnoy fixation time gives a decrease in the number of C+MC within 1 min, whereas the number of T+MC decreases only after 10 min. Within 1 min, the number of C+MC decreases to a level where continued fixation no longer gives further decreases in the number of cells. Two populations of mast cells can be distinguished here: one sensitive and the other insensitive to Carnoy's fluid. When double-staining is used, fixation with acetone gives three populations of mast cells: one positive for tryptase (T+C-MC), another positive for tryptase and chymase (T+C+MC), and a third one positive for chymase (T-C+MC). These three populations were found in lymph node, spleen, thymus, dermis, lung parenchyma, small intestinal submucosa, and nasal mucosa.

Acetic Acid↗

PubMed1996

Cloning of the cDNAs for mast-cell chymases from the jejunum of Mongolian gerbils, Meriones unguiculatus, and their sequence similarities with chymases expressed in the connective-tissue mast cells of mice and rats.

By using the combination of reverse-transcription CR and rapid amplification of cDNA ends methods, two distinct cDNAs encoding mast-cell proteases (chymases; MCPs), designated as gMCP-1 and -2, were successfully cloned and sequenced from the jejunum of Mongolian gerbil, Meriones unguiculatus, infected with Nippostrongylus brasiliensis. On the basis of a comparison of the deduced amino acid sequences with those of known rodent mast-cell chymases, gMCP-1 was found to be highly similar to mouse mast-cell protease (mMCP)-4 and rat mast-cell protease (rMCP)-1, while gMCP-2 was similar to mMCP-5 and rMCP-3. Alghough mMCP-4 and -5 and rMCP-1 and -3 were restrictedly or mainly expressed in connective-tissue mast cells and serosal mast cells, the gMCP-1 and -2 genes were mainly transcribed in the jejunal mucosa and to a lesser extent in the skin and tongue. Moreover, kinetic study after infection revealed that the amounts of the gMCP-1 and -2 mRNAs in jejunum paralleled well the degree of intestinal mastocytosis. The expression of gMCP-1 and -2 in mucosal mast cells of gerbil jejunum was also confirmed by in situ hybridization. Since a tryptase, another type of MCP, was also expressed in mucosal mast cells of gerbils but not in those of mice and rats, the expression of MPCs in mucosal mast cells of gerbils is different from those of mice and rats. The Mongolian gerbil would be a useful model with which to investigate the physiopathological role of MCPs.

Amino Acid Sequence↗