Detection Of Mast Cells Research Articles

Absence of protective role of afferent nerves in early intestinal mucosal alterations induced by abdominal irradiation in rats

Purpose : To assess the early effects of primary afferent nerve suppression by systemic treatment with the neurotoxin capsaicin in an acute model of abdominal irradiation in rats (10 Gy, gamma). Materials and methods : Changes in myeloperoxidase (MPO) activity, calcitonin gene-related peptide (CGRP) tissue content, number of mast cells and apoptotic cells were determined in jejunum and ileum in four groups of rat male Wistar (vehicle sham-irradiated, vehicle irradiated, capsaicin sham-irradiated and capsaicin irradiated) at 1 and 3 days post-irradiation. Results : In vehicle irradiated rats, CGRP was significantly increased from the first day after irradiation in jejunal mucosa; MPO activity increased in both segments at day 3 but not at day 1 after irradiation; the number of detectable mucosal mast cells dropped to nearly zero on days 1 and 3, while the apoptotic cells in the intestinal mucosa were significantly increased at day 1. Similar results were obtained for mast cells and apoptosis in capsaicin irradiated rats as compared to capsaicin sham-irradiated rats, while MPO activity was significantly increased and CGRP concentration in jejunal mucosa significantly decreased from the first day in these rats in comparison with capsaicin sham-irradiated rats. Conclusions : Intestinal sensory innervation seems not to have a major protective role against a radiation-induced intestinal inflammatory reaction.

Release of Mast Cell Tryptase from Human Colorectal Mucosa in Inflammatory Bowel Disease

Background: Histologic detection of mast cells cannot adequately reflect their function and state of activation, since degranulated mast cells may escape from histologic assessment. To better define the role of mast cells in inflammatory bowel disease, the spontaneous secretion of mast cell tryptase, a highly mast cell specific protease, was measured from colorectal samples. Methods: After detection of the initial basal tryptase release, gut mucosal samples were incubated in a modified Hanks/RPMI medium using a mucosa oxygenation system. Spontaneous tryptase secretion from 153 viable samples of 22 controls, 30 patients with Crohn disease (CD) and 19 with ulcerative colitis (UC) was followed over 4 h. Tryptase was measured by radioimmunoassay. Results: The rates of the initial basal tryptase release revealed that mast cell activation occurs during active inflammation in CD and UC. While the time course of tryptase release was similar in all three groups, spontaneous tryptase secretion (over 4 h) was found to be significantly enhanced and prolonged only in UC (P < 0.01 compared to controls), but not in CD. Conclusions: This study provides clear evidence from viable endoscopic colorectal samples that mast cell mediators were secreted during active inflammation in CD and UC. However, the extent of mast cell involvement and activation differs considerably between CD and UC. Significantly increased rates of tryptase secretion were found both in non-inflamed and inflamed tissue of UC, indicating that mast cell involvement is a typical feature of UC.

A role for the hepatobiliary system in IgE-mediated intestinal inflammation in the rat.

For many years the central focus of research into gastrointestinal hypersensitivity reactions has been the mast cell population of the intestinal lamina propria. Since bile is known to deliver immunological mediators to the gastrointestinal tract, the possibility arises that extra-intestinal populations of mast cells may also contribute to IgE-mediated intestinal damage. To characterize hepatic mast cells in the rat and to investigate the role of the hepatobiliary system in a model of IgE-mediated reactivity to dietary antigen. Wistar rats were passively sensitized with monoclonal antidinitrophenyl (DNP) IgE antibodies, and were later challenged orogastrically with DNP-HSA. Additional animals were sensitized, then bile duct-cannulated prior to antigen challenge. At various time points, liver and intestinal samples were collected for histological examination, and bile was collected and assayed for histamine and TNFalpha. Hepatic mast cells display a mucosal mast cell-like phenotype, and are closely associated with the vessels of the portal triads. Orogastric antigen challenge led to a rapid and significant decline (P<0.0001) in detectable mast cells as a result of anaphylactic degranulation. The median number of granulated mast cells associated with each portal triad in liver sections declined from six per portal triad to one per portal triad post-antigen challenge. After 15 min, biliary histamine concentrations rose above background levels (P<0.01). TNFalpha was also detectable in the majority (4/6) of bile samples within 15 min of challenge. Histological examination of the gastrointestinal mucosa revealed disruption to the villous epithelium ranging from oedematous changes to gross destruction. Such damage was not seen in animals in which bile had been externally drained. The data indicate that biliary products are major contributors to the gastrointestinal damage arising from IgE-mediated hypersensitivity reactions in the rat, and such hypersensitivity reactions may involve a population of mast cells which reside in the liver.

3] Ruthenium red as measure of mast cell activation

This chapter discusses Ruthenium red as measure of mast cell activation. Mast cells are found closely apposed to the vasculature in essentially all tissues, and as they are exquisitely sensitive to very subtle changes in the surrounding milieu, they may function as sentinels for potential disturbances within any given organ. Once activated, they are capable of releasing a myriad of inflammatory mediators, including preformed cytokines. For these reasons monitoring mast cell degranulation within tissues gives us clues about the potential involvement of these cells in trauma, infection, and inappropriate inflammation. A recently developed assay (ruthenium red staining) that measures mast cell activation online in a quantitative manner has been described in the chapter. The use of ruthenium red is already a quantitative and extremely sensitive approach to detect mast cell activation. Moreover, the assay may be useful as a screening process both for mast cell inhibitory compounds and for mediators that activate mast cells. However, application to other species and other tissues will be needed to further evaluate the usefulness of this technique in vivo.

Is radiation-induced degranulation of mast cells in salivary glands induced by substance P?

Although DNA is the critical target for the lethal effects of irradiation, the precise mechanisms by which irradiation causes damage in tissues and biological systems is not fully understood. In the present study, the number of mast cells and the expression of the neuropeptide substance P (SP) in salivary glands were examined 10 days after a regimen of irradiation. The irradiation was given as a single dose or 5 consecutive days with daily doses of 7 Gy up to a total dose of 35 Gy. In addition, the number of mast cells and the expression of SP were examined 2 and 24 h after a single dose of 7 Gy. Immunohistochemical staining for 5-hydroxytryptamine (5-HT) and staining with avidin peroxidase and toluidine blue were used to detect mast cells. At examination 2 and 24 h after irradiation treatment, no change in the number of mast cells and the pattern of SP expression was observed. Ten days after irradiation there was a remarkable reduction in the number of mast cells in all the three glands, but there was a marked increase in the number of nerve fibers showing SP-like immunoreactivity in the parenchyme. The results show that early time-dependent alterations in the density of mast cells occur in response to irradiation, and that these changes occur concomitantly with changes in the expression of SP. Since the peripheral nervous system is a main regulator of salivary gland function, it is tempting to speculate that the nervous system interacts with mast cells via SP in modulating irradiation provoked tissue responses in salivary glands.

Rapid mast cell activation causes leukocyte-dependent and -independent permeability alterations.

The major objective of this study was to systematically elucidate the mechanisms underlying microvascular permeability in rat mesenteric venules after the activation of perivascular mast cells. Intravital microscopy was used to assess polymorphonuclear leukocyte (PMN) infiltration and microvascular permeability alterations in single 25- to 40-micron diameter venules. Ruthenium red was used to detect mast cell activation on-line. Exposure of mast cells to compound 48/80 (CMP 48/80) caused a rapid mast cell activation and increase in microvascular permeability (within 15 min) that was maintained for the duration of the experiment. CMP 48/80 also increased PMN adhesion to the microvascular endothelium. Anti-PMN serum, as well as various antiadhesion therapies, including CL26 (anti-CD18 antibody) and fucoidan (selectin-immunoneutralizing carbohydrate), revealed that the early microvascular permeability was PMN independent. However, these regimens significantly reduced plasma protein leakage out of venules between 30 and 60 min. Methysergide (serotonin receptor antagonist), but not diphenhydramine (histamine receptor antagonist), inhibited the early PMN-independent microvascular permeability. Finally, a platelet-activating factor (PAF)-receptor antagonist did not affect the early phase of microvascular permeability but reversed the later phase, consistent with PAF's role as a proadhesive molecule for PMN during mast cell activation. These data demonstrate that, within the first hour of mast cell activation, a biphasic PMN-independent and -dependent response in microvascular permeability is observed. The data also raise the possibility that histamine's physiological role in this model may be unrelated to alterations in microvascular permeability.

2 MUCOSAL MAST CELLS DISTINGUISH ALLERGIC FROM GASTROESOPHAGEAL REFLUX-INDUCED ESOPHAGITIS.

BACKGROUND: Recent evidence suggests that allergy may play a role in a subgroup of infants and children with characteristic eosinophilic esophagitis. These patients may be indistinguishable from patients with esophagitis caused by gastroesophageal reflux (GER), both clinically and histologically. We have previously demonstrated increased mucosal mast cells (MMC) in biopsies from patients with esophagitis vs controls, with a small subgroup showing a marked increase in MMC. AIM: To investigate MMC in patients with allergic eosinophilic esophagitis vs GER-induced esophagitis and controls. METHODS: Formalin-fixed paraffin embedded esophageal tissue was obtained from archival material. Eosinophils and MMC were stained with a dual mast cell/eosinophil stain (astra blue/vital new red); immunohistochemistry for MMC was performed with anti-mast cell tryptase antibodies (Dako) and an avidin-biotin detection system. Biopsies were reviewed blindly (A.R.); MMC and eosinophils per hpf were quantitated. Tryptase was more sensitive at detecting mast cells than dual staining therefore tryptase was used to enumerate MMC. Control patients had normal pH probes and were histologically normal. GER/esophagitis was defined by histologic inflammation with an abnormal pH probe (> 5% time with pH<4). Allergic esophagitis was defined by eosinophilic esophagitis with a normal pH probe or concomitant eosinophilic inflammation in antral or duodenal biopsies. RESULTS: MMC were significantly increased in allergic esophagitis vs GER induced esophagitis and control tissue. Eosinophils paralleled mast cell increases, with considerable overlap. Table CONCLUSION: The recognition of increased MMC allows one to distinguish allergy-induced from GER-induced esophagitis. Mast cell staining should be part of the histopathologic evaluation of select cases of esophagitis in infants and children when allergy is being considered.

Mast cell TNF mRNA expression in nasal mucosa demonstrated by in situ hybridization: a comparison of mast cell detection methods

We have used 35S-labelled RNA probes to detect TNF cytokine gene expression in nasal mucosa derived from patients with perennial rhinitis. As mast cells comprise a minor component of the total cell population in nasal mucosa, additional methods are needed to determine whether mast cells contribute to the cytokine mRNA detected by in situ hybridization. We have combined in situ hybridization with alternate methods to detect mast cells (tryptase immunostaining or toluidine blue staining) and determined that in situ hybridization coupled with tryptase immunostaining provides optimal methods to detect mast cell cytokine gene expression in tissue sections. Using in situ hybridization and tryptase immunostaining, we demonstrate that mast cells in nasal mucosa can express TNF mRNA. However, the number of tryptase −, TNF + cells (1.99 ± 1.59 cells/mm 2) exceeded the number of tryptase +, TNF + mast cells (0.09 ± 0.21 cells/mm 2). Mast cells thus comprised a subpopulation of the total number of TNF mRNA positive cells in nasal mucosa.

Are mast cells involved in hypertensive heart disease?

To evaluate the potential relationship of mast cells with myocardial fibrosis in the cardiac ventricles of spontaneously hypertensive rats (SHR). Experiments were performed on hearts from 36-week-old SHR with established left ventricular hypertrophy (n = 12) and from 36-week-old normotensive Wistar-Kyoto (WKY) rats (n = 12). Furthermore, to evaluate whether antihypertensive treatment with the angiotensin converting enzyme inhibitor quinapril interferes with the potential relationship between mast cells and fibrosis in SHR, we treated 16-week-old SHR (n = 12) with oral quinapril (10 mg/kg body weight per day) for 20 weeks. Mast cells were counted in 25 high-power fields. Toluidine blue-stained sections and avidin staining were used to detect mast cells. The extent of myocardial fibrosis was analysed in samples stained with Masson's trichrome. The amount of collagen was evaluated morphometrically, using an automatic image analyser, and biochemically, using myocardial hydroxyproline concentration. In the left ventricle of untreated SHR compared with age- and sex-matched normotensive WKY rats we found more extensive interstitial and perivascular fibrosis, an increased collagen volume fraction, an increased hydroxyproline concentration and an increased number of mast cells. Similar but less intense abnormalities were observed in the right ventricles of untreated SHR compared with the left ventricles of the same rats. In the left ventricles of quinapril-treated SHR compared with those of untreated SHR we found a marked decrease in fibrosis, a lower collagen volume fraction, a lower hydroxyproline concentration and fewer mast cells. Treatment with quinapril was also accompanied by normalization in the myocardial structure of the right ventricles of SHR. A positive correlation was found between the density of mast cells and the collagen volume fraction in the left ventricles of all of the rats. The present findings suggest that mast cells can play a part in the development of the myocardial fibrosis that occurs in the cardiac ventricles with hypertensive cardiac hypertrophy. In addition, the present results suggest that the ability of quinapril to interfere with mast cells might be involved in its cardioreparative properties.

Reproductive Behavior, Endocrine State, and the Distribution of GnRH-like Immunoreactive Mast Cells in Dove Brain

Previous studies indicate that there is an increase in the number of detectable mast cells expressing gonadotropin-releasing hormone-like immunoreactivity (GnRH-ir) in the medial habenular region of the brain in ring doves after a period of 2 hr of courtship. In the present experiment the overall distribution of GnRH-ir mast cells in the brain is described, and the influence of behavioral and endocrine state on brain mast cell distribution and detectable mast cell number is explored. Four groups of male birds were studied: (1) paired with a female and displaying courtship behavior, (2) paired with a squab and displaying aggressive behavior, (3) visually isolated from other doves, and (4) long-term castrates housed communally. In all four groups, GnRH-ir mast cells were observed in the following areas: the medial habenula, circumventricular organs, organum vasculosum lamina terminalis and organum subseptale, the pia mater, and blood vessels. Cell counting revealed differences among groups in numbers of habenular mast cells. Courted doves had the most habenular mast cells, followed by doves paired with squabs, and then visually isolated doves, while castrates had the fewest. One-way ANOVA indicates significant differences among groups. All pairwise comparisons show that the courting animals had significantly more detectable mast cells in the habenular area than did visually isolated and castrated groups. These results suggest that the appearance and/or detectability of GnRH-ir mast cells in the habenula is related to the behavioral state and possibly to the endocrine state of the animal and suggest a novel mechanism for interactions among the nervous, endocrine, and immune systems.

Association between mast cells and the development of experimental autoimmune uveitis in different rat strains

To study the role of anterior uveal mast cells in experimental autoimmune uveitis (EAU), the mast cells in the iris and ciliary body of Lewis rats, Brown Norway (BN) rats, and their F1 hybrids (LBNF1) were quantitated in normal rats and during the induction period of EAU. The mean baseline mast cell number was 68.9 ± 10.8 per anterior uvea for Lewis rats, 0.3 ± 0.2 for BN rats, and 4.6 ± 0.6 for LBNF1 rats. Detectable mast cells in the anterior uvea of S-Ag-immunized Lewis rats decreased to 60% of control at 6 days postimmunization, recovered to 80% at 10 days, and dropped again to 16% at 13 days, with disease onset around 14 days. In Lewis rats that were adoptively transferred with a uveitogenic T-lymphocyte line, a profound drop in anterior uveal mast cell numbers occurred in the eyes with early signs of EAU, 3 days after the transfer. The decrease in detectable mast cells is consistent with mast cell degranulation. The data suggest that anterior mast cells participate in the immunopathogenesis of EAU and may influence the genetic susceptibility to EAU.

Resident mast cells are the main initiators of anaphylactic leukotriene production in the liver.

During anaphylaxis the sensitized liver can have substantial capacity for leukotriene production. However, the intrahepatic cellular source for these potent eicosanoid mediators has been unclear so far. We therefore analyzed the appropriate role of resident liver cells in organ-specific generation of leukotrienes by immunohistochemical localization of 5-lipoxygenase, by measurement of cysteinyl leukotriene production in animals or isolated livers and by histochemical monitoring of mast cells in rat, guinea pig and mouse livers, respectively. During anaphylaxis in vivo, these species all generated large amounts of leukotrienes. Immunohistochemistry with rat liver demonstrated resident mast cells as the predominant cell type in liver containing 5-lipoxygenase. Rat and guinea pig livers contained numerous mast cells and produced substantial amounts of leukotrienes on antigen challenge; in contrast, mouse livers neither showed detectable mast cells nor generated leukotrienes when stimulated analogously. Infusion of histamine or serotonin (1 mmol/L each) or of the degranulating substance P (8 mumol/L) did not elicit leukotriene generation in rat livers. Furthermore, substantial degranulation of liver mast cells by compound 48/80 (0.5 mg/kg body mass) was paralleled by only modest leukotriene formation (63 +/- 10 pmol in bile/kg body mass/30 min). These results indicate that during anaphylaxis mast cells are the main intrahepatic cells initiating leukotriene production and that such leukotriene generation is likely to be independent of mast cell degranulation or the release of histamine or serotonin.

Dynamics of mast cells in the nasal mucosa of patients with allergic rhinitis and non-allergic controls: a biopsy study.

Mast cell degranulation is thought to be an important component of the pathogenesis of allergic rhinitis. Quantitative studies on mast cells in nasal mucosa after allergen exposure have given widely divergent results, ranging from an overall decrease via redistribution to an overall increase. We investigated this problem by employing a combination of anti-IgE and toluidine blue staining of biopsy specimens. In allergic patients anti-IgE was found to identify all mast cells and toluidine blue to detect mast cells that were not (totally) degranulated. The study was composed of two parts done in different patient groups. In the first part of the study biopsies were performed in 23 patients with isolated grass-pollen allergy, once during natural provocation in the summer and once in the winter. Biopsies were also performed in 12 controls. Non-allergic controls were found to have the same number of mast cells in the lamina propria as asymptomatic allergic patients. The controls seldom have mast cells in the epithelium. The patients with isolated grass-pollen allergy showed an increase in the numbers of mast cells in the lamina propria during natural provocation and the same seemed to occur in the epithelium as well. During natural provocation almost all of the mast cells in the epithelium and half of those in the lamina propria were degranulated. In the second part of the study 17 patients with isolated grass-pollen allergy and four controls were challenged daily with allergen extract during a 2-week period in the winter. During this period biopsies were performed at eight different occasions, i.e. once before, six occasions during and once after the provocation period. The results of this part of the study showed that during provocation mast cells migrate to the surface of the nasal mucosa, where they become degranulated, and that the pool of mast cells in the lamina propria was apparently replenished by migration of mast cells from the vessels in the lamina propria. The total number of mast cells in the lamina propria remained approximately the same while the mast cells residing in an increasingly thick layer measured from the basal membrane into the lamina propria became degranulated. After 2 weeks, 82% of the mast cells in the lamina propria was degranulated and it was only in the deepest layers that some toluidine blue positive cells were found.(ABSTRACT TRUNCATED AT 400 WORDS)

An Analysis of Mast Cell Frequency in the Rodent Nervous System: Numbers Vary between Different Strains and Can Be Reconstituted in Mast Cell-Deficient Mice

There is evidence that nervous system mast cells may play a role in the pathogenesis of the experimental autoimmune demyelinating diseases, experimental allergic neuritis (EAN), and experimental allergic encephalomyelitis (EAE). We compared mast cell numbers in the peripheral nervous system (PNS) and central nervous system (CNS) of rodent strains that differed in their susceptibility to experimental demyelination. Mast cells were counted by toluidine blue staining of formalin-fixed tissue. Normal Lewis rats (susceptible to both EAN and EAE) had significantly greater numbers of mast cells in the dura mater (about 6x) of the meninges and the sciatic nerve (3x) than Brown Norway rats (resistant to EAE and EAN induction under normal circumstances). Similarly SJL/J mice (susceptible to EAE and EAN) had significantly greater numbers of CNS (3x) and PNS (8x) mast cells than C3H mice (more resistant to disease induction). Other mouse strains were also examined, and PNS mutant Trembler mice had high numbers of PNS mast cells, while the mast cell deficient W/Wv mice contained no detectable mast cells in either the CNS or PNS. Reconstitution of W/Wv mast cells was accomplished by intravenous injection of bone marrow cells from congenic littermates. After seven months, mast cells could be seen in both the CNS and PNS of reconstituted animals. The possibility that mast cells and mast cell precursors can migrate into the nervous system of animals, in the absence of inflammatory disease, may have implications for their role in the pathogenesis of experimental demyelinating diseases.

Attenuation of experimental autoimmune encephalomyelitis by compound 48/80 in Lewis rats

The appearance of increased levels of histamine in the central nervous system (CNS) concomitant with the development of clinically significant acute experimental autoimmune encephalomyelitis (EAE) in male Lewis rats suggests that CNS-associated mast cells may mediate acute EAE in Lewis rats. We now report that, compared to controls, rats with acute EAE exhibit fewer detectable mast cells in their dura mater and velum interpositum. In addition, intracisternal, but not intraperitoneal administration of Compound 48/80 just prior to the appearance of clinical signs of acute or recurrent EAE in male and female rats, respectively, significantly attenuates the clinical severity of both forms of EAE. These results further support the hypothesis that CNS-associated, but not peripheral mast cells are mediators or modulators of acute and recurrent EAE in Lewis rats.

Clinico-pathological study on denture stomatitis.

The clinical and pathological study was performed in order to determine the histopathological and cytoimmunological characteristics of denture stomatitis. All specimens were biopsy materials from seventeen patients with denture stomatitis. Normal palatal mucosae from ten patients served as the control. In addition to the usual staining methods, naphtol AS-D chloroacetate esterase stain and peroxidase-antiperoxidase method were used to detect mast cells and plasma cells. Denture stomatitis could be divided into atrophic and hyperplastic types. The former showed a smooth and atrophic mucosa. The latter showed a large number of exophytic projections which were composed of marked acanthosis and submucosal fibrosis, and was further subdivided into granular and papillary subtype according to the size of projections. In the present study, there were six cases of the atrophic type, and eleven cases of the hyperplastic type (consisting of seven granular and four papillary subtypes). The hyperplastic type was more frequently observed in patients with partial dentures compared with complete dentures and was associated frequently with ill fitting of the denture base as well as agglutination of denture plaque. Cytoimmunological study revealed that there was a pronounced increase of plasma cells, especially IgG- and IgA-producing cells, and a moderate increase of lymphocytes as well as mast cells in both types of denture stomatitis. Mast cells were always noted in the area with marked plasma cell infiltration, suggesting an intimate relation between both cells. These findings suggest that the immunological reactions play some role in the pathogenesis of denture stomatitis.

Dermal cellular responses of helminth-free and Ostertagia ostertagi-infected calves to intradermal injections of soluble extracts from O. ostertagi L 3 larvae

Twelve calves were raised helminth-free until 9 weeks of age when six were orally inoculated with 100 000 Ostertagia ostertagi infective stage larvae (L 3). Three uninfected and three experimentally infected calves received intradermal injections of sterile saline and soluble larval extract (SLE) from O. ostertagi L 3 with a protein concentration ranging from 1 to 200 μg ml −1. Biopsies were performed 48 h post-injection. A kinetic study was performed on the remaining six calves, three infected and three uninfected, using a 100 μg ml −1 concentration of SLE and taking biopsies 1, 4, 8, 12, 24, and 72 h post-injection at both the saline and SLE-injected sites. All calves had an immediate wheal and increase in skin thickness at the SLE-injected sites. The numbers of eosinophils infiltrating SLE-injected sites as comparred to saline-injected sites were significant in both uninfected and infected calves, but the infected calves had significant numbers to a wider range of SLE concentrations and had significantly higher numbers than uninfected calves in the kinetic study. Infected calves also had significant numbers of basophils in the dose response study at concentrations of 5 and 100 μg ml −1 SLE. Neutrophil infiltration was similar in both groups and was significant at SLE-injected sites early in the kinetic study. Detectable mast cells were decreased in SLE-injected sites of infected animals and perivascular accumulation of mononuclear and some polymorphonuclear cells was observed in the deep dermis of infected animals.

Reevaluation of reserpine-induced suppression of contact sensitivity. Evidence that reserpine interferes with T lymphocyte function independently of an effect on mast cells.

It has been suggested that reserpine blocks expression of delayed hypersensitivity (DH) by depleting tissue mast cells of serotonin (5-HT), thereby preventing a T cell-dependent release of mast cell 5-HT necessary to localize and to amplify the DH response. However, reserpine blocks expression of DH in mast cell-deficient mice. We therefore decided to reevaluate the mechanism by which reserpine abrogates expression of cellular immunity, and investigated whether the drug might interfere with T cell activity in vitro or in vivo. At concentrations as low as 4 microM, reserpine profoundly suppressed baseline or antigen-augmented levels of [3H]thymidine incorporation by immune lymph node cells obtained from mice sensitized to the contactant oxazolone [I-LNC(Ox)]. This effect was observed both with I-LNC derived from normal mice and with I-LNC derived from congenitally mast cell-deficient W/Wv mice, cell preparations that lacked detectable mast cells, histamine, and 5-HT. Furthermore, treatment of I-LNC with reserpine (20 microM) for 1 h in vitro virtually abolished the ability of these cells to transfer CS to naive mice. This was not a cytolytic effect, as the viability of the I-LNC treated with reserpine was not affected, and washing of the reserpine-treated I-LNC before transfer fully restored their ability to orchestrate a CS response. The action of the drug was not mediated by an effect on mast cells, since the experiment could be performed using mast cell-deficient W/Wv mice as both donors and recipients of I-LNC. In addition, the effect was specific for the treated cells: mice that received reserpine-treated I-LNC(Ox) intravenously together with untreated I-LNC(DNFB) did not develop CS to Ox but responded normally to DNFB; and local intradermal injection of reserpine-treated I-LNC(Ox) which failed to transfer reactivity to Ox, did not interfere with the development of CS to DNFB at the same site. Finally, cotransfer experiments indicated that the effect of reserpine on the transfer of CS was not due to activation of suppressor cells. Our findings strongly suggest that whatever effects reserpine might have on immunologically nonspecific host cells, the drug's effects on sensitized T cells are sufficient to explain its ability to block cell-mediated immune responses in vivo.

Quantitation of mast cells in experimental allergic neuritis.

The number and extent of degranulation of mast cells in the sciatic nerve have been determined in Lewis rats inoculated with bovine intradural root myelin to induce experimental allergic neuritis (EAN). No significant change in either the number of mast cells or the percentage of degranulated cells was observed before eight days post-inoculation (dpi). On day eight, however, there was a significant decrease in the number of detectable mast cells, and on day nine there was a significant increase in the percentage of degranulated cells. These changes immediately preceded the onset of clinical signs of EAN that occurred ten dpi. The number of detectable mast cells remained low throughout the acute stage of the disease, but by three weeks post-inoculation (wpi) there was a significant increase in the number of mast cells (or basophils) that persisted through 16 wpi. The increase in the percentage of degranulated mast cells also persisted through 16 wpi. These results suggest that mast cells participate in the development of EAN in the Lewis rat.

Microvascular heparin-like species with anticoagulant activity.

Calf microvasculature was isolated from retina and cerebral gray matter. These preparations contained 0.048-0.060 U of heparin-like anticoagulant activity per gram of wet tissue. The retinal microvascular material contained no detectable mast cells. The anticoagulant potency of this product was associated solely with endothelial cells. This property appears to be due to a heparinlike proteoglycan since molecular species with biologic activity are precipitated with 10% (wt/vol) trichloroacetic acid and are destroyed by incubation with Flavobacterium heparinase. Furthermore, the above component functions in a manner virtually identical to heparin since approximately 60% of these species with anticoagulant activity bind to antithrombin-concanavalin A-Sepharose 4B, and only 15% of their biologic potency is expressed in the presence of antithrombin modified near the mucopolysaccharide binding domain. The cerebral microvascular tissue contained a trace subpopulation of mast cells (approximately 0.3%). The anticoagulant activity of this preparation is most probably associated with both endothelial cells and mast cells. However, complete separation of these two cellular elements has proven difficult with current methodology.