Eosinophil granule major basic protein 1 deposition in eosinophilic esophagitis correlates with symptoms independent of eosinophil counts.

In patients with eosinophilic esophagitis (EoE), symptoms often do not correlate with peak eosinophil counts (PEC) determined on histopathological examination of biopsy specimens. This may be because eosinophils degranulate during active disease and lose their morphological identity as intact cells and, therefore, are not enumerated on microscopic examination. Eosinophil granule proteins that are released into tissues with degranulation, including major basic protein 1 (eMBP1), likely contribute to disease pathogenesis and, therefore, may correlate with symptoms better than PEC. We sought to determine whether symptoms in patients with EoE more closely relate to eosinophil granule protein deposition than to eosinophil enumeration, especially in patients with fewer than 15 eosinophils per high power field (HPF). Esophageal biopsy specimens from 34 patients diagnosed with EoE were obtained for histopathological examination and for evaluation of eMBP1 staining by indirect immunofluorescence. PEC by histopathology were compared to extracellular eMBP1 grades by immunostaining. PEC and eMBP1 grades also were analyzed for their relationship to symptoms and clinical course. Biopsy specimens from 19 of the 34 patients had fewer than 15 PEC on histopathological examination, and the other 15 patients had 15 or greater PEC. Positive eMBP1 immunostaining was found in all symptomatic patients. EoE symptoms were related to eMBP1 immunostaining grades (p = 0.0001), but not PEC (P = 0.14). Eosinophil granule protein deposition, specifically eMBP1, is increased in esophageal biopsy specimens from symptomatic patients with EoE and may be a marker of disease activity, including patients with EoE who have 'resolved' disease.

Repeat treatment of acute hereditary angioedema attacks with open-label icatibant in the FAST-1 trial.

Hereditary angioedema (HAE) is characterized by potentially life-threatening recurrent episodes of oedema. The open-label extension (OLE) phase of the For Angioedema Subcutaneous Treatment (FAST)-1 trial (NCT00097695) evaluated the efficacy and safety of repeated icatibant exposure in adults with multiple HAE attacks. Following completion of the randomized, controlled phase, patients could receive open-label icatibant (30 mg subcutaneously) for subsequent attacks. The primary end-point was time to onset of primary symptom relief, as assessed by visual analogue scale (VAS). Descriptive statistics were reported for cutaneous/abdominal attacks 1-10 treated in the OLE phase and individual laryngeal attacks. Post-hoc analyses were conducted in patients with ≥ 5 attacks across the controlled and OLE phases. Safety was evaluated throughout. During the OLE phase, 72 patients received icatibant for 340 attacks. For cutaneous/abdominal attacks 1-10, the median time to onset of primary symptom relief was 1·0-2·0 h. For laryngeal attacks 1-12, patient-assessed median time to initial symptom improvement was 0·3-1·2 h. Post-hoc analyses showed the time to onset of symptom relief based on composite VAS was consistent across repeated treatments with icatibant. One injection of icatibant was sufficient to treat 88·2% of attacks; rescue medication was required in 5·3% of attacks. No icatibant-related serious adverse events were reported. Icatibant provided consistent efficacy and was well tolerated for repeated treatment of HAE attacks.

Eosinophil major basic protein activates human cord blood mast cells primed with fibroblast membranes by integrin-ß1.

BACKGROUND: Mast cell (MC) - eosinophil (Eos) activating cross-talk might be critical for the severity and chronicity of allergy. Among soluble mediators, eosinophil major basic protein (MBP), a hallmark of allergy, is particularly important because it was shown to activate specific MC subtypes. We previously demonstrated that MBP activates IgE-desensitized rat MC and human lung and cord blood-derived MC (CBMC) after priming with fibroblast membranal stem cell factor. However, a distinct mechanism for this activation was missing. Therefore, we aimed to investigate it. METHODS: Major basic protein-1 activation of CBMC primed with fibroblast-derived membranes (FBM) was measured by ß-hexosaminidase and tryptase release. Chemical cross-linking followed by micrometric flow cytometry probed direct interactions. Antibodies neutralized integrin-ß1 and recognized its active form. Pertussis toxin (Ptx) was used to decrease integrin-ß1 active form expression. Hematopoietic cell kinase (Hck) was identified by immunoprecipitation (IP) and silenced by siRNA. RESULTS: Major basic protein-1-induced CBMC activation is mediated partly by MBP1-integrin-ß1 interaction on the MC surface. FBM prime CBMC via a G protein, as confirmed by Ptx, to shift integrin-ß1 to its active form. Following MBP1 binding, integrin-ß1 binds Hck that further transduces the activation signal. MC priming with FBM leads to up-regulation in Hck protein level. MC integrin-ß1 neutralization inhibits MBP1-induced activation and uptake. Hck silencing results with reduced MBP1-induced activation. CONCLUSIONS: Fibroblast-derived membranes, integrin-ß1, and Hck are involved in MBP1-induced activation of CBMC and therefore represent a distinct mechanism for this activation. This finding might implicate integrin-ß1 and Hck as targets for decreasing MC - Eos activating cross-talk in allergy.

The consequences of not having eosinophils.

Several lines of evidence suggest that deficiency of eosinophils is not associated with any characteristic abnormality. Patients lacking eosinophils, in the setting of immunodeficiency or as a consequence of IgG-mediated eosinophil precursor destruction, do not display any distinguishing abnormalities related to eosinophil reduction. The observation that eosinophil-deficient mice do not display any distinctive syndrome or failure of their health is evidence that, under ordinary laboratory conditions, the eosinophil does not play a critical role in the well-being of mammals. Observations that monoclonal antibodies to interleukin-5 (IL-5) are well tolerated appear unsurprising in light of these findings. For example, patients with the hypereosinophilic syndrome have received mepolizumab, an anti-IL-5 monoclonal antibody, for as long as 6 years and have not developed any characteristic set of adverse events. Safety data for reslizumab, another anti-IL-5 monoclonal antibody, and benralizumab, a monoclonal antibody to the IL-5 receptor α-chain, are comparatively limited, especially for benralizumab, although reports of administration of these antibodies to humans suggest that they are well tolerated. Thus, data to the present suggest that reduction of eosinophils appears to have no characteristic ill effects on normal health, and monoclonal antibodies that deplete eosinophils have the potential to be widely employed in the treatment of eosinophil-associated diseases.

Eosinophil peroxidase induces the expression and function of acid-sensing ion channel-3 in allergic rhinitis: in vitro evidence in cultured epithelial cells.

BACKGROUND: Acid-sensing ion channels (ASIC) are a family of acid-activated ligand-gated cation channels. As tissue acidosis is a feature of inflammatory conditions, such as allergic rhinitis (AR), we investigated the expression and function of these channels in AR. OBJECTIVES: The aim of the study was to assess expression and function of ASIC channels in the nasal mucosa of control and AR subjects. METHODS: Immunohistochemical localization of ASIC receptors and functional responses to lactic acid application were investigated. In vitro studies on cultured epithelial cells were performed to assess underlying mechanisms of ASIC function. RESULTS: Lactic acid at pH 7.03 induced a significant rise in nasal fluid secretion that was inhibited by pre-treatment with the ASIC inhibitor amiloride in AR subjects (n = 19). Quantitative PCR on cDNA isolated from nasal biopsies from control and AR subjects demonstrated that ASIC-1 was equally expressed in both populations, but ASIC-3 was significantly more highly expressed in AR (P < 0.02). Immunohistochemistry confirmed significantly higher ASIC-3 protein expression on nasal epithelial cells in AR patients than controls (P < 0.01). Immunoreactivity for EPO+ eosinophils in both nasal epithelium and submucosa was more prominent in AR compared with controls. A mechanism of induction of ASIC-3 expression relevant to AR was suggested by the finding that eosinophil peroxidase (EPO), acting via ERK1/2, induced the expression of ASIC-3 in epithelial cells. Furthermore, using a quantitative functional measure of epithelial cell secretory function in vitro, EPO increased the air-surface liquid depth via an ASIC-dependent chloride secretory pathway. CONCLUSIONS: This data suggests a possible mechanism for the observed association of eosinophils and rhinorrhoea in AR and is manifested through enhanced ASIC-3 expression.

The role of eosinophil major basic protein in angiogenesis.

BACKGROUND: Eosinophil-derived major basic protein (MBP) plays an active role in allergic inflammation and tissue remodelling. However, its role in angiogenesis has not been established as yet. Therefore our objective was to investigate whether MBP exhibits any direct pro-angiogenic effects. METHODS: Rat aortic endothelial cells and human umbilical vascular endothelial cells were cultured with different concentrations of MBP and their viability (Trypan blue exclusion test), proliferation (thymidine incorporation) and capillary-like structure formation (matrigel assay) were investigated in vitro. The angiogenic activity of MBP was then tested in vivo using the chick chorio allantoic membrane (CAM) assay. RESULTS: Subcytotoxic concentrations of MBP induce endothelial cell proliferation and enhance the pro-mitogenic effect of vascular endothelial growth factor (VEGF), but do not affect their VEGF release. MBP promotes capillarogenesis by endothelial cells seeded on matrigel and sprouting formation in the CAM assay. Furthermore, we have shown that the pro-angiogenic effect of MBP is not due to its cationic charge since stimulation of the CAMs with the synthetic polycation, poly-L-arginine does not induce any angiogenic effects. CONCLUSIONS: These data demonstrate that MBP has pro-angiogenic effects in vitro and in vivo, providing a novel mechanism whereby MBP can participate in tissue inflammation and remodelling in atopic diseases.

Effects of (R)- and (S)-isomers of beta-adrenergic agonists on eosinophil response to interleukin-5.

BACKGROUND: Racemic beta2-adrenergic receptor agonists (beta2-agonists) are used frequently to treat patients with asthma. Potential differences in the biological activities and clinical efficacies among racemic beta2-agonists and their isomers are controversial, and research into these possible differences is limited. OBJECTIVE: We hypothesized that the (S)- and the (R)-isomers of beta2-agonists have opposing effects on the activation of inflammatory cells. METHODS: Isolated human eosinophils were pretreated with 1:1 racemic (R,S)-, (R)- or (S)-albuterol, isobutyl methylxanthine (IBMX), and stimulated with IL-5. The kinetics of superoxide production were examined by reduction of cytochrome c, and the effects of pharmacological agents on superoxide production were monitored for 180 min. RESULTS: (R,S)-albuterol inhibited IL-5-induced superoxide production. This inhibition was enhanced by a cyclic adenosine monophosphate (cAMP) phosphodiesterase inhibitor, IBMX, and was reversed by the selective beta2-adrenergic receptor antagonist, ICI 118, 551, verifying the involvement of both cAMP and the beta2-adrenergic receptor. In addition, (R)-albuterol alone, similarly to (R,S)-albuterol, significantly inhibited IL-5-induced superoxide production up to 60 min (P<0.05, n=4), but the inhibition was lost with longer incubation. In contrast, (S)-albuterol with IBMX did not inhibit IL-5-induced superoxide production before 60 min, but it significantly enhanced IL-5-mediated superoxide production after 60 min (P<0.05, n=4). When both were present as racemic (R,S)-albuterol, the inhibitory effect of (R)-albuterol was not affected by (S)-albuterol. CONCLUSION: When incubated with IL-5-activated eosinophils, (R)-albuterol shows anti-inflammatory effects and (S)-albuterol shows pro-inflammatory effects in the presence of IBMX. The kinetics of these effects are different, and when used simultaneously, (R)-albuterol predominates. When marked usage of the (S)-isomer is anticipated, racemic (R,S)-albuterol should be used clinically with caution.

Screening for Down's syndrome in early and late first and second trimester using six maternal serum markers.

The efficiency of six maternal serum markers for Down's syndrome (DS), alpha fetoprotein (AFP), human chorionic gonadotropin (hCG), free beta-hCG, pregnancy-associated plasma protein-A (PAPP-A), the proform of eosinophil major basic protein (ProMBP), pregnancy-specific-beta-1-glycoprotein (SP(1)), and combinations thereof, was examined. Discriminant analysis in 156 DS pregnancies and 546 controls defined three effective combinations of serum marker logMoMs (multiples of the median in control samples) in three gestational age windows, i.e. Index I (weeks 7-9) = 0.52 logMoM ProMBP + 0.28 logMoM PAPP-A - logMoM SP(1); Index II (weeks 10-12) = 1.94 logMoM free beta-hCG - logMoM SP(1), and Index III (weeks 15-19) = 0.78 logMoM free beta-hCG + 1.12 logMoM ProMBP - logMoM AFP. The estimated detection rates of indices and age for a false-positive rate (FPR) of 5% were 73% for Index I, 69% for Index II, and 60% for Index III. Including the ultrasound marker nuchal translucency, using a DS at term risk of 1 : 400 as cut-off, the detection rates of the indices increased to 86, 83, and 82% for FPRs of 4.3, 4.1, and 5.8%, respectively. The indices are promising markers for screening for DS.

Removal of bronchoalveolar cells augments the late eosinophilic response to segmental allergen challenge.

BACKGROUND: In patients with quiescent asthma, macrophages are the most prevalent cells recovered by bronchoalveolar lavage (BAL). Through activation via their FcepsilonRII receptors or by acting as antigen-presenting cells, macrophages could, in theory, promote the late airway response to allergen. OBJECTIVE: In order to investigate the importance of macrophages and other airway luminal cells in inducing the late airway response, a novel washout experiment was designed. METHODS: Five patients with ragweed-allergic asthma underwent bronchoscopy and segmental bronchial challenge with either normal saline or short ragweed extract in two segments of one lung. In a third segment of the opposite lung, 12 successive BALs (25 mL each) were performed, followed by challenge with an identical dose of short ragweed (washed-challenged segment). After 24 h, all three challenged segments underwent BAL. RESULTS: Initially, in the washed-challenged segment, over 80% (mean 80.4%, range 68-88%) of the recoverable airway dwelling cells were removed. Unexpectedly, 24 h later these same washed-challenged segments contained more eosinophils in the BAL than the challenged segments from the opposite lung (P = 0.033). CONCLUSIONS: Removing the majority of airway luminal cells followed by allergen bronchoprovocation increased the number of eosinophils recovered 24 h after challenge. Our results suggest that in quiescent allergic asthma, the airway luminal cells are protective and attenuate the late eosinophilic response to allergen challenge.

Regulation of human eosinophil NADPH oxidase activity: a central role for PKCdelta.

Eosinophils play a primary role in the pathophysiology of asthma. In the lung, the activation state of the infiltrating eosinophils determines the extent of tissue damage. Interleukin-5 (IL-5) and leukotriene B4 (LTB4) are important signaling molecules involved in eosinophil recruitment and activation. However, the physiological processes that regulate these activation events are largely unknown. In this study we have examined the mechanisms of human eosinophil NADPH oxidase regulation by IL-5, LTB4, and phorbol ester (PMA). These stimuli activate a Zn2+-sensitive plasma membrane proton channel, and treatment of eosinophils with Zn2+ blocks superoxide production. We have demonstrated that eosinophil intracellular pH is not altered by IL-5 activation of NADPH oxidase. Additionally, PKCdelta inhibitors block PMA, IL-5 and LTB4 mediated superoxide formation. Interestingly, the PKCdelta-selective inhibitor, rottlerin, does not block proton channel activation by PMA indicating that the oxidase and the proton conductance are regulated at distinct phosphorylation sites. IL-5 and LTB4, but not PMA, stimulated superoxide production is also blocked by inhibitors of PI 3-kinase indicating that activation of this enzyme is an upstream event common to both receptor signaling pathways. Our results indicate that the G-protein-coupled LTB4 receptor and the IL-5 cytokine receptor converge on a common signaling pathway involving PI 3-kinase and PKCdelta to regulate NADPH oxidase activity in human eosinophils.

Increased detection of interleukin-5 in sputum by addition of protease inhibitors.

The measurement of interleukin (IL)-5 in sputum is problematic, with interfering factors affecting immunoassay. The authors investigated whether sputum proteases could be acting as interfering factors by studying the effect of protease inhibitors (PI) on sputum IL-5 measurement. Induced sputa from 20 subjects with asthma were divided into aliquots, processed with and without protease inhibitors (in low and high concentrations) and the levels of IL-5 (spiked and endogenous) measured by enzyme immunoassay were compared. The concentration of sputum IL-5 was significantly increased by PI, with median (interquartile range) levels processed with no, low and high PI concentrations being 0 (0), 41.8 (75.6) and 66.1 (124.4) pg x mL(-1), respectively. There was also a significant increase in percentage recovery of spiked IL-5. Although high concentrations of PI reduced cell viability, there was no effect on total or differential cell counts and low concentrations of PI had no effect on cell counts or viability. Levels of endogenous interleukin-5 in sputum of asthmatic subjects can be significantly increased by the addition of protease inhibitors, and samples which would be regarded as negative for interleukin-5 without protease inhibitors may instead have considerable amounts of interleukin-5 detected.

Eosinophil-associated inflammation and elaboration of eosinophil-derived proteins in 2 children with raccoon roundworm (Baylisascaris procyonis) encephalitis.

OBJECTIVE: Eosinophil-associated proteins, especially eosinophil-derived neurotoxin, may be important contributors to the neurologic pathology and symptoms caused by Baylisascaris procyonis infection. METHODS: Two cases of severe B procyonis encephalitis with evidence of marked eosinophil degranulation in the central nervous system are presented. Serial cerebrospinal fluid (CSF) specimens were collected from each patient during the course of their illness. Antibodies against B procyonis were measured in the patients' serum and CSF. Levels of the eosinophilopoietin interleukin-5 (IL-5) and 2 important eosinophil proteins, eosinophil-derived neurotoxin and major basic protein, were assayed in the CSF. RESULTS: Both patients had rapidly progressive central nervous system disease with evidence of eosinophilic meningoencephalitis. Both tested positive for antibodies to B procyonis in serum and CSF and had progressively worsening deep white matter changes on magnetic resonance images of the brain. CSF levels of IL-5, eosinophil-derived neurotoxin, and major basic protein were markedly elevated over controls. CONCLUSIONS: This is the first report of the measurement of IL-5, eosinophil-derived neurotoxin, and major basic protein in human CSF. In addition to traumatic damage and necrosis caused by migrating larvae, eosinophil-derived neurotoxin from associated eosinophilic inflammation may be an important contributory factor in the pathogenesis of B procyonis encephalitis. parasite, eosinophil-derived-neurotoxin, major basic protein, eosinophilia, hypereosinophilia, interleukin-5, encephalitis, child.

Peptide-based analysis of amino acid sequences important to the biological activity of eosinophil granule major basic protein.

Synthetic peptides corresponding to amino acid sequences in eosinophil granule major basic protein (MBP) were evaluated for cytotoxic activity toward K562 cells and for ability to stimulate basophil mediator release. Results obtained using 14 peptides spanning the 117-amino acid sequence of MBP in overlapping fashion indicated that the activities mapped to peptide sequences near the amino and carboxy termini of MBP. The activity of these regions was confirmed using two peptides corresponding to MBP residues 18-45 and 89-117. A 20-h incubation with 5 microM peptide 18-45 or peptide 89-117 caused approximately the same levels (>60%) of cytotoxicity in K562 cells as 5 microM MBP. Similarly, a 30-min incubation with peptides 18-44 and 89-117 stimulated basophil histamine release in a concentration-dependent manner over the range of 5-20 microM. The level of release stimulated by 20 microM peptide 89-117 approached that stimulated by 2 microM MBP. A 20 microM concentration of peptide 89-117 also stimulated leukotriene C4 (LTC4) production by the basophils. Neither peptide 18-45 nor peptide 89-117 was cytotoxic for basophils under the experimental conditions for histamine and LTC4 release, as determined by 51Cr release. These results indicate that two MBP peptide sequences, including one (89-117) that contains a unique carbohydrate-binding region, share the biologic activities of MBP.

Effects of human eosinophil granule-derived cationic proteins on C-fiber afferents in the rat lung.

Experiments were performed to test the hypothesis that human eosinophil granule-derived cationic proteins stimulate vagal C-fiber afferents in the lungs and elicit pulmonary chemoreflex responses in anesthetized Sprague-Dawley rats. Intratracheal instillation of eosinophil cationic protein (ECP; 1-2 mg/ml, 0.1 ml) consistently induced an irregular breathing pattern, characterized by tachypnea (change in breathing frequency of 44.7%) and small unstable tidal volume (VT). The tachypnea, accompanied by decreased heart rate and arterial blood pressure, started within 30 s after the delivery of ECP and lasted for >30 min. These ECP-induced cardiorespiratory responses were completely prevented by perineural capsaicin treatment of both cervical vagi, which selectively blocked C-fiber conduction, suggesting the involvement of these afferents. Indeed, direct recording of single-unit activities of pulmonary C-fibers further demonstrated that the same dose of ECP evoked a pronounced and sustained (>30-min) stimulatory effect on pulmonary C-fibers. Furthermore, the sensitivity of these afferents to lung inflation was also markedly elevated after the ECP instillation, whereas the vehicle of ECP administered in the same manner had no effect. Other types of eosinophil granule cationic proteins, such as major basic protein and eosinophil peroxidase, induced very similar respiratory and cardiovascular reflex responses. In conclusion, these results show that eosinophil granule-derived cationic proteins induce a distinct stimulatory effect on vagal pulmonary C-fiber endings, which may play an important role in the airway hyperresponsiveness associated with eosinophil infiltration in the airways.

Extensive eosinophil degranulation and peroxidase-mediated oxidation of airway proteins do not occur in a mouse ovalbumin-challenge model of pulmonary inflammation.

Paradigms of eosinophil effector function in the lungs of asthma patients invariably depend on activities mediated by cationic proteins released from secondary granules during a process collectively referred to as degranulation. In this study, we generated knockout mice deficient for eosinophil peroxidase (EPO) to assess the role(s) of this abundant secondary granule protein in an OVA-challenge model. The loss of EPO had no effect on the development of OVA-induced pathologies in the mouse. The absence of phenotypic consequences in these knockout animals extended beyond pulmonary histopathologies and airway changes, as EPO-deficient animals also displayed OVA-induced airway hyperresponsiveness after provocation with methacholine. In addition, EPO-mediated oxidative damage of proteins (e.g., bromination of tyrosine residues) recovered in bronchoalveolar lavage from OVA-treated wild-type mice was <10% of the levels observed in bronchoalveolar lavage recovered from asthma patients. These data demonstrate that EPO activities are inconsequential to the development of allergic pulmonary pathologies in the mouse and suggest that degranulation of eosinophils recruited to the lung in this model does not occur at levels comparable to those observed in humans with asthma.

Human eosinophils induce histamine release from antigen-activated rat peritoneal mast cells: a possible role for mast cells in late-phase allergic reactions.

BACKGROUND: Mast cells and eosinophils are believed to interact during the late and the chronic stages of allergic inflammation. OBJECTIVE: In this study we investigated whether eosinophils can cause activation and consequent histamine release of already challenged mast cells, a situation likely to take place during the allergic late-phase reaction. METHODS: Rat peritoneal mast cells presensitized with IgE anti-dinitrophenol-human serum albumin and challenged by dinitrophenol-human serum albumin or compound 48/80 were incubated with either eosinophil sonicate or major basic protein (MBP). Eosinophils were purified from the peripheral (>98%) blood of mildly allergic patients. Heparin and pertussis toxin and different extracellular Ca(2+) concentrations were used to modulate mast cell reactivation by MBP. Histamine release was assessed as a marker of mast cell activation. RESULTS: IgE-challenged mast cells were sensitive to reactivation induced by eosinophil sonicate and MBP. Reactivation was not cytotoxic for the mast cells. Mast cells previously challenged with compound 48/80 did not respond to subsequent MBP activation. Furthermore, heparin and pertussis toxin both inhibited mast cell reactivation induced by MBP. The ability of eosinophil sonicate and MBP to activate mast cells was not significantly affected at the different Ca(2+) concentrations. CONCLUSIONS: In summary, we have shown a direct activating activity of eosinophils, partially due to MBP, toward IgE-challenged and immunologically desensitized mast cells. This suggests that in vivo mast cells can be reactivated during a late-phase reaction to release histamine by a non-IgE-dependent mechanism.

Eosinophil infiltration and degranulation in normal human tissues: evidence for eosinophil degranulation in normal gastrointestinal tract.

Eosinophils play an important role in the pathogenesis of inflammatory diseases such as bronchial asthma and host immunity to parasitic infections. Deposition of eosinophil granule proteins and concomitant tissue damage have been documented in various diseases. Here, we review and summarize results of our immunofluorescence studies of eosinophil infiltration and degranulation in various normal human tissues. Furthermore, because eosinophil infiltration and degranulation are not normally present in healthy tissues, we examine whether eosinophil infiltration and degranulation normally occur in the small intestine and whether tissue procurement methods affect the extent of eosinophil infiltration and degranulation there. Hematopoietic and lymphatic tissues, including the thymus, showed eosinophil infiltration, but the only organ showing remarkable eosinophil infiltration and degranulation was the gastrointestinal (GI) tract. Eosinophil degranulation was significantly increased in specimens obtained by endoscopic forceps compared to those obtained by scalpel. These results suggest that tissue procurement methods affect the degree of eosinophil degranulation in the GI tract and that, among normal human body organs, both eosinophil infiltration and degranulation only occur in the GI tract.

Increased eosinophil infiltration and degranulation in colonic tissue from patients with collagenous colitis.

OBJECTIVE: Eosinophils infiltrate the colonic mucosa of patients with collagenous colitis (CC), although the pathogenetic implications are unknown, including whether these eosinophils are activated and degranulate in situ. We examined eosinophil infiltration and degranulation in the intestines of patients with CC by immunofluorescence for eosinophil granule major basic protein (MBP). METHODS: We used both conventional histology (hematoxylin and eosin) and indirect MBP immunofluorescence histochemistry on colon biopsy specimens from patients with CC (n = 21) and from healthy controls (n = 9). Scoring of histological features was performed on hematoxylin and eosin-stained sections on a 0 to 3 scale. Eosinophil infiltration and degranulation, as quantified by extracellular MBP staining, were scored in each specimen on a 0 to 4 scale. RESULTS: The inflammatory infiltrate of the lamina propria, the thickness of the collagen band, the numbers of intraepithelial lymphocytes, and degree of epithelial cell damage were all significantly increased in patients with CC as compared to controls (p < 0.0001). Scores for both eosinophil infiltration and degranulation were also significantly higher in the CC group compared to controls (p < 0.0001). The degree of infiltrating eosinophils by hematoxylin and eosin was correlated with eosinophil infiltration and degranulation by MBP immunostaining; however, no other correlations were found between eosinophil infiltration or degranulation by immunofluorescence and any of the histological parameters measured in the CC group. CONCLUSIONS: Eosinophil infiltration and degranulation are increased in the colonic mucosa of patients with CC compared to healthy controls. Eosinophils and their cytotoxic granule proteins could be involved in the pathogenesis of CC. Further studies will be necessary to elucidate the mechanisms of eosinophil activation in CC.