Increase in TPSB2 and TPSD1 Expression in Synovium of Hip Osteoarthritis Patients Who Are Overweight.

While research suggests that increasing body mass index (BMI) is a risk factor for hip osteoarthritis (HOA), the mechanisms of this effect are not fully understood. Tryptases are among the main proteases found in mast cells (MCs) and contribute to OA pathology. TPSB2, which encodes ß-tryptase, is increased in the synovium of overweight and obese knee OA patients. However, it remains unclear whether tryptase in the synovium of HOA is increased with increasing BMI. Here, we investigated tryptase genes (TPSB2 and TPSD1) in the synovium of overweight HOA patients. Forty-six patients radiographically diagnosed with HOA were allocated to two groups based on BMI, namely normal (<25 kg/m2) and overweight (25-29.99 kg/m2). TPSB2 and TPSD1 expression in the synovium of the two groups was compared using real-time polymerase chain reaction. To compare TPSB2 and TPSD1 expression in MCs between the groups, we isolated the MC-rich fraction (MC-RF) and MC-poor fraction (MC-PF), extracted using magnetic isolation. TPSB2 and TPSD1 expression was increased in the overweight group compared with the normal group. Expression of both genes in the MC-RF was significantly higher than that in MC-PF in both groups. However, TPSB2 and TPSD1 expression levels in the MC-RF did not differ between the groups. Tryptase genes were highly expressed in the synovium of overweight HOA patients. Further investigation to reveal the role of tryptase in the relationship between increasing BMI and HOA pathology is required.

Combined histochemical approach in assessing tryptase expression in the mast cell population.

To increase the efficiency of interpretation of mast cell's contribution to the state of a specific tissue microenvironment, it is necessary to detail the molecular composition of their secretome and analyze the pathways of degranulation. Developed method of combining immunomorphological and histochemical staining protocols contributes to the most objective detection of the integral level of tryptase expression in the intraorgan population of the skin mast cells. Novel technique for tryptase detection expands the possibilities of morphological analysis, provides researchers with additional data on the structure of the mast cell population and helps visualize the processing and cytological features and structural targets of tryptase during the development of adaptive and pathological reactions. Objective determination of the tryptase profile for organ-specific mast cell populations is in great demand in clinical practice for the interpretation of pathological processes, including inflammation and oncogenesis.

Respiratory Syncytial Virus Infection of Human Lung Fibroblasts Induces a Hyaluronan-Enriched Extracellular Matrix That Binds Mast Cells and Enhances Expression of Mast Cell Proteases.

Human lung fibroblasts (HLFs) treated with the viral mimetic polyinosine-polycytidylic acid (poly I:C) form an extracellular matrix (ECM) enriched in hyaluronan (HA) that avidly binds monocytes and lymphocytes. Mast cells are important innate immune cells in both asthma and acute respiratory infections including respiratory syncytial virus (RSV); however, the effect of RSV on HA dependent mast cell adhesion and/or function is unknown. To determine if RSV infection of HLFs leads to the formation of a HA-enriched ECM that binds and enhances mast cell activity primary HLFs were infected with RSV for 48 h prior to leukocyte binding studies using a fluorescently labeled human mast cell line (LUVA). Parallel HLFs were harvested for characterization of HA production by ELISA and size exclusion chromatography. In separate experiments, HLFs were infected as above for 48 h prior to adding LUVA cells to HLF wells. Co-cultures were incubated for 48 h at which point media and cell pellets were collected for analysis. The role of the hyaladherin tumor necrosis factor-stimulated gene 6 (TSG-6) was also assessed using siRNA knockdown. RSV infection of primary HLFs for 48 h enhanced HA-dependent LUVA binding assessed by quantitative fluorescent microscopy. This coincided with increased HLF HA synthase (HAS) 2 and HAS3 expression and decreased hyaluronidase (HYAL) 2 expression leading to increased HA accumulation in the HLF cell layer and the presence of larger HA fragments. Separately, LUVAs co-cultured with RSV-infected HLFs for 48 h displayed enhanced production of the mast cell proteases, chymase, and tryptase. Pre-treatment with the HA inhibitor 4-methylumbelliferone (4-MU) and neutralizing antibodies to CD44 (HA receptor) decreased mast cell protease expression in co-cultured LUVAs implicating a direct role for HA. TSG-6 expression was increased over the 48-h infection. Inhibition of HLF TSG-6 expression by siRNA knockdown led to decreased LUVA binding suggesting an important role for this hyaladherin for LUVA adhesion in the setting of RSV infection. In summary, RSV infection of HLFs contributes to inflammation via HA-dependent mechanisms that enhance mast cell binding as well as mast cell protease expression via direct interactions with the ECM.

Copper Regulates Maturation and Expression of an MITF:Tryptase Axis in Mast Cells.

Copper has previously been implicated in the regulation of immune responses, but the impact of this metal on mast cells is poorly understood. In this article, we address this issue and show that copper starvation of mast cells causes increased granule maturation, as indicated by higher proteoglycan content, stronger metachromatic staining, and altered ultrastructure in comparison with nontreated cells, whereas copper overload has the opposite effects. In contrast, copper status did not impact storage of histamine in mast cells, nor did alterations in copper levels affect the ability of mast cells to degranulate in response to IgER cross-linking. A striking finding was decreased tryptase content in mast cells with copper overload, whereas copper starvation increased tryptase content. These effects were associated with corresponding shifts in tryptase mRNA levels, suggesting that copper affects tryptase gene regulation. Mechanistically, we found that alterations in copper status affected the expression of microphthalmia-associated transcription factor, a transcription factor critical for driving tryptase expression. We also found evidence supporting the concept that the effects on microphthalmia-associated transcription factor are dependent on copper-mediated modulation of MAPK signaling. Finally, we show that, in MEDNIK syndrome, a condition associated with low copper levels and a hyperallergenic skin phenotype, including pruritis and dermatitis, the number of tryptase-positive mast cells is increased. Taken together, our findings reveal a hitherto unrecognized role for copper in the regulation of mast cell gene expression and maturation.

High infiltration of mast cells positive to tryptase predicts worse outcome following resection of colorectal liver metastases.

BACKGROUND: Accumulation of tumor-infiltrating mast cells (MCs) predicts poor survival in several cancers after resection. However, its effect on the prognosis of patients with colorectal liver metastases (CRLM) is not known. METHODS: Our retrospective study included 135 patients who underwent potentially curative resection for CRLM between 2001 and 2010. Expression of tryptase, MAC387, CD83, and CD31, which are markers for MCs, macrophages, mature dendritic cells, and vascular endothelial cells, respectively, was determined via immunohistochemistry of resected tumor specimens. The relationship between immune cell infiltration and long-term outcome was investigated. RESULTS: The median follow-up time was 48.4 months for all patients and 57.5 months for survivors. Overall survival (OS) rates at 1, 3, and 5 years were 91.0, 62.4, and 37.4 %, respectively. Five-year disease-free survival (DFS) and OS rates were 21.6 and 38.1 %, respectively, in patients with high MC infiltration, and 42.6 and 55.6 %, respectively, in patients with low MC infiltration (p < 0.01 for both DFS and OS). Infiltration of other types of immune cells did not correlate with survival. Multivariate analyses indicated that hypoalbuminemia and high peritumoral MC infiltration were significant predictors of unfavorable OS. CONCLUSION: High peritumoral MC infiltration predicts poor prognosis in patients who underwent hepatectomy for CRLM. The number of MCs in metastatic lesions is important for predicting the prognosis of CRLM patients and as an indication of therapy.

Ctr2 Regulates Mast Cell Maturation by Affecting the Storage and Expression of Tryptase and Proteoglycans.

Copper (Cu) is essential for multiple cellular functions. Cellular uptake of Cu(+) is carried out by the Ctr1 high-affinity Cu transporter. The mobilization of endosomal Cu pools is regulated by a protein structurally similar to Ctr1, called Ctr2. It was recently shown that ablation of Ctr2 caused an increase in the concentration of Cu localized to endolysosomes. However, the biological significance of excess endolysosomal Cu accumulation has not been assessed. In this study, we addressed this issue by investigating the impact of Ctr2 deficiency on mast cells, a cell type unusually rich in endolysosomal organelles (secretory granules). We show that Ctr2(-/-) mast cells have increased intracellular Cu concentrations and that the absence of Ctr2 results in increased metachromatic staining, the latter indicating an impact of Ctr2 on the storage of proteoglycans in the secretory granules. In agreement with this, the absence of Ctr2 caused a skewed ratio between proteoglycans of heparin and chondroitin sulfate type, with increased amounts of heparin accompanied by a reduction of chondroitin sulfate. Moreover, transmission electron microscopy analysis revealed a higher number of electron-dense granules in Ctr2(-/-) mast cells than in wild-type cells. The increase in granular staining and heparin content is compatible with an impact of Ctr2 on mast cell maturation and, in support of this, the absence of Ctr2 resulted in markedly increased mRNA expression, storage, and enzymatic activity of tryptase. Taken together, the present study introduces Ctr2 and Cu as novel actors in the regulation of mast cell maturation and granule homeostasis.

Low numbers of tryptase+ and chymase+ mast cells associated with reduced survival and advanced tumor stage in melanoma.

The role of mast cells in cutaneous melanoma remains unclear. Tryptase and chymase are serine proteinases and major proteins in mast cell secretory granules. Therefore, this study aimed to investigate the presence of tryptase and chymase mast cells in benign and malignant cutaneous melanocytic lesions and in lymph node metastases of melanomas. The presence of positively stained mast cells was correlated with clinicopathological characteristics in invasive melanomas. Paraffin-embedded sections of 28 benign (13 intradermal, 10 compound, and five junctional nevi) and 26 dysplastic nevi, 15 in-situ melanomas, 36 superficially (pT1, Breslow's thickness<1 mm), and 49 deeply (pT4, Breslow's thickness>4 mm) invasive melanomas and 30 lymph node metastases were immunohistochemically stained for mast cell tryptase and chymase, and immunopositive cells were counted using the hotspot counting method. The mean count of tryptase and chymase mast cells was lower in invasive melanomas compared with in-situ melanomas and dysplastic and benign nevi. In deeply invasive melanomas, the difference was statistically significant compared with dysplastic nevi (P=0.003 for tryptase and P=0.009 for chymase) and in-situ melanomas (0.043 for tryptase). Low numbers of tryptase mast cells were associated with poor overall survival (P=0.031) in deeply invasive melanomas and with a more advanced stage (T1b, P=0.008) in superficially invasive melanomas. Low numbers of chymase mast cells were associated with microsatellites (P=0.017) in deeply invasive melanomas. The results suggest that these serine proteinases of mast cells may be protective in the pathogenesis of melanoma.

High density of tryptase-positive mast cells in patients with multiple myeloma: correlation with parameters of disease activity.

Multiple myeloma (MM) is a plasma cell neoplasm characterized by bone marrow infiltration from malignant plasma cells. Mast cells play an important role in inflammation and angiogenesis in malignant diseases. The aim of the study was to evaluate the mast cell density in bone marrow of untreated MM patients with markers of disease activity such as serum interleukin-6 (IL-6), B2M, and C-reactive protein (CRP), the grade of bone marrow infiltration, and the levels of produced paraprotein. We studied 86 newly diagnosed MM patients (46 males, 40 females, mean age 59 ± 13.7 years). Thirty of them reached plateau phase after chemotherapy and 20 healthy volunteers. According to the criteria of International Staging System (ISS) staging system, 23 patients had stage I, 30 had stage II, and 33 had stage III. The serum concentrations of CRP, B2M, and IL-6, and the mast cell density (MCD) values were significantly higher in MM patients' group (1.6 ± 1.8, 4.3 ± 2.9, 7.1 ± 5.1, and 9 ± 4.8), in comparison with those found in control group (0.4 ± 0.1, 1.5 ± 0.6, 1.1 ± 0.5, and 1.9 ± 0.7; p < 0.001 in all the cases). Significant differences were found between the grade of infiltration in bone marrow, and the paraprotein values in patients' serum before and after chemotherapy. Furthermore, there was a significant correlation between the MCD values and the prognostic markers CRP (r = 0.452, p < 0.0001), IL-6 (r = 0.475, p < 0.0001), bone marrow infiltration (r = 0.333, p < 0.0002), and serum paraprotein levels(r = 0.221, p < 0.04). High MCD values strengthen the hypothesis that mast cells participate in the pathogenesis of disease progression and may be used as an indicator of the disease activity.

Annexin A2 participates in human skin keloid formation by inhibiting fibroblast proliferation.

Abnormal scarring results from the expression and composition of extracellular matrix molecules. The transcription and translation of collagens I and III, fibronectin, laminin, periostin, and tenascin are all increased in raised dermal scar tissue. However, human keloid development is not fully defined. In this study, we identified proteins expressed differentially between normal skin and keloid scar tissues and examined their function in keloid formation using fibroblasts. Skin specimens from normal volunteers and patients with keloids were obtained by skin biopsy. Whole proteins were isolated by two-dimensional electrophoresis, and differentially expressed proteins were identified by matrix-assisted laser desorption/ionization-time of flight/time of flight mass spectrometry. Protein function was determined by proliferation assay using annexin A2-overexpressing keloid fibroblasts. The expression of 11 protein spots was altered by at least 1.5-fold in patients with keloids than in normal volunteers. Of these proteins, annexin A2, a pre-serum amyloid P component, serum albumin precursor, and tryptase-I, were down-regulated in keloid tissue compared to normal skin. Collagen alpha 1(V) chain precursor, collagen alpha 1(I) chain precursor, ferritin light subunit, alpha 1(III) collagen, 6-phosphogluconolactonase, and calponin 2 were up-regulated. Diminished expression of annexin A2 was confirmed by immunoblotting and immunohistochemistry. Treatment with the recombinant human epidermal growth factor increased proliferation of keloid fibroblasts, which was more inhibited in annexin A2-overexpressing fibroblasts than in non-transfected control cells. These results imply that annexin A2 may participate in keloid formation by inhibiting keloid fibroblast proliferation. Therefore, it is concluded that annexin A2 may be a valuable therapeutic target for keloid lesions.

Absence of TLR4 reduces neurovascular unit and secondary inflammatory process after traumatic brain injury in mice.

BACKGROUND: Traumatic brain injury (TBI) initiates a neuroinflammatory cascade that contributes to neuronal damage and behavioral impairment. Toll-like receptors (TLRs) are signaling receptors in the innate immune system, although emerging evidence indicates their role in brain injury. We have therefore investigated the role played by TLR4 signaling pathway in the development of mechanisms of secondary inflammatory process in traumatic brain injury (TBI) differ in mice that lack a functional TLR4 signaling pathway. METHODS/PRINCIPAL FINDINGS: Controlled cortical impact injury was performed on TLR4 knockout (KO) mice (C57BL/10ScNJ) and wild-type (WT) mice (C57BL/10ScNJ). TBI outcome was evaluated by determination of infarct volume and assessment of neurological scores. Brains were collected at 24 h after TBI. When compared to WT mice, TLR4 KO mice had lower infarct volumes and better outcomes in neurological and behavioral tests (evaluated by EBST and rotarod test). Mice that lacked TLR4 had minor expression of TBI-induced GFAP, Chymase, Tryptase, IL-1ß, iNOS, PARP and Nitrotyrosine mediators implicated in brain damage. The translocation of expression of p-JNK, IκB-α and NF-κB pathway were also lower in brains from TLR4 KO mice. When compared to WT mice, resulted in significant augmentation of all the above described parameters. In addition, apoptosis levels in TLR4 KO mice had minor expression of Bax while on the contrary with Bcl-2. CONCLUSIONS/SIGNIFICANCE: Our results clearly demonstrated that absence of TLR4 reduces the development of neuroinflammation, tissues injury events associated with brain trauma and may play a neuroprotective role in TBI in mice.

Microvascular density and mast cells in benign and malignant pheochromocytomas.

Pheochromocytomas, uncommon adrenal tumors, have an uncertain behavior. Recently, PASS criteria were proposed for differentiating between benign and malignant cases. These are not perfect, however. The aim of the study was to investigate angiogenesis and mast cell density in context of the clinical behavior and morphologic characteristics of pheochromocytomas. Mean intratumoral chymase positive cell count was 14.50 for malignant, 15.73 for benign cases; mean subcapsular chymase positive cell count was 12.50 for malignant, 11.27 for benign cases. Mean intratumoral tryptase positive cell count was 17.50 for malignant and 17.91 for benign cases; mean subcapsular tryptase positive cell count was 15.25 for malignant and 15.73 for benign cases. Mean intratumoral CD31 positive vessel count was 46.98 for malignant and 51.02 for benign cases; mean subcapsular CD31 positive vessel count was 44.86 for malignant and 39.81 for benign cases. Mean intratumoral CD105 positive vessel count was 37.84 for malignant and 35.95 for benign cases; mean subcapsular CD105 positive vessel count was 26.36 for malignant and 22.03 for benign cases. The differences between benign and malignant cases were not significant. All the vascular counts were correlated with mast cells counts. PASS index was inversely correlated with mast cell counts.

High density of tryptase-positive mast cells in patients with renal cell carcinoma on hemodialysis: correlation with expression of stem cell factor and protease activated receptor-2.

Patients on hemodialysis are at higher risk of renal cell carcinoma probably because of inflammatory and immune system disorders. The aim of this study was to clarify the pathologic roles of 2 phenotypes of mast cells, mast cell tryptase and mast cell chymase, and their correlation with stem cell factor and protease-activated receptor 2 in patients with renal cell carcinoma on hemodialysis. The densities of mast cell tryptase and mast cell chymase and expressions of stem cell factor and protease-activated receptor 2 were examined in 35 patients with hemodialysis-renal cell carcinoma and 39 with non-hemodialysis-renal cell carcinoma who were diagnosed and treated in our hospital. Protein expression was examined by immunohistochemistry. The proliferation index represented the number of Ki-67-positive cells. There were no significant differences in clinicopathologic features between the 2 groups. Mast cell tryptase densities in intratumoral (8.3 per high-power field) and peritumoral areas (8.7 per high-power field) were higher in hemodialysis-renal cell carcinoma than non-hemodialysis-renal cell carcinoma (2.7 and 5.3 per high-power field). No such significant correlations were detected in mast cell chymase. In hemodialysis-renal cell carcinoma, intratumoral mast cell tryptase density correlated with the proliferation index (P = .039 and P = .008, respectively) and also with stem cell factor and protease-activated receptor 2 expression. Our results emphasize the important roles of mast cell tryptase in cancer cell proliferation and recurrence in hemodialysis-renal cell carcinoma. Stem cell factor and protease-activated receptor 2 seem to up-regulate mast cell tryptase functions in these patients. The results suggest collaborative effects of stem cell factor, mast cell tryptase, and protease-activated receptor 2 on the malignant potential of hemodialysis-renal cell carcinoma.

Higher intratumoral expression of CD1a, tryptase, and CD68 in a follicular variant of papillary thyroid carcinoma compared to adenomas: correlation with clinical and pathological parameters.

BACKGROUND: In a number of human malignancies, the presence of lymphocytic infiltration in or around tumor tissue is commonly considered to be part of the host tumor immune response. An association between thyroid carcinoma and chronic inflammation has been described. This relationship is not fully understood, so we performed a systematic study on a follicular variant of papillary thyroid carcinoma (FVPTC), to evaluate the type and distribution of certain immunological cells and their relationship with prognostic factors. METHODS: We selected 91 consecutive cases of FVPTC, in which we evaluated the presence of three different immunological cells: dendritic cells (DC), immature CD1a+ and mature DC-Lamp+; mast cells (MC), tryptase+; and macrophages (M), CD68+, in the intratumoral, peritumoral, and extratumoral areas. As a control we analyzed 44 cases of thyroid adenomas (A). RESULTS: In the intratumoral and peritumoral areas, the expression of CD1a, tryptase, and CD68 was significantly higher in FVPTC than in adenomas. Expression of CD1a and tryptase was comparable in the extratumoral compartment, whereas CD68 expression in the extratumoral area was significantly higher in FVPTC than in adenoma (p=0.0015). DC-Lamp expression was not significantly different among the intra-tumor, peri-tumor, and extra-tumor areas of FVPTC or adenoma. It was also very interesting that nonencapsulated FVPTC were more positive to tryptase. CONCLUSION: We highlight a higher presence of immunological cells in carcinomas than in adenomas. On this basis, it is possible to speculate that these inflammatory elements could be involved in tumor progression and invasion, as appears to be the case for MC and M.

Mast cell tryptase stimulates production of decorin by human testicular peritubular cells: possible role of decorin in male infertility by interfering with growth factor signaling.

BACKGROUND: Myofibroblastic, peritubular cells in the walls of seminiferous tubules produce low levels of the extracellular matrix (ECM) protein decorin (DCN), which has the ability to interfere with growth factor (GF) signaling. In men with impaired spermatogenesis, fibrotic remodeling of these walls and accumulation of tryptase-positive mast cells (MCs) occur. METHODS: Human testicular biopsies with normal and focally impaired spermatogenesis (mixed atrophy) were subjected to immunohistochemistry and laser micro-dissection followed by RT-PCR. Primary human testicular peritubular cells (HTPCs), which originate from normal and fibrotically altered testes (HTPC-Fs), were studied by qRT-PCR, western blotting, enzyme-linked immunosorbent assay measurements and Ca(2+) imaging. Phosphorylation and viability/proliferation assays were performed. RESULTS: Immunohistochemistry revealed DCN deposits in the walls of tubules with impaired spermatogenesis. Mirroring the situation in vivo, HTPC-Fs secreted more DCN than HTPCs (P < 0.05). In contrast to HTPCs, HTPC-Fs also responded to the main MC product, tryptase, and to a tryptase receptor (PAR-2) agonist by further increased production of DCN (P < 0.05). Several GF receptors (GFRs) are expressed by HTPCs and HTPC-Fs. DCN acutely increased intracellular Ca(2+)-levels and phosphorylated epidermal GF (EGFR) within minutes. Platelet-derived GF (PDGF) and EGF induced strong mitogenic responses in HTPC/-Fs, actions that were blocked by DCN, suggesting that DCN in the ECM interferes with GF/GFRs signaling of peritubular cells of the human testis. CONCLUSIONS: The data indicate that the increase in testicular DCN found in male infertility is a consequence of actions of MC-derived tryptase. We propose that the increases in DCN may consequently imbalance the paracrine signaling pathways in human testis.

Mast cells and neutrophils release IL-17 through extracellular trap formation in psoriasis.

IL-17 and IL-23 are known to be absolutely central to psoriasis pathogenesis because drugs targeting either cytokine are highly effective treatments for this disease. The efficacy of these drugs has been attributed to blocking the function of IL-17-producing T cells and their IL-23-induced expansion. However, we demonstrate that mast cells and neutrophils, not T cells, are the predominant cell types that contain IL-17 in human skin. IL-17(+) mast cells and neutrophils are found at higher densities than IL-17(+) T cells in psoriasis lesions and frequently release IL-17 in the process of forming specialized structures called extracellular traps. Furthermore, we find that IL-23 and IL-1ß can induce mast cell extracellular trap formation and degranulation of human mast cells. Release of IL-17 from innate immune cells may be central to the pathogenesis of psoriasis, representing a fundamental mechanism by which the IL-23-IL-17 axis mediates host defense and autoimmunity.

Immunohistochemical expression of mast cell tryptase in giant cell fibroma and inflammatory fibrous hyperplasia of the oral mucosa.

This study analysed the immunohistochemical expression of mast cell tryptase in giant cell fibromas (GCFs). In addition, the possible interaction of mast cells with stellate giant cells, as well as their role in fibrosis and tumour progression, was investigated. For this purpose, the results were compared with cases of inflammatory fibrous hyperplasia (IFH) and normal oral mucosa. Thirty cases of GCF, 30 cases of IFH and 10 normal mucosa specimens used as control were selected. Immunoreactivity of mast cells to the anti-tryptase antibody was analysed quantitatively in the lining epithelium and in connective tissue. In the epithelial component (p=0.250) and connective tissue (p=0.001), the largest mean number of mast cells was observed in IFHs and the smallest mean number in GCFs. In connective tissue, the mean percentage of degranulated mast cells was higher in GCFs than in IFHs and normal mucosa specimens (p<0.001). Analysis of the percentage of degranulated mast cells in areas of fibrosis and at the periphery of blood vessels also showed a larger mean number in GCFs compared to IFHs and normal mucosa specimens (p<0.001). The percent interaction between mast cells and stellate giant cells in GCFs was 59.62%. In conclusion, although mast cells were less numerous in GCFs, the cells exhibited a significant interaction with stellate giant cells present in these tumours. In addition, the results suggest the involvement of mast cells in the induction of fibrosis and modulation of endothelial cell function in GCFs.

Mast cells infiltrate the esophageal smooth muscle in patients with eosinophilic esophagitis, express TGF-ß1, and increase esophageal smooth muscle contraction.

BACKGROUND: Increased numbers of mast cells are present in the esophageal epithelium in patients with eosinophilic esophagitis (EE). However, mast cell infiltration into the esophageal lamina propria (LP) and smooth muscle (SM) and the effects of their products on SM function has not been determined. OBJECTIVE: We investigated mast cell localization and characterization in esophageal SM, the functional significance of mast cell TGF-ß1 expression to contraction of human esophageal smooth muscle (HESM) cells in vitro, and the effect of topical corticosteroids on the number of tryptase-positive (MC(T)) and chymase-positive (MC(C)) mast cells in patients with EE. METHODS: MC(T)- and MC(C)-positive mast cell numbers were quantitated in the epithelium, the LP before and after topical corticosteroid therapy, and the muscularis mucosa in patients with EE and control subjects by using immunohistology. Double immunofluorescence was used to assess mast cell production of TGF-ß1. The ability of TGF-ß1 to influence HESM cell contractility was assessed in vitro. RESULTS: In the SM in patients with EE, significantly increased numbers of MC(T)- and TGF-ß1-positive cells (but only low levels of eosinophils) were detected compared with those seen in control subjects. MC(T) expressed TGF-ß1, which increased the contractility of cultured primary HESM cells in vitro. Topical corticosteroid therapy in patients with EE significantly reduced epithelial MC(T) numbers but not LP tryptase-chymase-positive mast cell numbers. CONCLUSIONS: MC(T) numbers, rather than eosinophil numbers, are increased in the SM in patients with EE, express TGF-ß1, and increase the contractility of HESM cells in vitro. As such, mast cells localized to SM in patients with EE might modulate esophageal contractility.