Reduced Mast Cell Infiltration Research Articles

Cell-free fat extract regulates oxidative stress and alleviates Th2-mediated inflammation in atopic dermatitis.

Atopic dermatitis (AD) is a common inflammatory skin disease that significantly affects patients' quality of life. This study aimed to evaluate the therapeutic potential of cell-free fat extract (FE) in AD. In this study, the therapeutic effect of DNCB-induced AD mouse models was investigated. Dermatitis scores and transepidermal water loss (TEWL) were recorded to evaluate the severity of dermatitis. Histological analysis and cytokines measurement were conducted to assess the therapeutic effect. Additionally, the ability of FE to protect cells from ROS-induced damage and its ROS scavenging capacity both in vitro and in vivo were investigated. Furthermore, we performed Th1/2 cell differentiation with and without FE to elucidate the underlying therapeutic mechanism. FE reduced apoptosis and cell death of HaCat cells exposed to oxidative stress. Moreover, FE exhibited concentration-dependent antioxidant activity and scavenged ROS both in vitro and vivo. Treatment with FE alleviated AD symptoms in mice, as evidenced by improved TEWL, restored epidermis thickness, reduced mast cell infiltration, decreased DNA oxidative damage and lower inflammatory cytokines like IFN-γ, IL-4, and IL-13. FE also inhibited the differentiation of Th2 cells in vitro. Our findings indicate that FE regulates oxidative stress and mitigates Th2-mediated inflammation in atopic dermatitis by inhibiting Th2 cell differentiation, suggesting that FE has the potential as a future treatment option for AD.

The effects of a galectin-3 inhibitor on bladder pain syndrome in mice with cyclophosphamide-induced cystitis.

To explore the effect of blocking galectin-3 in the bladder pain syndrome associated with interstitial cystitis. A galectin-3 inhibitor was used to treat mice with cyclophosphamide-induced cystitis. The expression of galectin-3 in bladder tissues and urine was examined by immunohistochemistry and enzyme-linked immunosorbent assay (ELISA), respectively. Suprapubic-pelvic pain, bladder voiding, bladder pain-like nociceptive behavior, and referred hyperalgesia were assessed. The weights of the bladders were also measured, and inflammatory cell infiltration and inflammatory cytokine levels were examined by histopathological evaluation. The inflammatory cytokines interleukin 1β (IL-1β), nerve growth factor (NGF), IL-6, and tumor necrosis factor α (TNF-α) were measured by ELISA. Increases in galectin-3 levels, inflammation, bladder weight, and bladder pain-related symptoms were observed in bladders with cyclophosphamide-induced cystitis. Administration of the galectin-3 inhibitor significantly mitigated bladder pain-related symptoms and inflammatory response. In response to the 500 μM dose of the galectin-3 inhibitor, nociceptive behaviors, nociceptive score, and bladder-to-body weight ratios were reduced by 65.1%, 65.3%, and 40.3%, respectively, while 500 μM Gal-3 inhibitor increased pelvic pain threshold by 86.7%. Moreover, galectin-3 inhibitor treatment inhibited the inflammation. Compared to untreated CYP-induced mice, there were significant changes in the levels of IL-1β (41.72 ± 2.05 vs. 18.91 ± 2.26 pg/mg tissues), NGF (9.64 ± 0.38 vs. 1.88 ± 0.05 pg/mg tissues), IL-6 (42.67 + 1.51 vs. 21.26 + 2.78 pg/mg tissues, and TNF-α (22.02 ± 1.08 vs. 10.70 ± 0.80 pg/mg tissues) in response to the highest dose of the Gal-3 inhibitor subgroup (500 μM), and 500 μM Gal-3 inhibitor reduced mast cell infiltration ratios by 71.8%. The galectin-3 inhibitor relieved pelvic pain, urinary symptoms, and bladder inflammation in mice with cyclophosphamide-induced cystitis. Thus, galectin-3 inhibitors may be novel agents in interstitial cystitis treatment.

HucMSC-Ex alleviates DSS-induced colitis in mice by decreasing mast cell activation via the IL-33/ST2 axis.

Inflammatory bowel disease (IBD) is a chronic inflammatory disease that poses challenges in terms of treatment. The precise mechanism underlying the role of human umbilical cord mesenchymal stem cell-derived exosome (HucMSC-Ex) in the inflammatory repair process of IBD remains elusive. Mucosal mast cells accumulate within the intestinal tract and exert regulatory functions in IBD, thus presenting a novel target for addressing this intestinal disease. A mouse model of Dextran Sulfate Sodium (DSS)-induced colitis was established and hucMSC-Ex were administered to investigate their impact on the regulation of intestinal mast cells. An in vitro co-culture model using the human clonal colorectal adenocarcinoma cell line (Caco-2) and human mast cell line (LAD2) was also established for further exploration of the effect of hucMSC-Ex. We observed the accumulation of mast cells in the intestines of patients with IBD as well as mice. In colitis mice, there was an upregulation of mast cell-related tryptase, interleukin-33 (IL-33), and suppression of tumorigenicity 2 receptor (ST2 or IL1RL1), and the function of the intestinal mucosal barrier related to intestinal tight junction protein was weakened. HucMSC-Ex treatment significantly reduced mast cell infiltration and intestinal damage. In the co-culture model, a substantial number of mast cells interact with the epithelial barrier, triggering activation of the IL-33/IL1RL1 (ST2) pathway and subsequent release of inflammatory factors and trypsin. This disruption leads to aberrant expression of tight junction proteins, which can be alleviated by supplementation with hucMSC-Ex. Our results suggest that hucMSC-Ex may reduce the release of mast cell mediators via the IL-33/IL1RL1 (ST2) axis, thereby mitigating its detrimental effects on intestinal barrier function.

Tenebrio molitor Larva Trypsin Hydrolysate Ameliorates Atopic Dermatitis in C57BL/6 Mice by Targeting the TLR-Mediated MyD88-Dependent MAPK Signaling Pathway.

Atopic dermatitis (AD) is a widely researched chronic inflammatory skin disease with a complex etiology. The increased prevalence of AD necessitates exploration of natural sources as potential therapeutic agents with limited side effects. In the current study, a 1-chloro-2,4-dinitrobenzene (DNCB)-induced AD mouse model was used to examine the anti-AD effects of Tenebrio molitor trypsin hydrolysate (TMTH) and its underlying molecular mechanism. DNCB-treated mice were treated with TMTH (1 and 10 mg/kg), and prednisolone (3 mg/kg) was used as the positive control. Serum and skin tissue samples were collected for subsequent analyses. The expression levels of proteins linked to the myeloid differentiation primary response 88 (MyD88)-dependent mitogen-activated protein kinase (MAPK) signaling pathway and serum IgE levels were estimated via Western blotting technique and ELISA (enzyme-linked immunosorbent assay), respectively. Inflammatory cell infiltration and thickening of the dorsal skin were measured using toluidine blue and hematoxylin and eosin staining, respectively. Oral administration of TMTH significantly reduced mast cell infiltration and dermal and epidermal thickness. Moreover, TMTH treatment reduced serum IgE levels. Western blotting confirmed that TMTH treatment suppressed the MyD88-dependent MAPK signaling pathway. Therefore, TMTH substantially inhibited AD-like skin lesion formation via immunomodulation, showing considerable potential for AD treatment.

Dendrobium candidum polysaccharide reduce atopic dermatitis symptoms and modulate gut microbiota in DNFB-induced AD-like mice.

Atopic dermatitis (AD) is a chronic inflammatory skin disease with a high prevalence worldwide. Increasing evidence suggests that the gut microbiota plays an important role in the pathogenesis of AD. In this study, we sought to verify the effect of Dendrobium candidum polysaccharides (DCP) on AD induced by 2,4-Dinitrofluorobenzene (DNFB) in Balb/c mice regarding its impact on the intestinal microbiome. We found that 2-week oral administration of DCP improved AD-like symptoms and histological damage of skin, reduced mast cell infiltration, down-regulated the level of serum total IgE and the expression of pro-inflammatory cytokines such as TNF-α, IFN-γ, IL-4 and IL-6, and increased the expression level of anti-inflammatory cytokine IL-10. The beneficial effect of DCP was attributed to the restoration of the intestinal microbiome composition and the unbalance of the intestinal homeostasis. Our results indicated that DCP might be used as a promising novel microbiota-modulating agent for the treatment of AD.

7-O-Methylluteolin Suppresses the 2,4-Dinitrochlorobenzene-Induced Nrf2/HO-1 Pathway and Atopic Dermatitis-like Lesions.

7-O-methylluteolin (7-ML) is a flavonoid isolated from the aerial parts of Wikstroemia ganpi (W. ganpi). We describe the anti–atopic dermatitis (AD) effects of 7-ML in tert-butyl hydroperoxide (tBHP)-induced HepG2 cells and 2,4-dinitrochlorobenzene (DNCB)-induced SKH-1 hairless mice. Results demonstrated that 7-ML dose-dependently inhibited the activation of Nrf2 (nuclear factor-erythroid 2-related factor 2) in tBHP-induced HepG2 cells. 7-ML applied topically to our DNCB-induced mouse model upregulated the antioxidant protein expression (phosphorylated Nrf2 (pNrf2), Nrf2, and heme oxygenase-1 (HO-1)) in skin tissues, improved epidermal thickness, and reduced mast cell infiltration into the skin. In addition, 7-ML reduced the serum levels of immunoglobulin E (IgE) and interleukin-4 (IL-4) and improved skin barrier functions. These results suggest that 7-ML should be considered a novel antioxidant and anti-AD agent.

Ethanolic Garcinia mangostana extract and α-mangostin improve dextran sulfate sodium-induced ulcerative colitis via the suppression of inflammatory and oxidative responses in ICR mice

Ethnopharmacological relevanceUlcerative colitis (UC) is an inflammatory disorder of the colon. Garcinia mangostana Linn. (GM) has been traditionally used for its anti-inflammatory and antioxidant activities. Aim of the study:The effects of GM and its bioactive constituent α-mangostin on dextran sulfate sodium (DSS)-induced UC in mice were investigated. Materials and methodsAdult ICR mice (n = 63) were pretreated with ethanolic GM extract at 40, 200, and 1000 mg/kg/day (GM40, GM200, and GM1000), α-mangostin at 30 mg/kg/day, or sulfasalazine at 100 mg/kg/day (SA) for 7 consecutive days. On days 4–7, UC was induced in the mice by the oral administration of DSS (40 kDa, 6 g/kg/day), while control mice received distilled water. The UC disease activity index (DAI) and histological changes were recorded. The activities of myeloperoxidase, catalase, and superoxide dismutase, and the levels of reactive oxygen species (ROS), nitric oxide (NO), and malondialdehyde (MDA) were determined. The mRNA expression of inflammatory related genes including proinflammatory cytokine Tnf-α, Toll-like receptor (Tlr-2), adhesion molecules (Icam-1 and Vcam-1), and monocyte chemoattractant protein (Mcp-1) were evaluated. ResultsTreatment with GM or α-mangostin decreased the UC DAI and protected against colon shortening and spleen and kidney enlargement. GM and α-mangostin prevented histological damage, reduced mast cell infiltration in the colon, and decreased myeloperoxidase activity. GM and α-mangostin increased catalase and superoxide dismutase activity and decreased ROS, NO, and MDA production. GM downregulated mRNA expression of Tnf-α, Tlr-2, Icam-1, Vcam-1, and Mcp-1. ConclusionsGM and α-mangostin attenuated the severity of DSS-induced UC via anti-inflammatory and antioxidant effects. Therefore, GM is a promising candidate for development into a novel therapeutic agent for UC.

AR facilitates YAP-TEAD interaction with the AM promoter to enhance mast cell infiltration into cutaneous neurofibroma

Abundant mast cell infiltration and disease initiation at puberty are hallmark features of cutaneous neurofibroma (cNF). However, the association between mast cell infiltration and steroid hormones in cNF remains unclear. Here, we determined that androgen receptor (AR) expression is positively associated with mast cell density in cNF tissues. Moreover, both in vitro cell experiments and in vivo mouse models verified that activated AR promoted mast cell infiltration and that AR inhibition reduced mast cell infiltration. Analyses in cell models and xenograft tumours both demonstrated that AR upregulated Yes associate protein 1 (YAP)-adrenomedullin (AM) signalling. Clinical samples from cNF patients further verified that AR was positively related to YAP and AM. Mechanistic analysis revealed that AR accelerates AM transcription via enhancing YAP- TEA domain transcription factor (TEAD) binding to the AM promoter. Consequently, the upregulated AM enhanced mast cell recruitment. Interruption of the YAP-TEAD interaction or inhibition of AM could impair mast cell accumulation induced by active AR, which indicated that this newly found signalling pathway may provide novel targets for cNF treatment.

Intranasal nanoemulsion vaccine confers long-lasting immunomodulation and sustained unresponsiveness in a murine model of milk allergy.

Immunotherapy for food allergy requires prolonged treatment protocols and, in most cases, does not lead to durable modulation of the allergic immune response. We have demonstrated an intranasal (IN) nanoemulsion adjuvant that redirects allergen-specific Th2 responses toward Th1 and Th17 immunity, and protects from allergen challenge after only 2-4 monthly administrations. Here, we investigate the ability of this technology to provide long-term modulation of allergy in a murine model of cow's milk allergy. Six weeks after sensitization to bovine casein, mice received four, monthly IN immunizations with nanoemulsion formulated with casein. Protection from casein challenge was assessed at 4 and 16weeks after the final vaccine administration. The NE vaccine significantly blunted the physiological responses to allergen challenge, and this effect persisted for at least 16weeks. The protection from challenge was associated with the suppression of casein-specific Th2 immunity and induced Th1 and Th17 cytokines as well as induction of IL-10. Of interest, while immunized animals showed significantly decreased Th2 cytokine responses, cow's milk-specific IgE remained elevated in the serum at levels associated with reactivity in control sensitized animals. Protection was associated with suppressed mast cell activation and markedly reduced mast cell infiltration into the small intestine. The sustained unresponsiveness of at least 16weeks after vaccination suggests that the nanoemulsion vaccine alters the allergic phenotype in a persistent manner different from traditional desensitization, and this leads to long-term suppressive effects on allergic disease without eliminating serum IgE.

Involvement of the Negative Feedback of IL-33 Signaling in the Anti-Inflammatory Effect of Electro-acupuncture on Allergic Contact Dermatitis via Targeting MicroRNA-155 in Mast Cells.

In this study, we aimed to investigate the effect of electro-acupuncture (EA) at the Zusanli acupoint (ST36) on interleukin (IL)-33-mediated mast cell activation. Firstly, 2,4-dinitrofluorobenzene (DNFB)-induced allergic contact dermatitis (ACD) in rats was developed with or without EA treatment. Then, rat peritoneal mast cells (RPMCs) were obtained and cultured in the presence of IL-33. EA treatment relieved ear swelling and reduced mast cell infiltration in the local inflammation area with DNFB challenge, accompanying the decrement of IL-33 production. RPMCs isolated from ACD rats with EA treatment showed significant downregulation of IL-6, TNF-α, IL-13, and MCP-1 production following IL-33 stimulation. However, there was no obvious difference in surface ST2 receptor expression among different groups. In addition, EA selectively altered IL-33 signaling, suppressing p38 phosphorylation as well as NF-κB- and AP-1-mediated transcription but not Akt phosphorylation. Importantly, EA lowered microRNA (miR)-155 expression in the RPMCs, which presented a positive correlation with IL-33-induced IL-6 production. Furthermore, overexpression of miR-155 in the RPMCs was established following miR-155 mimic transfection. RPMCs with the overexpressed miR-155 displayed an obvious increment of inflammatory cytokine and abrogated the inhibitive effect of EA on NF-κB- and AP-1-regulated transcription in response to IL-33 compared with those without transfected-miR-155. These findings demonstrate EA treatment inhibits NF-κB and AP-1 activation as well as promotes the negative feedback regulation of IL-33 signaling via targeting miR-155 in mast cells, which contribute to the anti-inflammatory effect of EA on DNFB-induced ACD in rats.

Neutralization of Interleukin-9 Decreasing Mast Cells Infiltration in Experimental Autoimmune Encephalomyelitis.

Background:Th9 cells are a newly discovered CD4+ T helper cell subtype, characterized by high interleukin (IL)-9 secretion. Growing evidences suggest that Th9 cells are involved in the pathogenic mechanism of multiple sclerosis (MS). Mast cells are multifunctional innate immune cells, which are perhaps best known for their role as dominant effector cells in allergies and asthma. Several lines of evidence point to an important role for mast cells in MS and its animal models. Simultaneously, there is dynamic “cross-talk” between Th9 and mast cells. The aim of the present study was to examine the IL-9-mast cell axis in experimental autoimmune encephalomyelitis (EAE) and determine its interaction after neutralizing anti-IL-9 antibody treatment.Methods:Female C57BL/6 mice were randomly divided into three groups (n = 5 in each group): mice with myelin oligodendrocyte glycoprotein (MOG)-induced EAE (EAE group), EAE mice treated with anti-IL-9 antibody (anti-IL-9 Abs group), and EAE mice treated with IgG isotype control (IgG group). EAE clinical score was evaluated. Mast cells from central nervous system (CNS) were detected by flow cytometry. The production of chemokine recruiting mast cells in the CNS was explored by reverse transcription-polymerase chain reaction (RT-PCR). In mice with MOG-induced EAE, the expression of IL-9 receptor (IL-9R) complexes in CNS and spleen mast cells was also explored by RT-PCR, and then was repeating validated by immunocytochemistry. In vitro, spleen cells from EAE mice were cultured with anti-IL-9 antibody, and quantity of mast cells was counted by flow cytometry after co-culture.Results:Compared with IgG group, IL-9 blockade delayed clinical disease onset and ameliorated EAE severity (t = −2.217, P = 0.031), accompany with mast cells infiltration decreases (day 5: t = −8.005, P < 0.001; day 15: t = −11.857, P < 0.001; day 20: t = −5.243, P = 0.001) in anti-IL-9 Abs group. The messenger RNA expressions of C-C motif chemokine ligand 5 (t = −5.932, P = 0.003) and vascular cell adhesion molecule-1 (t = −4.029, P = 0.004) were significantly decreased after IL-9 neutralization in anti-IL-9 Abs group, compared with IgG group. In MOG-induced EAE, the IL-9R complexes were expressed in CNS and spleen mast cells. In vitro, splenocytes cultured with anti-IL-9 antibody showed significantly lower levels of mast cells in a dose-dependent manner, compared with splenocytes cultured with anti-mouse IgG (5 μg/ml: t = −0.894, P = 0.397; 10 μg/ml: t = −3.348, P = 0.019; 20 μg/ml: t = −7.639, P < 0.001).Conclusions:This study revealed that IL-9 neutralization reduced mast cell infiltration in CNS and ameliorated EAE, which might be relate to the interaction between IL-9 and mast cells.

TH9 cells are required for tissue mast cell accumulation during allergic inflammation

IL-9 is important for the growth and survival of mast cells. IL-9 is produced by T cells, natural killer T cells, mast cells, eosinophils, and innate lymphoid cells, although the cells required for mast cell accumulation during allergic inflammation remain undefined. We sought to elucidate the role of TH9 cells in promoting mast cell accumulation in models of allergic lung inflammation. Adoptive transfer of ovalbumin-specific TH2 and TH9 cells was used to assess the ability of each subset to mediate mast cell accumulation in tissues. Mast cell accumulation was assessed in wild-type mice and mice with PU.1-deficient T cells subjected to acute and chronic models of allergic inflammation. Adoptive transfer experiments demonstrated that recipients of TH9 cells had significantly higher mast cell accumulation and expression of mast cell proteases compared with control or TH2 recipients. Mast cell accumulation was dependent on IL-9, but not IL-13, a cytokine required for many aspects of allergic inflammation. In models of acute and chronic allergic inflammation, decreased IL-9 levels in mice with PU.1-deficient T cells corresponded to diminished tissue mast cell numbers and expression of mast cell proteases. Mice with PU.1-deficient T cells have defects in IL-9 production from CD4(+) T cells, but not natural killer T cells or innate lymphoid cells, suggesting a TH cell-dependent phenotype. Rag1(-/-) mice subjected to a chronic model of allergic inflammation displayed reduced mast cell infiltration comparable with accumulation in mice with PU.1-deficient T cells, emphasizing the importance of IL-9 produced by T cells in mast cell recruitment. TH9 cells are a major source of IL-9 in models of allergic inflammation and play an important role in mast cell accumulation and activation.

Pharmacologically Antagonizing the CXCR4-CXCL12 Chemokine Pathway with AMD3100 Inhibits Sunlight-Induced Skin Cancer

One way sunlight causes skin cancer is by suppressing anti-tumor immunity. A major mechanism involves altering mast cell migration via the C-X-C motif chemokine receptor 4-C-X-C motif chemokine ligand 12 (CXCR4-CXCL12) chemokine pathway. We have discovered that pharmacologically blocking this pathway with the CXCR4 antagonist AMD3100 prevents both UV radiation-induced immune suppression and skin cancer. The majority of control mice receiving UV-only developed histopathologically confirmed squamous cell carcinomas. In contrast, skin tumor incidence and burden was significantly lower in AMD3100-treated mice. Perhaps most striking was that AMD3100 completely prevented the outgrowth of latent tumors that occurred once UV irradiation ceased. AMD3100 protection from UV immunosuppression and skin cancer was associated with reduced mast cell infiltration into the skin, draining lymph nodes, and the tumor itself. Thus a major target of CXCR4 antagonism was the mast cell. Our results indicate that interfering with UV-induced CXCL12 by antagonizing CXCR4 significantly inhibits skin tumor development by blocking UV-induced effects on mast cells. Hence, the CXCR4-CXCL12 chemokine pathway is a novel therapeutic target in the prevention of UV-induced skin cancer.

Heparanase overexpression reduces carrageenan-induced mechanical and cold hypersensitivity in mice

Heparanase controls the structure and functions of extracellular matrix (ECM) by degrading heparan sulfate proteoglycans. Heparanase is involved in inflammatory process through modulating the functions of inflammatory cytokines. The present study aimed to find out whether overexpression of heparanase in mice affects carrageenan-induced localized inflammation and inflammatory hyperalgesia. Without challenge, the heparanase overexpression did not significantly affect the mice in response to mechanical, cold and heat stimulation. Unilateral subcutaneous administration of carrageenan produced hypersensitivity to mechanical and cold in both wildtype and the heparanase overexpression (Hpa-tg) mice 24h after treatment. In comparison to wildtype animals, the Hpa-tg mice showed significantly reduced mechanical and cold hypersensitivity. This may, at least partially, due to the reduced mast cell infiltration at the site of inflammation in Hpa-tg mice. These data support a role for heparanase that reduces localized inflammation and inflammatory hyperalgesia in mice.

A case of mast cell leukaemia with exon 9 KIT mutation and good response to imatinib

Mastocytosis is a myeloproliferative neoplasm characterized by the excessive proliferation of mast cells. Mast cell leukaemia (MCL), the aggressive form of this disease, requires cytoreductive therapy, such as cladribine, interferon-alpha-2b and, most recently, tyrosine kinase inhibitors - dasatinib or imatinib. We present a case of a 56-yr-old female patient with aleukaemic MCL in whom the typical KIT-D816V mutation was not detected. Sequencing of the entire coding sequence of KIT gene revealed a somatic mutation in exon 9 (p.A502_Y503dup). This mutation was previously reported in patients with gastrointestinal stromal tumours (GIST). Considering the good response to imatinib in such patients, therapy with imatinib was attempted in our patient. The treatment tolerance and outcomes were very good, with reduced mast cell infiltration of the bone marrow, normalization of the serum tryptase concentration and resolution of the clinical signs and symptoms. In the absence of the KIT-D816V in systemic forms of mast cell proliferation, a search for other mutations is indicated, preferably by sequencing the entire KIT gene, as this can influence the choice of treatment. The finding of the p.A502_Y503dup in exon 9, a mutation which has been observed in GIST but not previously reported in any form of aggressive mastocytosis, can be associated with a good response to imatinib in both diseases.

Mast cell leukaemia

In May 2003, a 65-year-old white male presented with weight loss, night sweats, pruritus, diarrhoea, and joint pain. Radiographic studies showed hepatosplenomegaly, abdominal lymphadenopathy and both osteoblastic and osteolytic bone lesions. Bone marrow biopsy showed extensive mast cell infiltration and effacement of marrow structure with reduction of fat cells and marked osteomyelosclerosis (top, Giemsa stain). We administered six courses of therapy with cladribine (0AE1 mg/kg body weight for 5 d, monthly), which reduced mast cell infiltration and restored marrow structure to a nearnormal state (bottom, Giemsa stain). Mediator-related symptoms (night sweats, pruritus, arthralgia, diarrhoea) disappeared completely. Two years after the diagnosis of mast cell leukaemia the patient has a 90–100% Karnofsky performance status.