OBJECTIVE: To compare the serum glycerophospholipid levels in the inflammatory subtypes of asthma by using targeted metabolomic analysis. METHODS: Demographic and clinical data were collected from 51 patients with asthma between January 2015 and December 2015. Routine blood and sputum induction tests were performed. Eosinophilic asthma was defined as induced sputum containing ⪖ 3% eosinophils, and neutrophilic asthma, as induced sputum containing ⪖ 71% neutrophils. Serum metabolic glycerophospholipid profile was determined by liquid chromatography-mass spectrometry. Differences in glycerophospholipid levels between eosinophilic and non-eosinophilic asthma and between neutrophilic and non-neutrophilic asthma were analyzed using partial least squares discriminant analysis. RESULTS: The serum lysophosphatidylglycerol level was significantly higher in the group with ⪖ 3% eosinophils in sputum than in the group with < 3% eosinophils in sputum. The area under the receiver-operating characteristic curve was ⪖ 70%. There was no significant difference in the serum metabolic glycerophospholipid profile between the group with sputum neutrophils ⪖ 71% and the group with sputum neutrophils < 71%. CONCLUSION Serum lysophosphatidylglycerol is produced abundantly in eosinophilic asthma and may be a biomarker of eosinophilic asthma. This information is helpful for identifying and tailoring treatment for the common asthma subtypes.
Adult ; Asthma ; blood ; immunology ; Eosinophils ; immunology ; Female ; Glycerophospholipids ; blood ; Humans ; Male ; Metabolomics ; Middle Aged ; Neutrophils ; immunology ; Sputum ; cytology ; immunology

OBJECTIVETo observed efficacy differences of acupuncture at "Zusanli" (ST 36) in rats with asthma and asthma with spleen-deficency, so as to investigate the therapeutic mechanism.

METHODSSixty SD rats were randomly divided into 5 groups according to their weight, named as an asthma with spleen-deficency group (group A), an acupuncture on asthma with spleen-deficency group (group B), an asthma group (group C), an acupuncture on asthma group (group D) and a control group. The rat models with spleen-deficiency in the first two groups were set up by TCM, then the rats of asthma model in the first four groups were induced by egg albumin, but the control group was treated by the same dose of saline. The group B and the group D were both treated with acupuncture at "Zusanli" (ST 36), once each day for 8 days, and the other groups remained unhandled. The mRNA expressions of Fas and Bcl-2 in the lung tissues were detected by hybridization in situ and apoptosis was detected by TUNEL (terminal dexynucleotidyl transferase-mediated dutp nick end labeling).

RESULTSCompared with the control group, in both the group A and the group C, the expression of Fas mRNA significantly decreased, but the expression of Bcl-2 mRNA significantly increased (all P < 0.01), and eosinophils (EOS) counts significantly increased, but EOS apoptosis rate significantly decreased (all P < 0.01). Compared with the group C, in the group A, the expressions of Fas mRNA significantly decreased, but the expressions of Bcl-2 mRNA and EOS counts significantly increased (all P < 0.01). At the same time, compared with the corresponding asthma groups, in both acupuncture groups, Fas mRNA expression obviously increased, Bcl-2 mRNA expression was significantly reduced (all P < 0.01), EOS counts remarkably decreased and EOS apoptosis rate significantly increased (all P < 0.01). There were no significant differences in the expressions of Fas mRNA and Bcl-2 mRNA between the two acupuncture groups (both P > 0.05), but compared with group B,in the group D, EOS counts significantly decreased and EOS apoptosis rate significantly increased (both P < 0.01).

CONCLUSIONAcupuncture at "Zusanli" (ST 36) can regulate the disorders of Fas mRNA and Bcl-2 mRNA expression in the state of both asthma and asthma with spleen-deficency, promote EOS apoptosis so as to inhibit the development of inflammatory reaction of asthma, showing that acupuncture at "Zusanli" (ST 36) has certain advantages on regulation of related gene of EOS in asthma with spleen-deficency.

Acupuncture Points ; Acupuncture Therapy ; Animals ; Apoptosis ; Asthma ; genetics ; immunology ; physiopathology ; therapy ; Eosinophils ; cytology ; immunology ; Gene Expression ; Humans ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; fas Receptor ; genetics ; immunology

It has been suggested that dendritic cells (DCs) are critical antigen presenting cells for eosinophilic airway inflammation in a mouse model of asthma, and cysteinyl leukotrienes may play a role in DC trafficking in asthmatics. We investigated whether the number of DCs is increased in the induced sputum of both atopic and nonatopic asthmatics and is related to activated eosinophil count in the sputum. Sputum was induced by inhalation of hypertonic saline in 9 atopic and 12 nonatopic asthmatics and 10 nonatopic normal controls, and differential cell counts were performed. DCs and activated eosinophils were identified by immunocytochemistry with monoclonal antibodies (anti-CD1a and EG2, respectively). There were significantly higher percentages of eosinophils, EG2+ cells, and CD1a+ DC in the sputum of atopic and nonatopic asthmatics compared with normal controls, respectively. In asthmatics, the percentage of CD1a+ DC was significantly correlated with that of EG2+ cells (Rs=0.62, p=0.004). We demonstrated that the increased number of DCs was evident in the induced sputum of both atopic and nonatopic asthmatics, and the DC number was related to the activated eosinophil count, which suggests that DCs may contribute to the ongoing eosinophilic inflammation in asthmatic airways, and vice versa.
Adult ; Aged ; Antigens, CD1/analysis ; Asthma/*immunology/pathology ; Comparative Study ; Dendritic Cells/*immunology ; Eosinophil Granule Proteins/analysis ; Eosinophils/cytology/*immunology ; Female ; Humans ; Immunohistochemistry ; Leukocyte Count ; Male ; Middle Aged ; Research Support, Non-U.S. Gov't ; Sputum/cytology/*immunology

OBJECTIVETo investigate the regulation of adipose-derived mesenchymal stem cells (ADSC) on helper T cells and regulatory T cells in allergic rhinitis(AR) mouse model and the underlying mechanisms.

METHODSUsing random number table, 60 Balb/c mice were divided into 6 groups (represented by: sensitized/challenged/treated ), they were the experimental group 1(OVA/OVA/high dose ADSC), the experimental group 2(OVA/OVA/low dose ADSC), the experimental group 3(OVA/OVA/PBS), the experimental group 4(OVA/OVA/0), the control group 1(PBS/PBS/0) and the control group 2(0/0/0). The mouse ADSC were isolated and cultured through conventional method, and AR mouse model was built with OVA and aluminum. The mice were injected with high (3×10(6)), low (1×10(6)) ADSC respectively labeled by CM-Dil for 3 consecutive days via tail-vein injection and sacrificed 48 hours later. Finally, levels of IL-4, IL-6, IL-10 and IFN -γ in serum were examined by ELISA; expressions of the four cytokines in spleen were examined by q RT-PCR; migration of ADSC to mouse model nasal mucosa were observed through fluorescence microscope; eosinophil infiltration were observed by the nasal HE staining.

RESULTSMouse ADSC was isolated, cultured and identified successfully. There was significant difference in symptom scores of AR models (compared with 0/0/0 group, P<0.01). The IL-4 and IL-6 levels of OVA/OVA/high ADSC group were significantly lower than OVA/OVA/0 group (group 1: (17.95±7.78), (27.51±5.93) pg/ml; group 4: (56.82±9.12), (70.03±7.22) pg/ml), the IFN-γ and IL-10 levels increased significantly (group 1: (367.74±13.79), (417.10±72.40) pg/ml; group 4: (199.46±11.25), (122.50±15.57) pg/ml) in serum. These differences were statistically significant(P<0.01). Compared with OVA/OVA/low ADSC group, the IL-4 and IL-6 levels decreased significantly (group 1: (17.95±7.78), (27.51±5.93) pg/ml; group 2: (41.57±12.27), (56.21±9.23)pg/ml) of OVA / OVA / high ADSC group, and the IFN-γ and IL-10 increased significantly (group 1: (367.74±13.79), (417.10±72.40)pg/ml; group 2: (281.77±30.41), (203.45±87.10) pg/ml). These differences were statistically significant(P<0.01). At the same time, the corresponding changes observed at the levels of the cytokines' mRNA. ADSC labeled by CM-Dil could migrate to the mouse nasal mucosa. OVA/OVA/high ADSC group showed the more red fluorescence than the OVA/OVA/low ADSC group. The eosinophils in nasal mucosa of the two groups reduced compared with the normal control.

CONCLUSIONADSC injected via tail-vein can migrate to nasal mucosa and play non-specific immune effects, that may to effect the releases of some cytokines then to regulate the Th1/Th2 imbalance and the function of Treg cell, finally that be dose-related in a certain extent.

Adipose Tissue ; cytology ; Animals ; Cytokines ; blood ; Disease Models, Animal ; Eosinophils ; immunology ; Inflammation ; Mesenchymal Stem Cell Transplantation ; Mice ; Mice, Inbred BALB C ; Nasal Mucosa ; immunology ; Rhinitis, Allergic ; immunology ; therapy ; T-Lymphocytes, Helper-Inducer ; immunology ; T-Lymphocytes, Regulatory ; immunology

OBJECTIVETo explore the feature of nasal mucosa remodeling in experimental allergic rhinitis.

METHODSTwenty-four male Hartley guinea pigs (4 weeks, 250 -300 g) were randomly divided into four groups (control group and allergen exposure groups 1 - 3), each group had 6 guinea pigs. Allergen exposure animals were sensitized by intraperitoneal (ip) injection of ovalbumin (OVA). Sensitized guinea pigs were subjected to either brief or prolonged exposure to allergen. Both brief exposure group (allergen exposure groups) and prolonged exposure group (allergen exposure group 2 and 3) received a daily intranasal challenge with 5% OVA in 0.9% saline from Day 22 to Day 28, the prolonged exposure group (allergen exposure group 2 and 3) followed by twice weekly exposure to 5% OVA intranasal for an additional 8 and 12 weeks respectively. Control animals were given saline only. At 24 h after the last intranasal challenge, the guinea pigs were killed and the heads of the animals were removed and fixed in 10% neutral buffered formalin for 24 hours, then decalcified in 5% trichloroacetic acid for 10 days. The tissue blocks were embedded in paraffin. The paraffin sections 3 microm thick were stained with hematoxylin and eosin (HE), alcian blue (pH, 2. 6)-periodic acid-Schiff (AB-PAS), and Masson's Trichrome (MT). The infiltrating eosinophils in nasal mucosa were examined, AB-PAS-positive cells in the surface epithelium in nasal septal mucosa were counted. The percentage area of MT stained extracellular matrix in septal mucosa and conchae and damage of epithelium were determined by an image analyzer.

RESULTSThe control group only presented a few eosinophils. Significant eosinophil infiltration was observed in the sensitized groups. Compared with control group (intact epithelium 87.7% +/- 11.1%), there was no significant epithelial damage in 1 week exposure group. Significant epithelial damage were observed in 8 and 12 weeks groups (intact epithelium 36.7% +/- 16.9%, 37.9% +/- 12.9%, respectively). An increase in AB-PAS-positive cells was observed in the mucosa of nasal septum in the prolonged allergen exposure groups, but not in the brief allergic inflammation group in comparison with the control. The brief OVA exposure group did not show increased collagen fibrils within the mucosa of nasal septum and conchae. In contrast, after prolonged OVA exposure an increase in matrix was observed. Furthermore, in both the nasal septum and conchae, significant increasing of ECM deposition was found in a further prolonged exposure for 12 weeks compared to 8 weeks.

CONCLUSIONSEpithelial damage, goblet cells hyperplasia and extracellular matrix deposition were observed as the features of remodeling in this guinea pig model of allergic rhinitis.

Animals ; Disease Models, Animal ; Eosinophils ; immunology ; Epithelial Cells ; pathology ; Extracellular Matrix ; pathology ; Goblet Cells ; pathology ; Guinea Pigs ; Male ; Mice ; Nasal Mucosa ; cytology ; pathology ; Rhinitis, Allergic, Perennial ; immunology ; pathology

BACKGROUNDAsthma is a chronic airway disease with inflammation characterized by physiological changes (airway hyper-responsiveness, AHR) and pathological changes (inflammatory cells infiltration and mucus production). Eosinophils play a key role in the allergic inflammation. But the causative relationship between eosinophils and airway inflammation is hard to prove. One of the reasons is lack of activation marker of murine eosinophils. We investigated the expression of CD69 on murine eosinophils in vitro, the relationship between the expression of CD69 on eosinophils from peripheral blood and bronchoalveolar lavage fluid and on airway inflammation in asthmatic mice.

METHODSEosinophils from peripheral blood of IL-5 transgenic mice (NJ.1638) were purified. Mice were divided into five groups: wild type mice sensitized and challenged with saline (WS group), wild type mice sensitized and challenged with ovalbumin (WO group), IL-5(-/-) mice sensitized and challenged with saline and transferred with purified eosinophils (ISE group), IL-5(-/-) mice sensitized and challenged with OVA and transferred with purified eosinophils (IOE group), IL-5(-/-) mice sensitized and challenged with OVA and transferred with purified eosinophils, pretreated with anti CD4 monoclonal antibody (IOE+antiCD4mAb group). IL-5(-/-) mice were sensitized with OVA at day 0 and day 14, then challenged with OVA aerosol. On days 24, 25, 26 and 27 purified eosinophils were transferred intratracheally to IL-5(-/-) mice. On day 28, blood and BALF were collected and CD69 expression on eosinophils measured by flowcytometry.

RESULTSPurified eosinophils did not express CD69. But eosinophils cultured with PMA + MA, IFN-gamma, IL-5 or GM-CSF expressed CD69 strongly. Eosinophils from blood of WO, WS group did not express CD69 at all. The numbers of eosinophils in BALF of WO group, IOE group, ISE group and IOE + antiCD4mAb group were significantly higher than in mice of WS group which did not have eosinophils at all. CD69 expression on eosinophils in BALF of IOE and WO groups was strong. Eosinophils in BALF of ISE and IOE + antiCDmAb groups did not express CD69. The mucus production result was similar to CD69 expression. There were eosinophils infiltration in lung slides of all groups except WS group.

CONCLUSIONActivation in airway of eosinophils could directly lead to airway inflammation.

Animals ; Antigens, CD ; analysis ; Antigens, Differentiation, T-Lymphocyte ; analysis ; Asthma ; immunology ; physiopathology ; Bronchoalveolar Lavage Fluid ; cytology ; Eosinophils ; immunology ; Inflammation ; physiopathology ; Lectins, C-Type ; Lung ; physiopathology ; Mice ; Mice, Transgenic

OBJECTIVE: To investigate the relationship between airborne allergen and patients with nasal polyps (NP). METHOD: Eighty-three cases of patients with NP were classified into the eosinophilic group(ENP) and non eosinophilic group (nENP) in terms of the EOS count in NP specimen, all patients underwent specific-IgE test with standardized allergens and the results were analyzed. RESULT: The specific-IgE positive rate was 59.26% in ENP group and 23.21% in the nENP group respectively (P < 0.01), and the positive rate of ENP was statistical significance compared with nENP. The dust mite was the most prevalent airborne allergen in ENP group. CONCLUSION The perennial airborne allergens may play a certain role in the etiology and pathogenesis of NP.
Adult ; Allergens ; immunology ; Animals ; Eosinophils ; cytology ; Female ; Humans ; Immunoglobulin E ; blood ; Leukocyte Count ; Male ; Middle Aged ; Nasal Polyps ; blood ; immunology ; Young Adult

Development of alternatively activated (M2) macrophage phenotypes is a complex process that is coordinately regulated by a plethora of pathways and factors. Here, we report that RBP-J, a DNA-binding protein that integrates signals from multiple pathways including the Notch pathway, is critically involved in polarization of M2 macrophages. Mice deficient in RBP-J in the myeloid compartment exhibited impaired M2 phenotypes in vivo in a chitin-induced model of M2 polarization. Consistent with the in vivo findings, M2 polarization was partially compromised in vitro in Rbpj-deficient macrophages as demonstrated by reduced expression of a subset of M2 effector molecules including arginase 1. Functionally, myeloid Rbpj deficiency impaired M2 effector functions including recruitment of eosinophils and suppression of T cell proliferation. Collectively, we have identified RBP-J as an essential regulator of differentiation and function of alternatively activated macrophages.
Animals ; Cell Polarity ; drug effects ; genetics ; immunology ; Cell Proliferation ; drug effects ; genetics ; Chitin ; immunology ; pharmacology ; Eosinophils ; cytology ; immunology ; Gene Expression Regulation ; drug effects ; immunology ; Immunoglobulin J Recombination Signal Sequence-Binding Protein ; genetics ; immunology ; Macrophage Activation ; drug effects ; genetics ; Macrophages ; cytology ; immunology ; Mice ; Mice, Transgenic ; T-Lymphocytes ; cytology ; immunology

Corticosteroids are considered to be one of the most effective medicine for asthma by suppressing airway inflammation. This study was carried out to investigate the effects of prednisolone in the sputum of exacerbated asthmatics. Clinical severity, cell differentials, levels of interleukin (IL)-5, eosinophil cationic protein (ECP), EG2+ eosinophils, and nitric oxide (NO) metabolites were measured. Sputum was examined 2 weeks apart in 13 exacerbated asthmatics before and after prednisolone treatment, and once in 12 stable asthmatics. We used a sandwich ELISA for IL-5, fluoroimmunoassay for ECP, immunohistochemical staining for EG2+ eosinophils, a NO metabolites assay using modified Griess reaction. Exacerbated asthmatics, in comparison with stable asthmatics, had significantly higher proportion of eosinophils, higher level of ECP, higher percentage of EG2+ eosinophils, and NO metabolites. Exacerbated asthmatics after treatment with prednisolone had reduced the proportions of eosinophils, reduced level of IL-5, ECP and percentage of EG2+ eosinophils. FEV1 was correlated with the proportion of eosinophils, ECP, and IL-5 respectively. These findings suggest that prednisolone is considered to be effective medicine for asthma by suppressing eosinophil activation through IL-5.
Administration, Oral ; Adolescence ; Adrenal Cortex/metabolism ; Adult ; Aged ; Anti-Inflammatory Agents, Steroidal/administration & dosage* ; Asthma/metabolism ; Asthma/immunology* ; Asthma/drug therapy* ; Biological Markers ; Blood Proteins/metabolism* ; Eosinophils/metabolism ; Eosinophils/immunology ; Eosinophils/drug effects* ; Female ; Human ; Interleukin-5/metabolism* ; Leukocyte Count ; Male ; Middle Age ; Nitric Oxide/metabolism ; Prednisolone/administration & dosage* ; Sputum/immunology ; Sputum/cytology

OBJECTIVEAllergic asthma is thought to be mediated by CD4+ T lymphocytes producing the Th2-associated cytokines, which play a critical role in the development of the airway hyper-responsiveness and the eosinophilic inflammatory response. The costimulatory pathway CD28/B7 has been shown to play an important role in CD4+ T cell activation in allergic asthma. This study was conducted to evaluate the role of another costimulatory pathway OX40/OX40 ligand (L) in the pathogenesis of allergic asthma in BALB/c mice.

METHODSAn allergic asthma model in BALB/c mice was established. Thirty-six BALB/c mice were randomly divided into three groups with 12 in each. Mice in treatment group (group B) were treated with neutralizing anti-OX40L monoclonal antibody (mAb, 300 microg per mouse) during the sensitization period. Mice in two control groups, asthma model group (group A) and IgG antibody group (group C) were treated with normal saline (NS) and control IgG respectively instead of anti-OX40L mAb. Bronchoalveolar lavage fluid (BALF) was collected from the mice of each group for counting the total number of white blood cells (including neutrophil granulocyte, lymphocyte, monocyte and eosinophil granulocyte) and the proportions of these cells. The levels of IL-4 and INF-gamma in BALF were measured by ELISA. Lungs were removed for morphological examination after HE and PAS staining, and expression of OX40 in lungs was evaluated by immunohistochemical method.

RESULTS(1) The count of total number of white blood cells in BALF (x10(6)/ml) was lower in group B than that of group A and group C (26.6 +/- 4.6 vs. 36.8 +/- 5.2 and 34.3 +/- 6.9, respectively), the difference between the treatment group (group B) and two control groups (groups A and C) was significant; The proportions of eosinophils and lymphocytes in the BALF (%) were lower in group B than those in group A and group C (eosinophils 15.1 +/- 2.6 vs. 20.0 +/- 4.1 and 19.9 +/- 3.9, respectively; lymphocytes 7.0 +/- 0.9 vs. 8.9 +/- 1.6 and 8.6 +/- 1.8, respectively), the difference between the treatment group and two control groups was significant. (2) The IL-4 level in BALF (pg/ml) was lower in group B than that in group A and group C (672 +/- 58 vs. 809.57 +/- 106.00 and 784 +/- 58, respectively), but the INF-gamma levels in BALF (pg/ml) were higher than those in group A and group C (0.86 +/- 0.09 vs. 0.69 +/- 0.15 and 0.67 +/- 0.13 respectively), and all the differences were statistically significant. (3) The expression of OX40 in the lungs of mice in group B were at a lower level than that of group A and group C, and the morphological changes of asthma were ameliorated in the mice of the treatment group. The signs of mice in treatment group were obviously ameliorated as compared to the two control groups.

CONCLUSIONBlocking the costimulatory pathway by administering the neutralizing anti-OX40L mAb during the sensitization period of allergic asthma model could balance the Th1/Th2 responses, inhibit lung inflammation and ameliorate the signs of mice model of asthma.

Animals ; Antibodies, Monoclonal ; administration & dosage ; immunology ; pharmacology ; Antigens, Differentiation ; immunology ; metabolism ; Asthma ; immunology ; metabolism ; pathology ; therapy ; Bronchoalveolar Lavage Fluid ; cytology ; immunology ; Disease Models, Animal ; Enzyme-Linked Immunosorbent Assay ; Eosinophils ; immunology ; Female ; Immunoglobulin G ; administration & dosage ; immunology ; pharmacology ; Immunohistochemistry ; Interferon-gamma ; analysis ; immunology ; Interleukin-4 ; analysis ; immunology ; Leukocyte Count ; Leukocytes ; immunology ; Lung ; drug effects ; immunology ; pathology ; Lymphocytes ; immunology ; Membrane Glycoproteins ; immunology ; Mice ; Mice, Inbred BALB C ; Ovalbumin ; toxicity ; Tumor Necrosis Factors ; immunology