Inhibition of IL-13-induced periostin in airway epithelium attenuates cellular protein expression of MUC5AC.

BACKGROUND AND OBJECTIVE: Serum periostin is increased in asthma and serves as a surrogate marker for IL-13 activity in the lung. Serum levels of periostin are the most robust biomarker predicting a favourable response to the anti-IL-13 drug, lebrikizumab. We investigated the mechanisms of IL-13 stimulation of periostin, the polarized secretion of periostin and whether periostin would have a direct effect on mucin secretion by airway cells. METHODS: Normal human bronchial epithelial (NHBE) cells were cultured at air-liquid interface (ALI) in the presence of IL-13, and we evaluated the effect of the specific inhibitors, leflunomide (Janus kinase (JAK)/signal transducer and activator of transcription factor 6 (STAT6) inhibitor) or PD98059 (MEK/extracellular regulated protein kinase (ERK) inhibitor), on periostin production. We examined MUC5AC secretion from NHBE cells exposed to recombinant human (rh) periostin or IL-13 in the presence and absence of OC-20, a periostin-neutralizing antibody. RESULTS: IL-13 induced periostin protein which was predominantly secreted towards the basal surface of the cells. Periostin production was much greater from goblet cells than ciliated cells (P < 0.001). Periostin production after exposure to IL-13 was attenuated by both leflunomide (P < 0.001) and PD98059 (P < 0.001). The addition of exogenous periostin modestly increased MUC5AC secretion (P < 0.01), but did not visibly change cell morphology. IL-13-induced MUC5AC secretion was attenuated by OC-20 (P < 0.01). CONCLUSION: Periostin production in differentiated airway cells is mediated by JAK/STAT6 and MEK/ERK pathways. Periostin secretion is much greater from immunologically active goblet cells. IL-13-driven mucin production is partially inhibited by OC-20.

Suppression of neuropeptide production by quercetin in allergic rhinitis model rats.

BACKGROUND: Quercetin, a dietary flavonoid found in many fruits, red wine and onion, among others, has been reported to have potent anti-oxidant, anti-viral and anti-cancer effects. Although quercetin is also reported to have anti-inflammatory and anti-allergic effects, the precise mechanisms by which quercetin favorably modify the clinical conditions of allergic diseases such as allergic rhinitis (AR). The present study was designed to examine the influence of quercetin on the development of AR by using AR model rats. METHODS: Sprague-Dawley (SD) rats were sensitized with toluene 2,4-diisocyanate (TDI) by intranasal instillation of a 10 % TDI in ethyl acetate in a volume of 5 µl once a day for 5 consecutive days. This sensitization procedure was repeated after a 2-day interval. After 5 days of the second sensitization, rats were treated with various doses of quercetin once a day for 2 to 7 days. Nasal allergy-like symptoms, which were induced by bilateral application of 5 µl of 10 % TDI in ethyl acetate, were assessed by counting sneezing and nasal rubbing behaviors for 10 min just after TDI nasal challenge. The levels of substance P (SP), calcitonin gene-related peptide (CGRP) and nerve growth factor (NGF) in nasal lavage fluids obtained 6 h after TDI nasal challenge was examined by ELISA. RESULTS: Oral administration of quercetin for 5 and 7 days, but not 2 and 3 days, could inhibit sneezing and nasal rubbing movements, which were increased by TDI nasal challenge. The minimum dose that caused significant inhibition was 25 mg/kg. Oral administration of quercetin at more than 25 mg/kg for 5 days significantly inhibited the increase in SP, CGRP and NGF contents in nasal lavage fluids induced by TDI nasal challenge. CONCLUSION: The present results strongly suggested that quercetin will be a good candidate for the supplement on the management and treatment of allergic diseases, especially AR.

Club cell 10-kDa protein attenuates airway mucus hypersecretion and inflammation.

Bacterial lipopolysaccharide (LPS) and interleukin (IL)-13 increase mucus secretion and inflammatory cytokine production in normal human bronchial epithelial (NHBE) cells. We evaluated the effect of club cell 10-kDa protein (CC10), an anti-inflammatory protein produced by epithelial cells, on mucus secretion, cell morphology and inflammatory cytokine production. NHBE cells were cultured at an air-liquid interface with CC10 or vehicle and exposed to LPS on day 14. Mucin MUC5AC, IL-8 and granulocyte-macrophage colony-stimulating factor were measured in cell supernatants. MUC5AC and IL-8 mRNA expression were measured by real-time PCR. Western blotting was used to evaluate nuclear factor (NF)-κB and extracellular signal-regulated kinase (ERK) activation. Cells were evaluated histologically. Additionally, NHBE cells were exposed to IL-13 and CC10 for 14 days, and secretion of the mucins MUC5AC and MUC5B was measured. MUC5AC secretion stimulated either by LPS or by IL-13 was attenuated by CC10 at 20 ng·mL(-1) (p<0.05). CC10 at 20 ng·mL(-1) also attenuated IL-8 secretion (p<0.05). MUC5AC and IL-8 mRNA expression were also decreased by CC10 (p<0.05). CC10 attenuated phosphorylation of NF-κB (p<0.05) and ERK1/2 (p<0.05). CC10 attenuates LPS-induced mucus secretion in airway cells, in part due to inhibition of NF-κB and ERK phosphorylation.

Quercetin-Induced Enhancement of Nasal Epithelial Cells' Ability to Produce Clara Cell 10-kD Protein In Vitro and In Vivo.

Background: Quercetin, a polyphenolic flavonoid found in various plants and foods, is known to have antioxidant, antiviral and anticancer effects. Although quercetin is well known to exert anti-inflammatory and anti-allergic effects, the precise mechanisms by which quercetin favorably modifies the clinical status of allergic diseases, such as allergic rhinitis (AR), remain unclear. The present study examined whether quercetin could modulate the production of the endogenous anti-inflammatory molecule, Clara cell 10-kD protein (CC10), in vitro and in vivo. Methods: Human nasal epithelial cells (1 × 105 cells/mL) were stimulated with 20 ng/mL of tumor necrosis factor-alpha (TNF) in the presence of quercetin for 24 h. CC10 levels in culture supernatants were examined by ELISA. Sprague Dawley rats were sensitised with toluene 2,4-diisocyanate (TDI) by intranasal instillation of 10% TDI in ethyl acetate at a volume of 5.0 µL once daily for five days. This sensitisation procedure was repeated after an interval of two days. The rats were treated with different dosages of quercetin once daily for five days starting on the 5th day following the second sensitization. Nasal allergy-like symptoms induced by the bilateral application of 5.0 µL of 10% TDI were assessed by counting sneezing and nasal-rubbing behaviours for 10 min immediately after the TDI nasal challenge. The levels of CC10 in nasal lavage fluids obtained 6 h after TDI nasal challenge were examined using ELISA. Results: The treatment of cells with low doses of quercetin (<2.5 µM) scarcely affected TNF-induced CC10 production from nasal epithelial cells. However, the ability of nasal epithelial cells to produce CC10 after TNF stimulation significantly increased on treatment with quercetin doses (>5.0 µM). The oral administration of quercetin (>25 mg/kg) for five days significantly increased the CC10 content in nasal lavage fluids and attenuated the nasal symptoms induced by the TDI nasal challenge. Conclusions: Quercetin inhibits AR development by increasing the ability of nasal epithelial cells to produce CC10.

Suppressive effect of juzentaihoto on vascularization induced by b16 melanoma cells in vitro and in vivo.

Juzentaihoto (JTT) is well known to be one of Japanese herbal medicines, and used for the supplemental therapy of cancer patients with remarkable success. The present study, therefore, was undertaken to examine the possible therapeutic mechanisms of JTT on cancer using B16 melanoma cell (B16 cell)/experimental mouse system. JTT was well mixed with rodent chow at 3.0% concentrations, and was administered orally ad libitum. Administration of JTT was started one week before tumor cell injection and continued throughout the experiment. Administration of JTT into mice significantly inhibited tumor metastasis in lungs after intravenous injection of 2 × 10(5) B16 cells in a volume of 50 µL. JTT also significantly suppressed enlargement of tumor size in hind footpad after the subcutaneous injection of 2 × 10(5) (50 µL) B16 cells. In the second part of experiments, the chamber that containing B16 cells was buried in the murine back. In JTT administrated group, vascular endothelial growth factor (VEGF) of chamber internal fluid significantly decreased, and vascularization of chamber circumference was also inhibited. These results strongly suggest that oral administration of JTT caused decrease in the generation of VEGF, which is responsible for vascularization, and results in inhibition of B16 cell metastasis.

Enhancement of clara cell 10-kD protein (CC10) production from nasal epithelial cells by fexofenadine hydrochloride.

BACKGROUND: Clara cell 10-kD protein (CC10) is well known to be an immuno-suppressive protein secreted from airway epithelial cells after inflammatory stimulation and is involved in the development of allergic disorders. Although histamine H1 receptor antagonists are used for the treatment of allergic disorders, the influence of the agents on CC10 production is not well understood. In the present study, we examined the influence of a histamine H1 receptor antagonist, fexofenadine hydrochloride (FEX) on CC10 production in vitro and in vivo. METHODS: Nasal epithelial cells (5 x 10(6) cells/ml) were stimulated with 20 ng/ml TNF-alpha in the presence of various concentrations of FEX for 24 hours. CC10 levels in culture supernatants were examined by ELISA. Patients with Japanese cedar pollinosis were treated orally with FEX twice a day at a single dose of 60 mg for two weeks during Japanese cedar pollen season (February 2011 to April 2011). CC10 levels in nasal secretions were also examined by ELISA. RESULTS: The addition of FEX into cell cultures caused increase in CC10 production induced by TNF-alpha stimulation, and the minimum concentration that caused significant increase was 200 ng/ml. Oral administration of FEX also increased CC10 levels in nasal secretions from pollinosis patients along with attenuation of clinical symptoms. CONCLUSION: The ability of FEX to enhance CC10 production may account, at least in part, for the clinical efficacy of the agent in allergic disorders, including allergic rhinitis.

Effect of histamine H1 receptor antagonists on TARC/CCL17 and MDC/CCL22 production from CD14+ cells induced by antigenic stimulation in vitro.

BACKGROUND: Thymus- and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC) are accepted to be important molecules in the development and maintenance of allergic diseases. Although several types of histamine H(1) receptor antagonist (antihistamine) have been developed and used for the treatment of allergic diseases, the influence of antihistamines on TARC and MDC production is not well understood. OBJECTIVE: The present study was undertaken to examine the influence of antihistamines on TARC and MDC production from CD14+ cells after antigenic stimulation in vitro. METHODS: CD14+ cells prepared from patients with pollinosis to Japanese cedar pollen were stimulated with specific allergen extracted from Japanese cedar pollen (Cry j 1) in the presence of azelastine (AZE), ketotifen (KET), fexofenadine (FEX) and oxatomide (OXA) for 6 days. TARC and MDC levels in culture supernatants were examined by ELISA. We also examined the influence of FEX on TARC and MDC mRNA expression, phosphorylation of mitogen-activated protein kinases (MAPKs) and transcription factor activation in CD14+ cells after Cry j 1 stimulation. RESULTS: FEX at 250 ng/ml, which is almost equal to therapeutic blood levels, caused a significant inhibition of TARC and MDC production.However, AZE, OXA and KET required higher concentrations than their therapeutic blood levels to suppress production of these factors. FEX at 250 ng/ml also suppressed NF-κB activation, phosphorylation of p38 MAPK and extracellular signal-regulated kinases 1 and 2 and expression of mRNA for TARC and MDC. CONCLUSIONS: These results suggest that antihistamines, especially FEX, suppress CC chemokine production from CD14+ cells through interference with antigen-mediated signaling and result in favorable modification of allergic disease states or conditions.

Suppressive effect of juzen-taiho-to on lung metastasis of b16 melanoma cells in vivo.

Juzen-Taiho-To (JTT) is well known to be one of Kampo (Japanese herbal) medicine consisted of 10 component herbs and used for the supplemental therapy of cancer patients with remarkably success. However, the precise mechanisms by which JTT could favorably modify the clinical conditions of cancer patients are not well defined. The present study, therefore, was undertaken to examine the possible mechanisms of JTT on prevention of cancer metastasis using experimental mouse model. JTT was well mixed with rodent chow at concentrations of either 0.2 or 1.0%, and administered orally ad libitum, which was started 1 week before tumor cell injection and continue throughout the experiment. Oral administration of JTT at concentration 0.2 and 1.0% into C57BL/6 male mice significantly inhibited tumor metastasis in lungs, which was induced by the intravenous injection of 2 × 10(5) B16 melanoma cell. JTT at a concentration of 1.0% also significantly suppressed lung metastasis of B16 melanoma cell from hind footpad in C57BL/6 mice. In the second part of experiments, the influence of the depression of natural killer (NK) cell, natural killer T (NKT) cell and several types of cytokines on JTT-mediated inhibition of tumor cell metastasis. Intraperitoneal injection of anti asialo-GM1 antibody against NK cells and anti NK-1.1 monoclonal antibody (mAb) to NKT cells abrogated the inhibitory action of JTT on lung metastasis of B16 melanoma cells. Although intraperitoneal administration of anti-IFN-γ mAb scarcely affected the inhibitory action of JTT on tumor cell metastasis, injection of amrinone, which used for IL-12 suppression, significantly decreased the ability of JTT to prevent tumor cell metastasis. These results strongly suggest that oral administration of JTT caused increase in the production of IL-12, which is responsible for the activation of both NK cell and NKT cell, in the lungs and results in inhibition of B16 melanoma cell metastasis in the lungs.

Inhibition of Angiogenic Factor Productions by Quercetin In Vitro and In Vivo.

Background: Angiogenesis is well known to be an important event in the tissue remodeling observed in allergic diseases. Although there is much evidence that quercetin, one of the most abundant dietary flavonoids, exerts anti-allergic effects in both human and experimental animal models of allergic diseases, the action of quercetin on angiogenesis has not been defined. Therefore, in this study, we first examined the action of quercetin on the secretion of angiogenic factors from murine mast cells in vitro. We also examined the action of quercetin on angiogenic factor secretion in the murine allergic rhinitis model in vivo. Methods: Mast cells (1 × 105 cells/mL) sensitized with ovalbumin (OVA)-specific murine IgE were stimulated with 10.0 ng/mL OVA in the presence or the absence of quercetin for 24 h. The concentrations of angiogenic factors, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), tumor necrosis factor-α, IL-6 and IL-8 in the supernatants were examined by ELISA. BALB/c male mice immunized with OVA were challenged intranasally with OVA every other day, starting seven days after the final immunization. These mice were then orally administered quercetin once a day for five days, starting seven days after the final immunization. Clinical symptoms were assessed by counting the number of sneezes and nasal rubbing behaviors during the 10 min period just after OVA nasal provocation. The angiogenic factor concentrations in the nasal lavage fluids obtained 6 h after nasal antigenic provocation were examined by ELISA. Results: Quercetin significantly inhibited the production of angiogenetic factors induced by IgE-dependent mechanisms at 5.0 µM or more. Oral administration of 25.0 mg/kg quercetin into the mice also suppressed the appearance of angiogenetic factors in nasal lavage fluids, along with the attenuation of nasal symptoms. Conclusions: These results strongly suggest that the inhibitory action of quercetin on angiogenic factor secretion may be implicated in the therapeutic action of quercetin on allergic diseases, especially allergic rhinitis.

Modulation of Th1/Th2 Cytokine Balance by Quercetin In Vitro.

Background: Allergic rhinitis (AR) is well known to be an IgE-mediated chronic inflammatory disease in the nasal wall, which is primarily mediated by Th2-type cytokines such as IL-4, IL-5, and IL-13. Although quercetin is also accepted to attenuate the development of allergic diseases such as AR, the influence of quercetin on Th2-type cytokine production is not well understood. The present study was designed to examine whether quercetin could attenuate the development of AR via the modulation of Th2-type cytokine production using an in vitro cell culture technique. Methods: Human peripheral-blood CD4+ T cells (1 × 106 cells/mL) were cultured with 10.0 ng/mL IL-4 in the presence or absence of quercetin. The levels of IL-5, IL-13, and INF-γ in 24 h culture supernatants were examined by ELISA. The influence of quercetin on the phosphorylation of transcription factors NF-κB and STAT6, and mRNA expression for cytokines were also examined by ELISA and RT-PCR, respectively. Results: Treatment of cells with quercetin at more than 5.0 µM inhibited the production of IL-5 and IL-13 from CD4+ T cells induced by IL-4 stimulation through the suppression of transcription factor activation and cytokine mRNA expression. On the other hand, quercetin at more than 5.0 µM abrogated the inhibitory action of IL-4 on INF-γ production from CD4+ T cells in vitro. Conclusions: The immunomodulatory effects of quercetin, especially on cytokine production, may be responsible, in part, for the mode of therapeutic action of quercetin on allergic diseases, including AR.

Evidence for passing down of postnasal drip into respiratory organs.

UNLABELLED: Postnasal drip is believed to be one of the main sources of serious respiratory diseases, such as sinobronchial syndrome. However, there is little direct evidence showing that postnasal drip flows into the trachea and results in the development of inflammatory responses in the lower airway. In the present study, whether postnasal drip entered the respiratory organs and whether material in the trachea moved toward the lungs and the digestive organs were examined by using an experimental model with mice. MATERIALS AND METHODS: In the first set of experiments, 1.0 microL of 51Cr-labeled pseudo-postnasal drip in a normal saline or a glycerin solution was instilled into the nasal cavity of male ICR mice anesthetized with sodium barbital by intraperitoneal injection. In the second set of experiments, the destination of 51Cr-labeled red blood cells (RBCs) after intratracheal instillation was examined in the anesthetized mice. The lungs, the stomach and the intestines were removed from mice killed under anesthesia at various intervals after instillation, and measured for radioactivity. RESULTS: When glycerin solution containing 51Cr (but not normal saline) was instilled in mice, the presence of much higher levels of 51Cr was observed in the lungs. Although the presence of high levels of 51Cr-labeled RBCs was observed in the lungs one hour after instillation radioactivity in the lungs gradually decreased as time went by. On the other hand, radioactivity in the digestive organs gradually increased and peaked three hours after instillation with 51Cr-labeled RBC. CONCLUSION: These results suggest that thicker viscous postnasal drip can flow into the respiratory organs when the host is asleep. In addition, postnasal drip which flows into the trachea can move gradually to the oral side by mucociliary transportation of the tracheal mucosa and thus be swallowed.

Influence of epinastine hydrochloride, an H1-receptor antagonist, on the function of mite allergen-pulsed murine bone marrow-derived dendritic cells in vitro and in vivo.

There is established concept that dendritic cells (DCs) play essential roles in the development of allergic immune responses. However, the influence of H(1) receptor antagonists on DC functions is not well defined. The aim of the present study was to examine the effect of epinastine hydrochloride (EP), the most notable histamine H(1) receptor antagonists in Japan, on Dermatophagoides farinae (Der f)-pulsed mouse bone marrow-derived DCs in vitro and in vivo. EP at more than 25 ng/mL could significantly inhibit the production of IL-6, TNF-alpha and IL-10 from Der f-pulsed DCs, which was increased by Der f challenge in vitro. On the other hand, EP increased the ability of Der f-pulsed DCs to produce IL-12. Intranasal instillation of Der f-pulsed DCs resulted in nasal eosinophilia associated with a significant increase in IL-5 levels in nasal lavage fluids. Der f-pulsed and EP-treated DCs significantly inhibited nasal eosinophila and reduced IL-5. These results indicate that EP inhibits the development of Th2 immune responses through the modulation of DC functions and results in favorable modification of clinical status of allergic diseases.

Suppressive Activity of Adiponectin on the Development of Allergic Rhinitis in Mice.

BACKGROUND/AIM: Adiponectin is accepted as playing pivotal roles in the development of allergic rhinitis (AR) through modulation of production of inflammatory mediators. Although it is also well known that neuropeptides, especially substance P (SP), function in the development and persistence of clinical conditions of AR, the influence of adiponectin on neuropeptide production is not well understood. The present study was designed to examine the influence of adiponectin on the production of SP both in vivo and in vitro. MATERIALS AND METHODS: PC-12 cells (1×104 cells) were stimulated with 10.0 ng/ml nerve growth factor (NGF) for 2 h and then with 10.0 ng/ml capsaicin in the presence of different concentrations of adiponectin. After 72 h, culture supernatants were obtained, and SP levels were measured with enzyme-linked immunosorbent assay (ELISA). The influence of adiponectin on the total number of neurites developed per PC-12 cell and on the percentage of PC-12 cells with outgrowing neurites was also examined 24 and 72 h after the start of culture, respectively. In the second part of the study, BALB/c mice were sensitized intraperitoneally with 1.0 µg of ovalbumin and then challenged with intranasal ovalbumin. At 7 days following sensitization, these mice were treated with different doses of adiponectin intranasally in a volume of 5.0 µl. Nasal allergy-like symptoms, which were induced by bilateral application of 0.1 % OVA (5.0 µl), were assessed by counting sneezing and nasal rubbing behavior for 10 min immediately after nasal ovalbumin challenge. SP levels in nasal lavage fluid obtained 6 h after nasal ovalbumin challenge were examined by ELISA. RESULTS: Treatment of NGF-stimulated PC-12 cells with adiponectin suppressed SP production, which was induced by capsaicin stimulation. The minimum concentration of adiponectin that caused significant suppression was 7.5 ng/ml. On the other hand, adiponectin did not affect the total number of neurites and the percentage of PC-12 cells with outgrowing neurites, even at 1,000 ng/ml. Intranasal instillation of adiponectin into ovalbumin-sensitized mice at more than 10.0 ng/ml, but not 5.0 ng/ml, significantly inhibited the appearance of SP in nasal secretions, which was increased by intranasal challenge with ovalbumin. Adiponectin also suppressed the development of nasal allergic-like symptoms, sneezing and rubbing behavior, when ovalbumin-sensitized mice were treated intranasally with adiponectin at more than 10.0 ng/ml. The present results strongly suggested that adiponectin suppresses the production of SP and results in improvement of the clinical conditions of AR.

Modulation of eosinophil survival by epinastine hydrochloride, an H1 receptor antagonist, in vitro.

The influence of epinastine hydrochloride (EP) on eosinophil survival was examined by an in vitro cell culture technique. Nasal epithelial cells (NECs) were stimulated with 25 ng/ml TNF-alpha in the presence of EP (10 to 30 ng/ml). After 24 h, the culture supernatants were obtained and used as conditioned media of NECs (CM). Eosinophils (1 x 10(3) cells/ml) prepared from healthy human peripheral blood were incubated with 25% CM and eosinophil survival was assessed by trypan blue dye exclusion test 48 h later. CM prepared from NEC cultures pre-treated with TNF-alpha and EP caused a decrease in eosinophil survival as compared with that from NEC cells pre-treated with TNF-alpha alone. The minimum concentration of EP that caused a significant decrease in eosinophil survival was 25 ng/ml. The addition of EP into eosinophil cultures did not cause inhibition of eosinophil survival, which was prolonged by stimulation with granulocyte-macrophage colony-stimulating factor (GM-CSF), even when 40 ng/ml EP was added to cell cultures. We then examined the levels of GM-CSF in CM by ELISA. Treatment of NECs with EP at more than 25 ng/ml, reduced the ability of NECs to produce GM-CSF in response to TNF-alpha stimulation. These results may suggest that EP suppresses eosinophil survival through the suppression of GM-CSF production from NECs induced by inflammatory stimulation and that this suppressive effect contributes, in part, to the therapeutic mode of action of EP on allergic diseases.

Suppressive activity of epinastine hydrochloride on eosinophil activation in vitro.

The influence of a histamine H1 receptor antagonist, epinastine hydrochloride (EP), on eosinophil functions was examined in vitro and in vivo. The first set of experiments was undertaken to examine whether EP could suppress eosinophilia and IgE hyperproduction induced by Mesocestoides cortii infection in BALB/c mice. The number of peripheral blood eosinophils and levels of IgE were examined 21 days after infection. Oral administration of EP at a daily dose of 0.3 mg/kg, which is the recommended human therapeutic dose, for 21 days was not able to suppress either peripheral blood eosinophilia or IgE hyperproduction, which was observed in mice infected with M. cortii. The second part of the experiment was designed to examine the influence of EP on eosinophil activation induced by stem cell factor (SCF) stimulation in vitro. Eosinophils were obtained from M. cortii-infected mice and stimulated with SCF in the presence of different concentrations of EP for 24 h. The addition of EP into cell cultures suppressed eosinophil activation induced by SCF stimulation as assessed by measuring the contents of acronym for Regulated upon Activation, Normal T cell Expressed and presumably Secreted (RANTES), macrophage inflammatory protein-1beta (MIP-1beta) and leukotriene C4 (LTC4) levels in culture supernatants. The minimum concentration of EP which caused significant suppression of factor productions was 25 ng/ml, which is similar to the concentration in plasma after oral administration of the therapeutic dose in humans. These results may suggest that EP exerts inhibitory effects on eosinophil activation and results in favorable modification of the clinical status of allergic patients.

Quercetin Enhances the Thioredoxin Production of Nasal Epithelial Cells In Vitro and In Vivo.

Background: Thioredoxin (TRX) acts as both a scavenger of reactive oxygen species (ROS) and an immuno-modulator. Although quercetin has been shown to favorably modify allergic rhinitis (AR) symptoms, its influence on TRX production is not well defined. The present study was designed to examine whether quercetin could favorably modify AR symptoms via the TRX production of nasal epithelial cells in vitro and in vivo. Methods: Human nasal epithelial cells (HNEpCs) were stimulated with H2O2 in the presence of quercetin. TRX levels in 24-h culture supernatants were examined with ELISA. BALB/c male mice were intraperitoneally sensitized to ovalbumin (OVA) and intranasally challenged with OVA every other day, beginning seven days after the final sensitization. The mice were orally administered quercetin once a day for five consecutive days, beginning seven days after the final sensitization. Nasal symptoms were assessed by counting the number of sneezes and nasal rubbing behaviors during a 10-min period immediately after the challenge. TRX levels in nasal lavage fluids obtained 6 h after the challenge were examined by ELISA. Results: Treatment with 1.0 nM quercetin increased H2O2-induced TRX levels. The oral administration of 20.0 mg/kg of quercetin significantly inhibited nasal symptoms after the challenge. The same dose of quercetin significantly increased TRX levels in nasal lavage fluids. Conclusions: Quercetin's ability to increase TRX production may account, at least in part, for its clinical efficacy toward AR.

Suppressive Effect of Quercetin on Nitric Oxide Production from Nasal Epithelial Cells In Vitro.

Nitric oxide (NO) is known to play pivotal roles as one of the final effector molecules in the development of allergic diseases, including allergic rhinitis (AR). Although quercetin has been reported to attenuate the clinical conditions of AR, its influence on NO production is not well defined. The present study aimed to examine the influence of quercetin on in vitro NO production from nasal epithelial cells after interleukin- (IL-) 4 stimulation. Human nasal epithelial cells (HNEpCs) at a concentration of 1 x 105 cells/ml were stimulated with 10.0 ng/ml of IL-4 in the presence and absence of quercetin. After 48 hours, the culture supernatants were collected and assayed for NO (NO2 and NO3) using the Griess method. The influences of quercetin on the transcription factor, STAT6, activation, and iNOS mRNA expression were also examined using ELISA and real-time quantitative RT-PCR, respectively. Addition of quercetin to cell cultures caused suppression of NO production from HNEpCs after IL-4 stimulation. The minimum concentration of quercetin that caused significant suppression was 1.0 nM. Treatment of cells with quercetin at more than 1.0 nM suppressed STAT6 activation and iNOS mRNA expression induced by IL-4 stimulation. The present results strongly suggested that quercetin favorably modified the clinical condition of AR through the suppression of NO production from nasal epithelial cells after IL-4 stimulation.

Suppressive activity of fexofenadine hydrochloride on nitric oxide production in-vitro and in-vivo.

The aim of this study was to examine the effect of fexofenadine hydrochloride (FEX), a histamine H1-receptor antagonist, on nitric oxide (NO) production in-vitro and in-vivo. Nasal fibroblasts (5 x 10(5) cells per mL) were stimulated with 25 ng mL(-1) tumour necrosis factor-alpha in the presence of various concentrations of FEX. NO levels in 24-h-culture supernatants were measured by the Griess method and levels of inducible nitric oxide synthase (iNOS) mRNA levels in 12-h-cultured cells were measured by ELISA. FEX at more than 0.5 microg mL(-1) suppressed NO production from fibroblasts by inhibiting expression of iNOS mRNA. We also examined whether FEX could suppress NO production induced by lipopolysaccharide (LPS) stimulation in-vivo. BALB/c mice were treated with 5.0 mg kg(-1) LPS i.p. after daily oral doses of FEX, 1.0 mg kg(-1), for 1-3 weeks. Plasma was obtained 6 h later and NO levels measured by the Griess method. Expression of iNOS mRNA in lung tissues was measured by ELISA 6 h after LPS injection. Oral administration of FEX for 2 and 3 weeks, but not 1 week, significantly suppressed NO levels in plasma through the inhibition of iNOS mRNA expression, which were enhanced by LPS stimulation. These results suggest that the attenuating effect of FEX on NO production may be of therapeutic benefit in allergic diseases.

Anti-stress, anti-HIV and vitamin C-synergized radical scavenging activity of mulberry juice fractions.

Anti-stress and anti-HIV activity of mulberry juice were separated by centrifugation. The anti-stress activity was enriched in the supernatant fraction whereas the anti-HIV activity in the precipitate fraction. Oral administration of the supernatant fraction significantly reduced the elevated plasma level of lipid peroxide in mice loaded with water immersion restraint stress. The kinetic study revealed that the anti-stress activity was maintained for 4 hours after cessation of the administration of mulberry juice. The lignin fraction in the precipitate fraction scavenged superoxide and hydroxyl radicals more efficiently than other fractions, in a synergistic fashion with sodium ascorbate. Anti-HIV activity of mulberry juice was concentrated in the lignin fraction, whereas blueberry juice, which has no precipitating fibrous materials, did not show anti-HIV activity. The present study suggests the functionality of mulberry juice as an alternative medicine.