Undaria pinnatifida extract attenuates combined allergic rhinitis and asthma syndrome by the modulation of epithelial cell dysfunction and oxidative stress

Abstract

Undaria pinnatifida ( U. pinnatifida) has long been a part of the human diet and medicine. Although U. pinnatifida has been reported to have immunomodulatory, anti-inflammatory, anti-diabetic and antibacterial activities, its specific effect on patients with combined allergic rhinitis and asthma syndrome (CARAS) has not been clarified. In this study, the anti-allergic and anti-inflammatory effects of U. pinnatifida extract (UPE) are investigated in a mouse model of ovalbumin (OVA)-induced CARAS. The oral administration of UPE inhibits allergic responses by reducing OVA-specific immunoglobulin levels. As a result, the symptoms of early reactions are also improved. UPE inhibits the accumulation of inflammatory cells and attenuates the expression of Th2 cytokines in both nasal and bronchoalveolar lavage fluid. Furthermore, UPE treatment inhibits the NF-κB/MAPK signaling pathway in lung homogenates. Additionally, UPE prevents shedding of the nasal mucosal epithelium, protects the integrity of the epithelium, enhances the expression of E-cadherin at the junction of epithelial cells, and inhibits the degradation of ZO-1 and occludin in the airway epithelium. In addition, UPE ameliorates dysfunction of the nasal epithelial barrier by enhancing antioxidant properties and downregulating the expression of the inflammatory factor IL-33. These results suggest that UPE may treat CARAS by modulating epithelial cell dysfunction and oxidative stress.

Keywords: IL-33; antioxidant; combined allergic rhinitis and asthma syndrome; nasal epithelial barrier dysfunction.

Figure 1

Reduction in allergic nasal symptoms caused by UPE in OVA-induced CARAS mice (A) Rubbing and (B) nasal sneezing were evaluated for 15 min after the last OVA challenge. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6.

Figure 2

Reduction in the number of inflammatory cells in the NALF and BALF caused by UPE in OVA-induced CARAS mice (A,D) Total number of cells in the NALF and BALF. (B,E) Differential cells were isolated via cytospin. (C,F) Cytospin preparation was stained with Diff-Quik. The red arrows indicate eosinophils. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6. Scale bars: 50 μm.

Figure 3

Alleviation of airway inflammation in lung tissues by UPE in OVA-induced CARAS mice (A) The inflammatory response in the lungs was examined by hematoxylin and eosin (H&E) staining. (B) Histological changes were examined via periodic acid-Schiff (PAS) staining. Scale bars: 100 μm. The values (n = 6) represent the means ± SEM of triplicate experiments. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, **P < 0.01, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6.

Figure 4

Alleviation of nasal inflammation in the nasal mucosa by UPE in OVA-induced CARAS mice (A) Submucosal thickness was examined by hematoxylin and eosin (H&E) staining. (B) Clobet cell numbers were examined via periodic acid-Schiff (PAS) staining. Scale bars: 50 μm. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, **P < 0.01, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6.

Figure 5

Reduction in OVA-specific antibody levels in the serum caused by UPE in OVA-induced CARAS mice (A) The level of OVA-specific IgE in the serum was analyzed by ELISA kit. (B) The level of OVA-specific IgG1 in the serum was analyzed by ELISA kit. (C) The level of OVA-specific IgG2a in the serum was analyzed by ELISA kit. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6.

Figure 6

Regulation of Th1/Th2-related cytokines by UPE in OVA-induced CARAS mice (A–D) IFN-γ, IL-4, IL-5, and IL-13 levels in NALF were examined via ELISA kits. (E–H) IFN-γ, IL-4, IL-5, and IL-13 levels in the BALF were examined via ELISA kits. Data were presented as the mean±SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6.

Figure 7

Modulation of oxidative stress mediator expression in NALF and BALF by UPE in OVA-induced CARAS mice (A‒D) MDA, SOD, Nrf2, and HO-1 levels in NALF were examined via ELISA kits. (E‒H) MDA, SOD, Nrf2, and HO-1 levels in the BALF were examined via ELISA kits. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6.

Figure 8

Regulation of tight junction protein expression by UPE in OVA-induced CARAS mice (A) Immunohistochemical labelling revealed E-cadherin expression in the epithelial layer. (B,C) ZO-1 and occludin levels in NALF were examined via ELISA kits. (D,E) ZO-1 and occludin levels in the BALF were examined by ELISA kits. (F‒H) ZO-1 and occludin protein levels in lung tissues were examined by western blot analysis. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6. Scale bars: 50 μm.

Figure 9

Inhibition of the NF-κB/MAPK signaling pathway by UPE in OVA-induced CARAS mice (A,B) NF-κB and NF-κB phosphorylation levels in NALF were examined via ELISA kits. (C,D) NF-κB and NF-κB phosphorylation levels in the BALF were examined via ELISA kits. (E) Western blot analysis of MAPKs. (F‒H) Quantification of the p-p38/p38 ratio, p-Erk/Erk ratio, and p-JNK/JNK ratio. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6.

Figure 10

Inhibition of IL-33/ST2 expression by UPE in OVA-induced CARAS mice (A) IL-33 levels in the NALF in different groups were detected by an ELISA kit. (B) IL-33 in the BALF of different groups was detected with an ELISA kit. (C) ST2 in the nasal tissue was examined by immunohistochemistry. (D) ST2 in the lung was examined by immunohistochemical staining. Data were presented as the mean ± SEM of triplicate experiments. One-way ANOVA was used to compare the differences among multiple groups. *P < 0.05, ***P < 0.001 vs OVA group; ### P < 0.001 vs control group. n = 6. Scale bars: 50 or 100 μm.