Codium fragile Extract Ameliorates Respiratory Function by Controlling Allergic Inflammation in Ovalbumin-Induced Bronchial Disorders in Mice

Abstract

This study investigated the effect of Codium fragile (WCF) water extract in reducing allergic inflammation in ovalbumin (OVA)-induced mice. Mice were sensitized to OVA + aluminum hydroxide, administered WCF for one week, and exposed to 1% aerosolized OVA. As a result, WCF intake reduced the OVA-induced increase in CD4⁺ T cells, CD8⁺ T cells, the T helper type 2 (Th2)/T helper type 1 (Th1) cell ratio, and inflammatory cells such as eosinophils and lymphocytes. Furthermore, WCF reduced Th2 cytokines such as interleukin (IL)-5, IL-13, and IL-33 and inflammatory cytokines such as tumor necrosis factor α (TNF-α) and IL-1β in lung tissues. A histological analysis showed that WCF intake decreases OVA-induced pulmonary inflammation, bronchial wall thickness, and mucus score and increases pulmonary alveolar area. Moreover, WCF inhibited the nuclear factor κB (NF-κB) pathway, the transforming growth factor β (TGF-β)/Smad pathway, and apoptosis-related proteins in lung tissues that OVA excessively activated. The oleamide (9-octadecenamide) content, representing a physiologically active component of WCF, was analyzed and validated using a high-performance liquid chromatography-photodiode array (HPLC-PDA) system. These results demonstrate that WCF may serve as a potential preventive agent for respiratory dysfunction such as allergic asthma by suppressing NF-κB and TGF-β/Smad pathways.

Keywords: Codium fragile; Th2 cytokine; allergic inflammation; fibrosis; oleamide; pulmonary function.

Figure 1

The HPLC chromatograms of oleamide standard (a) and water extract of Codium fragile (WCF) (b). The calibration curve of oleamide (c).

Figure 2

Effect of water extract of Codium fragile (WCF) on activation of T cells in whole blood of ovalbumin (OVA)-induced mice. Representative FACS plots (a), proportion of CD3⁺CD4⁺ as Th cells (b), and CD3⁺CD8⁺ as Tc cells (c). Th cell identification was performed by detecting intracellular cytokines IFN-ɣ and IL-4. Proportion of CD3⁺CD4⁺ IFN-γ⁺ as Th1 cells (d) and CD3⁺CD4⁺ IL-4⁺ as Th2 cells (e), and Th2/Th1 ratio (f). Data are presented as mean ± SD (n = 5). * p < 0.05, ** p < 0.01, and *** p < 0.001: OVA group vs. control group; # p < 0.05, ## p < 0.01, and ### p < 0.001: WCF groups vs. OVA group.

Figure 3

Effect of water extract of Codium fragile (WCF) on allergic inflammatory cytokines in lung tissues of ovalbumin (OVA)-induced mice. Western blot images (a) and relative expression levels of IL-33, IL-5, IL-13, IL-1β, and TNF-α (b). Data are presented as mean ± SD (n = 3). * p < 0.05, ** p < 0.01, and *** p < 0.001: OVA group vs. control group; # p < 0.05 and ## p < 0.01: WCF groups vs. OVA group.

Figure 4

Effect of water extract of Codium fragile (WCF) on ovalbumin (OVA)-specific immunoglobulin (Ig)E levels in bronchoalveolar lavage fluid (BALF) (a) and serum (b) of OVA-induced mice. N.D. means not detected. Data are presented as mean ± SD (n = 5). ** p < 0.01 and *** p < 0.001: OVA group vs. control group; # p < 0.05, ## p < 0.01, and ### p < 0.001: WCF groups vs. OVA group.

Figure 5

Effect of water extract of Codium fragile (WCF) on histopathological changes in lung tissue of ovalbumin (OVA)-induced mice. Representative hematoxylin and eosin (H&E)-stained sections of bronchiole (a) and alveoli (b), and periodic acid–Schiff (PAS)-stained sections (c) of bronchiole in lung tissues. Inflammation score (d), bronchial thickness (e), pulmonary alveolar area (f), and mucus score (g). N.D. means not detected. Data are presented as mean ± SD (n = 3). ** p < 0.01 and *** p < 0.001: control group vs. OVA group; ## p < 0.01, and ### p < 0.001: OVA group vs. WCF groups.

Figure 6

Effect of water extract of Codium fragile (WCF) on levels of antioxidant system in lung tissues of ovalbumin (OVA)-induced mice. Malondialdehyde (MDA) content (a), reduced glutathione (GSH) (b), and superoxide dismutase (SOD) level (c). Data are presented as mean ± SD (n = 5). ** p < 0.01 and *** p < 0.001: OVA group vs. control group; # p < 0.05, ## p < 0.01, and ### p < 0.001: WCF groups vs. OVA group.

Figure 7

Effect of water extract of Codium fragile (WCF) on activation of NF-κB pathway in lung tissues of ovalbumin (OVA)-induced mice. Western blot images (a) and relative expression levels of TLR-4, p-IκB-α, p-NF-κB, iNOS, and COX-2 (b). Data are presented as mean ± SD (n = 3). ** p < 0.01 and *** p < 0.001: OVA group vs. control group; # p < 0.05 and ## p < 0.01: WCF groups vs. OVA group.

Figure 8

Effect of water extract of Codium fragile (WCF) on activation of TGF-β/Smad pathway in lung tissues of ovalbumin (OVA)-induced mice. Western blot images (a) and relative expression levels of MMP-2, MMP-9, TGF-β1, p-Smad-2, and p-Smad-3 (b). Data are represent mean ± SD (n = 3). * p < 0.05, ** p < 0.01, and *** p < 0.001: OVA group vs. control group; # p < 0.05: WCF groups vs. OVA group.

Figure 9

Effect of water extract of Codium fragile (WCF) on activation of apoptosis pathway in lung tissues of ovalbumin (OVA)-induced mice. Western blot images (a) and relative expression levels of p-JNK, BAX, BCL-2, BAX/BCL-2 ratio, and CAS-3 (b). Data are presented as represent mean ± SD (n = 3). ** p < 0.01, and *** p < 0.001: OVA group vs. control group; # p < 0.05 and ## p < 0.01: WCF groups vs. OVA group.

Figure 10

Experimental procedure for ovalbumin (OVA)-induced asthma model and treatment with water extract of Codium fragile (WCF).