Luteolin Attenuates Allergic Nasal Inflammation via Inhibition of Interleukin-4 in an Allergic Rhinitis Mouse Model and Peripheral Blood From Human Subjects With Allergic Rhinitis

Abstract

Objectives: Luteolin is the active component of Perilla frutescens, an herb for the treatment of allergy in Asia. In this study, we aimed to investigate the effects and mechanisms of luteolin treatment. Methods: BALB/c mice sensitized with house dust mite (HDM) to induce allergic rhinitis (AR), and treated with dexamethasone or luteolin. In addition, mononuclear cells from peripheral blood (PBMC) of AR patients were co-cultured with dexamethasone or luteolin, and were re-stimulated with HDM. Results: Luteolin-treated mice had decreased allergic symptoms, and serum HDM-specific IgE when compared to the untreated group. Flow cytometric analyses of splenocytes and nasal lymphoid tissues from AR mice found that luteolin decreased CD4+ IL-4-secreting T cells when compared to those from vehicle treated AR mice. Histopathology sections showed reduced infiltration of eosinophils and decreased mucus secretion of mouse nasal epithelium. In the in vitro study, the results showed that luteolin reduced the percentage of CD4+ IL-4-secreting splenocytes expression was through reducing expression of pSTAT6 and GATA3. PBMCs from AR patients pretreated with luteolin could decrease percentage of CD4+ IL-4-secreting cells. Conclusion: Our study identified that luteolin attenuates allergic nasal inflammation via inhibition of IL-4 production, which supports the potential pharmaceutical application of luteolin treatment for AR.

Keywords: IL-4; allergic rhinitis; luteolin; traditional Chinese medicine; type 2 helper T cells.

Figure 1

(A) Sensitization and treatment protocols for the experimental mice. (B) Nasal symptoms of allergic mice decreased after luteolin or dexamethasone treatment. (C) Levels of immunoglobulin (Ig) E and IgG of experimental mice. HDM = house dust mite; Blank = mice with null sensitization and treatment; AR = allergic mice with vehicle treatment; Dex = allergic mice treated with 1 mg/kg dexamethasone; LO10 = allergic mice treated with 10 mg/kg luteolin; LO30 = allergic mice treated with 30 mg/kg luteolin; ^#P < 0.05 vs. blank group; *P < 0.05 vs. AR group.

Figure 2

Cytokines and FOXP3 expression of CD4+ splenocytes (A) or nasal lymphoid tissues (B) from experimental mice. AR = allergic rhinitis group; Dex = allergic mice treated with 1 mg/kg dexamethasone; LO10 = allergic mice treated with 10 mg/kg luteolin; LO30 = allergic mice treated with 30 mg/kg luteolin; ^#P < 0.05 vs. blank group; *P < 0.05 vs. AR group.

Figure 3

(A) Hematoxylin and eosin staining (400×) of tissue sections from experimental mice. The arrowheads mark the eosinophils in laminar propria. (B) Periodic acid-Schiff staining of the nasal mucosa from experimental mice (400×). The arrowheads mark the PAS staining cells.

Figure 4

Levels of cytokines in the supernatants of culture medium from splenocytes harvested from experimental mice re-stimulated with house dust mite. ^#P < 0.05 vs. blank group; *P < 0.05 vs. AR group.

Figure 5

(A) Sensitization and treatment protocols for the experimental mice, nasal symptom and immunoglobulin levels. (B,C) Cytokines expression of CD4+ splenocytes and nasal lymphoid tissues. (D) Decreased eosinophil infiltration in Hematoxylin and eosin staining of tissue sections from experimental mice. AR = allergic rhinitis group; Dex = allergic mice treated with 1 mg/kg dexamethasone; LO10 = allergic mice treated with 10 mg/kg luteolin; LO30 = allergic mice treated with 30 mg/kg luteolin; ^#P < 0.05 vs. blank group; *P < 0.05 vs. AR group.

Figure 6

(A) Experimental protocols for evaluating the efficacy of luteolin on murine CD4+ cell polarization. (B) Pre-treatment of luteolin reduced the percentage of CD4+ IL-4-secreting cells. *P < 0.05 vs. T_H2. (C) Experimental protocols for evaluating the efficacy of luteolin on murine naïve CD4+ cell polarization. (D) STAT6 and GATA3 protein expression of mouse naïve CD4+ cells were detected by western blot. pSTAT6 = phosphorylated (active) STAT6 protein; tSTAT6 = total STAT6 protein.

Figure 7

Luteolin inhibited CD4+IL-4+ cells expression of PBMCs isolated from HDM-AR or non-AR patients. (A) The protocol for human PBMCs pre-treatment with serial concentrations of luteolin and dexamethasone. Decreased percentage of CD4+ IL-4-secreting cells in PBMCs of (B) HDM allergic and (C) HDM non-allergic subjects after pretreatment with luteolin or dexamethasone. NC = negative control (medium without HDM); PC = positive control (medium with HDM); LO & Dex = pretreated with luteolin or dexamethasone, cultured in medium with HDM. ^#P < 0.05 vs. NC group. *P < 0.05 vs. PC group.

Figure 8

Schematic diagram depicting how luteolin inhibits airway allergic responses. (A) Luteolin inhibits IL-4 induced airway inflammatory responses. (B) Luteolin acts through the IL-4/STAT6/GATA3 signaling pathway.