TRPV1 Blocking Alleviates Airway Inflammation and Remodeling in a Chronic Asthma Murine Model

Abstract

Purpose: Asthma is a chronic inflammatory airway disease characterized by airway hyperresponsiveness (AHR), inflammation, and remodeling. There is emerging interest in the involvement of the transient receptor potential vanilloid 1 (TRPV1) channel in the pathophysiology of asthma. This study examined whether TRPV1 antagonism alleviates asthma features in a murine model of chronic asthma.

Methods: BALB/c mice were sensitized to and challenged by ovalbumin to develop chronic asthma. Capsazepine (TRPV1 antagonist) or TRPV1 small interfering RNA (siRNA) was administered in the treatment group to evaluate the effect of TPV1 antagonism on AHR, airway inflammation, and remodeling.

Results: The mice displayed increased AHR, airway inflammation, and remodeling. Treatment with capsazepine or TRPV1 siRNA reduced AHR to methacholine and airway inflammation. Type 2 T helper (Th2) cytokines (interleukin [IL]-4, IL-5, and IL-13) were reduced and epithelial cell-derived cytokines (thymic stromal lymphopoietin [TSLP], IL-33, and IL-25), which regulate Th2 cytokine-associated inflammation, were also reduced. Airway remodeling characterized by goblet cell hyperplasia, increased α-smooth muscle action, and collagen deposition was also alleviated by both treatments.

Conclusions: Treatment directed at TRPV1 significantly alleviated AHR, airway inflammation, and remodeling in a chronic asthma murine model. The TRPV1 receptor can be a potential drug target for chronic bronchial asthma.

Keywords: TRPV1 receptor; airway remodeling; asthma; capsazepine.

Fig. 1. Effect of capsazepine and TRPV1 siRNA on AHR to Mch. AHR was measured 24 hours after the final OVA challenge with the flexiVent system. Mch concentration was increased from 6.25 to 100 mg/mL. The values are expressed as mean±SEM (n=4–8/group). TRPV1, transient receptor potential vanilloid 1; siRNA, small interfering RNA; AHR, airway hyperresponsiveness; Mch, methacholine; OVA, ovalbumin; SEM, standard error of the mean. ^*P<0.01, ^†P<0.001 compared to control, ^‡P<0.01, ^§P<0.001 compared to the OVA group.

Fig. 2. Effect of capsazepine and TRPV1 siRNA on total/differential cell counts in BAL fluid. Mice were sacrificed after measurement of AHR, and BAL was analyzed immediately. The values are expressed as mean±SEM (n=4–8/group). TRPV1, transient receptor potential vanilloid 1; siRNA, small interfering RNA; BAL, bronchoalveolar lavage; AHR, airway hyperresponsiveness; SEM, standard error of the mean; OVA, ovalbumin. ^*P<0.001 compared to control, ^†P<0.001 compared to the OVA group.

Fig. 3. Effect of capsazepine and TRPV1 siRNA on levels of Th2 cytokines and epithelial-derived cytokines. (A) The concentration of IL-4, IL-5, and IL-13 were measured in the BAL fluid with and ELISA. (B) Levels of IL-17E, IL-33, and TSLP in the lung homogenates were measured. The values are expressed as mean±SEM (n=4–8/group). TRPV1, transient receptor potential vanilloid 1; siRNA, small interfering RNA; Th2, type 2 T helper; IL, interleukin; BAL, bronchoalveolar lavage; ELISA, enzyme-linked immunosorbent assay; TSLP, thymic stromal lymphopoietin; SEM, standard error of the mean; OVA, ovalbumin. ^*P<0.01, ^†P<0.001 compared to control, ^‡P<0.05, ^§P<0.01, ^∥P<0.001 compared to the OVA group for (A). ^¶P<0.05, ^**P<0.001 compared to control, ^††P<0.05, ^‡‡P<0.001 compared to the OVA group for (B).

Fig. 4. Effect of capsazepine and TRPV1 siRNA on perbronchial inflammation and TRPV1 receptor expression in lung tissues. (A) Lung tissues were fixed with 4% paraformaldehyde and embedded in paraffin. Paraffin block were sectioned (4-µm thick) and stained with H&E (×200). (B) The TRPV1 receptor was immunostained in paraffin-embedded lung section (blue). TRPV1, transient receptor potential vanilloid 1; siRNA, small interfering RNA; H&E, hematoxylin and eosin; Br, bronchus; Bm, basement membrane; Eo, eosinophil; Ep, epithelium; Bl, blood vessel.

Fig. 5. Effect of capsazepine and TRPV1 siRNA on goblet cell hyperplasia in lung tissues. (A) The paraffin-embedded tissues were cut into 5- to 6-µm-thick sections and stained with PAS. (B) Goblet cell hyperplasia was quantified using a modified 5-point scoring system, based on the percentage of goblet cells in the epithelium: grade 0 (no goblet cells), grade 1 (<25%), grade 2 (25%–50%), grade 3 (51%–75%), and grade 4 (>75%). The values are expressed as mean±SEM (n=4–8/groups). TRPV1, transient receptor potential vanilloid 1; siRNA, small interfering RNA; PAS, periodic acid-Schiff; OVA, ovalbumin. ^*P<0.001 compared to control, ^†P<0.001 compared to the OVA group.

Fig. 6. Effect of capsazepine and TRPV1 siRNA on the area of peribronchial airway smooth muscle. (A) Peribronchial α-SMA was immunostained in paraffine-embedded lung section. (B) The immunostained area was quantified by using a light microscope attached to BX50. The values are expressed as mean±SEM (n=4–8/group). TRPV1, transient receptor potential vanilloid 1; siRNA, small interfering RNA; α-SMA, α-smooth muscle actin; SEM, standard error of the mean; OVA, ovalbumin. ^*P<0.01 compared to control, ^†P<0.01 compared to the OVA group.

Fig. 7. Effect of capsazepine and TRPV1 siRNA on total collagen levels. Lung tissue was collected from each mouse for the hydroxyproline assay. The values are expressed as mean±SEM (n=4–8/groups). TRPV1, transient receptor potential vanilloid 1; siRNA, small interfering RNA; SEM, standard error of the mean; OVA, ovalbumin; HYP, hydroxyproline. ^*P<0.01 compared to control, ^†P<0.01 compared to the OVA group.