Am J Respir Cell Mol Biol. 2014 Dec;51(6):783-92.
Fenretinide prevents inflammation and airway hyperresponsiveness in a mouse model of allergic asthma.
Kanagaratham C, Kalivodová A, Najdekr L, Friedecký D, Adam T, Hajduch M, De Sanctis JB, Radzioch D.
Arachidonic acid (AA) and docosahexaenoic acid (DHA) play important roles in inflammation and disease progression, where AA is viewed as proinflammatory and DHA as antiinflammatory. We observe in our model of allergic asthma that the AA/DHA ratio is significantly skewed in a proinflammatory direction. Fenretinide, a vitamin A derivative, has been shown to correct fatty acid imbalances in other diseases. Therefore, we explored if fenretinide can have a protective effect in allergic asthma. To accomplish this, we measured the levels of AA and DHA in the lungs of nonallergic, ovalbumin-induced allergic, and fenretinide-treated allergic mice.
We also investigated the effect of allergic asthma and fenretinide treatment on markers of oxidative stress, levels of metabolites, IgE production, airway hyperresponsiveness, and histological changes. Our data demonstrate that treatment of allergen-sensitized mice with fenretinide before allergen challenge prevents ovalbumin-induced changes in the AA/DHA ratio. The levels of several metabolites, such as serotonin, and markers of cellular stress, which are increased after ovalbumin challenge, are also controlled by fenretinide treatment. We observed the protective effect of fenretinide against ovalbumin-induced airway hyperresponsiveness and inflammation in the lungs, illustrated by a complete block in the infiltration of inflammatory cells to the airways and dramatically diminished goblet cell proliferation, even though IgE remained high. Our results demonstrate that fenretinide is an effective agent targeting inflammation, oxidation, and lung pathology observed in allergic asthma.
KEYWORDS: allergic asthma; arachidonic acid; docosahexaenoic acid; fenretinide; inflammation
Allergic asthma is a chronic respiratory immune disorder that affects over 300 million people worldwide (1). The burden of disease characterized by disability and premature death is greatest in early adolescents of 10-14 years of age and the elderly population of greater than 75 years. Asthma is not generally viewed as a deadly disease but it has a heavy cost on society due to frequent emergency room visits, and absenteeism from school and work.
Symptoms of allergic asthma include cough and wheezing, and excessive mucous production causing airway obstruction. Asthmatics also experience rapid and reversible airway constriction in the presence of low doses of irritants, a condition known as airway hyperresponsiveness. Allergic asthmatics become sensitized to allergens and have high levels of circulating IgE levels in the blood that could make them more at risk to allergic reactions. Pathologies of allergic asthma include recruitment of inflammatory cells, such as neutrophils and eosinophils, to the airways. Some studies have described allergic asthmatics as having higher levels of pro-inflammatory omega-6 fatty acids, such as arachidonic acid (AA), and lower levels of anti-inflammatory omega-3 fatty acids, such as docosahexaenoic acid (DHA), compared to healthy individuals (2).
Figure 1: FEN prevents increase in AA/DHA ratio (A), cyclooxygenase-2 (COX-2) expression (B), and markers of lipid and protein oxidation, malondialdehyde and nitrotyrosine respectively (C and D), in allergic asthmatic mice. Data are presented as mean ± SEM. **p < 0.01, and ***p < 0.001.
In our laboratory we are familiar with the methods for studying the multitude of allergic asthma phenotypes in animal models due to our extensive experience in the field (3-8). In addition, we continue to thoroughly study the beneficial effect of fenretinide (FEN), a vitamin A derivative, in various inflammatory diseases. Our most expansive research is on the potential of using FEN as a treatment for cystic fibrosis (CF). Our research has shown that FEN is capable of improving lung infection and can protect against osteoporosis in CF mice (9;10). In addition, we have shown that FEN can dampen the inflammation and improve the process of recovery following spinal cord injury (11). In both models, we observe that FEN can correct the imbalances in fatty acid levels (11;12). FEN is also being explored as a treatment for cancers and arthritis (13;14). Seeing the versatility of this drug and the overlapping phenotypes in asthma and other inflammatory diseases, we hypothesized that FEN can have a protective effect in treating allergic asthma.
Our experimental model was a combination of our allergic asthma protocol (three weekly intraperitoneal sensitizations with ovalbumin (OVA), followed by three aerosol challenges with OVA, with a FEN treatment protocol similar to what we use in CF mice (four weeks of daily oral treatment with 60mg/ml of FEN). The treatment was incorporated in between the antigen sensitizations and challenges. This model is most representative of the sequence of events which might happen in clinical setting as it describes treating subjects who are antigen sensitized (like most allergic asthmatics) and who would aim to prevent future allergic asthmatic reactions by taking prophylactic treatments. All of our animals were initially sensitized to OVA, subsequently they were either treated with FEN or drug vehicle (ethanol, ETOH) and either challenged with PBS or OVA. Overall, our study had three groups, our negative control that was treated with ETOH and challenged with PBS (PBS), our positive control group for allergic asthma that was treated with ETOH and challenged with OVA (OVA) and our experimental group that was treated with FEN and challenged with OVA (FEN-OVA).
Figure 2: FEN prevents airway hyperresponsiveness (AHR) (A) but not IgE production (B) in allergic asthmatic mice. Data are presented as mean ± SEM. *p < 0.05, **p < 0.01, and ***p < 0.001.
We observed that in our model of allergic asthma, OVA group mice had a greater ratio of AA/DHA compared to PBS group mice. This is observed in association with a greater increase in cyclooxygenase-2, an enzyme involved in the conversion of AA into bonchoconstricting mediators. Markers of lipid and protein oxidation, malondialdehyde and nitrotyrosine respectively, were also elevated in antigen challenged animals. Mice from the FEN-OVA group did not display the increase in lipid markers as observed in untreated and OVA-challenged animals (Figure 1). In addition, we also explored the protective effect of FEN on hallmark features of allergic asthma such as airway hyperresponsiveness and IgE production (Figure 2), and changes in lung histology (eosinophilia and goblet cell hyperplasia) (Figure 3), all of which are elevated in our positive control group compared to the negative control group. We observed that airway responsiveness to methacholine exposure is significantly reduced in FEN treated allergic animals compared to vehicle treated allergic animals (Figure 2A). Interestingly, we did not see any effect of FEN on IgE production after antigen challenge. FEN treated animals had high levels of circulating IgE in the plasma, similar to vehicle treated animals (Figure 2B). Our most remarkable finding was at the level of lung histology. Treatment with FEN significantly decreased the recruitment of inflammatory cells to the airways (as observed by hematoxylin and eosin (H and E) staining) (Figure 3A-C). Similarly, FEN prevented the increase in mucous producing goblet cells caused by OVA challenge (as observed by periodic acid-Schiff (PAS) staining) (Figures 3D-F).
Figure 3: FEN prevents recruitment of inflammatory cells to the airways (A-C) and goblet cell hyperplasia (D-F). Panels A and D are from vehicle treated mice challenged with PBS; panels B and E are from vehicle treated mice challenged with OVA; and panels C and F are from FEN treated mice challenged with OVA. (H and E: hematoxylin and eosin; PAS: periodic acid-Schiff)
From our data, we were able to conclude that daily treatment with FEN can prevent or at least mitigate the inflammatory phenotypes caused by antigen challenge in antigen sensitized subjects. The absence of an effect on IgE production may have been due to the fact that the treatment was started after antigen sensitization was completed. Perhaps a longer treatment or treatment during the antigen sensitization period would prevent the increase in IgE circulating in the blood. FEN strengthens the immune system of allergen sensitized mice in order to balance between the pro- and anti-inflammatory pathways making the response to antigen sensitization not as severe as observed in allergic and untreated animals. Based on our results FEN is a good candidate for asthmatic patients with low IgE, who would not benefit from anti-IgE treatments such as omalizumab. Many questions regarding the molecular mechanism of FEN still remain to be studied; however, our promising results highlight a novel and multifaceted candidate drug for asthma treatment.
- Imbalance in the AA/DHA ratio seen in asthmatic patients is reproducible in an ovalbumin induced mouse model of allergic asthma.
- This imbalance observed in allergic asthmatic mice can be corrected by treatment with FEN.
- Treatment with FEN prior to antigen challenge has a protective effect against most allergic asthma phenotypes.
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