Synthetic siRNA molecules suppress Il4 and Il13 gene expression and reduce allergic inflammation and remodeling of upper airways in mouse model

• Igor P. Shilovskiy
• Ekaterina D. Timotievich
• Mariya M. Kaganova
• Valeriia I. Kovchina
• Kirill V. Yumashev
• Aleksandr A. Nikolskii
• Liudmila I. Vishniakova
• Mariya S. Sundukova
• Vadlen V. Tyulyubaev
• Vera E. Brylina
• Tatyana E. Rusak
• Oksana M. Kurbacheva
• Miramgul E. Dyneva
• Olga A. Petukhova
• Georgii O. Gudima
• Dmitry A. Kudlay
• Musa R. Khaitov

Abstract

Introduction. Inflammation of the upper airways is usually associated with allergic rhinitis (AR), chronic rhinosinusitis with nasal polyps (CRSwNP) etc. AR is one of the most common inflammatory diseases. Its pathogenesis includes increased production of allergen-specific IgE antibodies, infiltration of the nasal mucosa with inflammatory cells, and remodeling of the respiratory tract. It has been shown that Th2-cytokines (IL-4 and IL-13) play a key role in this pathology, which makes them promising targets for therapy. Drugs based on monoclonal antibodies targeted to these cytokines have been developed and demonstrated clinical efficacy, while its broad use is limited by high cost. Currently, new technologies for regulation of gene expression are developed. For example, small interfering RNA (siRNA) molecules can suppress the expression of genes encoding pro-inflammatory cytokines.

The aim of this study was to develop siRNAs suppressing the expression of Il4 and Il13 genes and to reveal their biological effects on the inflammation of the upper respiratory tract in a mouse model.

Material and methods. The design of siRNA molecules was carried out using specialized software. Screening of siRNA biological activity was performed by in vitro experiments. The production of IL-4 and IL-13 was evaluated by ELISA and the expression of the corresponding genes by real-time polymerase chain reaction (RT-PCR). Induction of upper respiratory tract inflammation in mice was performed by subcutaneous and intranasal administration of ovalbumin chicken egg allergen (OVA). Experimental therapy was carried out by intranasal administration of siRNA molecules in complex with the LTP carrier peptide. A corticosteroid drug, budesonide, was used as a positive control. Nasal hyperreactivity was evaluated. Levels of allergen-specific IgE, IgG1 and IgG2a antibodies in mice blood sera, as well as levels of IL-4 and IL-13 production by submandibular lymph nodes were determined by ELISA. Infiltration of the nasal mucosa by proinflammatory cells was assessed by histological methods.

Results. Intranasal administration of siRNAs targeted to genes encoding IL-4 and IL-13 reduced the production of these cytokines in local (submandibular) lymph nodes. Suppression IL-4 and IL-13 production resulted in decrease in the levels of allergen-specific IgE- and IgG1-antibodies in the serum of mice and nasal hyperreactivity. A decrease in inflammation and upper respiratory tract remodeling was also observed after siRNA mediated suppression of IL-4 and IL-13 production.

Conclusion. Experimental therapy with siRNAs suppressing the production of pro-inflammatory cytokines IL-4 and IL-13 significantly alleviated the symptoms of allergic inflammation of the upper respiratory tract. Synthetic siRNAs have significant potential in the treatment of inflammatory diseases.

Keywords:allergy; allergic rhinitis; inflammation; cytokines; siRNA; peptide

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