Experimental model of equine alveolar macrophage stimulation with TLR ligands

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• 作者：Ingrid Waldschmidt ; Dimitri Pirottin ; Tatiana Art ; Fabrice Audigi¨¦ ; Fabrice Bureau ; Irene Tosi ; Sophie El Abbas ; Fr¨¦d¨¦ric Farnir ; Eric Richard ; Mari-Capucine Dupuis
• 关键词：Horse ; Alveolar macrophages ; Innate immunity ; mRNA ; Cytokines
• 刊名：Veterinary Immunology and Immunopathology
• 出版年：2013
• 出版时间：1 September, 2013
• 年：2013
• 卷：155
• 期：1-2
• 页码：30-37
• 全文大小：651 K

文摘

Pulmonary diseases are common in horses and have a major economic impact on the equine industry. Some of them could be associated with an inadequate immune response in the lung, but methods to evaluate this response in horses are lacking.

The aim of this study was to develop and validate an experimental model that could be applied in several physiological and pathological conditions to assess the innate immune response of equine pulmonary cells.

Equine alveolar macrophages (AMs) obtained from bronchoalveolar lavages were isolated from other cells by adhesion. TLR2, 3, and 4 expression in AMs was studied and their responses to commercial ligands (respectively FSL-1, Poly(I:C), and LPS) were evaluated after determination of the appropriate dose and time of incubation. TLR responses were assessed by measuring cytokine production using (1) gene expression of TNF¦Á, IFN¦Â, Il-1¦Â, and IFN¦Á by qPCR (indirect method); and (2) cytokine production for TNF¦Á and IFN¦Â by ELISA (direct method).

TLR 2, 3, and 4 were expressed by AMs. TLR 2 stimulation with 10 ng/mL of FSL-1 during 3 h significantly increased IL-1¦Â and TNF¦Á gene expression. TLR 3 stimulation with 1000 ng/mL of Poly(I:C) during 1 h increased IFN¦Â, IFN¦Á, Il-1¦Â and TNF¦Á expression. TLR 4 stimulation with 100 ng/mL of LPS during 3 h increased TNF¦Á, IFN¦Â, and Il-1¦Â expression. Results obtained by ELISA quantification of TNF¦Á and IFN¦Â produced by AMs following stimulation during 6 h were similar: FSL-1 increased TNF¦Á production but not IFN¦Â, Poly(I:C) and LPS increased production of IFN¦Â and TNF¦Á. In conclusion, pulmonary innate immunity of horses can be assessed ex vivo by measuring cytokine production following stimulation of AMs with TLR agonists.

This experimental model could be applied under several conditions especially to improve the understanding of equine respiratory disease pathogenesis, and to suggest novel therapeutic opportunities.