Cyclosporine and glucocorticoids are powerful immunosuppressive agents used to treat many inflammatory diseases in dogs. Cyclosporine inhibits calcineurin-dependent pathways of T cell activation and resultant T cell cytokine production, and glucocorticoids directly inhibit genes coding for cytokines. Little work has been done comparing the effects of these agents on T cell cytokine production in dogs. Our study measured T cell interleukin-2 (IL-2) and interferon-gamma (IFN-γ) production using flow cytometry and T cell IL-2 and IFN-γ gene expression using quantitative reverse transcription polymerase chain reaction (qRT-PCR) in activated canine T cells incubated with cyclosporine and dexamethasone in vitro. For flow cytometric assays, diluted whole blood was cultured for 7 h in the presence of cyclosporine (10, 100, 500, and 1000 ng/mL) or dexamethasone (10 ng/mL, 100 ng/mL, 1 μg/mL, and 10 μg/mL). For qRT-PCR, whole blood was cultured for 5 h with the same drugs at the same concentrations, and RNA was then extracted from leukocytes. Flow cytometry and qRT-PCR both demonstrated inhibition of IL-2 and IFN-γ that was concentration-dependent in response to cyclosporine, and was more variable for dexamethasone. Quantitative RT-PCR but not flow cytometry documented significant reduction of IL-2 expression after dexamethasone treatment, while both methods showed concentration-dependent suppression of IFN-γ. Quantitative RT-PCR also revealed additional cytokine suppression at higher cyclosporine concentrations, an effect not found using flow cytometry, and may therefore be the preferred method for cytokine determination in dogs. Suppression of IL-2 and IFN-γ in activated T cells may have potential as an indicator of the efficacy of cyclosporine and glucocorticoids in suppressing canine T cell function in vivo, and may therefore be of value for characterizing the immunosuppression induced by these drugs in clinical patients.

中文翻译：

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环孢素和地塞米松对犬白细胞介素2和干扰素-γ的T细胞表达的影响。

环孢素和糖皮质激素是有效的免疫抑制剂，可用于治疗犬的许多炎症性疾病。环孢菌素抑制钙调神经磷酸酶依赖性的T细胞活化途径和由此产生的T细胞细胞因子产生，而糖皮质激素直接抑制编码细胞因子的基因。比较这些试剂对犬T细胞细胞因子产生的影响的工作还很少。我们的研究使用流式细胞仪测量了T细胞白介素2（IL-2）和干扰素-γ（IFN-γ）的产生，并使用定量逆转录聚合酶链反应（qRT-PCR）测量了T细胞IL-2和IFN-γ的基因表达。在体外与环孢霉素和地塞米松一起孵育的活化犬T细胞中 对于流式细胞分析，将稀释的全血在环孢菌素（10，100，500，和1000 ng / mL）或地塞米松（10 ng / mL，100 ng / mL，1μg/ mL和10μg/ mL）。对于qRT-PCR，将全血与相同浓度的相同药物培养5小时，然后从白细胞中提取RNA。流式细胞仪和qRT-PCR均显示IL-2和IFN-γ的抑制作用是对环孢霉素的浓度依赖性，并且对地塞米松的影响更大。定量RT-PCR而非流式细胞术记录了地塞米松治疗后IL-2表达的显着降低，而两种方法均显示出浓度依赖性地抑制IFN-γ。定量RT-PCR还显示了在较高环孢菌素浓度下细胞因子的抑制作用，使用流式细胞仪未发现这种作用，因此可能是犬中确定细胞因子的首选方法。