人白细胞介素17(IL-17)的克隆及其在大肠杆菌中的表达

树突状细胞(DC)是一类能特异性启动T淋巴细胞介导的免疫反应的抗原提呈细胞,与其它抗原提呈细胞不同的是,树突状细胞能激活初始型T淋巴细胞(naive T cells).近年来,人们试图通过刺激血液中的前体细胞以提高DC的量.最近,有学者通过GM-CSF、IL-4及MCM的培养单个核细胞中获得了DC.本研究对此法加以改良并从恒河猴血中获得了形态、表型及功能均与人成熟DC非常类似的DC.去除了T细胞的单个核细胞在含有1%人血清及GM-CSF和IL-4的培养基中培养,在第7天,50%的培养基换成单核细胞的条件培养基MCM.此培养基是获得成熟的具有免疫刺激作用的DC所必需的,这样可从2Oml血中获得0.5-1.0×10~6DC.对这些DC与在同样培养基中培养产生的巨噬细胞的比较表明,这些DC刺激同种异体T细胞的能力较巨噬细胞明显增强,而且这些DC的形态和表型很典型且能表达高水平的p55及与细胞内CD68.对比之下,贴壁的巨噬细胞表达非常低水平的p55及高水平的弥漫性CD68,用此方法制备的恒河猴

Cloning of Human Interleukin-17 and Its Expression in E. coli

Interleukin-17 (IL-17) is a newly found cytokine secreted by activated T lymphocytes. From preliminary study, IL-17 was found to play a role in the relationship between T lymphocytes and hematopoiesis, but many of its characteristics remain unknown up to now. To investigate its biological functions and related mechanisms, we cloned the complete coding region of human IL-17(hIL-17) from activated human peripheral blood mononuclear cells (PBMC) by RT-PCR and constructed an integrative vector pLCM182-hIL-17 for cloning, expressing and sequencing the hIL-17 gene. By DNA sequencing, the cloned hIL-17 is identical to the reptorted, and with temperature inducing, the hEL-17 was highly expressed in E. coli up to 30% of bacterial protein. The expressed hIL-17 protein could stimulate primary human fibrob-lasts to secrete IL-6 and GM-CSF after initial purification. This results indicate that the expressed hIL-17 has biological activity.