Cytokine의 in vitro 대체 시험법 (II) = Establishment of in vitro cytokine assay-II

다국어 초록 (Multilingual Abstract)

This study is focused on the establishment of in vitro assays for the biologically active cytokines such as IL-1 and IL-18 which enhance or recover the reduced immune responses of patients administered with anticancer drugs. IL-1 and IL-18 are being used as immunomodulating agents. In vivo assays for these biological response modifiers are time consuming, expensive due to animal experiment and inaccurate depending on animals. In order to replace in vivo test, Th2 helper cell line DlOS, a subclone of Type 2 (Th2) DIOG4.1, which is responded and proliferated by only IL-1 even in the absence of co-mitogen, has been used. DlOS cells expressing a lots of IL-1R type I (IL-IRI) and IL-1R accessory protein (IL-1RAcP) were cloned and maintained. DlOS cells were proliferated at the minimal concentration of IL-1β(2.5 + 0.5 pg/ml) in the absence of co-mitogen. In addition, in order to establish in vitro test for bioactive IL-18, NKO. a subclone of natural killer (NK) cells has been used. The surface IL-18 receptor on NKO cells were increased by IL-12 and NKO cells produced IFN-γ by IL-18 coincubated with IL-12. Mammalian expression pcDNA3.1 vector or pLXN retroviral vector were used for the expression of IL-12p40 (pcDNA3.1/IL-12p40. pLXN/IL-12p40). Those expression vectors were transfected into NKO cells in order to replace the addition of IL-12 for the activation of NKO cells in vitro assay. These NKO/IL-12 cells were used in vitro test for IL-18 because these NKOIIL-12 cells produced IFN-Y by IL-18 in a dose response manner even in the absence of IL-12. In this study, we have been used the established those cell lines D10S and NKO cells which express a lots of IL-1R and IL-18R, respectively. Those cell lines are being used for the assays of IL-1- or IL-18 modulating factors. Some of those IL-1- or IL-18 modulating factors have been screened and identified. Their efficacy is being evaluated and characterized for their application.

국문 초록 (Abstract)

Ⅰ . 제 목 : Cytokine의 in vitro 대체 시험법 II
Ⅱ. 연구개발의 목적 및 필요성
본 연구에서는 면역세포 등을 활성화 시켜 GM-CSF 생성이나 IFN-Y 생산에 관여하는 interleukin-1 (IL-1)과 IL-18의 in vitro 생물학적 활성 시험법을 확립하여 이들 생물학적 제제의 생물학적 효능을 효율적으로 검증하는데 이용하고자 한다. 이들 싸이토카인들은 항암치료 시 체내 면역 능이 떨어진 환자의 면역력을 회복시키거나 면역력을 높이는 효능이 있는 일종의 면역 기능 조절제 (immunomodulating agents)로 사용되기도 한다. 이들 생물학적 제제의 in vivo 시험법은 동물의 체내에 주사한 후 그 효과를 측정해야 하므로 시간과 비용이 많이 소요되고 실험동물의 개체에 따라 시험오차범위가 큰 단점이 있다. 따라서 이들 사이토카인의 in vivo 시험법을 대체하기 위해, IL-1 에 특이적으로 분화하는 Type 2 helper T cell (Th2)인 D10G.4.1 세포 중에서도 co-mitogen없이 IL-1 만을 처리하여도 분화하는 D10S를 유지 • 확립하여 이를 IL-1 의 활성을 검정하는데 이용하고자 한다.
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