新设计的引物组可用于通过逆转录聚合酶链反应（RT-PCR）评估豚鼠肺组织中各种细胞因子和诱导型一氧化氮合酶（iNOS）的mRNA表达。

Newly designed primer sets available for evaluating various cytokines and iNOS mRNA expression in guinea pig lung tissues by RT-PCR.

作者信息

Yamada Hiroyuki, Udagawa Tadashi, Mizuno Satoru, Hiramatsu Kumiko, Sugawara Isamu

机构信息

Pathology Division, Mycobacterium Reference Center, The Research Institute of Tuberculosis, Japan Anti-Tuberculosis Association, Tokyo.

出版信息

Exp Anim. 2005 Apr;54(2):163-72. doi: 10.1538/expanim.54.163.

DOI:10.1538/expanim.54.163

PMID:15897626

Abstract

Guinea pigs are often used as an animal model of human tuberculosis (TB). However, there are few methods available for pursuing the immunological processes involved in guinea pig TB. In this study, we developed for the first time systematic reverse transcription (RT)-PCR for evaluation of guinea pig mRNA expression. RT-PCR primer sets were newly designed for detection of cytokines and inducible nitric oxide synthase (iNOS) mRNA in guinea pig TB. Interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, transforming growth factor (TGF)-beta, interleukin (IL)-1beta, IL-2, IL-10, IL-12p40, granulocyte-macrophage-colony stimulating factor (GM-CSF) and iNOS mRNA expression were detected significantly and reproducibly when these primer sets were used. The data by real-time PCR were comparable with those of RT-PCR. We showed that these RT-PCR primer sets could be used to examine mRNA expression semi-quantitatively in guinea pig tissues, and conclude that these newly designed primer sets for conventional RT-PCR will be useful for studying the immunological processes in guinea pig tuberculosis experiments to investigate and evaluate efficacy of new vaccines or anti-mycobacterial drugs.

摘要

豚鼠常被用作人类结核病（TB）的动物模型。然而，用于探究豚鼠结核病相关免疫过程的方法很少。在本研究中，我们首次开发了用于评估豚鼠mRNA表达的系统性逆转录（RT）-PCR。新设计了RT-PCR引物组，用于检测豚鼠结核病中细胞因子和诱导型一氧化氮合酶（iNOS）的mRNA。当使用这些引物组时，可显著且可重复地检测到干扰素（IFN）-γ、肿瘤坏死因子（TNF）-α、转化生长因子（TGF）-β、白细胞介素（IL）-1β、IL-2、IL-10、IL-12p40、粒细胞-巨噬细胞集落刺激因子（GM-CSF）和iNOS的mRNA表达。实时PCR的数据与RT-PCR的数据具有可比性。我们表明，这些RT-PCR引物组可用于半定量检测豚鼠组织中的mRNA表达，并得出结论，这些新设计的常规RT-PCR引物组将有助于在豚鼠结核病实验中研究免疫过程，以调查和评估新疫苗或抗分枝杆菌药物的疗效。