Consequences of chronic Interferon-gamma expression on the host

我们正在利用慢性IFN-γ表达的小鼠模型来确定对宿主的生物学后果以及这种表型与人类疾病的相关性。我们已经使用生物信息学方法来确定干扰素-γ mRNA的3'非翻译部分的保守区域。据信，这些保守区域代表基因结构中的重要调控元件，因为除非mRNA的非编码区提供一些进化优势，否则在进化中不存在用于保守的固有区域。基于该分析，我们靶向了鼠干扰素-γ 3'非翻译区的160-bp区域进行缺失，因为该区域富含AUUA序列，并且这些区域先前已被证明在细胞因子基因表达的调节中是重要的。在C57 BL/6遗传背景下成功地建立了敲除（KO）小鼠，我们的数据表明，与野生型小鼠相比，该小鼠在基础水平和用IL-12治疗后产生显著更多的干扰素-γ。此外，淋巴结、脾脏和胸腺的结构被破坏，肝脏表现出慢性炎症的迹象。由于存在增加的CD 4+和CD 8 + T细胞，并且小鼠中的T reg细胞具有更有效的抑制活性，T细胞稳态已被破坏。对抗原刺激也有增加的TH 1应答和减少的TH 2应答。B细胞群也发生了改变，基线抗体产生也发生了偏移。在胸腺中也观察到B细胞频率增加，因此表明IFN-γ可能改变B细胞运输。除了表型结果外，B细胞对抗原的应答也被破坏，因为观察到IgM和Ig 2a ab应答增加，而IgG 1应答降低。还观察到强的抗DNA和抗核抗原抗体应答，表明慢性IFN-γ表达可能在狼疮的发展中起作用。雌性小鼠也会发生原发性胆汁性手病，这是一种病因不明的疾病。这是第一个在性别偏见方面重现人类疾病的小鼠模型。奇怪的是，在Balb/c小鼠中没有看到狼疮样和PBC症状，其中160 bp的缺失也被交叉到遗传背景上。这些小鼠的脾脏尺寸增大，淋巴细胞向淋巴结的运输存在明显缺陷。这导致一个表型非常相似的再生障碍性贫血，我们正在阐明IFN-γ在这种疾病的发展中的作用。这些作用似乎是干扰素-γ抑制该小鼠品系造血祖细胞的直接结果。此外，缺失的Balb/c小鼠似乎对肾癌细胞系RENCA的攻击更具抗性。总之，我们的方法对阐明干扰素-γ的生物学中涉及的多种机制表明干扰素-γ基因表达改变宿主体内平衡的复杂性。此外，我们现在已经开发了一种小鼠模型，用于理解和阐明长期慢性IFN-γ基因表达的系统生物学效应。

We are utilizing mouse models of chronic IFN-gamma expression to determine the biological consequences to the host and the relevance of this phenotype to human disease. We have used a bioinformatics approach to identify conserved regions of the 3' untranslated portion of the interferon-gamma mRNA. It is believed that these conserved regions represent important regulatory elements in the gene structure, as there would be no inherent region for conservation through evolution unless the non-coding regions of the mRNA provided some evolutionary advantage. Based on this analysis, we have targeted a 160-bp region of the murine interferon-gamma 3' untranslated region for deletion as this region is rich in AUUA sequences, and such regions have been previously shown to be important in the regulation of cytokine gene expression. The knockout (KO) mouse has been successfully created on the C57 BL/6 genetic background and our data indicates that this mouse produces significantly more interferon-gamma at a basal level and upon treatment with IL-12 as compared to the wild-type mouse. Furthermore, the architecture of lymph nodes, spleen, and thymus is disrupted and the liver exhibits signs of chronic inflammation. T cell homeostasis has been disrupted as increased CD4+ and CD8+ T cells are present and the T reg cells in the mouse have more potent suppressor activity. There is also an increased TH1 response and a decreased TH2 response to antigenic stimulation. The B cell population is also altered and baseline antibody production is skewed. B cells are also observed in the thymus at increased frequency, thus indicating that IFN-gamma may alter B cell trafficking. In addition to the phenotypic consequences, the B cell response to antigen is also disrupted as increased IgM and Ig2a ab responses are seen with a decrease in the IgG1 response. Strong anti-DNA and anti-nuclear antigen antibody responses are also observed suggesting that chronic IFN-gamma expression may play a role in the development of lupus. The female mice also develop primary biliary chirosis, a disease that has no known etiology. This is the first mouse model to recapitulate the human disease with respect to sex bias. Curiously, lupus-like and PBC symptoms are not seen in the Balb/c mouse, where the 160-bp deletion has also been crossed onto that genetic background. These mice have increased spleen sizes and an apparent defect in lymphocyte trafficking to the lymph nodes. This results in a phenotype very similar to aplastic anemia and we are elucidating the role of IFN-gamma in the development of this disease. These effects appear to be a direct consequence of interferon-gamma inhibition of hematopoietic progenitors in this mouse strain. In addition, the Balb/c mice with the deletion appear to be more resistant to challenge with the renal carcinoma line RENCA. In summary, our approach towards elucidating the multiple mechanisms involved in the biology of interferon-gamma demonstrates the complexity by which interferon-gamma gene expression alters host homeostasis. Furthermore, we now have developed a mouse model for understanding and elucidating the systems biology effects of long term chronic IFN-gamma gene expression.