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。我们使用生物信息学方法来鉴定干扰素γ mRNA的3'非翻译部分的保守区域。据认为，这些保守区域代表了基因结构中重要的调控元件，因为除非mRNA的非编码区域提供了一些进化优势，否则通过进化就没有固有的保护原因。基于这一分析，我们针对小鼠干扰素- γ 3'非翻译区域的160 bp区域进行删除，因为该区域富含AUUA序列，并且该区域在之前被证明对细胞因子基因mRNA不稳定性的调节很重要。在C57 BL/6基因背景下成功构建了敲除（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刺激下产生更多的干扰素- 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。随着CD4+和CD8+ T细胞的增加，T细胞的稳态被破坏，小鼠体内的T细胞具有更强的抑制活性。对抗原刺激，TH1反应增加，TH2反应减少。B细胞群也被改变，基线抗体产生是倾斜的。在胸腺中也观察到B细胞的频率增加，从而表明ifn - γ可能改变B细胞的运输。除了表型后果外，由于IgM和Ig2a抗体反应增加而IgG1反应减少，B细胞对抗原的反应也被破坏。强烈的抗dna和抗核抗原抗体反应也被观察到，这表明慢性ifn - γ表达可能在狼疮的发展中起作用。雌性小鼠也会患上原发性胆管炎，这是一种病因不明的疾病。这是第一个概括人类疾病与性别偏见的小鼠模型因为90%的人类病例是女性。奇怪的是，在Balb/c小鼠中没有发现狼疮样症状和PBC症状，而Balb/c小鼠的160 bp缺失也被交叉到该遗传背景上。这些小鼠的脾脏增大，淋巴细胞转运到淋巴结有明显缺陷。这导致了与再生障碍性贫血非常相似的表型。这些影响似乎是干扰素γ抑制造血祖细胞的直接结果。此外，缺失的Balb/c小鼠似乎更能抵抗肾癌RENCA细胞系的攻击。总之，我们阐明干扰素- γ生物学中涉及的多种机制的方法表明了干扰素- γ基因表达改变宿主稳态的复杂性。除了这些影响，我们现在已经观察到慢性IFN-g表达影响小鼠的行为和心脏功能。因此，我们现在已经建立了一个小鼠模型来理解和阐明长期慢性ifn - γ基因表达的系统生物学效应，从而导致宿主的慢性炎症。

We are utilizing a mouse model 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 reason for conservation through evolution unless the non-coding regions of the mRNA provided some evolutionary advantage. Based on this analysis, we 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 mRNA instability. 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 stimulation 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 cholangitis, a disease that has no known etiology. This is the first mouse model to recapitulate the human disease with respect to sex bias as 90% of the human cases are in women. 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. 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. In addeition to these effects, we have now observed that chronic IFN-g expression affects mouse behavior and heart function. Thus, we now have developed a mouse model for understanding and elucidating the systems biology effects of long term chronic IFN-gamma gene expression, resulting in chronic inflammation in the host.