Response of Major Histocompatibility Complex (MHC) to DEX & PCN in Non-APC(pilot)

合成类固醇，双烯醇酮-16-α腈（PCN），几十年来一直用于探测在环境“压力”情况下激活的一系列假定的肝脏防御。以前的研究进行探索的全域基因的控制下的PCN采用差异显示检测大鼠肝脏mRNA的物种选择性诱导PCN或地塞米松（DEX）。这些研究揭示了RT1.B（I）β的诱导，RT1.B（I）β是通常仅在抗原呈递细胞（APC）中发现的主要组织相容性II类（MHC）基因家族的成员。从大鼠肝脏或培养的肝细胞中获得的初步数据证实，在DEX或PCN处理后3-6 h内，RT1.B（I）β mRNA及其伴随基因RT1.B（I）α的量容易诱导，而在脾脏（APC部位）中未观察到诱导。使用糖皮质激素的经典剂量反应实验总是 显示免疫组织中MHCII基因的抑制。然而，γ干扰素IFN γ在许多组织和细胞中是MHCU基因的最有效的诱导剂之一，但在肝细胞中不是。糖皮质激素在大鼠肝脏中对MHCII基因的意外诱导，在导致MHCII基因表达的可能机制方面提出了一系列问题。 这样的效果。在毒性应激期间，一组协调控制以维持实质组织内稳态的基因的概念现在必须扩展到包括免疫系统。由于大鼠和小鼠MHC-II组分之间的高度相似性（cDNA序列中91%的同源性），我们认为这种大鼠肝诱导现象也会在小鼠中发生。由于MHCII小鼠启动子是众所周知的，具有相当好的特征，并且非常相似 对于人类，它将是研究类固醇治疗引起的MHCII调节的极好和优选的模型。II类反式激活因子（CIITA）已被认为是MHCII基因转录的关键调节因子。已经描述了三种不同类型的启动子（I、III和IV）来调节小鼠中的CIITA转录。这项研究旨在确定是否可以通过类固醇治疗在小鼠中实现MHC-II基因的诱导，就像在大鼠中一样。如果 如果达到了这一点，那么我们就有了一个极好的模型来研究非APC中MHCII诱导所涉及的机制。我们将立即确定MHC II诱导是否依赖于或独立于CIITA，或者是否是通过新的机制。我们还将讨论CIITA启动子参与肝和肺的特定组织中的这种诱导。在小鼠中MHC II基因对类固醇诱导不敏感的情况下，我们计划继续我们的研究。 大鼠模型，并充分表征该物种中的启动子，以阐明与该现象相关的机制。这些实验的结果对于理解类固醇对MHC调节的作用至关重要，因为许多类固醇如地塞米松被用于治疗某些自身免疫性疾病（其特征在于： MHCII基因的过表达）。该提议旨在阐明DEX和PCN是否通过其在小鼠和大鼠中已知的反式激活途径或通过独立于CIITA的不同途径诱导非APC中的MHC II类基因。

The synthetic steroid, pregnenolone-16-alpha carbonitrile (PCN), has served for decades to probe for a postulated series of hepatic defenses activated under situations of environmental "stress". Previous research conducted to explore the full domain of genes under the control of PCN employed differential display to detect rat liver mRNA species selectively induced by PCN or by Dexamethasone(DEX). These studies revealed the induction of RT1.B (I)beta a member of the major histocompatability class II (MHC) gene family usually found only in antigen presenting cells (APC). Preliminary data obtained from rat liver or from cultured hepatocytes confirmed that amounts of RT1.B (I)beta mRNA and also of its companion gene, RT1.B (I)alpha, became readily induced within 3-6 h following DEX or PCN treatment, whereas no induction was observed in spleen (site of APC). Classical dose response experiments using glucocorticoids have always revealed inhibition of MHCII genes in immune tissues. However, gamma interferon IFNgamma is one of the most potent inducers of MHCU genes in many tissues and cells but not in hepatocytes. This unexpected induction of MHCII genes in the rat liver by glucocorticoids has opened a series of questions in terms of the possible mechanisms responsible for such effect. The concept of a set of genes coordinately controlled to maintain homeostasis in parenchymal tissues during toxic stress must now be extended to include the immune system. Due to the high similarities between rat and mice MHC-II components (91% homology in cDNA sequence) we believe that this rat liver induction phenomenon would also take place in mice. Since the MHCII mouse promoter is well known, fairly well characterized, and is very similar to humans, it would be an excellent and preferred model for studying MHCII regulation due to steroid treatment. The class II transactivator (CIITA) has been accepted as the key regulator for the transcription of MHCII genes. Three different types of promoters (I, III, and IV) have been described to regulate CIITA transcription in mice. This research is designed to determine if induction of MHC-II genes can be achieved by steroid treatment in mice as it is in rats. If this is achieved then we have an excellent model to study the mechanisms involved in MHCII induction in non-APC. We will immediately establish whether MHC II induction is dependent or independent on CIITA or whether it is through a new mechanism. We will also address which CIITA promoter is involved in this induction in the specific tissues of liver and lung. In the event that MHC II genes are refractory to steroid induction in mice we plan to continue with our rat model and fully characterize the promoter in this species in order to elucidate the mechanism associated with this phenomenon. The results of these experimentsare of utmost importance for understanding the effect of steroids on MHC regulation since many steroids such as dexamethasone are used to treat certain autoimmune diseases (characterized by over-expression of MHCII genes) at considerably high doses of up to 1000mg/day. This proposal is designed to elucidate if DEX and PCN are inducing the MHC class II genes in non-APC through its known transactivation pathway in mice and rat or through a different route independent of CIITA.