The Role of KCNMB1 and the Large Conductance Potassium (BK) Channel in Myofibroblast Differentiation and Pulmonary Fibrosis

项目总结 尽管在理解肺纤维化的发病机制方面取得了重大进展，但许多治疗方法 针对研究最充分的基因和途径的研究并未在逆转或 甚至可以阻止疾病的发展。这与特发性疾病患者的临床异质性有关。 肺纤维化(IPF)，提示在疾病发病机制模型中考虑其他基因可能是 很有用。特发性肺纤维化患者成纤维细胞许多基因的表达与正常相比不同 成纤维细胞，这个实验室长期以来一直对识别表观遗传变化感兴趣，这解释了 对于这些差异。KCNMB1编码大电导的β亚基(BK、Maxi-K、KCa1.1) 钾通道，并在我们先前的微阵列研究中被确定为最高差异甲基化基因 在IPF成纤维细胞中。BK通道调节钾电流，众所周知在血管中起重要作用 肌张力和平滑肌生物学，但其在纤维化中的重要性从未被研究过。我们最近展示了 在一篇文献中，1)KCNMB1在IPF患者的成纤维细胞中表达增加，2)KCNMB1 有助于增加BK通道的活性，以及3)BK通道功能的增强促进肌成纤维细胞 差异化，IPF的一个标志。它是如何做到这一点的，这是否足以促进或恶化肺脏 体内的纤维化尚不清楚。这笔赠款的目标是确定BK渠道的机制 促进肌成纤维细胞分化并建立BK通道在动物模型中的重要性 肺纤维化。我们的中心假设是KCNMB1的表观遗传学上调和BK的增加 IPF成纤维细胞中的通道活性通过促进成纤维细胞中的钙信号转导而参与肺纤维化。 从而导致肌成纤维细胞分化。第一个目标是确定BK渠道对 体内肺纤维化的发生，并定位其对肺成纤维细胞的致病作用。第二个 目的是阐明BK通道促进肌成纤维细胞分化的机制。 假设BK通道促进细胞内钙信号，这是分化为 肌成纤维细胞。第三个目的是确定KCNMB1在肺成纤维细胞中的表达是如何调节的 促纤维化刺激如何调节BK通道的开放和关闭。这项建议意义重大，因为它 使BK通道成为肺纤维化发病机制中一种新的但重要的驱动因素。 实现这些目标还将确定BK通道在分化中的机制和作用 以前从未被描述过的肌成纤维细胞。最终，这些研究将有助于确定新的 未来IPF治疗的靶点。

PROJECT SUMMARY Despite major advances in the understanding of the pathogenesis of pulmonary fibrosis, many of the therapies that target the most well-studied genes and pathways have not achieved universal success in reversing or even halting disease progression. This, along with the clinical heterogeneity of patients with idiopathic pulmonary fibrosis (IPF), suggest that consideration of other genes in models of disease pathogenesis may be useful. Fibroblasts from patients with IPF differ in the expression of many genes compared to normal fibroblasts, and this laboratory has had a longstanding interest in identifying epigenetic changes that account for these differences. KCNMB1 codes for the beta subunit of the large conductance (BK, Maxi-K, KCa1.1) potassium channel and was identified in our previous microarray study as the top differentially methylated gene in IPF fibroblasts. BK channels modulate potassium current and are well known to be important in vascular tone and smooth muscle biology, but its importance in fibrosis has never been examined. We recently showed in a publication that 1) KCNMB1 expression is increased in fibroblasts from IPF patients, 2) KCNMB1 contributes to increased BK channel activity, and 3) increased function of BK channels promote myofibroblast differentiation, a hallmark of IPF. How it does so and whether this is sufficient to promote or worsen pulmonary fibrosis in vivo is unknown. The objectives of this grant are to determine the mechanism of how BK channels contribute to myofibroblast differentiation and establish the importance of BK channels to animal models of pulmonary fibrosis. Our central hypothesis is that the epigenetic upregulation of KCNMB1 and increased BK channel activity in IPF fibroblasts contribute to pulmonary fibrosis by promoting calcium signaling in fibroblasts, which lead to myofibroblast differentiation. The First Aim is to establish the importance of BK channels to the development of pulmonary fibrosis in vivo, and localize its pathogenic actions to lung fibroblasts. The Second Aim is to delineate the mechanism by which BK channels contribute to myofibroblast differentiation, with the hypothesis that BK channels promote intracellular calcium signaling, which is necessary for differentiation into myofibroblasts. The Third Aim is to determine how expression of KCNMB1 is regulated in lung fibroblasts and how profibrotic stimuli modulates opening and closing of BK channels. This proposal is significant because it establishes BK channels as a novel, but important driver in the pathogenesis of pulmonary fibrosis. Accomplishing these aims will also identify a mechanism and role for BK channels in the differentiation of myofibroblasts that has never been previously described. Ultimately, these studies will serve to identify new targets for future IPF therapeutics.