Gene expression regulation mediating the activation and differentiation of cardiac fibroblasts and pulmonary fibroblasts after myocardial infarction

在组织损伤后，成纤维细胞被迅速激活，然后分化为肌成纤维细胞，这有助于愈合过程。我们最近发现，在小鼠心肌梗死(ML)模型中，肌成纤维细胞进一步分化为基质细胞，这是一种新发现的成纤维细胞分化状态，可能与慢性纤维化组织的进行性僵硬和功能降低有关。除了对经常发生ML的左心室的直接影响外，左心室功能下降还会导致左心房的压力超负荷，然后转移到肺中。肺内压力积聚会激活肺成纤维细胞(PFS)，导致间质纤维化，从而对心力衰竭的预后产生负面影响。有趣的是，我们的初步研究发现，与ML后心脏成纤维细胞(CFs)的一过性MF状态不同，PFS甚至在ML后4周仍处于MF状态，这表明两种成纤维细胞对ML的反应存在差异，这可能涉及到尚不清楚的不同机制。我们假设CFS和PFS对ML的反应的激活和分化与差异基因表达和染色质可及性有关，并涉及特定的转录因子。由于成纤维细胞在心脏和肺重建中的重要而复杂的作用，研究成纤维细胞在CFs和PFS中的基因表达调控将是至关重要的，因为它可以微调成纤维细胞的活性，这将有助于开发新的治疗策略。在目标1中，将进行单细胞RNA-seq(scRNA-seq)和scATAC-seq在单细胞水平上研究CFS的基因表达谱和染色质可及性。结果将与我们初步研究中产生的大量RNA-SEQ和ATAC-SEQ数据相结合，揭示染色质可及性在ML后CFS基因表达调控中的作用。RNA-SEQ和ATAC-SEQ数据的基序浓缩分析将通过CUT&RUN实验进行验证。 验证的结果将被用于构建一个基因调控网络，介导ML后CF的激活和分化。在目标2中，将使用相同的策略来研究染色质可及性在心肌梗死后PFS基因表达调控中的作用。此外，还将构建基因调控网络，以探索介导心肌梗死后PF激活和分化的信号通路。将对两个AIMS产生的数据进行比较，以确定共同和独特的特征。

Upon tissue injury, fibroblasts are rapidly activated and then differentiate into myofibroblasts, which facilitate the healing process. We recently identified that in a mouse model of myocardial infarction (Ml), myofibroblasts further differentiated to matribrocytes, a newly discovered fibroblasts differentiation state which may contribute to the progressive stiffening and function reduction in tissue suffering from chronic fibrosis. In addition to the direct impact on the left ventricle, where Ml often occurs, the reduced function of the left ventricle also causes pressure overload in the left atrium, which is then transferred into the lung. Pressure buildup in the lung activates pulmonary fibroblasts (PFs) and leads to interstitial fibrosis, which negatively impacts the prognosis of heart failure. Interestingly, our preliminary study identified that unlike the transient MF state of cardiac fibroblasts (CFs) after Ml, PFs remain in the MF state even 4 weeks after Ml, suggesting a difference in the response of two fibroblast populations to Ml, which may involve distinct mechanisms that are still unknown. We hypothesize that the activation and differentiation of CFs and PFs in response to Ml are associated with differential gene expression and chromatin accessibilities and involve specific transcription factors. Due to the important and complex role of fibroblasts in post-Ml cardiac and pulmonary remodeling, the study of the gene expression regulation in CFs and PFs will be critical as it may allow the fine-tuning of fibroblast activities, which can contribute to the development of novel treatment strategies. In Aim #1, single-cell RNA-seq (scRNA-seq) and scATAC-seq will be performed to study the gene expression profile and chromatin accessibility in CFs at the single-cell level. Results will be combined with bulk RNA-seq and ATAC-seq data generated in our preliminary study to reveal the role of chromatin accessibility in gene expression regulation in CFs after Ml. The motif enrichment analysis of RNA-seq and ATAC-seq data will be validated by CUT&RUN experiments. Validated results will be used in the construction of a gene regulatory network mediating CF activation and differentiation after Ml. In Aim #2, the same strategy will be used to study the role of chromatin accessibility in the gene expression regulations of PFs after MI. Gene regulatory network construction will also be performed to explore the signaling pathways mediating PF activation and differentiation after MI. Data generated in the two aims will be compared to identify common and unique features.