Pulmonary hypertension (PH) is a debilitating disease in which a remodeled pulmonary vasculature augments pulmonary vascular resistance with increased right ventricular afterload and ultimately right ventricular failure. Current PH therapies provide symptomatic relief, but fall short as to reestablishment of structural and functional lung vascular integrity, as a basis for handicap-free long-term survival. Thus, the restoration of physiological vascular structure and function is the challenge in current PH research (Pullamsetti et al, Am J Respir Crit Care Med 195:425, 2017; Humbert et al, Eur Respir J 53:1801887,2019). Over the past several years, the Pullamsetti lab substantially contributed to progress in this field. This group delineated growth factor and inflammatory networks and their downstream effectors like FOXO, HIF and NFkB and demonstrated their importance in PH onset and development (Pullamsetti et al, Circulation 123:1194, 2011; Savai et al, Am J Respir Crit Care Med 186:897, 2012; Weiss et al, Nat Commun 10:2204, 2019; Berghausen et al, J Clin Invest, 2021). In seminal studies, smooth muscle cell FoxO1 was identified as critical integrator of multiple signaling pathways, driving a phenotypic switch of these cells with subsequent lung vascular remodeling (Savai et al, Nat Med 20:1289, 2014). Notably, reconstitution of FoxO1 activity reversed lung vascular remodeling, a concept carried forward to the current (pre)-clinical development of nanoparticle-based inhaled paclitaxel as novel approach for the treatment of PH. In addition, this group has identified a hitherto unknown feed-forward loop between RASSF1A (scaffold protein) and HIF-1α, promoting metabolic reprogramming and glycolytic shift in hypoxia-exposed lung vascular cells, a mechanism overlapping with central features in cancer biology (Dabral et al, Nat Commun 10:2130, 2019; Pullamsetti et al, J Clin Invest 30:5638, 2020). Furthermore, when delineating the epigenetic landscape underlying human pulmonary vascular cell phenotypic switching in PH, strong differential expression of > 2000 genes (paired-end RNA-seq) as compared to healthy controls was documented, along with massive alterations in DNA methylation (Illumina Arrays) and changes in the chromatin landscape (histone acetylation, methylation by ChIP-seq). Importantly, the “integromic” approaches, i.e. the combination of epigenetic and transcriptional changes, allowed to comprehensively elucidate the disease-specific networks and potential targets for the reversal of disease phenotypes. Completing this approach, the Pullamsetti lab has delineated abnormalities of non-coding RNAs and of histone- and non-histone mediated functions of histone modifiers linked with the pathobiology of PH, leading to the evaluation of the therapeutic efficacy of histone acetylation/deacetylation inhibitors in PH (Pullamsetti et al, Am J Respir Crit Care Med 185:409, 2012; Zehendner et al, Am J Respir Crit Care Med 202:1445, 2020; Qi et al, Circulation, 2021; Chelladurai et al, Sci Transl Med, in revision).
Extending the studies in gene regulatory networks and epigenomics to further lung diseases, the Pullamsetti lab documented an important role of nitric oxide signaling in alveolar epithelial cells and FoxO3 as critical integrator of multiple signaling pathways in parenchymal fibroblasts from experimental models of lung fibrosis and from tissue samples of patients with IPF (idiopathic lung fibrosis) (Pullamsetti et al, Sci Transl Med 3:87ra53, 2011; Al-Tamari et al, EMBO Mol Med 10:276, 2018) and identified histone deacetylases (HDAC9) as a master regulator of lung fibrogenesis (Korfei et al, Thorax 70:1022, 2015; Sarvari et al, in submission). This group also identified an association of clonal hematopoiesis of indeterminate potential (ChIP) and DNA methylation with inflammatory gene expression in patients with COPD (Kuhnert et al, in submission). Furthermore, zooming into the molecular heterogeneity of the microenvironmental niche in lung cancer, transcriptional and epigenomic profiling of macrophages and T cells was accomplished (Pullamsetti et al, Sci Transl Med 9:eaai9048, 2017; Salazar et al, J Clin Invest 130:3560, 2020; Sarode et al, Sci Adv 6:eaaz6105, 2020).
Epigenetic mechanisms, including DNA accessibility, histone modifications, DNA methylation and non-coding RNAs, regulate gene expression and translation and thereby cell phenotype and function. Moreover, epigenetic changes act as mediators of the interaction between environmental factors (hypoxia, infection, smoke, physical activity etc.) and genetic factors. Epigenome-focused therapies thus represent a novel arena for the development of lung health maintaining and restoring therapies. Against this background, the Pullamsetti lab will focus on epigenomic profiling of different lung cells to understand their phenotypic/metabolic/functional heterogeneity in pathological vascular and parenchymal remodeling events, as occurring in various lung diseases (see overview figure). Importantly, generating cell-type specific transcriptomic and epigenomic maps at single-cell resolution is crucial to understand the cell-state transitions and the cellular cross talk between epithelial, endothelial, fibroblast-type and smooth muscle cells, including their resident stem/progenitor cells, as well as infiltrating bone marrow-derived immune and progenitor cells. To this end, this group has established and will further extend single-cell and single-nuclei preparation protocols for experimental and human lung tissue. Specifically, to define the cellular composition and phenotypic heterogeneity of novel subpopulations, isolated cells will be sorted by flow activated cell sorting and subjected to gene expression profiling using single-cell or single-nuclei RNA sequencing (scRNA-seq, snRNA-seq, 10x Genomics), and chromatin accessibility using microfluidics-based single-cell assays for transposase-accessible chromatin (scATAC-seq, 10x Genomics). Multi-omic analysis of corresponding scRNA-seq and scATAC-seq datasets will be performed to map cellular identities and subclusters and their respective molecular signatures. Results of this “integromic” approach will be linked with architectural and functional pulmonary abnormalities in the various lung diseases, profiting from the rich source of experimental models and biobanked human tissue available via ILH and DZL. Moreover, the studies will be extended to nucleosome epigenetic features including histone and DNA modifications as well as histone variants or nucleosome adducts, representing a further level of complexity. We expect that these studies will substantially advance our knowledge of the “deep” genomic/gene regulatory events governing lung health maintenance versus disease development, that new classes of diagnostic and prognostic biomarkers will be identified, and that “epigenome-focused therapies” will offer new approaches for preserving and restoring lung health beyond currently available therapies. Against this background, a large number of cooperative projects within the ILH have been and will be pursued in the forthcoming years, as listed in the subsequent section.
Prof. Dr. Soni Savai Pullamsetti
Lung Vascular Epigenetics
Center for Infection and Genomics of the Lung (CIGL),
Justus Liebig University Giessen
Tel.: +49 (0) 641 99 36452
Fax: +49 (0) 641 99 36519
Max-Planck-Institute for Heart and Lung Research
61231 Bad Nauheim
Tel.: +49 (0) 6032 705 380
Fax: +49 (0) 6032 705 471
Tel: +49 (0) 641 99 36541
Fax: +49 (0) 641 99 36519
Tel: +49 (0) 6032 705 249
Fax: +49 (0) 6032 705 471
Prakash Chelladurai, Giovanni Maroli, Manjupadma Nandigama, Anoop Cherian, Chanil Valasarajan
Sreenath Nayakanti, Edibe Avci, Golnaz Hesami, Samuel Olapoju, Leili Jafari, Fatemeh Khassafi, Zahraa Msheik
Michéle Ebeling, Jonas Lund
Katharina Leib, Laura Kahnke, Uta Eule, Jana Rostkovius