Publications

2021
Lindsay T Fourman, Takara L Stanley, James M Billingsley, Shannan Ho J Sui, Meghan N Feldpausch, Autumn Boutin, Isabel Zheng, Colin M McClure, Kathleen E Corey, Martin Torriani, David E Kleiner, Colleen M. Hadigan, Raymond T. Chung, and Steven K Grinspoon. 2021. “Delineating tesamorelin response pathways in HIV-associated NAFLD using a targeted proteomic and transcriptomic approach.” Scientific Reports, 11, 1, Pp. 1-8.Abstract
NAFLD is a leading comorbidity in HIV with an exaggerated course compared to the general population. Tesamorelin has been demonstrated to reduce liver fat and prevent fibrosis progression in HIV-associated NAFLD. We further showed that tesamorelin downregulated hepatic gene sets involved in inflammation, tissue repair, and cell division. Nonetheless, effects of tesamorelin on individual plasma proteins pertaining to these pathways are not known. Leveraging our prior randomized-controlled trial and transcriptomic approach, we performed a focused assessment of 9 plasma proteins corresponding to top leading edge genes within differentially modulated gene sets. Tesamorelin led to significant reductions in vascular endothelial growth factor A (VEGFA, log2-fold change - 0.20 ± 0.35 vs. 0.05 ± 0.34, P = 0.02), transforming growth factor beta 1 (TGFB1, - 0.35 ± 0.56 vs. - 0.05 ± 0.43, P = 0.05), and macrophage colony stimulating factor 1 (CSF1, - 0.17 ± 0.21 vs. 0.02 ± 0.20, P = 0.004) versus placebo. Among tesamorelin-treated participants, reductions in plasma VEGFA (r = 0.62, P = 0.006) and CSF1 (r = 0.50, P = 0.04) correlated with a decline in NAFLD activity score. Decreases in TGFB1 (r = 0.61, P = 0.009) and CSF1 (r = 0.64, P = 0.006) were associated with reduced gene-level fibrosis score. Tesamorelin suppressed key angiogenic, fibrogenic, and pro-inflammatory mediators. CSF1, a regulator of monocyte recruitment and activation, may serve as an innovative therapeutic target for NAFLD in HIV. Clinical Trials Registry Number: NCT02196831.
Takara L Stanley, Lindsay T Fourman, Lai Ping Wong, Ruslan Sadreyev, James M Billingsley, Meghan N Feldpausch, Isabel Zheng, Chelsea S Pan, Autumn Boutin, Hang Lee, Kathleen E Corey, Martin Torriani, David E Kleiner, Raymond T Chung, Colleen M Hadigan, and Steven K Grinspoon. 2021. “Growth Hormone Releasing Hormone Reduces Circulating Markers of Immune Activation in Parallel with Effects on Hepatic Immune Pathways in Individuals with HIV-Infection and Nonalcoholic Fatty Liver Disease. 2021 Apr 14:Epub ahead of print.” Clin Infect Dis.
2020
Rushdia Zareen Yusuf, Borja Saez, Azeem Sharda, Nick van Gastel, Vionnie WC Yu, Ninib Baryawno, Elizabeth W Scadden, Sanket Acharya, Shrikanta Chattophadhyay, Cherrie Huang, Vasanthi Viswanathan, Dana S'aulis, Julien Cobert, David B Sykes, Mark A Keibler, Sudeshna Das, John N Hutchinson, Michael Churchill, Siddhartha Mukherjee, Dongjun Lee, Francois Mercier, John Doench, Lars Bullinger, David J Logan, Stuart Schreiber, Gregory Stephanopoulos, William B Rizzo, and David T Scadden. 2020. “Aldehyde dehydrogenase 3a2 protects AML cells from oxidative death and the synthetic lethality of ferroptosis inducers.” Blood, 136, 11, Pp. 1303-1316.Abstract
Metabolic alterations in cancer represent convergent effects of oncogenic mutations. We hypothesized that a metabolism-restricted genetic screen, comparing normal primary mouse hematopoietic cells and their malignant counterparts in an ex vivo system mimicking the bone marrow microenvironment, would define distinctive vulnerabilities in acute myeloid leukemia (AML). Leukemic cells, but not their normal myeloid counterparts, depended on the aldehyde dehydrogenase 3a2 (Aldh3a2) enzyme that oxidizes long-chain aliphatic aldehydes to prevent cellular oxidative damage. Aldehydes are by-products of increased oxidative phosphorylation and nucleotide synthesis in cancer and are generated from lipid peroxides underlying the non-caspase-dependent form of cell death, ferroptosis. Leukemic cell dependence on Aldh3a2 was seen across multiple mouse and human myeloid leukemias. Aldh3a2 inhibition was synthetically lethal with glutathione peroxidase-4 (GPX4) inhibition; GPX4 inhibition is a known trigger of ferroptosis that by itself minimally affects AML cells. Inhibiting Aldh3a2 provides a therapeutic opportunity and a unique synthetic lethality to exploit the distinctive metabolic state of malignant cells.
Julie A Dragon, Chris Gates, Shannan Ho Sui, John N Hutchinson, Krishna Murthy R Karuturi, Alper Kucukural, Shawn Polson, Alberto Riva, Matthew Lee Settles, Jyothi Thimmapuram, and Stuart S Levine. 2020. “Bioinformatics Core Survey Highlights the Challenges Facing Data Analysis Facilities.” J Biomol Tech, 31, 2, Pp. 66-73.Abstract
Over the last decade, the cost of -omics data creation has decreased 10-fold, whereas the need for analytical support for those data has increased exponentially. Consequently, bioinformaticians face a second wave of challenges: novel applications of existing approaches (, single-cell RNA sequencing), integration of -omics data sets of differing size and scale (, spatial transcriptomics), as well as novel computational and statistical methods, all of which require more sophisticated pipelines and data management. Nonetheless, bioinformatics cores are often asked to operate under primarily a cost-recovery model, with limited institutional support. Seeing the need to assess bioinformatics core operations, the Association of Biomolecular Resource Facilities Genomics Bioinformatics Research Group conducted a survey to answer questions about staffing, services, financial models, and challenges to better understand the challenges bioinformatics core facilities are currently faced with and will need to address going forward. Of the respondent groups, we chose to focus on the survey data from smaller cores, which made up the majority. Although all cores indicated similar challenges in terms of changing technologies and analysis needs, small cores tended to have the added challenge of funding their operations largely through cost-recovery models with heavy administrative burdens.
Eri Katsuyama, Abel Suarez-Fueyo, Sean J Bradley, Masayuki Mizui, Ana V Marin, Lama Mulki, Suzanne Krishfield, Fabio Malavasi, Joon Yoon, Shannan Ho J Sui, Vasileios C Kyttaris, and George C Tsokos. 2020. “The CD38/NAD/SIRTUIN1/EZH2 Axis Mitigates Cytotoxic CD8 T Cell Function and Identifies Patients with SLE Prone to Infections.” Cell Rep, 30, 1, Pp. 112-123.e4.Abstract
Patients with systemic lupus erythematosus (SLE) suffer frequent infections that account for significant morbidity and mortality. T cell cytotoxic responses are decreased in patients with SLE, yet the responsible molecular events are largely unknown. We find an expanded CD8CD38 T cell subset in a subgroup of patients with increased rates of infections. CD8CD38 T cells from healthy subjects and patients with SLE display decreased cytotoxic capacity, degranulation, and expression of granzymes A and B and perforin. The key cytotoxicity-related transcription factors T-bet, RUNX3, and EOMES are decreased in CD8CD38 T cells. CD38 leads to increased acetylated EZH2 through inhibition of the deacetylase Sirtuin1. Acetylated EZH2 represses RUNX3 expression, whereas inhibition of EZH2 restores CD8 T cell cytotoxic responses. We propose that high levels of CD38 lead to decreased CD8 T cell-mediated cytotoxicity and increased propensity to infections in patients with SLE, a process that can be reversed pharmacologically.
Ruei-Jiun Hung, Yanhui Hu, Rory Kirchner, Yifang Liu, Chiwei Xu, Aram Comjean, Sudhir Gopal Tattikota, Fangge Li, Wei Song, Shannan Ho Sui, and Norbert Perrimon. 2020. “A cell atlas of the adult midgut.” Proc Natl Acad Sci U S A, 117, 3, Pp. 1514-1523.Abstract
Studies of the adult midgut have led to many insights in our understanding of cell-type diversity, stem cell regeneration, tissue homeostasis, and cell fate decision. Advances in single-cell RNA sequencing provide opportunities to identify new cell types and molecular features. We used single-cell RNA sequencing to characterize the transcriptome of midgut epithelial cells and identified 22 distinct clusters representing intestinal stem cells, enteroblasts, enteroendocrine cells (EEs), and enterocytes. This unbiased approach recovered most of the known intestinal stem cells/enteroblast and EE markers, highlighting the high quality of the dataset, and led to insights on intestinal stem cell biology, cell type-specific organelle features, the roles of new transcription factors in progenitors and regional variation along the gut, 5 additional EE gut hormones, EE hormonal expression diversity, and paracrine function of EEs. To facilitate mining of this rich dataset, we provide a web-based resource for visualization of gene expression in single cells. Altogether, our study provides a comprehensive resource for addressing functions of genes in the midgut epithelium.
Lina Driouk, Joanina K Gicobi, Yusuke Kamihara, Kayleigh Rutherford, Glenn Dranoff, Jerome Ritz, and Susanne HC Baumeister. 2020. “Chimeric Antigen Receptor T Cells Targeting NKG2D-Ligands Show Robust Efficacy Against Acute Myeloid Leukemia and T-Cell Acute Lymphoblastic Leukemia.” Front Immunol, 11, Pp. 580328.Abstract
CAR T cell approaches to effectively target AML and T-ALL without off-tumor effects on healthy myeloid or T cell compartments respectively are an unmet medical need. NKG2D-ligands are a promising target given their absence on healthy cells and surface expression in a wide range of malignancies. NKG2D-ligand expression has been reported in a substantial group of patients with AML along with evidence for prognostic significance. However, reports regarding the prevalence and density of NKG2D-ligand expression in AML vary and detailed studies to define whether low level expression is sufficient to trigger NKG2D-ligand directed CART cell responses are lacking. NKG2D ligand expression in T-ALL has not previously been interrogated. Here we report that NKG2D-ligands are expressed in T-ALL cell lines and primary T-ALL. We confirm that NKG2D-ligands are frequently surface expressed in primary AML, albeit at relatively low levels. Utilizing CAR T cells incorporating the natural immune receptor NKG2D as the antigen binding domain, we demonstrate striking activity of CAR T cells targeting NKG2D-ligands against AML and T-ALL cell lines and show that even low-level ligand expression in primary AML targets results in robust NKG2D-CAR activity. We found that NKG2D-ligand expression can be selectively enhanced in low-expressing AML cell lines and primary AML blasts pharmacologic HDAC inhibition. Such pharmacologic NKG2D-ligand induction results in enhanced NKG2D-CAR anti-leukemic activity without affecting healthy PBMC, thereby providing rationale for the combination of HDAC-inhibitors with NKG2D-CAR T cell therapy as a potential strategy to achieve clinical NKG2D-CAR T cell efficacy in AML.
Lindsay T Fourman, James M Billingsley, George Agyapong, Shannan J Ho Sui, Meghan N Feldpausch, Julia Purdy, Isabel Zheng, Chelsea S Pan, Kathleen E Corey, Martin Torriani, David E Kleiner, Colleen M Hadigan, Takara L Stanley, Raymond T Chung, and Steven K Grinspoon. 2020. “Effects of tesamorelin on hepatic transcriptomic signatures in HIV-associated NAFLD.” JCI Insight, 5, 16.Abstract
Nonalcoholic fatty liver disease (NAFLD) is a common comorbidity among people living with HIV that has a more aggressive course than NAFLD among the general population. In a recent randomized placebo-controlled trial, we demonstrated that the growth hormone-releasing hormone analog tesamorelin reduced liver fat and prevented fibrosis progression in HIV-associated NAFLD over 1 year. As such, tesamorelin is the first strategy that has shown to be effective against NAFLD among the population with HIV. The current study leveraged paired liver biopsy specimens from this trial to identify hepatic gene pathways that are differentially modulated by tesamorelin versus placebo. Using gene set enrichment analysis, we found that tesamorelin increased hepatic expression of hallmark gene sets involved in oxidative phosphorylation and decreased hepatic expression of gene sets contributing to inflammation, tissue repair, and cell division. Tesamorelin also reciprocally up- and downregulated curated gene sets associated with favorable and poor hepatocellular carcinoma prognosis, respectively. Notably, among tesamorelin-treated participants, these changes in hepatic expression correlated with improved fibrosis-related gene score. Our findings inform our knowledge of the biology of pulsatile growth hormone action and provide a mechanistic basis for the observed clinical effects of tesamorelin on the liver.
Cory J Smith, Oscar Castanon, Khaled Said, Verena Volf, Parastoo Khoshakhlagh, Amanda Hornick, Raphael Ferreira, Chun-Ting Wu, Marc Güell, Shilpa Garg, Alex HM Ng, Hannu Myllykallio, and George M Church. 2020. “Enabling large-scale genome editing at repetitive elements by reducing DNA nicking.” Nucleic Acids Res, 48, 9, Pp. 5183-5195.Abstract
To extend the frontier of genome editing and enable editing of repetitive elements of mammalian genomes, we made use of a set of dead-Cas9 base editor (dBE) variants that allow editing at tens of thousands of loci per cell by overcoming the cell death associated with DNA double-strand breaks and single-strand breaks. We used a set of gRNAs targeting repetitive elements-ranging in target copy number from about 32 to 161 000 per cell. dBEs enabled survival after large-scale base editing, allowing targeted mutations at up to ∼13 200 and ∼12 200 loci in 293T and human induced pluripotent stem cells (hiPSCs), respectively, three orders of magnitude greater than previously recorded. These dBEs can overcome current on-target mutation and toxicity barriers that prevent cell survival after large-scale genome engineering.
Zhihui Wang, Mulong Du, Qianyu Yuan, Yichen Guo, John N Hutchinson, Li Su, Yinan Zheng, Jun Wang, Lorelei A Mucci, Xihong Lin, Lifang Hou, and David C Christiani. 2020. “Epigenomic analysis of 5-hydroxymethylcytosine (5hmC) reveals novel DNA methylation markers for lung cancers.” Neoplasia, 22, 3, Pp. 154-161.Abstract
BACKGROUND: DNA methylation at the fifth position of cytosine (5mC) is a common epigenetic alteration affecting a range of cellular processes. In recent years, 5-hydroxymethylcytosine (5hmC), an oxidized form of 5mC, has risen broad interests as a potential biomarker for lung cancer diagnosis and survival. METHODS: We analyzed the epigenome-wide 5hmC profiles of paired lung tumor and adjacent normal tissues, using the TET-Assisted Bisulfite (TAB) array - Infinium MethylationEPIC BeadChip (EPIC) approach. The differentially methylated CpG sites were identified, and the biological relevance of 5hmC was assessed by differential methylation regions (DMR) analysis and gene set analysis (GSA). RESULTS: We observed global hypomethylation of 5hmC comparing tumor to normal tissues, and hypermethylated 5hmC were enriched in CpG islands and gene upstream. Comparison of 5hmC and 5modC (total methylation: 5mC + 5hmC) profiling showed low correlation, and only a small proportion of the significant 5hmC loci overlapped with the significant total methylation loci. GSA analysis suggested that 5hmC was mainly involved in biological processes such as cellular process, biological regulation, and metabolic process. CONCLUSION: This is the first study to analyze the epigenome-wide 5hmC profiles among paired lung tumor and normal tissues. We observed global hypomethylation of 5hmC in lung cancers, and hypermethylated 5hmC enriched in CpG islands and gene upstream. We found that the genome-wide 5hmC levels do not correlate with the total methylation, and the GSA suggested different biological functions of 5hmC compared to 5modC. Overall, our results demonstrate the potential of 5hmC as a novel biomarker for lung cancer.
Misty R Riddle, Ariel C Aspiras, Fleur Damen, John N Hutchinson, Daniel J-F Chinnapen, Julius Tabin, and Clifford J Tabin. 2020. “Genetic architecture underlying changes in carotenoid accumulation during the evolution of the blind Mexican cavefish, Astyanax mexicanus.” J Exp Zool B Mol Dev Evol, 334, 7-8, Pp. 405-422.Abstract
Carotenoids are lipid-soluble yellow to orange pigments produced by plants, bacteria, and fungi. They are consumed by animals and metabolized to produce molecules essential for gene regulation, vision, and pigmentation. Cave animals represent an interesting opportunity to understand how carotenoid utilization evolves. Caves are devoid of light, eliminating primary production of energy through photosynthesis and, therefore, limiting carotenoid availability. Moreover, the selective pressures that favor carotenoid-based traits, like pigmentation and vision, are relaxed. Astyanax mexicanus is a species of fish with multiple river-adapted (surface) and cave-adapted populations (i.e., Tinaja, Pachón, Molino). Cavefish exhibit regressive features, such as loss of eyes and melanin pigment, and constructive traits, like increased sensory neuromasts and starvation resistance. Here, we show that, unlike surface fish, Tinaja and Pachón cavefish accumulate carotenoids in the visceral adipose tissue. Carotenoid accumulation is not observed in Molino cavefish, indicating that it is not an obligatory consequence of eye loss. We used quantitative trait loci mapping and RNA sequencing to investigate genetic changes associated with carotenoid accumulation. Our findings suggest that multiple stages of carotenoid processing may be altered in cavefish, including absorption and transport of lipids, cleavage of carotenoids into unpigmented molecules, and differential development of intestinal cell types involved in carotenoid assimilation. Our study establishes A. mexicanus as a model to study the genetic basis of natural variation in carotenoid accumulation and how it impacts physiology.
Dmitry Prokopenko, Julian Hecker, Rory Kirchner, Brad A Chapman, Oliver Hoffman, Kristina Mullin, Winston Hide, Lars Bertram, Nan Laird, Dawn L DeMeo, Christoph Lange, and Rudolph E Tanzi. 2020. “Identification of Novel Alzheimer's Disease Loci Using Sex-Specific Family-Based Association Analysis of Whole-Genome Sequence Data.” Sci Rep, 10, 1, Pp. 5029.Abstract
With the advent of whole genome-sequencing (WGS) studies, family-based designs enable sex-specific analysis approaches that can be applied to only affected individuals; tests using family-based designs are attractive because they are completely robust against the effects of population substructure. These advantages make family-based association tests (FBATs) that use siblings as well as parents especially suited for the analysis of late-onset diseases such as Alzheimer's Disease (AD). However, the application of FBATs to assess sex-specific effects can require additional filtering steps, as sensitivity to sequencing errors is amplified in this type of analysis. Here, we illustrate the implementation of robust analysis approaches and additional filtering steps that can minimize the chances of false positive-findings due to sex-specific sequencing errors. We apply this approach to two family-based AD datasets and identify four novel loci (GRID1, RIOK3, MCPH1, ZBTB7C) showing sex-specific association with AD risk. Following stringent quality control filtering, the strongest candidate is ZBTB7C (P = 1.83 × 10), in which the minor allele of rs1944572 confers increased risk for AD in females and protection in males. ZBTB7C encodes the Zinc Finger and BTB Domain Containing 7C, a transcriptional repressor of membrane metalloproteases (MMP). Members of this MMP family were implicated in AD neuropathology.
Georgios Theocharidis, Dimitrios Baltzis, Matthieu Roustit, Ana Tellechea, Seema Dangwal, Radhika S Khetani, Bin Shu, Wanni Zhao, Jianfang Fu, Swati Bhasin, Antonios Kafanas, Daniel Hui, Shannan Ho Sui, Nikolaos A Patsopoulos, Manoj Bhasin, and Aristidis Veves. 2020. “Integrated Skin Transcriptomics and Serum Multiplex Assays Reveal Novel Mechanisms of Wound Healing in Diabetic Foot Ulcers.” Diabetes, 69, 10, Pp. 2157-2169.Abstract
Nonhealing diabetic foot ulcers (DFUs) are characterized by low-grade chronic inflammation, both locally and systemically. We prospectively followed a group of patients who either healed or developed nonhealing chronic DFUs. Serum and forearm skin analysis, both at the protein expression and the transcriptomic level, indicated that increased expression of factors such as interferon-γ (IFN-γ), vascular endothelial growth factor, and soluble vascular cell adhesion molecule-1 were associated with DFU healing. Furthermore, foot skin single-cell RNA sequencing analysis showed multiple fibroblast cell clusters and increased inflammation in the dorsal skin of patients with diabetes mellitus (DM) and DFU specimens compared with control subjects. In addition, in myeloid cell DM and DFU upstream regulator analysis, we observed inhibition of interleukin-13 and IFN-γ and dysregulation of biological processes that included cell movement of monocytes, migration of dendritic cells, and chemotaxis of antigen-presenting cells pointing to an impaired migratory profile of immune cells in DM skin. The and genes, which were upregulated at the forearm of nonhealers, were mainly expressed by the vascular endothelial cell cluster almost exclusively in DFU, indicating a potential important role in wound healing. These results from integrated protein and transcriptome analyses identified individual genes and pathways that can potentially be targeted for enhancing DFU healing.
Pritha Sen, Adrian R Wilkie, Fei Ji, Yiming Yang, Ian J Taylor, Miguel Velazquez-Palafox, Emilia AH Vanni, Jean M Pesola, Rosio Fernandez, Han Chen, Liza M Morsett, Erik R Abels, Mary Piper, Rebekah J Lane, Suzanne E Hickman, Terry K Means, Eric S Rosenberg, Ruslan I Sadreyev, Bo Li, Donald M Coen, Jay A Fishman, and Joseph El Khoury. 2020. “Linking indirect effects of cytomegalovirus in transplantation to modulation of monocyte innate immune function.” Sci Adv, 6, 17, Pp. eaax9856.Abstract
Cytomegalovirus (CMV) is an important cause of morbidity and mortality in the immunocompromised host. In transplant recipients, a variety of clinically important "indirect effects" are attributed to immune modulation by CMV, including increased mortality from fungal disease, allograft dysfunction and rejection in solid organ transplantation, and graft-versus-host-disease in stem cell transplantation. Monocytes, key cellular targets of CMV, are permissive to primary, latent and reactivated CMV infection. Here, pairing unbiased bulk and single cell transcriptomics with functional analyses we demonstrate that human monocytes infected with CMV do not effectively phagocytose fungal pathogens, a functional deficit which occurs with decreased expression of fungal recognition receptors. Simultaneously, CMV-infected monocytes upregulate antiviral, pro-inflammatory chemokine, and inflammasome responses associated with allograft rejection and graft-versus-host disease. Our study demonstrates that CMV modulates both immunosuppressive and immunostimulatory monocyte phenotypes, explaining in part, its paradoxical "indirect effects" in transplantation. These data could provide innate immune targets for the stratification and treatment of CMV disease.
Stefan Haemmig, Dafeng Yang, Xinghui Sun, Debapria Das, Siavash Ghaffari, Roberto Molinaro, Lei Chen, Yihuan Deng, Dan Freeman, Norman Moullan, Yevgenia Tesmenitsky, Khyrul AKM Wara, Viorel Simion, Eugenia Shvartz, James F Lee, Tianlun Yang, Galina Sukova, Jarrod A Marto, Peter H Stone, Warren L Lee, Johan Auwerx, Peter Libby, and Mark W Feinberg. 2020. “Long noncoding RNA integrates a DNA-PK-mediated DNA damage response and vascular senescence.” Sci Transl Med, 12, 531.Abstract
Long noncoding RNAs (lncRNAs) are emerging regulators of biological processes in the vessel wall; however, their role in atherosclerosis remains poorly defined. We used RNA sequencing to profile lncRNAs derived specifically from the aortic intima of mice on a high-cholesterol diet during lesion progression and regression phases. We found that the evolutionarily conserved lncRNA small nucleolar host gene-12 () is highly expressed in the vascular endothelium and decreases during lesion progression. knockdown accelerated atherosclerotic lesion formation by 2.4-fold in mice by increased DNA damage and senescence in the vascular endothelium, independent of effects on lipid profile or vessel wall inflammation. Conversely, intravenous delivery of protected the tunica intima from DNA damage and atherosclerosis. LncRNA pulldown in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis showed that interacted with DNA-dependent protein kinase (DNA-PK), an important regulator of the DNA damage response. The absence of reduced the DNA-PK interaction with its binding partners Ku70 and Ku80, abrogating DNA damage repair. Moreover, the anti-DNA damage agent nicotinamide riboside (NR), a clinical-grade small-molecule activator of NAD, fully rescued the increases in lesional DNA damage, senescence, and atherosclerosis mediated by knockdown. expression was also reduced in pig and human atherosclerotic specimens and correlated inversely with DNA damage and senescent markers. These findings reveal a role for this lncRNA in regulating DNA damage repair in the vessel wall and may have implications for chronic vascular disease states and aging.
Congli Zeng, Gabriel C Motta-Ribeiro, Takuga Hinoshita, Marcos Adriano Lessa, Tilo Winkler, Kira Grogg, Nathan M Kingston, John N Hutchinson, Lynette Marie Sholl, and Xiangming Fang. 2020. “Lung atelectasis promotes immune and barrier dysfunction as revealed by transcriptome sequencing in female sheep.” Anesthesiology, 133, 5, Pp. 1060-1076.
Zaniar Ghazizadeh, Tuomas Kiviniemi, Sigurast Olafsson, David Plotnick, Manu E Beerens, Kun Zhang, Leah Gillon, Michael J Steinbaugh, Victor Barrera, Shannan Ho Sui, Andreas A Werdich, Sunil Kapur, Antti Eranti, Jarmo Gunn, Juho Jalkanen, Juhani Airaksinen, Andre G Kleber, Maija Hollmén, and Calum A MacRae. 2020. “Metastable Atrial State Underlies the Primary Genetic Substrate for MYL4 Mutation-Associated Atrial Fibrillation.” Circulation, 141, 4, Pp. 301-312.Abstract
BACKGROUND: Atrial fibrillation (AF) is the most common clinical arrhythmia and is associated with heart failure, stroke, and increased mortality. The myocardial substrate for AF is poorly understood because of limited access to primary human tissue and mechanistic questions around existing in vitro or in vivo models. METHODS: Using an knock-in reporter line, we developed a protocol to generate and highly purify human pluripotent stem cell-derived cardiomyocytes displaying physiological and molecular characteristics of atrial cells. We modeled human mutants, one of the few definitive genetic causes of AF. To explore non-cell-autonomous components of AF substrate, we also created a zebrafish knockout model, which exhibited molecular, cellular, and physiologic abnormalities that parallel those in humans bearing the cognate mutations. RESULTS: There was evidence of increased retinoic acid signaling in both human embryonic stem cells and zebrafish mutant models, as well as abnormal expression and localization of cytoskeletal proteins, and loss of intracellular nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide + hydrogen. To identify potentially druggable proximate mechanisms, we performed a chemical suppressor screen integrating multiple human cellular and zebrafish in vivo endpoints. This screen identified Cx43 (connexin 43) hemichannel blockade as a robust suppressor of the abnormal phenotypes in both models of MYL4 (myosin light chain 4)-related atrial cardiomyopathy. Immunofluorescence and coimmunoprecipitation studies revealed an interaction between MYL4 and Cx43 with altered localization of Cx43 hemichannels to the lateral membrane in mutants, as well as in atrial biopsies from unselected forms of human AF. The membrane fraction from MYL4-/- human embryonic stem cell derived atrial cells demonstrated increased phospho-Cx43, which was further accentuated by retinoic acid treatment and by the presence of risk alleles at the Pitx2 locus. PKC (protein kinase C) was induced by retinoic acid, and PKC inhibition also rescued the abnormal phenotypes in the atrial cardiomyopathy models. CONCLUSIONS: These data establish a mechanistic link between the transcriptional, metabolic and electrical pathways previously implicated in AF substrate and suggest novel avenues for the prevention or therapy of this common arrhythmia.
Antonella FM Dost, Aaron L Moye, Marall Vedaie, Linh M Tran, Eileen Fung, Dar Heinze, Carlos Villacorta-Martin, Jessie Huang, Ryan Hekman, Julian H Kwan, Benjamin C Blum, Sharon M Louie, Samuel P Rowbotham, Julio Sainz de Aja, Mary E Piper, Preetida J Bhetariya, Roderick T Bronson, Andrew Emili, Gustavo Mostoslavsky, Gregory A Fishbein, William D Wallace, Kostyantyn Krysan, Steven M Dubinett, Jane Yanagawa, Darrell N Kotton, and Carla F Kim. 2020. “Organoids Model Transcriptional Hallmarks of Oncogenic KRAS Activation in Lung Epithelial Progenitor Cells.” Cell Stem Cell, 27, 4, Pp. 663-678.e8.Abstract
Mutant KRAS is a common driver in epithelial cancers. Nevertheless, molecular changes occurring early after activation of oncogenic KRAS in epithelial cells remain poorly understood. We compared transcriptional changes at single-cell resolution after KRAS activation in four sample sets. In addition to patient samples and genetically engineered mouse models, we developed organoid systems from primary mouse and human induced pluripotent stem cell-derived lung epithelial cells to model early-stage lung adenocarcinoma. In all four settings, alveolar epithelial progenitor (AT2) cells expressing oncogenic KRAS had reduced expression of mature lineage identity genes. These findings demonstrate the utility of our in vitro organoid approaches for uncovering the early consequences of oncogenic KRAS expression. This resource provides an extensive collection of datasets and describes organoid tools to study the transcriptional and proteomic changes that distinguish normal epithelial progenitor cells from early-stage lung cancer, facilitating the search for targets for KRAS-driven tumors.
Arpan C Ghosh, Sudhir Gopal Tattikota, Yifang Liu, Aram Comjean, Yanhui Hu, Victor Barrera, Shannan J Ho Sui, and Norbert Perrimon. 2020. “ PDGF/VEGF signaling from muscles to hepatocyte-like cells protects against obesity.” Elife, 9.Abstract
PDGF/VEGF ligands regulate a plethora of biological processes in multicellular organisms via autocrine, paracrine, and endocrine mechanisms. We investigated organ-specific metabolic roles of PDGF/VEGF-like factors (Pvfs). We combine genetic approaches and single-nuclei sequencing to demonstrate that muscle-derived Pvf1 signals to the hepatocyte-like cells/oenocytes to suppress lipid synthesis by activating the Pi3K/Akt1/TOR signaling cascade in the oenocytes. Functionally, this signaling axis regulates expansion of adipose tissue lipid stores in newly eclosed flies. Flies emerge after pupation with limited adipose tissue lipid stores and lipid level is progressively accumulated via lipid synthesis. We find that adult muscle-specific expression of increases rapidly during this stage and that muscle-to-oenocyte Pvf1 signaling inhibits expansion of adipose tissue lipid stores as the process reaches completion. Our findings provide the first evidence in a metazoan of a PDGF/VEGF ligand acting as a myokine that regulates systemic lipid homeostasis by activating TOR in hepatocyte-like cells.
Daniela M Santos, Lorena Pantano, Gina Pronzati, Paula Grasberger, Clemens K Probst, Katharine E Black, Jillian J Spinney, Lida P Hariri, Royale Nichols, Yufei Lin, Michael Bieler, Peter Seither, Paul Nicklin, David Wyatt, Andrew M Tager, and Benjamin D Medoff. 2020. “Screening for YAP Inhibitors Identifies Statins as Modulators of Fibrosis.” Am J Respir Cell Mol Biol, 62, 4, Pp. 479-492.Abstract
Idiopathic pulmonary fibrosis is a lung disease with limited therapeutic options that is characterized by pathological fibroblast activation and aberrant lung remodeling with scar formation. YAP (Yes-associated protein) is a transcriptional coactivator that mediates mechanical and biochemical signals controlling fibroblast activation. In this study, we developed a high-throughput small-molecule screen for YAP inhibitors in primary human lung fibroblasts. Multiple HMG-CoA (hydroxymethylglutaryl-coenzyme A) reductase inhibitors (statins) were found to inhibit YAP nuclear localization via induction of YAP phosphorylation, cytoplasmic retention, and degradation. We further show that the mevalonate pathway regulates YAP activation, and that simvastatin treatment reduces fibrosis markers in activated human lung fibroblasts and in the bleomycin mouse model of pulmonary fibrosis. Finally, we show that simvastatin modulates YAP in mouse lung fibroblasts. Our results highlight the potential of small-molecule screens for YAP inhibitors and provide a mechanism for the antifibrotic activity of statins in idiopathic pulmonary fibrosis.

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