Publications by authors named "Sara Ranjbarvaziri"

12 Publications

  • Page 1 of 1

Deep learning detects cardiotoxicity in a high-content screen with induced pluripotent stem cell-derived cardiomyocytes.

Elife 2021 08 2;10. Epub 2021 Aug 2.

Tenaya Therapeutics, South San Francisco, United States.

Drug-induced cardiotoxicity and hepatotoxicity are major causes of drug attrition. To decrease late-stage drug attrition, pharmaceutical and biotechnology industries need to establish biologically relevant models that use phenotypic screening to detect drug-induced toxicity in vitro. In this study, we sought to rapidly detect patterns of cardiotoxicity using high-content image analysis with deep learning and induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). We screened a library of 1280 bioactive compounds and identified those with potential cardiotoxic liabilities in iPSC-CMs using a single-parameter score based on deep learning. Compounds demonstrating cardiotoxicity in iPSC-CMs included DNA intercalators, ion channel blockers, epidermal growth factor receptor, cyclin-dependent kinase, and multi-kinase inhibitors. We also screened a diverse library of molecules with unknown targets and identified chemical frameworks that show cardiotoxic signal in iPSC-CMs. By using this screening approach during target discovery and lead optimization, we can de-risk early-stage drug discovery. We show that the broad applicability of combining deep learning with iPSC technology is an effective way to interrogate cellular phenotypes and identify drugs that may protect against diseased phenotypes and deleterious mutations.
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http://dx.doi.org/10.7554/eLife.68714DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8367386PMC
August 2021

Hypertrophic cardiomyopathy β-cardiac myosin mutation (P710R) leads to hypercontractility by disrupting super relaxed state.

Proc Natl Acad Sci U S A 2021 Jun;118(24)

Department of Pediatrics (Cardiology), Stanford University School of Medicine, Palo Alto, CA 94304;

Hypertrophic cardiomyopathy (HCM) is the most common inherited form of heart disease, associated with over 1,000 mutations, many in β-cardiac myosin (MYH7). Molecular studies of myosin with different HCM mutations have revealed a diversity of effects on ATPase and load-sensitive rate of detachment from actin. It has been difficult to predict how such diverse molecular effects combine to influence forces at the cellular level and further influence cellular phenotypes. This study focused on the P710R mutation that dramatically decreased in vitro motility velocity and actin-activated ATPase, in contrast to other MYH7 mutations. Optical trap measurements of single myosin molecules revealed that this mutation reduced the step size of the myosin motor and the load sensitivity of the actin detachment rate. Conversely, this mutation destabilized the super relaxed state in longer, two-headed myosin constructs, freeing more heads to generate force. Micropatterned human induced pluripotent derived stem cell (hiPSC)-cardiomyocytes CRISPR-edited with the P710R mutation produced significantly increased force (measured by traction force microscopy) compared with isogenic control cells. The P710R mutation also caused cardiomyocyte hypertrophy and cytoskeletal remodeling as measured by immunostaining and electron microscopy. Cellular hypertrophy was prevented in the P710R cells by inhibition of ERK or Akt. Finally, we used a computational model that integrated the measured molecular changes to predict the measured traction forces. These results confirm a key role for regulation of the super relaxed state in driving hypercontractility in HCM with the P710R mutation and demonstrate the value of a multiscale approach in revealing key mechanisms of disease.
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http://dx.doi.org/10.1073/pnas.2025030118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8214707PMC
June 2021

4HNE Impairs Myocardial Bioenergetics in Congenital Heart Disease-Induced Right Ventricular Failure.

Circulation 2020 10 18;142(17):1667-1683. Epub 2020 Aug 18.

Department of Pediatrics (Cardiology) (HT.V.H., N.S., S.L.P., S. Ranjbarvairi, D-Q.H., M.Z., G.F., D.B., S. Reddy), Stanford University, Palo Alto, CA.

Background: In patients with complex congenital heart disease, such as those with tetralogy of Fallot, the right ventricle (RV) is subject to pressure overload stress, leading to RV hypertrophy and eventually RV failure. The role of lipid peroxidation, a potent form of oxidative stress, in mediating RV hypertrophy and failure in congenital heart disease is unknown.

Methods: Lipid peroxidation and mitochondrial function and structure were assessed in right ventricle (RV) myocardium collected from patients with RV hypertrophy with normal RV systolic function (RV fractional area change, 47.3±3.8%) and in patients with RV failure showing decreased RV systolic function (RV fractional area change, 26.6±3.1%). The mechanism of the effect of lipid peroxidation, mediated by 4-hydroxynonenal ([4HNE] a byproduct of lipid peroxidation) on mitochondrial function and structure was assessed in HL1 murine cardiomyocytes and human induced pluripotent stem cell-derived cardiomyocytes.

Results: RV failure was characterized by an increase in 4HNE adduction of metabolic and mitochondrial proteins (16 of 27 identified proteins), in particular electron transport chain proteins. Sarcomeric (myosin) and cytoskeletal proteins (desmin, tubulin) also underwent 4HNE adduction. RV failure showed lower oxidative phosphorylation (moderate RV hypertrophy, 287.6±19.75 versus RV failure, 137.8±11.57 pmol/[sec×mL]; =0.0004), and mitochondrial structural damage. Using a cell model, we show that 4HNE decreases cell number and oxidative phosphorylation (control, 388.1±23.54 versus 4HNE, 143.7±11.64 pmol/[sec×mL]; <0.0001). Carvedilol, a known antioxidant did not decrease 4HNE adduction of metabolic and mitochondrial proteins and did not improve oxidative phosphorylation.

Conclusions: Metabolic, mitochondrial, sarcomeric, and cytoskeletal proteins are susceptible to 4HNE-adduction in patients with RV failure. 4HNE decreases mitochondrial oxygen consumption by inhibiting electron transport chain complexes. Carvedilol did not improve the 4HNE-mediated decrease in oxygen consumption. Strategies to decrease lipid peroxidation could improve mitochondrial energy generation and cardiomyocyte survival and improve RV failure in patients with congenital heart disease.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.120.045470DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7606813PMC
October 2020

Metabolic Maturation Media Improve Physiological Function of Human iPSC-Derived Cardiomyocytes.

Cell Rep 2020 07;32(3):107925

Cardiovascular Institute and Department of Medicine, Stanford University, Stanford, CA 94305, USA; Sanford-Burnham-Prebys Medical Discovery Institute, La Jolla, CA, USA; Department of Bioengineering, University of California, San Diego, San Diego, CA, USA. Electronic address:

Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) have enormous potential for the study of human cardiac disorders. However, their physiological immaturity severely limits their utility as a model system and their adoption for drug discovery. Here, we describe maturation media designed to provide oxidative substrates adapted to the metabolic needs of human iPSC (hiPSC)-CMs. Compared with conventionally cultured hiPSC-CMs, metabolically matured hiPSC-CMs contract with greater force and show an increased reliance on cardiac sodium (Na) channels and sarcoplasmic reticulum calcium (Ca) cycling. The media enhance the function, long-term survival, and sarcomere structures in engineered heart tissues. Use of the maturation media made it possible to reliably model two genetic cardiac diseases: long QT syndrome type 3 due to a mutation in the cardiac Na channel SCN5A and dilated cardiomyopathy due to a mutation in the RNA splicing factor RBM20. The maturation media should increase the fidelity of hiPSC-CMs as disease models.
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http://dx.doi.org/10.1016/j.celrep.2020.107925DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7437654PMC
July 2020

Cardiac Fibrosis Is Associated With Decreased Circulating Levels of Full-Length CILP in Heart Failure.

JACC Basic Transl Sci 2020 May 15;5(5):432-443. Epub 2020 Apr 15.

Division of Cardiology, Department of Medicine, David Geffen School of Medicine, University of California, Los Angeles (UCLA), Los Angeles, California.

Cardiac fibrosis is a pathological process associated with various forms of heart failure. This study identified latent transforming growth factor-β binding protein 2, cartilage oligomeric matrix protein, and cartilage intermediate layer protein 1 as potential biomarkers for cardiac fibrosis. All 3 encoded proteins showed increased expression in fibroblasts after transforming growth factor-β stimulation in vitro and localized specifically to fibrotic regions in vivo. Of the 3, only the full-length cartilage intermediate layer protein 1 showed a significant decrease in circulating levels in patients with heart failure compared with healthy volunteers. Further studies on these 3 proteins will lead to a better understanding of their biomarker potential for cardiac fibrosis.
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http://dx.doi.org/10.1016/j.jacbts.2020.01.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7251193PMC
May 2020

Multiscale light-sheet for rapid imaging of cardiopulmonary system.

JCI Insight 2018 08 23;3(16). Epub 2018 Aug 23.

Department of Medicine, David Geffen School of Medicine at UCLA, and.

The ability to image tissue morphogenesis in real-time and in 3-dimensions (3-D) remains an optical challenge. The advent of light-sheet fluorescence microscopy (LSFM) has advanced developmental biology and tissue regeneration research. In this review, we introduce a LSFM system in which the illumination lens reshapes a thin light-sheet to rapidly scan across a sample of interest while the detection lens orthogonally collects the imaging data. This multiscale strategy provides deep-tissue penetration, high-spatiotemporal resolution, and minimal photobleaching and phototoxicity, allowing in vivo visualization of a variety of tissues and processes, ranging from developing hearts in live zebrafish embryos to ex vivo interrogation of the microarchitecture of optically cleared neonatal hearts. Here, we highlight multiple applications of LSFM and discuss several studies that have allowed better characterization of developmental and pathological processes in multiple models and tissues. These findings demonstrate the capacity of multiscale light-sheet imaging to uncover cardiovascular developmental and regenerative phenomena.
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http://dx.doi.org/10.1172/jci.insight.121396DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6141183PMC
August 2018

Genetic Regulation of Fibroblast Activation and Proliferation in Cardiac Fibrosis.

Circulation 2018 09;138(12):1224-1235

Division of Cardiology, Department of Medicine, David Geffen School of Medicine (S.P., S.R., P.Z., M.J.M., J.S.D., A.H.-V., X.W., R.Q., J.M.S., A.J.L., R.A.), University of California, Los Angeles.

Background: Genetic diversity and the heterogeneous nature of cardiac fibroblasts (CFbs) have hindered characterization of the molecular mechanisms that regulate cardiac fibrosis. The Hybrid Mouse Diversity Panel offers a valuable tool to examine genetically diverse cardiac fibroblasts and their role in fibrosis.

Methods: Three strains of mice (C57BL/6J, C3H/HeJ, and KK/HlJ) were selected from the Hybrid Mouse Diversity Panel and treated with either isoproterenol (ISO) or saline by an intraperitoneally implanted osmotic pump. After 21 days, cardiac function and levels of fibrosis were measured by echocardiography and trichrome staining, respectively. Activation and proliferation of CFbs were measured by in vitro and in vivo assays under normal and injury conditions. RNA sequencing was done on isolated CFbs from each strain. Results were analyzed by Ingenuity Pathway Analysis and validated by reverse transcription-qPCR, immunohistochemistry, and ELISA.

Results: ISO treatment in C57BL/6J, C3H/HeJ, and KK/HlJ mice resulted in minimal, moderate, and extensive levels of fibrosis, respectively (n=7-8 hearts per condition). Isolated CFbs treated with ISO exhibited strain-specific increases in the levels of activation but showed comparable levels of proliferation. Similar results were found in vivo, with fibroblast activation, and not proliferation, correlating with the differential levels of cardiac fibrosis after ISO treatment. RNA sequencing revealed that CFbs from each strain exhibit unique gene expression changes in response to ISO. We identified Ltbp2 as a commonly upregulated gene after ISO treatment. Expression of LTBP2 was elevated and specifically localized in the fibrotic regions of the myocardium after injury in mice and in human heart failure patients.

Conclusions: This study highlights the importance of genetic variation in cardiac fibrosis by using multiple inbred mouse strains to characterize CFbs and their response to ISO treatment. Our data suggest that, although fibroblast activation is a response that parallels the extent of scar formation, proliferation may not necessarily correlate with levels of fibrosis. In addition, by comparing CFbs from multiple strains, we identified pathways as potential therapeutic targets and LTBP2 as a marker for fibrosis, with relevance to patients with underlying myocardial fibrosis.
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http://dx.doi.org/10.1161/CIRCULATIONAHA.118.035420DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6202226PMC
September 2018

Light-Sheet Imaging to Elucidate Cardiovascular Injury and Repair.

Curr Cardiol Rep 2018 03 24;20(5):35. Epub 2018 Mar 24.

Department of Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA, 90095, USA.

Purpose Of Review: Real-time 3-dimensional (3-D) imaging of cardiovascular injury and regeneration remains challenging. We introduced a multi-scale imaging strategy that uses light-sheet illumination to enable applications of cardiovascular injury and repair in models ranging from zebrafish to rodent hearts.

Recent Findings: Light-sheet imaging enables rapid data acquisition with high spatiotemporal resolution and with minimal photo-bleaching or photo-toxicity. We demonstrated the capacity of this novel light-sheet approach for scanning a region of interest with specific fluorescence contrast, thereby providing axial and temporal resolution at the cellular level without stitching image columns or pivoting illumination beams during one-time imaging. This cutting-edge imaging technique allows for elucidating the differentiation of stem cells in cardiac regeneration, providing an entry point to discover novel micro-circulation phenomenon with clinical significance for injury and repair. These findings demonstrate the multi-scale applications of this novel light-sheet imaging strategy to advance research in cardiovascular development and regeneration.
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http://dx.doi.org/10.1007/s11886-018-0979-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5987244PMC
March 2018

Analysis of cardiomyocyte clonal expansion during mouse heart development and injury.

Nat Commun 2018 02 21;9(1):754. Epub 2018 Feb 21.

Division of Cardiology, Department of Internal Medicine, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA, 90095, USA.

The cellular mechanisms driving cardiac tissue formation remain poorly understood, largely due to the structural and functional complexity of the heart. It is unclear whether newly generated myocytes originate from cardiac stem/progenitor cells or from pre-existing cardiomyocytes that re-enter the cell cycle. Here, we identify the source of new cardiomyocytes during mouse development and after injury. Our findings suggest that cardiac progenitors maintain proliferative potential and are the main source of cardiomyocytes during development; however, the onset of αMHC expression leads to reduced cycling capacity. Single-cell RNA sequencing reveals a proliferative, "progenitor-like" population abundant in early embryonic stages that decreases to minimal levels postnatally. Furthermore, cardiac injury by ligation of the left anterior descending artery was found to activate cardiomyocyte proliferation in neonatal but not adult mice. Our data suggest that clonal dominance of differentiating progenitors mediates cardiac development, while a distinct subpopulation of cardiomyocytes may have the potential for limited proliferation during late embryonic development and shortly after birth.
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http://dx.doi.org/10.1038/s41467-018-02891-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5821855PMC
February 2018

Generation of Nkx2-5/CreER transgenic mice for inducible Cre expression in developing hearts.

Genesis 2017 08 15;55(8). Epub 2017 Jun 15.

Division of Cardiology, Department of Internal Medicine, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, California, 90095.

Nkx2-5 is a homeobox-containing transcriptional regulator that serves as one of the earliest markers of cardiac lineage commitment. To study the role of Nkx2-5-expressing progenitors at specific time points in cardiac development, we have generated a novel and inducible NKX2-5 mouse line by knocking in a CreER cassette into the Nkx2-5 genomic locus, while preserving the endogenous Nkx2-5 gene to avoid haploinsufficiency. We evaluated the specificity and efficiency of CreER activity after 4-OHT injection by crossing Nkx2-5 mice with a Rosa26 reporter strain. Our immunohistochemistry results confirmed Cre-induced tdTomato expression specifically in cells expressing Nkx2-5. These cells were mainly cardiomyocytes and were observed in the embryonic heart as early as day 9.5. Additionally, quantitative polymerase chain reaction on postnatal hearts showed enriched expression of Nkx2-5 in isolated tdTomato-expressing cells. No tdTomato expression was observed in Nkx2-5 ;Rosa26 mice in the absence of 4-OHT, confirming the inducible nature of CreER activity. The Nkx2-5/CreER mouse model described in this article will serve as an invaluable tool to trace myocardial lineage and to temporally induce genetic manipulation in a selective population of cardiac progenitors during embryonic development and in the adult heart.
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http://dx.doi.org/10.1002/dvg.23041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5568924PMC
August 2017

Developmental heterogeneity of cardiac fibroblasts does not predict pathological proliferation and activation.

Circ Res 2014 Sep 18;115(7):625-35. Epub 2014 Jul 18.

From the Departments of Pathology and Developmental Biology, Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, CA (S.R.A.); Department of Internal Medicine, Division of Cardiology, and Broad Stem Cell Research Center, University of California Los Angeles School of Medicine (S.R., M.T., P.Z., A.S., A.H., P.K., Z.T., R.A.); and Division of Blood and Marrow Transplantation, Department of Medicine (A.M. S.M.) and Department of Biology (K.S.V., K.R.-H.), Stanford University, CA.

Rationale: Fibrosis is mediated partly by extracellular matrix-depositing fibroblasts in the heart. Although these mesenchymal cells are reported to have multiple embryonic origins, the functional consequence of this heterogeneity is unknown.

Objective: We sought to validate a panel of surface markers to prospectively identify cardiac fibroblasts. We elucidated the developmental origins of cardiac fibroblasts and characterized their corresponding phenotypes. We also determined proliferation rates of each developmental subset of fibroblasts after pressure overload injury.

Methods And Results: We showed that Thy1(+)CD45(-)CD31(-)CD11b(-)Ter119(-) cells constitute the majority of cardiac fibroblasts. We characterized these cells using flow cytometry, epifluorescence and confocal microscopy, and transcriptional profiling (using reverse transcription polymerase chain reaction and RNA-seq). We used lineage tracing, transplantation studies, and parabiosis to show that most adult cardiac fibroblasts derive from the epicardium, a minority arises from endothelial cells, and a small fraction from Pax3-expressing cells. We did not detect generation of cardiac fibroblasts by bone marrow or circulating cells. Interestingly, proliferation rates of fibroblast subsets on injury were identical, and the relative abundance of each lineage remained the same after injury. The anatomic distribution of fibroblast lineages also remained unchanged after pressure overload. Furthermore, RNA-seq analysis demonstrated that Tie2-derived and Tbx18-derived fibroblasts within each operation group exhibit similar gene expression profiles.

Conclusions: The cellular expansion of cardiac fibroblasts after transaortic constriction surgery was not restricted to any single developmental subset. The parallel proliferation and activation of a heterogeneous population of fibroblasts on pressure overload could suggest that common signaling mechanisms stimulate their pathological response.
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http://dx.doi.org/10.1161/CIRCRESAHA.115.303794DOI Listing
September 2014
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