Scientists from the Broad Institute of MIT and Harvard have developed a first-of-its-kind cross-tissue cell atlas, and in collaboration with researchers at Mass Eye and Ear, have uncovered new clues for specific cell types and genes involved in complex diseases.
Genetic studies of common human diseases have linked many genetic variants to disease risk, but understanding the implicated genes and the cell types through which the genes affect disease is challenging. Complex diseases are often caused by dysfunction of more than one cell type or tissue. Profiling the cell types and genes active in each cell type across multiple tissues or organs is needed to tackle this problem for a range of diseases.
Previous research has primarily focused on single-cell atlases derived from one particular healthy or diseased tissue. In a new study published May 12 in Science, researchers described for the first time how their novel cross-tissue cell atlas derived from an analysis of nuclei from 25 frozen samples from 8 tissue types may increase understanding of the cellular and genetic underpinnings of complex diseases, including heart disease and cancers.
“Having a better understanding of the genes and cell types involved in complex diseases from different tissues can help us better understand what predisposes people to these diseases and may ultimately lead to the development of better, more targeted therapeutics,” said co-senior and co-corresponding author Ayellet V. Segrè, PhD. Dr. Segrè is a genetic biostatistician and member of the Harvard Ophthalmology Ocular Genomics Institute at Mass Eye and Ear, an assistant professor of Ophthalmology at Harvard Medical School and an associate member of the Broad Institute of MIT and Harvard.
Developing maps to better understand health and disease
This study is part of the international Human Cell Atlas (HCA) consortium, which aims to map every cell type in the human body as a basis for both understanding human health and for diagnosing, monitoring, and treating disease. An open, global, scientist-led consortium, HCA is a collaborative effort of researchers, institutes and funders worldwide, with more than 2,300 members from 83 countries across the globe. The paper is one of four major collaborative studies for the HCA published in Science this week.
A cell atlas is a map that characterizes and catalogs different cell types in a single tissue and the genes expressed in each cell type. This has often been accomplished using a fresh sample of a single tissue, such as from heart or lung, whose cells are dissociated and then analyzed. The resulting maps have been coupled with advances in human genetics to better understand what is happening on a molecular and cellular level during disease.
However, many age-related diseases are complex and caused by dysfunctional processes in more than one tissue. Fully understanding the way in which genetic variation affects disease requires generating atlases from diverse tissues across the body and from numerous individuals. However, access to fresh tissue can be a challenge for scientists, and some cell types, such as neurons or fat cells, are not easily dissociated into single cell suspensions. New approaches in experimental technology and computational techniques have been needed to accomplish this goal.
In a collaborative effort led by Aviv Regev, MS, PhD; Orit Rozenblatt-Rosen, MSc, PhD; and Kristin G. Ardlie, PhD, at the Broad Institute, the researchers developed new experimental techniques and a single-nucleus sequencing pipeline to construct a cross-tissue cell atlas from 25 frozen healthy tissue samples, archived as part of the Genotype-Tissue Expression (GTEx) project, that span three to four samples from each of eight tissue sites—breast, esophagus mucosa, esophagus muscularis, heart, lung, prostate, skeletal muscle, and skin—taken from 16 donors. Since 2010, GTEx project researchers have analyzed dozens of tissue types from hundreds of donors using methods that process tissue into a bulk mixture, but they wanted to see how genetic variation altered individual cells.
“We needed a more precise look at cells within tissues, because the cell is where biology happens, both in health and disease,” said institute scientist Kristin Ardlie, co-senior author on the new study and director of the GTEx Laboratory Data Analysis and Coordination Center at the Broad.
The researchers dissociated the cells and extracted their nuclei from the tissue, then extracted their RNA and sequenced it. Then, they aligned the RNA sequencing reads from each cell to the genes in the genome and obtained a quantification for the expression of each gene in each cell in each tissue. This culminated in over 200,000 nuclei profiles across the eight tissues, with an average of about 900 genes and 1,500 transcripts detected per nucleus. The researchers next applied a deep-learning artificial intelligence approach to harmonize the data across protocols and tissues correcting for batch effects.
Dr. Segrè and her team from Mass Eye and Ear, including co-authors, John Rouhana, MSc, and Jiali Wang, MSc, PhD, developed a new computational method that analyzes large population-based genome-wide association studies (GWAS) that implicated multiple regions in the genome involved in complex disease together with the cross-tissue cell atlas. They first mapped genes to genetic risk factors associated with disease based on gene expression changes associated with genetic variants in GTEx tissues. They then tested whether the disease genes show specific expression in particular cell types and tissues. They applied this computational technique to about 20 different complex diseases and traits, including atrial fibrillation, coronary artery disease, breast cancer, prostate cancer, skin cancers (melanoma and non-melanoma), and autoimmune diseases.
“We essentially asked whether these genes mapped to disease showed specific expression in a given cell type,” explained Dr. Segrè. “If so, that would suggest that cell type was important to the pathogenesis of the disease.”
The researchers have shared the software of their method with the scientific community.
Unexpected findings for common complex diseases
Using their novel methodology, the researchers identified known and unknown cell types that may affect complex diseases, and proposed cell type-specific genes that contribute to disease, which Dr. Segrè says demonstrated the value of the atlas.
The cross-tissue cell atlas confirmed several known cell type links to disease, such as T cells and natural killer cells in autoimmune and inflammatory diseases, luminal epithelial cells in prostate cancer, heart myonuclei in atrial fibrillation, lung fibroblasts in chronic obstructive pulmonary disease and skeletal muscle adipocytes in type 2 diabetes. Their work proposed less well-established cell types for disease, such as pericytes for coronary artery disease and heart rate, and lymphatic endothelial cells in multiple tissues for type 2 diabetes, which may explain the increased rate of vascular disease in patients with type 2 diabetes.
In addition, their analysis revealed new and noteworthy findings across tissues. In particular, genes mapped to atrial fibrillation GWAS loci were enriched in myonuclei—not only in the relevant tissue of action (heart)—but also in uninvolved tissues, such as skeletal muscle, esophagus and prostate. Coronary artery disease and heart rate loci were enriched in pericytes in five or six different tissues in addition to heart; and prostate cancer loci were enriched in luminal epithelial cells in both the prostate and breast. This is because gene programs in a given cell type are largely shared across tissues.
Taken together, the findings suggest that scientists can still learn about cell types that might be involved in disease processes from non-relevant tissues, if single cell data are not available for the relevant tissue.
“These studies represent a key moment for single-cell research and the Human Cell Atlas,” said Dr. Regev, co-senior author of the study who was a core institute member at the Broad when the study began and is currently head of Genentech Research and Early Development. “In our study, we’ve shown that this approach can generate crucial insights about the role of cells and tissues in many diseases, which will spark new scientific and biomedical inquiries aimed at a shared goal of revolutionizing medicine.”
Future studies will look at applying these experimental and computational methods to a diverse range of frozen tissue samples.
In ophthalmology, Dr. Segrè’s team is applying similar approaches to primary open-angle glaucoma, the most common form of the irreversible eye disease. Their early findings have revealed some known and novel cell types for glaucoma. They also plan to apply these methods to other common eye diseases like age-related macular degeneration, in the hopes of learning more about these blinding diseases and one day developing better strategies for prevention and therapies.
This research was funded in part by the Manton Foundation, Klarman Family Foundation, National Institutes of Health, and Chan Zuckerberg Initiative.
Eraslan G, et al. Single-nucleus cross-tissue molecular reference maps to decipher disease gene function. Science. DOI: 10.1126/science.abl4290.
About Mass Eye and Ear
Massachusetts Eye and Ear, founded in 1824, is an international center for treatment and research and a teaching hospital of Harvard Medical School. A member of Mass General Brigham, Mass Eye and Ear specializes in ophthalmology (eye care) and otolaryngology–head and neck surgery (ear, nose and throat care). Mass Eye and Ear clinicians provide care ranging from the routine to the very complex. Also home to the world’s largest community of hearing and vision researchers, Mass Eye and Ear scientists are driven by a mission to discover the basic biology underlying conditions affecting the eyes, ears, nose, throat, head and neck and to develop new treatments and cures. In the 2021–2022 “Best Hospitals Survey,” U.S. News & World Report ranked Mass Eye and Ear #4 in the nation for eye care and #2 for ear, nose and throat care. For more information about life-changing care and research at Mass Eye and Ear, visit our blog, Focus, and follow us on Instagram, Twitter and Facebook.
About Harvard Medical School Department of Ophthalmology
The Harvard Medical School Department of Ophthalmology is one of the leading and largest academic departments of ophthalmology in the nation. Composed of nine affiliates (Massachusetts Eye and Ear, which is home to Schepens Eye Research Institute; Massachusetts General Hospital; Brigham and Women’s Hospital; Boston Children’s Hospital; Beth Israel Deaconess Medical Center; Joslin Diabetes Center/Beetham Eye Institute; Veterans Affairs Boston Healthcare System; Veterans Affairs Maine Healthcare System; and Cambridge Health Alliance) and several international partners, the department draws upon the resources of a global team to pursue a singular goal—eradicate blinding diseases so that all children born today will see throughout their lifetimes. Formally established in 1871, the department is committed to its three-fold mission of providing premier clinical care, conducting transformational research, and providing world-class training for tomorrow’s leaders in ophthalmology.
Method of Research
Subject of Research
Single-nucleus cross-tissue molecular reference maps toward understanding disease gene function
Article Publication Date
A.R. is a co-founder and equity holder of Celsius Therapeutics, is an equity holder in Immunitas, and was a scientific advisory board member of ThermoFisher Scientific, Syros Pharmaceuticals, Neogene Therapeutics, and Asimov until 1 July 2020. Since 1 August 2020, A.R. is an employee of Genentech with equity in Roche. G.G. was partially funded by the Paul C. Zamecnik Chair in Oncology at the Massachusetts General Hospital Cancer Center. G.G. receives research funds from IBM and Pharmacyclics and is an inventor on patent applications related to MSMuTect, MSMutSig, MSIDetect, POLYSOLVER, and TensorQTL. G.G. is a founder of and consultant to, and holds privately held equity in, Scorpion Therapeutics. F.A. is an inventor on a patent application related to TensorQTL. F.A. has been an employee of Illumina, Inc., since 8 November 2021. E.D. is an employee of Bristol Myers Squibb. G.E. has been an employee of Genentech since 4 April 2022. O.R.-R. has been an employee of Genentech since 19 October 2020. She has given numerous lectures on the subject of single-cell genomics to a wide variety of audiences and, in some cases, has received remuneration to cover time and costs. O.R.-R. and A.R. are co-inventors on patent applications filed at the Broad Institute of MIT and Harvard related to single-cell genomics. G.E., E.D., O.R.-R., and A.R. are co-applicants on patent WO 2020/232271 relating to the work in this manuscript.