The AIMS network is working to generate a cellular ‘roadmap’ of ancestrally diverse tissues of the healthy adult musculoskeletal system for the Human Cell Atlas (HCA).
The Human Cell Atlas
The Human Cell Atlas (HCA) is an international consortium that uses single-cell sequencing, spatial and computational approaches to catalogue each of the 37 trillion cells in the human body. Efforts are initially focused on identifying cell types and their locations within healthy tissues across lifetime – generating cellular reference maps of all tissues from development to adulthood.
Studying healthy tissues provides key insights into how cells function and interact during growth and homeostasis, informs mechanistic understanding of pathological changes when compared to future maps of diseased tissues and delivers metrics for evaluating potential disease diagnostics and treatments.
Critically, donors from a broad range of ancestries reference maps of tissues must be built from a broad range of ancestries. Ancestral diversity in tissue atlassing ensures advances in understanding of tissue health and disease are applicable to all populations, particularly in light of historical underrepresentation of many ancestral groups in biomedical research.
Musculoskeletal tissues pose unique challenges in single cell atlasing due to their rich extracellular matrix and cell paucity. The AIMS network is addressing these challenges to initially deliver spatially resolved single nuclei maps of four healthy adult MSK tissues (muscle, tendon, bone and capsule) across a minimum of five ancestries that can be integrated with ongoing developmental, paediatric and adult atlases.
The Oxford hub is dedicated to training our international network to empower data collection and analysis across our network sites. This is to enable capacity building and ensure all sites are empowered to collect and analyse their own data. We aim to help generate local infrastructure and train future scientists in countries currently under-presented in the HCA to facilitate grant applications and establishing of research programmes well into the future.
To support the generation of an ancestrally inclusive atlas of musculoskeletal tissues, we will collect healthy tissues from donors at several centres, including Argentina, The Gambia, India, Oman, Turkey and Zimbabwe. Donors will be from ancestrally under-represented populations with the majority from low-and middle-income countries. Given the complex ethical approval process associated with incorporating so many different countries within out network, a major goal in the first year of this project has been to establish ethical approvals at each site. Currently, we have ethical approval for fully consented collection of musculoskeletal tissues in several of our centres and we hope to have finalised all approvals by Q1 of 2023.
Donated tissues will be snap frozen prior to processing for sequencing and imaging. The Zimbabwe site will soon have capability to undertake single-nuclei extraction and library preparation for sequencing. Sites that currently do not have single-cell capacity will ship frozen samples to our Oxford hub or Zimbabwe.
Our current protocol for snap freezing of tissue prior to sequencing and imaging is openly available on the protocols.io.
Single nuclei RNA sequencing (snRNAseq) For successful sequencing that represents ground truth it is essential that cells can be isolated from tissues. The cells of musculoskeletal tissues are embedded within a rich ECM which has previously hindered application of single-cell enzymatic digestion approaches to the MSK system. To overcome this we have optimised single-nuclei extraction methods that work across a range of MSK tissues. SnRNAseq provides several advantages for the AIMS network. These advantages include reduced dissociation bias and elimination of stress response to prolonged digestions. Furthermore, due to the compatibility of snRNAseq with frozen samples, tissues can be collected and stored prior to processing for sequencing therefore removing batch effects associated with both cell isolation and library preparation.
A UMAP of hamstring tissue showing the different cell types found within the tissue. Single nucleus and spatial transcriptomic profiling of human healthy hamstring tendon, Mimpen et al, BioRxiv
Cell DIVE multiplex imaging is being used to spatially map tissues through high-dimensional quantitative analysis of up to 60 biomarkers on a single tissue section. Cell Dive validates the cell signatures identified using our single-nuclei sequencing approaches and allows mapping of cell locations within a tissue with reference to other cell types and anatomical landmarks. Understanding the spatial organisation of cells within MSK tissues provides greater depth to our cellular atlases and critical clues to cell interactions and functions both within and across tissue types.
Community engagement is integral to the AIMS network. We seek to understand the public attitude towards tissue atlasing studies and establish the site-specific requirements and barriers to such studies. We aim to establish adaptable resources that ensure informed consent for tissue donation for tissue atlasing and for the discussion and dissemination of research outcomes.
We aim to empower data analysis within our network. Our goal is to ‘train the trainers’, helping to establish global centres of genomic research within ancestrally under-represented countries.
An innovative computational biology training module was launched in November 2022 at the African Institute of Biomedical Science and Technology (AiBST) in Harare, Zimbabwe, where students were trained in single-cell RNA sequencing data analysis, as part of their MSc in Genomics and Precision Medicine.
Ongoing informal training in single-cell analysis is available to all AIMS sites for the duration of the project, to equip and upskill the wider teams.