A multimodal and integrated approach to interrogate human kidney biopsies with rigor and reproducibility: guidelines from the Kidney Precision Medicine Project

Abstract

Comprehensive and spatially mapped molecular atlases of organs at a cellular level are a critical resource to gain insights into pathogenic mechanisms and personalized therapies for diseases. The Kidney Precision Medicine Project (KPMP) is an endeavor to generate 3-dimensional (3D) molecular atlases of healthy and diseased kidney biopsies using multiple state-of-the-art OMICS and imaging technologies across several institutions. Obtaining rigorous and reproducible results from disparate methods and at different sites to interrogate biomolecules at a single cell level or in 3D space is a significant challenge that can be a futile exercise if not well controlled. We describe a "follow the tissue" pipeline for generating a reliable and authentic single cell/region 3D molecular atlas of human adult kidney. Our approach emphasizes quality assurance, quality control, validation and harmonization across different OMICS and imaging technologies from sample procurement, processing, storage, shipping to data generation, analysis and sharing. We established benchmarks for quality control, rigor, reproducibility and feasibility across multiple technologies through a pilot experiment using common source tissue that was processed and analyzed at different institutions and different technologies. A peer review system was established to critically review quality control measures and the reproducibility of data generated by each technology before being approved to interrogate clinical biopsy specimens. The process established economizes the use of valuable biopsy tissue for multi-OMICS and imaging analysis with stringent quality control to ensure rigor and reproducibility of results and serves as a model for precision medicine projects across laboratories, institutions and consortia.

An atlas of healthy and injured cell states and niches in the human kidney

“…If present they were tagged as possible altered states, if absent they were assessed as the final cluster. Annotations of clusters were based on known positive and negative cell type markers 10,11,[55][56][57] (also see Supplementary Table 5), regional distribution of the clusters across the corticomedullary axis and altered state (including cell cycle) features. For separation of EC-DVR from EC-AEA, the combined population was independently clustered using pagoda2 and clusters associated with medullary sampling were annotated as EC-DVR.…”

“…Annotating snCv3 Clusters. To overcome the challenge of disparate nomenclature for kidney cell annotations, we leveraged a cross-consortium effort to use the extensive knowledge base from human and rodent single-cell gene expression data sets, as well as the domain expertise from pathologists, biologists, nephrologists and ontologists 10,11,19,[55][56][57][58] (also see Supplementary Table 4, 5 and the HuBMAP ASCT+B Reporter: hubmapconsortium.github.io/ccf-asct-reporter). This allowed the adoption of a standardized anatomical and cell type nomenclature for major and minor cell types and their subclasses (Supplementary Table 4), showing distinct and consistent expression profiles of known markers and absence of specific segment markers for some of the cell types (Extended Data Fig.…”

“…To this end, we report a next-generation multimodal single cell and 3D atlas that leverages integrated transcriptomic, epigenomic and imaging data over three major consortia: the Human Biomolecular Atlas Program (HuBMAP) 7 , the Kidney Precision Medicine Project (KPMP) 8 , and the Human Cell Atlas (HCA) 9 . To ensure robust cell state profiles, reference tissues were obtained from multiple sources, and biopsies were collected from AKI and CKD patients under rigorous quality assurance and control procedures 7,8,10 . We define micro niches for healthy and altered states across different regions of the human kidney spanning the cortex and medulla, to the papillary tip, and identify gene expression and regulatory modules in altered states associated with worsening kidney function.…”

Profiling Immune Cells in the Kidney Using Tissue Cytometry and Machine Learning

“…We previously showed that this approach could be used in archived frozen human kidney biopsies, whereby quantitation of immune cell densities could be potentially linked with other molecular or clinical features (51,53). Large scale 3D imaging and tissue cytometry of human kidney biopsy is currently being used as a main imaging technology in the Kidney Precision Medicine Project (KPMP) consortium (55,56,60).…”

Autofluorescence microscopy as a label-free tool for renal histology and glomerular segmentation

“…Recent kidney atlas programs have emerged with the goal of characterizing normal human tissues and discovering drivers of disease using cellular and molecular phenotypes spanning wide spatial scales and biomolecular classes. 1,2 Functional tissue units (FTUs), multicellular structures that perform vital functions, including components of the nephron in the kidney, such as glomeruli, proximal tubules, distal tubules, and collecting ducts, are an important intermediate between whole organ and individual cells. 3 Indeed, kidney diseases like diabetic nephropathy, acute kidney injury, and chronic kidney disease are a direct result of damage to the aforementioned FTUs.…”