Biology subjects

Long, D. A.

Publications and source records attributed to Long, D. A..

3 recordsLinked to original sources

bioRxiv2021

Kidney organoids: A system to study human basement membrane assembly in health and disease

Basement membranes (BMs) are complex macromolecular networks underlying all continuous layers of cells. Essential components include type IV collagen and laminins, which are affected by human genetic defects leading to a range of debilitating conditions including kidney, muscle, and cerebrovascular phenotypes. We investigated the dynamics of BM assembly in human pluripotent stem cell-derived kidney organoids. We resolved their global BM composition and discovered a conserved temporal sequence in BM assembly that paralleled mammalian fetal kidneys. We identified the emergence of key BM isoforms, which were altered by a pathogenic variant in COL4A5. Integrating organoid, fetal and adult kidney proteomes we found dynamic regulation of BM composition through development to adulthood, and with single-cell transcriptomic analysis we mapped the cellular origins of BM components. Overall, we define the complex and dynamic nature of vertebrate BM assembly and provide a platform for understanding its wider relevance in human development and disease.

cell biology↗

bioRxiv2021

Systemic gene therapy with thymosin β4 alleviates glomerular injury in mice

Plasma ultrafiltration in the kidney occurs across glomerular capillaries, which are surrounded by epithelial cells called podocytes. Podocytes have a unique shape maintained by a complex cytoskeleton, which becomes disrupted in glomerular disease resulting in defective filtration and albuminuria. Lack of endogenous thymosin {beta}4 (TB4), an actin sequestering peptide, exacerbates glomerular injury and disrupts the organisation of the podocyte actin cytoskeleton, however, the effect of exogenous TB4 therapy on podocytopathy is unknown. Here, through interrogating single-cell RNA-sequencing data of isolated glomeruli we demonstrate that Adriamycin, a toxin which injures podocytes and leads to leakage of albumin in the urine of mice, results in reduced levels of podocyte TB4. Systemic administration of an adeno-associated virus vector encoding TB4 prevented Adriamycin-induced podocyte loss and albuminuria. Adriamycin injury was associated with disorganisation of the actin cytoskeleton in vitro, which was ameliorated by exogenous TB4. Furthermore, Adriamycin administration in mice was associated with increased prevalence of podocyte vesicles, a mechanism by which albumin may leak into the urine, which was also prevented by TB4. Collectively, we propose that TB4 gene therapy prevents podocyte injury and maintains glomerular filtration via modulation of the podocyte cytoskeleton thus presenting a novel treatment strategy for glomerular disease.

physiology↗

bioRxiv2021

Multiscale three-dimensional imaging of intact human organs down to the cellular scale using hierarchical phase-contrast tomography

Human organs are complex, three-dimensional and multiscale systems. Spatially mapping the human body down through its hierarchy, from entire organs to their individual functional units and specialised cells, is a major obstacle to fully understanding health and disease. To meet this challenge, we developed hierarchical phase-contrast tomography (HiP-CT), an X-ray phase propagation technique utilising the European Synchrotron Radiation Facilitys Extremely Brilliant Source: the worlds first high-energy 4th generation X-ray source. HiP-CT enabled three-dimensional and non-destructive imaging at near-micron resolution in soft tissues at one hundred thousand times the voxel size whilst maintaining the organs structure. We applied HiP-CT to image five intact human parenchymal organs: brain, lung, heart, kidney and spleen. These were hierarchically assessed with HiP-CT, providing a structural overview of the whole organ alongside detail of the organs individual functional units and cells. The potential applications of HiP-CT were demonstrated through quantification and morphometry of glomeruli in an intact human kidney, and identification of regional changes to the architecture of the air-tissue interface and alveolar morphology in the lung of a deceased COVID-19 patient. Overall, we show that HiP-CT is a powerful tool which can provide a comprehensive picture of structural information for whole intact human organs, encompassing precise details on functional units and their constituent cells to better understand human health and disease.

physiology↗