Biology subjects

Kolatsi-Joannou, M.

Publications and source records attributed to Kolatsi-Joannou, M..

7 recordsLinked to original sources

bioRxiv2025

Genetic lineage tracing identifies intermediate mesoderm as a novel contributor to mammalian kidney lymphatics

The lymphatic vasculature is essential for fluid homeostasis, immune regulation and possesses diverse organ-specific functions. During development, lymphatic endothelial cells (LEC) arise from multiple progenitor sources that form organ-specific lymphatic networks. While the origins of LECs in the heart, skin, and mesentery have been studied, those in the kidney remain unresolved. Here, we combined genetic lineage tracing in mouse embryos with optical clearing and high-resolution three-dimensional imaging to identify two distinct progenitor sources of kidney lymphatics. The majority of kidney LECs originate from a Tie2 endothelial lineage previously linked to venous or capillary vessels. Approximately 15% derive from Osr1 intermediate mesoderm, a lineage that generates kidney nephrons and stroma. Osr1-derived LECs were absent from the heart, mesentery, and skin, indicating a kidney-specific contribution, and arose independently of nephron and stromal lineages. Both Tie2 and Osr1 lineages contributed to vessel sprouting and de novo formation of lymphatic clusters. Revealing a novel cellular origin of LECs and identifying a dual origin for kidney lymphatics, we demonstrate that de novo lymphatic formation can occur from both shared and organ-specific progenitors. This work advances our understanding of how lymphatics assemble during development and provides a framework for targeting kidney lymphatics in disease.

developmental biology↗

bioRxiv2024

Reversal of elevated Gli3 in Autosomal Recessive Polycystic Kidney Disease does not alter cystogenesis

Polycystic kidney diseases (PKD) are genetic disorders characterised by the formation of fluid-filled cysts, which disrupt kidney architecture and function. Autosomal recessive PKD (ARPKD) is a rare form of PKD, caused by mutations in PKHD1, and clinically more severe than the more common autosomal dominant PKD (ADPKD). Prior studies have implicated the ciliary-located Hedgehog (Hh) pathway in ADPKD, with increased levels of Hh components in experimental ADPKD models, and reduced cystogenesis following pharmacological Hh inhibition. In contrast, the role of the Hh pathway in ARPKD is poorly understood. We hypothesised that Hh pathway activity would be elevated during ARPKD pathogenesis, and its modulation may inhibit cystogenesis, akin to prior findings in ADPKD. To test this, we utilised Cpk mice, a model which replicates the pathophysiology of ARPKD, and generated a human cellular ARPKD 3-dimensional cystogenesis model by mutating PKHD1 in human collecting duct cells through CRISPR-Cas9 technology. We found significantly elevated levels of the Hh transcriptional effector Gli3 in the Cpk mouse, a finding replicated in our human cellular ARPKD model. In the Cpk mouse, we also observed an increase in total GLI3 and GLI3 repressor protein levels. However, reduction of increased Gli3 levels via genetic deletion in the Cpk mouse did not affect cyst formation. Similarly, lowering GLI3 transcript to wildtype levels, did not influence cyst size in our human cellular ARPKD model. Collectively, these data show that elevated Gli3 does not modulate cyst progression in the context of ARPKD, highlighting the complexity of the Hh pathway in PKD. New and NoteworthyThe role of the Hedgehog pathway in autosomal recessive polycystic kidney disease (ARPKD) is poorly understood. Here, we describe elevated levels of Gli3, the Hedgehog transcriptional effector, in murine and human ARPKD models. However, reversal of the increase in Gli3 did not significantly affect cystogenesis in a human cell model of ARPKD or disease progression in a mouse model which replicates ARPKD pathophysiology. Collectively, our data indicates that Gli3 does not modulate ARPKD progression.

physiology↗

bioRxiv2024

Nanobody immunolabelling and three-dimensional imaging reveals spatially restricted LYVE1 expression by kidney lymphatic vessels in mice

Lymphatic vessels are complex three-dimensional (3D) structures that facilitate tissue fluid clearance and regulate immune responses in health and inflammatory contexts. Recent advances in wholemount immunolabelling and 3D imaging have provided insights into organ-specific heterogeneity of lymphatic structure and function. However, the visualisation of lymphatic vessels deep within an intact organ remains a challenge. We hypothesised that nanobodies, single-domain antibodies raised in camelid species, would result in improved labelling of lymphatics in intact mouse organs, without loss of information due to organ sectioning or inadequate penetration. We generated and characterised nanobody clones targeting lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1), a marker of lymphatic capillaries. Nanobodies were superior at penetrating whole mouse organs and enhanced labelling of lymphatics compared with a conventional anti-LYVE1 polyclonal antibody. Utilising this new tool, we found that kidney lymphatics; an organ in which labelling of lymphatics is challenging, have spatially restricted LYVE1 expression compared with lymphatics of skin, heart, and lung. The timing of this LYVE1 spatial restriction coincides with the early postnatal period in mice. Our findings highlight an unexpected, organ-specific characteristic of kidney lymphatic vessels, whilst providing a novel experimental tool for characterisation, isolation, or perturbation of lymphatic vessels in health and disease.

immunology↗

bioRxiv2024

A unique subset of pericystic endothelium associates with aberrant microvascular remodelling and impaired blood perfusion early in polycystic kidney disease

Hallmarks of autosomal dominant polycystic kidney disease (ADPKD), the most common hereditary kidney anomaly, include expanding fluid-filled epithelial cysts, inflammation, and fibrosis. Despite previous work showing the potential of vascular-based therapies, renal microvascular alterations in ADPKD, and their timing, are poorly understood. Using single-cell transcriptomics of human kidney microvasculature, we identify a population of endothelial cells adjacent to cysts in ADPKD. This pericystic endothelium, distinguishable by its expression of osteopontin (SPP1), has a distinct molecular profile compared to the common endothelial cell injury signature in other kidney diseases. SPP1+ pericystic endothelium was also present in an orthologous mouse model of ADPKD before overt kidney functional decline. By interrogating geometric, topological and fractal properties from three-dimensional imaging of early ADPKD mouse kidneys, we show that pericystic endothelium associates with disorganisation and non-uniformity of the renal cortical microvasculature. Concurrently, we detected region-specific reductions in cortical blood flow within ADPKD murine kidneys using arterial spin labelling. We conclude that ADPKD kidneys contain a unique subset of endothelium manifesting with aberrant remodelling and impaired blood perfusion. Its detection, prior to renal functional decline, advocates the vasculature as a therapeutic target to modulate or preserve renal function in early ADPKD.

pathology↗

bioRxiv2022

Three-dimensional imaging and single-cell transcriptomics of the human kidney implicate perturbation of lymphatics in alloimmunity

Studies of the structural and molecular features of the lymphatic vasculature, which clears fluid, macromolecules and leukocytes from the tissue microenvironment, have largely relied on animal models, with limited information in human organs beyond traditional immunohistochemical assessment. Here, we use three-dimensional imaging and single-cell RNA-sequencing to study lymphatics in the human kidney. We found a hierarchical arrangement of lymphatic vessels within human kidneys, initiating along specialised nephron epithelium in the renal cortex and displaying a distinct, kidney-specific transcriptional profile. In chronic transplant rejection we found kidney allograft lymphatic expansion alongside a loss of structural hierarchy, with human leukocyte antigen-expressing lymphatic vessels infiltrating the medulla, presenting a putative target for alloreactive antibodies. This occurred concurrently with lymphatic vessels invading and interconnecting tertiary lymphoid structures at early stages of lymphocyte colonisation. Analysis of intercellular signalling revealed upregulation of co-inhibitory molecule-mediated CD4+ T cell-lymphatic crosstalk in rejecting kidneys, potentially acting to limit local alloimmune responses. Overall, we delineate novel structural and molecular features of human kidney lymphatics and reveal perturbations to their phenotype and transcriptome in the context of alloimmunity. SUMMARYLymphatics regulate fluid balance and immune cell accumulation but are under-studied in human organs such as the kidney. Jafree and colleagues profiled human kidney lymphatics using three-dimensional imaging and single-cell RNA-sequencing, revealing structural and transcriptional perturbations in rejecting kidney transplants.

immunology↗

bioRxiv2022

Exploration of single-cell transcriptomic landscape identifies aberrant glomerular crosstalk in a murine model of WT1 kidney disease

The glomerulus mediates kidney ultrafiltration through specialised epithelial cells called podocytes which line a basement membrane shared with blood capillary endothelium. Cell-cell crosstalk is critical for glomerular function, but its investigation in childhood glomerular diseases has received little attention. WT1 encodes a transcription factor expressed in podocytes, whose heterozygous variants cause devastating kidney disease in childhood. We used single-cell RNA sequencing and ligand-receptor interaction analysis to resolve the glomerular transcriptional landscape of mice that carry an orthologous human mutation in WT1 (Wt1R394W/+). Podocytes were the most dysregulated cell type in early disease, with disrupted angiogenic signalling preceding glomerular capillary loss. Comparative analyses with additional murine and human glomerular disease datasets identified unique transcriptional changes in WT1 glomerular disease, reflecting a non-immunological pathology, whilst revealing a common injury signature across multiple glomerular diseases. Collectively, this work advocates vascular-based therapies over immunosuppressive drugs in the treatment of WT1 glomerular 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↗