Biology subjects

Chandler, J. C.

Publications and source records attributed to Chandler, J. C..

9 recordsLinked to original sources

bioRxiv2026

PDGFRα-directed fibroblast-homing peptides for targeted delivery of nucleic acid therapeutics

Efficient and cell type-specific delivery remains a major barrier to the clinical translation of nucleic acid therapeutics (NATs), particularly for fibroblast-driven diseases in which target cells are poorly accessible in vivo. By integrating single-cell transcriptomics with cell-surface protein profiling, we identified platelet-derived growth factor receptor alpha (PDGFR) as a fibroblast-enriched surface receptor. A panel of PDGFA-derived short peptides was designed and screened across multiple human and mouse cell types, leading to the identification of two fibroblast-homing peptides, Pep3 and Pep5, which displayed rapid and preferential uptake in fibroblasts. PDGFR; knockdown reduced peptide internalisation as measured by live-cell imaging, confirming receptor-mediated uptake. To evaluate their potential as delivery vehicles for NATs, a fibroblast-based cellular model of collagen VI-related congenital muscular dystrophy (COL6-CMD) was used for further validation. Lead peptides were conjugated to a splice-switching phosphorodiamidate morpholino oligomer (PMO) targeting the recurrent COL6A1 c.930+189C>T variant. Peptide-PMO conjugates retained exon-skipping activity and Pep3-PMO enhanced collagen VI protein deposition in patient-derived fibroblasts following repeated treatment compared to unconjugated PMO. Transcriptomic profiling demonstrated no activation of inflammatory, oncogenic, or PDGFR; receptor-associated signalling pathways following peptide-PMO treatment. Collectively, these findings establish PDGFR-targeting fibroblast-peptides as a promising platform for selective delivery of NATs to fibroblasts.

molecular biology↗

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

Vincristine treatment reverses podocyte damage in focal segmental glomerulosclerosis

IntroductionFocal segmental glomerulosclerosis (FSGS) is a significant cause of chronic kidney disease and triggered by podocyte damage which can result in cytoskeletal alterations leading to foot process effacement. Vincristine is a chemoprotective drug which alters cytoskeletal microtubules and has been used clinically to reverse FSGS. However, the mechanisms underlying the beneficial effect of vincristine are not understood. MethodsWe exposed immortalised human podocytes to serum obtained from an FSGS patient before, during, and after vincristine treatment. Using RNA-sequencing we determined the effect on the podocyte transcriptome alongside impacts on cytoskeletal structure and filtration barrier integrity using a glomerulus-on-a-chip model. ResultsWe describe an adult index FSGS patient successfully treated on multiple occasions by vincristine. Podocytes exposed to serum obtained during or after vincristine treatment contained lower levels of genes associated with microtubule function compared with cells stimulated with serum collected before treatment during disease presentation. Presentation serum altered the patterning of two key podocyte cytoskeletal components, tubulin and F-actin and increased albumin permeability, changes prevented by vincristine treatment. Immunoglobulin depletion experiments revealed that the podocyte damage initiated by the presentation serum was not due to circulating autoantibodies. Defects in tubulin patterning were observed when podocytes were exposed to serum from other FSGS patients, suggestive of a common disease mechanism. ConclusionVincristine therapy produces a milieu that protects against pathological changes induced by FSGS serum, associated with preservation of tubulin and F-actin organisation. The functional role of vincristine warrants further investigation, to advance our understanding of this alternative FSGS therapeutic.

cell 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

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↗

bioRxiv2023

SARS-CoV-2 occurrence in white-tailed deer throughout their range in the conterminous United States

Broad-scale data show SARS-CoV-2 occurrence in white-tailed deer throughout much of their range in the conterminous United States and reinforce findings of considerable SARS-CoV-2 infection and exposure. Results shed light on both current infections and prior exposure, with prevalence decreasing over time and seroprevalence increasing. One-Sentence SummaryWhite-tailed deer are infected with, and have been exposed to, SARS-CoV-2 throughout their range in the conterminous US.

ecology↗

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

SARS-CoV-2 exposure in wild white-tailed deer (Odocoileus virginianus)

Widespread human SARS-CoV-2 infections combined with human-wildlife interactions create the potential for reverse zoonosis from humans to wildlife. We targeted white-tailed deer (Odocoileus virginianus) for serosurveillance based on evidence these deer have ACE2 receptors with high affinity for SARS-CoV-2, are permissive to infection, exhibit sustained viral shedding, can transmit to conspecifics, and can be abundant near urban centers. We evaluated 624 pre- and post-pandemic serum samples from wild deer from four U.S. states for SARS-CoV-2 exposure. Antibodies were detected in 152 samples (40%) from 2021 using a surrogate virus neutralization test. A subset of samples was tested using a SARS-CoV-2 virus neutralization test with high concordance between tests. These data suggest white-tailed deer in the populations assessed have been exposed to SARS-CoV-2. One-Sentence SummaryAntibodies to SARS-CoV-2 were detected in 40% of wild white-tailed deer sampled from four U.S. states in 2021.

microbiology↗