Cao, K. X. | Search bioRxiv

bioRxiv2025

A simple technique to create pre-vascularised delivery scaffolds for organoid therapies

The growth in complex cellular constructs such as organoids for disease-modelling or regenerative therapies creates a need to overcome the problem of nutrition-delivery and surgical delivery. This logically requires a pre-vascularised scaffold, something that can embed and sustain the growth of these constructs and minimise the time needed for blood supply integration on implantation. We present here a series of experiments and their techniques that demonstrate a low-cost fabrication pipeline to create biologically-compatible scaffolds alongside evaluations of their optimal parameters in-vivo, on-CAM and in-vitro using mice and the chick chorioallantoic membrane (CAM) assay as a rapid throughput testbed of scaffolds. Utilising polylactic-glycolic acid (PLGA) scaffolds, we found an optimal range of porosity (300-500um, variable pore size), optimal lactide to glycolide ratio (82:18) and an approximation of the optimal cell seeding density using reset-vascular endothelial cells (R-VECs). Finally, we demonstrate a method to delineate the internal vascular network of these scaffolds with micro-computer-tomography (MicroCT), a low-cost analytical method that may become useful for network analysis in regenerative medicine research.

bioengineering↗

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↗