Spatiotemporal dynamics and heterogeneity of renal lymphatics in mammalian development and cystic kidney disease

Abstract

Heterogeneity of lymphatic vessels during embryogenesis is critical for organ-specific lymphatic function. Little is known about lymphatics in the developing kidney, despite their established roles in pathology of the mature organ. We performed three-dimensional imaging to characterize lymphatic vessel formation in the mammalian embryonic kidney at single-cell resolution. In mouse, we visually and quantitatively assessed the development of kidney lymphatic vessels, remodeling from a ring-like anastomosis under the nascent renal pelvis; a site of VEGF-C expression, to form a patent vascular plexus. We identified a heterogenous population of lymphatic endothelial cell clusters in mouse and human embryonic kidneys. Exogenous VEGF-C expanded the lymphatic population in explanted mouse embryonic kidneys. Finally, we characterized complex kidney lymphatic abnormalities in a genetic mouse model of polycystic kidney disease. Our study provides novel insights into the development of kidney lymphatic vasculature; a system which likely has fundamental roles in renal development, physiology and disease.

Beyond a Passive Conduit: Implications of Lymphatic Biology for Kidney Diseases

“…By the end of mouse gestation, lymphatics are present in both the hilum and the cortex. A similar pattern of lymphatic vessels is established by the end of the first trimester in humans 16 .…”

“…A combination of immunohistochemical studies [12][13][14][15] and more recently, three-dimensional (3D) imaging coupled with quantitative analysis have characterized lymphatic development in mouse and human embryonic kidneys (Figure 1) 16 . In mouse, the metanephros, the precursor to the adult kidney, begins to form around embryonic day (E)10.5 17 .…”

“…One factor responsible for immune cell mobilization is CD9 + CD63 + exosome-rich endothelial vesicles, which are identified surrounding lymphatic vessels in chronic allograft nephropathy. Exposure of cultured LECs to TNF-α; which is released from damaged renal cells 78,79 , increases the chemokine and growth factor content Using wholemount immunofluorescence, optical clearing and high-resolution 3D imaging 16,118 we examined the kidney lymphatics in mice homozygous for a p.R3277C allele (Pkd1 RC ), a slow progressing model of ADPKD. We found that lymphatic vessels and corticomedullary cysts in this model sat in close proximity, suggestive of fluid transport 31 or molecular crosstalk between LECs and cyst epithelium.…”

Cellular Origins of the Lymphatic Endothelium: Implications for Cancer Lymphangiogenesis

“…The kidney provides another example of how different LEC populations assemble to generate an organ-specific lymphatic plexus. Mouse and human kidneys were reported to bear one "traditional" network of lymphatic capillaries that forms from a ring-like anastomosis in the renal hilum, and a separate population of PROX1+/LYVE1+ isolated LEC clusters similar to those described in the heart (Gancz et al, 2019b), suggesting that both lymphangiogenesis and lymphvasculogenesis play a role in renal lymphatic development (Jafree et al, 2019). Although the isolated LECs were assumed to stem from non-endothelial structures, based on their low PECAM and endomucin expression, the identity of the specific progenitors was not determined by lineage-tracing experiments.…”

Multi-Fluorescence High-Resolution Episcopic Microscopy (MF-HREM) for Three-Dimensional Imaging of Adult Murine Organs

“…Optical imaging techniques that rely on whole-mount staining (such as MF-HREM), require homogenous and rapid stain penetration, which can be enabled by increasing tissue permeability. We investigated four methods to increase the permeability of tissue samples for use with MF-HREM based on the literature (6,10,37,38): freeze-thaw, proteinase K (P[K]) digestion, iDISCO (which combines several mild detergents (6)) and saponin (N=4). Figure 5 shows the comparison of the four methods and reveals that saponin-treated samples showed significantly greater stain penetration, compared with the control case (p=0.04).…”