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

Daniyal J Jafree,Christiana Ruhrberg,Karen L Price,Adrian S Woolf,Peter J Scambler,Paul R Riley,Peter C Harris,Claire L Walsh,Dale Moulding,Rosa Maria Correra,Nuria Perretta Tejedor,David A Long,Maria Kolatsi-Joannou,Paul Jd Winyard,Simon Walker-Samuel,Natalie J Milmoe

doi:10.7554/elife.48183

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.

Highlights

The lymphatic vasculature is a blind-ended network of vessels that is essential for tissue fluid balance, absorbance of dietary lipids and surveillance of the immune system in vertebrate homeostasis and has been implicated in cardiovascular disease (Vieira et al, 2018), neurodegenerative disorders (Da Mesquita et al, 2018), autoimmunity (Louveau et al, 2018) and cancer metastasis (Le et al, 2016)
To observe lymphatic vessels during kidney development, we isolated metanephroi from CD-1 outbred wildtype mouse embryos at a range of developmental stages, and wholemount immunolabelled them for the early lymphatic endothelial cell markers (Banerji et al, 1999; Wigle and Oliver, 1999) prospero homeobox protein 1 (PROX1) and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1)
We optically cleared and imaged entire labelled kidneys using confocal microscopy. 3D reconstructions and z-stacks of confocal images revealed a cellular network of PROX1+/LYVE-1+ lymphatic endothelial cells in the E14.5 mouse kidney (Figure 1A)

Summary

Introduction

The lymphatic vasculature is a blind-ended network of vessels that is essential for tissue fluid balance, absorbance of dietary lipids and surveillance of the immune system in vertebrate homeostasis and has been implicated in cardiovascular disease (Vieira et al, 2018), neurodegenerative disorders (Da Mesquita et al, 2018), autoimmunity (Louveau et al, 2018) and cancer metastasis (Le et al, 2016). Each organ has a ‘waste disposal system’; a set of specialized tubes called lymphatic vessels, to clear away this excess fluid and keep a check on inflammation Defects in these tubes have been linked to a wide range of diseases including heart attacks, obesity, dementia and cancer. We have characterized the spatiotemporal development of the kidney lymphatic vasculature using a combination of wholemount immunofluorescence, optical clearing and high-resolution confocal microscopy to obtain three-dimensional (3D) images of intact mouse and human fetal kidneys at single-cell resolution. Using this approach, we provide novel insights into lymphatic vessel formation in mammalian kidney development, including a first description of the forming lymphatics in human fetal kidneys. We identified lymphatic anomalies in a mouse model of polycystic kidney disease (PKD), the most common genetic cause of kidney failure

Results and discussion

Conclusion

Materials and methods

Funding Funder