Branching morphogenesis of the urinary collecting system in the human embryonic metanephros

Abstract

An elaborate system of ducts collects urine from all nephrons, and this structure is known as the urinary collecting system (UCS). This study focused on how the UCS is formed during human embryogenesis. Fifty human embryos between the Carnegie stage (CS) 14 and CS23 were selected from the Kyoto Collection at the Congenital Anomaly Research Center of Kyoto University, Japan. Metanephroses, including the UCS, were segmented on serial digital virtual histological sections. Three-dimensional images were computationally reconstructed for morphological and quantitative analyses. A CS timeline was plotted. It consisted of the 3-D structural morphogenesis of UCS and quantification of the total amount of end-branching, average and maximum numbers of generations, deviation in the metanephros, differentiation of the urothelial epithelium in the renal pelvis, and timing of the rapid expansion of the renal pelvis. The first UCS branching generation occurred by CS16. The average branching generation reached a maximum of 8.74 ± 1.60 and was already the twelfth in CS23. The total end-branching number squared between the start and the end of the embryonic period. UCS would reach the fifteenth branching generation soon after CS23. The number of nephrons per UCS end-branch was low (0.21 ± 0.14 at CS19, 1.34 ± 0.49 at CS23), indicating that the bifid branching occurred rapidly and that the formation of nephrons followed after. The renal pelvis expanded mainly in CS23, which was earlier than that reported in a previous study. The number of nephrons connected to the UCS in the expanded group (246.0 ± 13.2) was significantly larger than that of the pre-expanded group (130.8 ± 80.1) (P < 0.05). The urothelial epithelium differentiated from the zeroth to the third generations at CS23. Differentiation may have continued up until the tenth generation to allow for renal pelvis expansion. The branching speed was not uniform. There were significantly more branching generations in the polar- than in the interpolar regions (P < 0.05). Branching speed reflects the growth orientation required to form the metanephros. Further study will be necessary to understand the renal pelvis expansion mechanism in CS23. Our CS-based timeline enabled us to map UCS formation and predict functional renal capacity after differentiation and growth.

Nascent nephrons during human embryonic development: Spatial distribution and relationship with urinary collecting system

“…The sample size was determined; therefore, there were at least five at each CS for statistical examinations (CS18, n = 5; CS19, n = 5; CS20, n = 5; CS21, n = 5; CS22, n = 6; CS23, n = 5). Our previous study indicated that the nephron may first appear at CS19 (Ishiyama et al ., 2018). Samples with overt damage and anomalies were excluded.…”

“…Histological sections of the metanephros were digitalized and three‐dimensional (3D) reconstructions were generated as described previously (Ishiyama et al ., 2018, 2019). Briefly, serial transverse sections (thickness, 10 µm) of whole embryos were digitalized using an Olympus virtual slide system (VS120‐S5‐J; Olympus Corp., Tokyo, Japan) for histological observations and 3D reconstructions.…”

“…Our previous study indicated that the nephron may first appear at CS19 (Ishiyama et al, 2018). Samples with overt damage and anomalies were excluded.…”

“…The branching pattern was categorized as deviated with EB-AP(+) and non-EB, as even with EB-AP(+) and EB-AP(−), and as even with EB-AP(+) and EB-AP(+). When the paired branch was non-EB, further branching generations were indicated Histological sections of the metanephros were digitalized and threedimensional (3D) reconstructions were generated as described previously (Ishiyama et al, 2018(Ishiyama et al, , 2019. Briefly, serial transverse sections (thickness, 10 µm) of whole embryos were digitalized using an Olympus virtual slide system (VS120-S5-J; Olympus Corp., Tokyo, Japan) for histological observations and 3D reconstructions.…”

“…The nephron number influences adult health because it affects the reserve capacity of renal function and susceptibility to renal disease. Furthermore, the development of nascent nephrons, their connection to the UCS, and the initiation of urinary excretion may contribute to the proximal UCS expansion (Ishiyama et al ., 2018). Therefore, the initial development and distribution of the nascent nephrons warrant investigation.…”